Fructose-1,6-bisphosphate and aldolase mediate glucose sensing by AMPK

PubMed Central

Wang, Zhichao; Gray, Alexander; Ma, Teng; Cui, Jiwen; Feng, Jin-Wei; Zhu, Mingjiang; Wu, Yu-Qing; Li, Terytty Yang; Ye, Zhiyun; Lin, Shu-Yong; Yin, Huiyong; Piao, Hai-Long; Hardie, D. Grahame; Lin, Sheng-Cai

2017-01-01

The major energy source for most cells is glucose, from which ATP is generated via glycolysis and/or oxidative metabolism. Glucose deprivation activates AMP-activated protein kinase (AMPK)1, but it has been unclear whether this occurs solely via changes in AMP or ADP, the classical activators of AMPK2–5. Here, we uncover a mechanism that triggers AMPK activation via an AMP/ADP-independent mechanism sensing absence of FBP, with AMPK being progressively activated as extracellular glucose and intracellular FBP decrease. When unoccupied by FBP, aldolases promote the formation of lysosomal complexes containing the v-ATPase, Ragulator, AXIN, LKB1 and AMPK, previously shown to be required for AMPK activation6,7. Knockdown of aldolases activates AMPK even in cells with abundant glucose, while the catalysis-defective D34S aldolase mutant, which still binds FBP, blocks AMPK activation. Cell-free reconstitution assays show that addition of FBP disrupts association of AXIN/LKB1 with v-ATPase/Ragulator. Importantly, in some cell types AMP:ATP/ADP:ATP ratios remain unchanged during acute glucose starvation, and intact AMP-binding sites on AMPK are not required for AMPK activation. These results establish that aldolase, as well as a glycolytic enzyme, is a sensor of glucose availability that regulates AMPK. PMID:28723898

Carbohydrate metabolism during starvation in the silkworm Bombyx mori.

PubMed

Satake, S; Kawabe, Y; Mizoguchi, A

2000-06-01

The effect of starvation on carbohydrate metabolism in the last instar larvae of the silkworm Bombyx mori was examined. Trehalose concentration in the hemolymph increased slightly during the first 6 h of starvation and decreased thereafter, whereas glucose concentration decreased rapidly immediately after diet deprivation. Starvation-induced hypertrehalosemia was completely inhibited by neck ligation, suggesting that starvation stimulates the release of a hypertrehalosemic factor(s) from the head. The percentage of active glycogen phosphorylase in the fat body increased within 3 h of starvation and its glycogen content decreased gradually. These observations suggest that production of trehalose from glycogen is enhanced in starved larvae. However, hypertrehalosemia during starvation cannot be explained by the increased supply of trehalose into hemolymph alone, as similar changes in phosphorylase activity and glycogen content in the fat body were observed in neck-ligated larvae, in which hemolymph trehalose concentration did not increase but decreased gradually. When injected into larvae, trehalose disappeared from hemolymph at a rate about 40% lower in starved larvae than neck-ligated larvae. The hemolymph lipid concentration increased during starvation, suggesting that an increased supply of lipids to tissues suppresses the consumption of hemolymph trehalose and this is an important factor in hypertrehalosemia. Copyright 2000 Wiley-Liss, Inc.

ATP-driven and AMPK-independent autophagy in an early branching eukaryotic parasite.

PubMed

Li, Feng-Jun; Xu, Zhi-Shen; Soo, Andy D S; Lun, Zhao-Rong; He, Cynthia Y

2017-04-03

Autophagy is a catabolic cellular process required to maintain protein synthesis, energy production and other essential activities in starved cells. While the exact nutrient sensor(s) is yet to be identified, deprivation of amino acids, glucose, growth factor and other nutrients can serve as metabolic stimuli to initiate autophagy in higher eukaryotes. In the early-branching unicellular parasite Trypanosoma brucei, which can proliferate as procyclic form (PCF) in the tsetse fly or as bloodstream form (BSF) in animal hosts, autophagy is robustly triggered by amino acid deficiency but not by glucose depletion. Taking advantage of the clearly defined adenosine triphosphate (ATP) production pathways in T. brucei, we have shown that autophagic activity depends on the levels of cellular ATP production, using either glucose or proline as a carbon source. While autophagosome formation positively correlates with cellular ATP levels; perturbation of ATP production by removing carbon sources or genetic silencing of enzymes involved in ATP generation pathways, also inhibited autophagy. This obligate energy dependence and the lack of glucose starvation-induced autophagy in T. brucei may reflect an adaptation to its specialized, parasitic life style.

Nitrogen Starvation Induced Oxidative Stress in an Oil-Producing Green Alga Chlorella sorokiniana C3

PubMed Central

He, Chen-Liu; Wang, Qiang

2013-01-01

Microalgal lipid is one of the most promising feedstocks for biodiesel production. Chlorella appears to be a particularly good option, and nitrogen (N) starvation is an efficient environmental pressure used to increase lipid accumulation in Chlorella cells. The effects of N starvation of an oil-producing wild microalga, Chlorella sorokiniana C3, on lipid accumulation were investigated using thin layer chromatography (TLC), confocal laser scanning microscopy (CLSM) and flow cytometry (FCM). The results showed that N starvation resulted in lipid accumulation in C. sorokiniana C3 cells, oil droplet (OD) formation and significant lipid accumulation in cells were detected after 2 d and 8 d of N starvation, respectively. During OD formation, reduced photosynthetic rate, respiration rate and photochemistry efficiency accompanied by increased damage to PSII were observed, demonstrated by chlorophyll (Chl) fluorescence, 77K fluorescence and oxygen evolution tests. In the mean time the rate of cyclic electron transportation increased correspondingly to produce more ATP for triacylglycerols (TAGs) synthesis. And 0.5 d was found to be the turning point for the early stress response and acclimation of cells to N starvation. Increased level of membrane peroxidation was also observed during OD formation, and superoxide dismutase (SOD), peroxide dismutase (POD) and catalase (CAT) enzyme activity assays suggested impaired reactive oxygen species (ROS) scavenging ability. Significant neutral lipid accumulation was also observed by artificial oxidative stress induced by H2O2 treatment. These results suggested coupled neutral lipid accumulation and oxidative stress during N starvation in C. sorokiniana C3. PMID:23874918

Effect of starvation on free histidine and amino acids in white muscle of milkfish Chanos chanos.

PubMed

Shiau, C Y; Pong, Y P; Chiou, T K; Tin, Y Y

2001-03-01

Milkfish (Chanos chanos) decreased their body weight from 47 to 28 g over the 60-day period of starvation. Starvation also resulted in the reduction of muscle lipid and protein, and hepatosomatic index. The predominant free amino acid (FAA) in white muscle of milkfish was histidine, followed by taurine and glycine. In the first 25 days of starvation, no significant change in histidine was found. After 40 days of starvation, however, the histidine concentration was significantly decreased by 46%, and remained unchanged thereafter. As compared to control group fish, the 60-day-starved fish possessed only half the amount of histidine. Taurine and glycine, on the other hand, showed no significant changes throughout starvation. Taurine became the most predominant in the FAA pool after 40 days of starvation, and the concentration of 60-day-starved fish was two times higher than that of control group fish without starvation. The ratios of histidine, taurine, and glycine to total FAAs remained approximately the same although the individual contributions varied considerably to the total FAAs during starvation. The results of this study suggested that a good strategy would be to keep taurine and glycine in milkfish muscle at relatively high levels for physiological function as histidine decreased drastically for energy source under conditions of food deprivation.

Effects of food deprivation on expression of growth hormone receptor and proximate composition in liver of black seabream Acanthopagrus schlegeli.

PubMed

Deng, L; Zhang, W M; Lin, H R; Cheng, Christopher H K

2004-04-01

The effects of food deprivation on the hepatic level growth hormone receptor (GHR) were investigated in black seabream (Acanthopagrus schlegeli) both at the protein level (by radioreceptor assay) and at the mRNA level (by ribonuclease protection assay). Serum levels of growth hormone (GH) and triiodothyronine (T(3)) were also measured. Condition factor and hepatic proximate composition of the fish were also assessed. Significant decrease in hepatic GHR binding was recorded as early as on day 2 of starvation. On day 30 this decrease was even more pronounced, with the level in the starved fish reaching less than 20% the fed control level. A concomitant decrease in the hepatic GHR mRNA content was also noted during this period, with a progressive decrease from day 2 to day 30 of starvation. The extent of decrease in the mRNA content was less pronounced than the decrease in receptor binding, with the hepatic GHR mRNA content in the day 30 starved fish representing approximately 30% of the level in the fed control. In large contrast, serum GH level increased progressively during starvation. After 30 days of starvation, serum GH levels in the starved fish were more than three times the concentration found in the fed control. Serum T(3) levels, on the other hand, decreased during starvation, with the difference reaching significance on day 15 and day 30. After 30 days of starvation, serum T(3) levels in the starved fish were only approximately 40% the concentration found in the fed control. The hepatic lipid content exhibited an increasing trend during starvation. On day 30 the hepatic lipid content of the starved fish had doubled the level found in the fed control. However, the hepatic protein content did not exhibit much change during starvation. There was also a minor decrease in the moisture content of the liver during starvation, but the condition factor of the fish as a whole registered a gradual decrease during the course of food deprivation.

Drosophila larvae lacking the bcl-2 gene, buffy, are sensitive to nutrient stress, maintain increased basal target of rapamycin (Tor) signaling and exhibit characteristics of altered basal energy metabolism

PubMed Central

2012-01-01

Background B cell lymphoma 2 (Bcl-2) proteins are the central regulators of apoptosis. The two bcl-2 genes in Drosophila modulate the response to stress-induced cell death, but not developmental cell death. Because null mutants are viable, Drosophila provides an optimum model system to investigate alternate functions of Bcl-2 proteins. In this report, we explore the role of one bcl-2 gene in nutrient stress responses. Results We report that starvation of Drosophila larvae lacking the bcl-2 gene, buffy, decreases survival rate by more than twofold relative to wild-type larvae. The buffy null mutant reacted to starvation with the expected responses such as inhibition of target of rapamycin (Tor) signaling, autophagy initiation and mobilization of stored lipids. However, the autophagic response to starvation initiated faster in larvae lacking buffy and was inhibited by ectopic buffy. We demonstrate that unusually high basal Tor signaling, indicated by more phosphorylated S6K, was detected in the buffy mutant and that removal of a genomic copy of S6K, but not inactivation of Tor by rapamycin, reverted the precocious autophagy phenotype. Instead, Tor inactivation also required loss of a positive nutrient signal to trigger autophagy and loss of both was sufficient to activate autophagy in the buffy mutant even in the presence of enforced phosphoinositide 3-kinase (PI3K) signaling. Prior to starvation, the fed buffy mutant stored less lipid and glycogen, had high lactate levels and maintained a reduced pool of cellular ATP. These observations, together with the inability of buffy mutant larvae to adapt to nutrient restriction, indicate altered energy metabolism in the absence of buffy. Conclusions All animals in their natural habitats are faced with periods of reduced nutrient availability. This study demonstrates that buffy is required for adaptation to both starvation and nutrient restriction. Thus, Buffy is a Bcl-2 protein that plays an important non-apoptotic role to promote survival of the whole organism in a stressful situation. PMID:22824239

Suppressed translation and ULK1 degradation as potential mechanisms of autophagy limitation under prolonged starvation.

PubMed

Allavena, Giulia; Boyd, Caroline; Oo, Kyaw Soe; Maellaro, Emilia; Zhivotovsky, Boris; Kaminskyy, Vitaliy O

2016-11-01

Macroautophagy/autophagy is a well-organized process of intracellular degradation, which is rapidly activated under starvation conditions. Recent data demonstrate a transcriptional upregulation of several autophagy genes as a mechanism that controls autophagy in response to starvation. Here we report that despite the significant upregulation of mRNA of the essential autophagy initiation gene ULK1, its protein level is rapidly reduced under starvation. Although both autophagic and proteasomal systems contribute to the degradation of ULK1, under prolonged nitrogen deprivation, its level was still reduced in ATG7 knockout cells, and only initially stabilized in cells treated with the lysosomal or proteasomal inhibitors. We demonstrate that under starvation, protein translation is rapidly diminished and, similar to treatments with the proteosynthesis inhibitors cycloheximide or anisomycin, is associated with a significant reduction of ULK1. Furthermore, it was found that inhibition of the mitochondrial respiratory complexes or the mitochondrial ATP synthase function that could also take place in the absence of substrates, promote upregulation of ULK1 mRNA and protein expression in an AMPK-dependent manner in U1810 lung cancer cells growing in complete culture medium. These inhibitors could also drastically increase the ULK1 protein in U1810 cells with knockout of ATG13, where the ULK1 expression is significantly diminished. However, such upregulation of ULK1 protein is negligible under starvation conditions, further signifying the contribution of translation and suggesting that transcriptional upregulation of ULK1 protein will be diminished under such conditions. Thus, we propose a model where inhibition of protein translation, together with the degradation systems, limit autophagy during starvation.

Transcriptional and Proteomic Profiling of Aspergillus flavipes in Response to Sulfur Starvation.

PubMed

El-Sayed, Ashraf S A; Yassin, Marwa A; Ali, Gul Shad

2015-01-01

Aspergillus flavipes has received considerable interest due to its potential to produce therapeutic enzymes involved in sulfur amino acid metabolism. In natural habitats, A. flavipes survives under sulfur limitations by mobilizing endogenous and exogenous sulfur to operate diverse cellular processes. Sulfur limitation affects virulence and pathogenicity, and modulates proteome of sulfur assimilating enzymes of several fungi. However, there are no previous reports aimed at exploring effects of sulfur limitation on the regulation of A. flavipes sulfur metabolism enzymes at the transcriptional, post-transcriptional and proteomic levels. In this report, we show that sulfur limitation affects morphological and physiological responses of A. flavipes. Transcription and enzymatic activities of several key sulfur metabolism genes, ATP-sulfurylase, sulfite reductase, methionine permease, cysteine synthase, cystathionine β- and γ-lyase, glutathione reductase and glutathione peroxidase were increased under sulfur starvation conditions. A 50 kDa protein band was strongly induced by sulfur starvation, and the proteomic analyses of this protein band using LC-MS/MS revealed similarity to many proteins involved in the sulfur metabolism pathway.

IF1, a natural inhibitor of mitochondrial ATP synthase, is not essential for the normal growth and breeding of mice.

PubMed

Nakamura, Junji; Fujikawa, Makoto; Yoshida, Masasuke

2013-09-17

IF1 is an endogenous inhibitor protein of mitochondrial ATP synthase. It is evolutionarily conserved throughout all eukaryotes and it has been proposed to play crucial roles in prevention of the wasteful reverse reaction of ATP synthase, in the metabolic shift from oxidative phosphorylation to glycolysis, in the suppression of ROS (reactive oxygen species) generation, in mitochondria morphology and in haem biosynthesis in mitochondria, which leads to anaemia. Here, we report the phenotype of a mouse strain in which IF1 gene was destroyed. Unexpectedly, individuals of this IF1-KO (knockout) mouse strain grew and bred without defect. The general behaviours, blood test results and responses to starvation of the IF1-KO mice were apparently normal. There were no abnormalities in the tissue anatomy or the autophagy. Mitochondria of the IF1-KO mice were normal in morphology, in the content of ATP synthase molecules and in ATP synthesis activity. Thus, IF1 is not an essential protein for mice despite its ubiquitous presence in eukaryotes.

Glucose starvation impairs DNA repair in tumour cells selectively by blocking histone acetylation.

PubMed

Ampferl, Rena; Rodemann, Hans Peter; Mayer, Claus; Höfling, Tobias Tim Alexander; Dittmann, Klaus

2018-03-01

Tumour cells are characterized by aerobic glycolysis and thus have high glucose consumption. Because repairing radiation-induced DNA damage is an energy-demanding process, we hypothesized that glucose starvation combined with radiotherapy could be an effective strategy to selectively target tumour cells. We glucose-starved tumour cells (A549, FaDu) in vitro and analysed their radiation-induced cell responses compared to normal fibroblasts (HSF7). Irradiation depleted intracellular ATP levels preferentially in cancer cells. Consequently, glucose starvation impaired DNA double-strand break (DSB) repair and radiosensitized confluent tumour cells but not normal fibroblasts. In proliferating tumour cells glucose starvation resulted in a reduction of proliferation, but failed to radiosensitize cells. Glucose supply was indispensable during the late DSB repair in confluent tumour cells starting approximately 13 h after irradiation, and glucose starvation inhibited radiation-induced histone acetylation, which is essential for chromatin relaxation. Sirtinol - an inhibitor of histone deacetylases - reverted the effects of glucose depletion on histone acetylation and DNA DSB repair in tumour cells. Furthermore, a glucose concentration of 2.8 mmol/L was sufficient to impair DSB repair in tumour cells and reduced their clonogenic survival under a fractionated irradiation regimen. In resting tumour cells, glucose starvation combined with irradiation resulted in the impairment of late DSB repair and the reduction of clonogenic survival, which was associated with disrupted radiation-induced histone acetylation. However, in normal cells, DNA repair and radiosensitivity were not affected by glucose depletion. Copyright © 2017 Elsevier B.V. All rights reserved.

Evaluating death and activity decay of Anammox bacteria during anaerobic and aerobic starvation.

PubMed

Wang, Qilin; Song, Kang; Hao, Xiaodi; Wei, Jing; Pijuan, Maite; van Loosdrecht, Mark C M; Zhao, Huijun

2018-06-01

The decreased activity (i.e. decay) of anaerobic ammonium oxidation (Anammox) bacteria during starvation can be attributed to death (i.e. decrease in the amount of viable bacteria) and activity decay (i.e. decrease in the specific activity of viable bacteria). Although they are crucial for the operation of the Anammox process, they have never been comprehensively investigated. This study for the first time experimentally assessed death and activity decay of the Anammox bacteria during 84 days' starvation stress based on ammonium removal rate, Live/Dead staining and fluorescence in-situ hybridization. The anaerobic and aerobic decay rates of Anammox bacteria were determined as 0.015 ± 0.001 d -1 and 0.028 ± 0.001 d -1 , respectively, indicating Anammox bacteria would lose their activity more quickly in the aerobic starvation than in the anaerobic starvation. The anaerobic and aerobic death rates of Anammox bacteria were measured at 0.011 ± 0.001 d -1 and 0.025 ± 0.001 d -1 , respectively, while their anaerobic and aerobic activity decay rates were determined at 0.004 ± 0.001 d -1 and 0.003 ± 0.001 d -1 , respectively. Further analysis revealed that death accounted for 73 ± 4% and 89 ± 5% of the decreased activity of Anammox bacteria during anaerobic and aerobic starvations, and activity decay was only responsible for 27 ± 4% and 11 ± 5% of the decreased Anammox activity, respectively, over the same starvation periods. These deeply shed light on the response of Anammox bacteria to the starvation stress, which would facilitate operation and optimization of the Anammox process. Copyright © 2018 Elsevier Ltd. All rights reserved.

[Effects of starvation on the consumption of energy sources and swimming performance in juvenile Gambusia affinis and Tanichthys albonubes].

PubMed

Li, Jiang-tao; Lin, Xiao-tao; Zhou, Chen-hui; Zeng, Peng; Xu, Zhong-neng; Sun, Jun

2016-01-01

To explore the consumption of energy sources and swimming performance of juvenile Gambusia affinis and Tanichthys albonubes after starvation, contents of glycogen, lipid and protein, burst swimming speeds (Uburst), and critical swimming speeds (Ucrit) at different starvation times (0, 10, 20, 30 and 40 days) were evaluated. The results showed that, at 0 day, contents of glycogen and lipid were significantly lower in G. affinis than those in T. albonubes, whereas no significant difference in content of protein between two experimental fish was found. Swimming speeds in G. affinis were significantly lower than those in T. albonubes for all swimming performances. After different starvation scenarios, content of glycogen both in G. affinis and T. albonubes decreased significantly in power function trend with starvation time and were close to zero after starvation for 10 days, whereas the contents of lipid and protein were linearly significantly decreased. The slope of line regression equation between content of lipid and starvation time in G. affinis was significantly lower than that in T. albonubes, whereas there was a significantly higher slope of line equation between content of protein and starvation time in G. affinis. 40 days later, the consumption rate of glycogen both in G. affinis and T. albonubes were significantly higher than that of lipid, while the consumption rate of protein was the least. Consumption amounts of glycogen in all experimental fish were the least, G. affinis consumed more protein than lipid, and T. albonubes consumed more lipid than protein. Uburst and Ucrit decreased significantly linearly with starvation time for all experimental fish. Slope of linear equation between Uburst and starvation time was not significantly different between G. affinis and T. albonubes. However, the straight slope between Ucrit and starvation time was significantly lower in G. affinis than that in T. albonubes. These findings indicated that there was close relationship between the consumption of energy sources and swimming performance in starvation. Although the store amounts of energy sources and swimming performance were lower in G. affinis than those in T. albonubes, G. affinis mainly used protein during starvation. The result of more stable lipid content and Ucrit in G. affinis in starvation compared with that in T. albonubes indicated that G. affinis had a fair endurance to starvation, which helped them to adapt to the poor nutrition environment in stream habitat.

Critical Involvement of Extracellular ATP Acting on P2RX7 Purinergic Receptors in Photoreceptor Cell Death

PubMed Central

Notomi, Shoji; Hisatomi, Toshio; Kanemaru, Takaaki; Takeda, Atsunobu; Ikeda, Yasuhiro; Enaida, Hiroshi; Kroemer, Guido; Ishibashi, Tatsuro

2011-01-01

Stressed cells release ATP, which participates in neurodegenerative processes through the specific ligation of P2RX7 purinergic receptors. Here, we demonstrate that extracellular ATP and the more specific P2RX7 agonist, 2′- and 3′-O-(4-benzoylbenzoyl)-ATP, both induce photoreceptor cell death when added to primary retinal cell cultures or when injected into the eyes from wild-type mice, but not into the eyes from P2RX7−/− mice. Photoreceptor cell death was accompanied by the activation of caspase-8 and -9, translocation of apoptosis-inducing factor from mitochondria to nuclei, and TUNEL-detectable chromatin fragmentation. All hallmarks of photoreceptor apoptosis were prevented by premedication or co-application of Brilliant Blue G, a selective P2RX7 antagonist that is already approved for the staining of internal limiting membranes during ocular surgery. ATP release is up-regulated by nutrient starvation in primary retinal cell cultures and seems to be an initializing event that triggers primary and/or secondary cell death via the positive feedback loop on P2RX7. Our results encourage the potential application of Brilliant Blue G as a novel neuroprotective agent in retinal diseases or similar neurodegenerative pathologies linked to excessive extracellular ATP. PMID:21983632

Transcriptional and Proteomic Profiling of Aspergillus flavipes in Response to Sulfur Starvation

PubMed Central

El-Sayed, Ashraf S. A.; Yassin, Marwa A.; Ali, Gul Shad

2015-01-01

Aspergillus flavipes has received considerable interest due to its potential to produce therapeutic enzymes involved in sulfur amino acid metabolism. In natural habitats, A. flavipes survives under sulfur limitations by mobilizing endogenous and exogenous sulfur to operate diverse cellular processes. Sulfur limitation affects virulence and pathogenicity, and modulates proteome of sulfur assimilating enzymes of several fungi. However, there are no previous reports aimed at exploring effects of sulfur limitation on the regulation of A. flavipes sulfur metabolism enzymes at the transcriptional, post-transcriptional and proteomic levels. In this report, we show that sulfur limitation affects morphological and physiological responses of A. flavipes. Transcription and enzymatic activities of several key sulfur metabolism genes, ATP-sulfurylase, sulfite reductase, methionine permease, cysteine synthase, cystathionine β- and γ-lyase, glutathione reductase and glutathione peroxidase were increased under sulfur starvation conditions. A 50 kDa protein band was strongly induced by sulfur starvation, and the proteomic analyses of this protein band using LC-MS/MS revealed similarity to many proteins involved in the sulfur metabolism pathway. PMID:26633307

Effect of fasting on the urinary excretion of water-soluble vitamins in humans and rats.

PubMed

Fukuwatari, Tsutomu; Yoshida, Erina; Takahashi, Kei; Shibata, Katsumi

2010-01-01

Recent studies showed that the urinary excretion of the water-soluble vitamins can be useful as a nutritional index. To determine how fasting affects urinary excretion of water-soluble vitamins, a human study and an animal experiment were conducted. In the human study, the 24-h urinary excretion of water-soluble vitamins in 12 healthy Japanese adults fasting for a day was measured. One-day fasting drastically decreased urinary thiamin content to 30%, and increased urinary riboflavin content by 3-fold. Other water-soluble vitamin contents did not show significant change by fasting. To further investigate the alterations of water-soluble vitamin status by starvation, rats were starved for 3 d, and water-soluble vitamin contents in the liver, blood and urine were measured during starvation. Urinary excretion of thiamin, riboflavin, vitamin B(6) metabolite 4-pyridoxic acid, nicotinamide metabolites and folate decreased during starvation, but that of vitamin B(12), pantothenic acid and biotin did not. As for blood vitamin levels, only blood vitamin B(1), plasma PLP and plasma folate levels decreased with starvation. All water-soluble vitamin contents in the liver decreased during starvation, whereas vitamin concentrations in the liver did not decrease. Starvation decreased only concentrations of vitamin B(12) and folate in the skeletal muscle. These results suggest that water-soluble vitamins were released from the liver, and supplied to the peripheral tissues to maintain vitamin nutrition. Our human study also suggested that the effect of fasting should be taken into consideration for subjects showing low urinary thiamin and high urinary riboflavin.

Effects of Starvation on Lipid Metabolism and Gluconeogenesis in Yak

PubMed Central

Yu, Xiaoqiang; Peng, Quanhui; Luo, Xiaolin; An, Tianwu; Guan, Jiuqiang; Wang, Zhisheng

2016-01-01

This research was conducted to investigate the physiological consequences of undernourished yak. Twelve Maiwa yak (110.3±5.85 kg) were randomly divided into two groups (baseline and starvation group). The yak of baseline group were slaughtered at day 0, while the other group of yak were kept in shed without feed but allowed free access to water, salt and free movement for 9 days. Blood samples of the starvation group were collected on day 0, 1, 2, 3, 5, 7, 9 and the starved yak were slaughtered after the final blood sample collection. The liver and muscle glycogen of the starvation group decreased (p<0.01), and the lipid content also decreased while the content of moisture and ash increased (p<0.05) both in Longissimus dorsi and liver compared with the baseline group. The plasma insulin and glucose of the starved yak decreased at first and then kept stable but at a relatively lower level during the following days (p<0.01). On the contrary, the non-esterified fatty acids was increased (p<0.01). Beyond our expectation, the ketone bodies of β-hydroxybutyric acid and acetoacetic acid decreased with prolonged starvation (p<0.01). Furthermore, the mRNA expression of lipogenetic enzyme fatty acid synthase and lipoprotein lipase in subcutaneous adipose tissue of starved yak were down-regulated (p<0.01), whereas the mRNA expression of lipolytic enzyme carnitine palmitoyltransferase-1 and hormone sensitive lipase were up-regulated (p<0.01) after 9 days of starvation. The phosphoenolpyruvate carboxykinase and pyruvate carboxylase, responsible for hepatic gluconeogenesis were up-regulated (p<0.01). It was concluded that yak derive energy by gluconeogenesis promotion and fat storage mobilization during starvation but without ketone body accumulation in the plasma. PMID:26954191

Effects of Starvation on Lipid Metabolism and Gluconeogenesis in Yak.

PubMed

Yu, Xiaoqiang; Peng, Quanhui; Luo, Xiaolin; An, Tianwu; Guan, Jiuqiang; Wang, Zhisheng

2016-11-01

This research was conducted to investigate the physiological consequences of undernourished yak. Twelve Maiwa yak (110.3±5.85 kg) were randomly divided into two groups (baseline and starvation group). The yak of baseline group were slaughtered at day 0, while the other group of yak were kept in shed without feed but allowed free access to water, salt and free movement for 9 days. Blood samples of the starvation group were collected on day 0, 1, 2, 3, 5, 7, 9 and the starved yak were slaughtered after the final blood sample collection. The liver and muscle glycogen of the starvation group decreased (p<0.01), and the lipid content also decreased while the content of moisture and ash increased (p<0.05) both in Longissimus dorsi and liver compared with the baseline group. The plasma insulin and glucose of the starved yak decreased at first and then kept stable but at a relatively lower level during the following days (p<0.01). On the contrary, the non-esterified fatty acids was increased (p<0.01). Beyond our expectation, the ketone bodies of β-hydroxybutyric acid and acetoacetic acid decreased with prolonged starvation (p<0.01). Furthermore, the mRNA expression of lipogenetic enzyme fatty acid synthase and lipoprotein lipase in subcutaneous adipose tissue of starved yak were down-regulated (p<0.01), whereas the mRNA expression of lipolytic enzyme carnitine palmitoyltransferase-1 and hormone sensitive lipase were up-regulated (p<0.01) after 9 days of starvation. The phosphoenolpyruvate carboxykinase and pyruvate carboxylase, responsible for hepatic gluconeogenesis were up-regulated (p<0.01). It was concluded that yak derive energy by gluconeogenesis promotion and fat storage mobilization during starvation but without ketone body accumulation in the plasma.

Effect of Thymine Starvation on Messenger Ribonucleic Acid Synthesis in Escherichia coli

PubMed Central

Luzzati, Denise

1966-01-01

Luzzati, Denise (Institut de Biologie Physico-Chimique, Paris, France). Effect of thymine starvation on messenger ribonucleic acid synthesis in Escherichia coli. J. Bacteriol. 92:1435–1446. 1966.—During the course of thymine starvation, the rate of synthesis of messenger ribonucleic acid (mRNA, the rapidly labeled fraction of the RNA which decays in the presence of dinitrophenol or which hybridizes with deoxyribonucleic acid) decreases exponentially, in parallel with the viability of the thymine-starved bacteria. The ability of cell-free extracts of starved bacteria to incorporate ribonucleoside triphosphates into RNA was determined; it was found to be inferior to that of extracts from control cells. The analysis of the properties of cell-free extracts of starved cells shows that their decreased RNA polymerase activity is the consequence of a modification of their deoxyribonucleic acid, the ability of which to serve as a template for RNA polymerase decreases during starvation. PMID:5332402

Critical involvement of extracellular ATP acting on P2RX7 purinergic receptors in photoreceptor cell death.

PubMed

Notomi, Shoji; Hisatomi, Toshio; Kanemaru, Takaaki; Takeda, Atsunobu; Ikeda, Yasuhiro; Enaida, Hiroshi; Kroemer, Guido; Ishibashi, Tatsuro

2011-12-01

Stressed cells release ATP, which participates in neurodegenerative processes through the specific ligation of P2RX7 purinergic receptors. Here, we demonstrate that extracellular ATP and the more specific P2RX7 agonist, 2'- and 3'-O-(4-benzoylbenzoyl)-ATP, both induce photoreceptor cell death when added to primary retinal cell cultures or when injected into the eyes from wild-type mice, but not into the eyes from P2RX7(-/-) mice. Photoreceptor cell death was accompanied by the activation of caspase-8 and -9, translocation of apoptosis-inducing factor from mitochondria to nuclei, and TUNEL-detectable chromatin fragmentation. All hallmarks of photoreceptor apoptosis were prevented by premedication or co-application of Brilliant Blue G, a selective P2RX7 antagonist that is already approved for the staining of internal limiting membranes during ocular surgery. ATP release is up-regulated by nutrient starvation in primary retinal cell cultures and seems to be an initializing event that triggers primary and/or secondary cell death via the positive feedback loop on P2RX7. Our results encourage the potential application of Brilliant Blue G as a novel neuroprotective agent in retinal diseases or similar neurodegenerative pathologies linked to excessive extracellular ATP. Copyright © 2011 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Membrane potential independent transport of NH3 in the absence of ammonium permeases in Saccharomyces cerevisiae.

PubMed

Cueto-Rojas, Hugo F; Milne, Nicholas; van Helmond, Ward; Pieterse, Mervin M; van Maris, Antonius J A; Daran, Jean-Marc; Wahl, S Aljoscha

2017-04-17

Microbial production of nitrogen containing compounds requires a high uptake flux and assimilation of the N-source (commonly ammonium), which is generally coupled with ATP consumption and negatively influences the product yield. In the industrial workhorse Saccharomyces cerevisiae, ammonium (NH 4 + ) uptake is facilitated by ammonium permeases (Mep1, Mep2 and Mep3), which transport the NH 4 + ion, resulting in ATP expenditure to maintain the intracellular charge balance and pH by proton export using the plasma membrane-bound H + -ATPase. To decrease the ATP costs for nitrogen assimilation, the Mep genes were removed, resulting in a strain unable to uptake the NH 4 + ion. Subsequent analysis revealed that growth of this ∆mep strain was dependent on the extracellular NH 3 concentrations. Metabolomic analysis revealed a significantly higher intracellular NH X concentration (3.3-fold) in the ∆mep strain than in the reference strain. Further proteomic analysis revealed significant up-regulation of vacuolar proteases and genes involved in various stress responses. Our results suggest that the uncharged species, NH 3 , is able to diffuse into the cell. The measured intracellular/extracellular NH X ratios under aerobic nitrogen-limiting conditions were consistent with this hypothesis when NH x compartmentalization was considered. On the other hand, proteomic analysis indicated a more pronounced N-starvation stress response in the ∆mep strain than in the reference strain, which suggests that the lower biomass yield of the ∆mep strain was related to higher turnover rates of biomass components.

NDRG2 overexpression suppresses hepatoma cells survival during metabolic stress through disturbing the activation of fatty acid oxidation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pan, Tao; Xi'an Medical University, Xi'an, Shaanxi Province; Zhang, Mei

Because of the high nutrient consumption and inadequate vascularization, solid tumor constantly undergoes metabolic stress during tumor development. Oncogenes and tumor suppressor genes participated in cancer cells' metabolic reprogramming. N-Myc downstream regulated gene 2 (NDRG2) is a recently identified tumor suppressor gene, but its function in cancer metabolism, particularly during metabolic stress, remains unclear. In this study, we found that NDRG2 overexpression significantly reduced hepatoma cell proliferation and enhanced cell apoptosis under glucose limitation. Moreover, NDRG2 overexpression aggravated energy imbalance and oxidative stress by decreasing the intracellular ATP and NADPH generation and increasing ROS levels. Strikingly, NDRG2 inhibited the activationmore » of fatty acid oxidation (FAO), which preserves ATP and NADPH purveyance in the absence of glucose. Finally, mechanistic investigation showed that NDRG2 overexpression suppressed the glucose-deprivation induced AMPK/ACC pathway activation in hepatoma cells, whereas the expression of a constitutively active form of AMPK abrogated glucose-deprivation induced AMPK activation and cell apoptosis. Thus, as a negative regulator of AMPK, NDRG2 disturbs the induction of FAO genes by glucose limitation, leading to dysregulation of ATP and NADPH, and thus reduces the tolerance of hepatoma cells to glucose limitation. - Highlights: • NDRG2 overexpression reduces the tolerance of hepatoma cells to glucose limitation. • NDRG2 overexpression aggravates energy imbalance and oxidative stress under glucose deprivation. • NDRG2 overexpression disturbs the activation of FAO in hepatoma cells under glucose limitation. • NDRG2 overexpression inhibits the activation of AMPK/ACC pathway in hepatoma cells during glucose starvation.« less

Effect of starvation and refeeding on the hepatopancreas of whiteleg shrimp Penaeus vannamei (Boone) using computer-assisted image analysis.

PubMed

Cervellione, F; McGurk, C; Berger Eriksen, T; Van den Broeck, W

2017-11-01

Under normal farming conditions, shrimp can experience starvation periods attributable to disease outbreaks or adverse environmental conditions. Starvation leads to significant morphological changes in the hepatopancreas (HP), being the main organ for absorption and storage of nutrients. In the literature, limited research has described the effect on the HP of periods of starvation followed by refeeding and none in whiteleg shrimp (Penaeus vannamei) using computer-assisted image analysis (CAIA). This study describes the effect of starvation and starvation followed by refeeding on the HP of whiteleg shrimp using CAIA. Visiopharm ® software was used to quantify the following morphological parameters, measured as ratio to the total tissue area (TLA): total lumen area (TLA:TTA), haemocytic infiltration area in the intertubular spaces (HIA:TTA), B-cell vacuole area (VBA:TTA), lipid droplet area within R cells (LDA:TTA) and F-cell area (FCA:TTA). Significant changes were measured for HIA:TTA and LDA:TTA during starvation (increase in HIA:TTA associated with decrease in LDA:TTA) and starvation followed by refeeding (decrease in HIA:TTA associated with increase in LDA:TTA). In the future, HIA:TTA and LDA:TTA have the potential to be used in a pre-emptive manner to monitor the health of the HP, facilitate early diagnosis of diseases and study the pathophysiology of the organ. © 2017 John Wiley & Sons Ltd.

Impact of starvation on survival, meat condition and metabolism of Chlamys farreri

NASA Astrophysics Data System (ADS)

Yang, Hong-Sheng; Wang, Jian; Zhou, Yi; Wang, Ping; He, Yi-Chao; Zhang, Fu-Sui

2001-03-01

The effects of 60-day starvation on survival rate, condition index (CI), changes of nutrient composition of different tissues, respiration and excretion of scallop Chlamys farreri were studied in laboratory from Oct. 17 to Dec. 15, 1997. Two groups (control and starvation with 200 individuals each) were cultured in two 2 m3 tanks, with 31 to 32 salinity water at 17°C. Starvation effects were measured after 10, 20, 40 and 60 days. There was no mass mortality of scallops of the two tanks and survival rates of the control and starvation groups were 93.5% and 92.0%, respectively. Starvation had strong effect on the meat condition of the scallops, especially after 10 days; when relative lipid percentage dropped sharply while relative protein percentage increased. The impact of starvation on the oxygen consumption rate (OCR) and the ammonia-N excretion rate (AER) was obvious. The OCR increased rapidly after 10 days but decreased after 20 days. The AER increased after 10 days and 20 days, but decreased obviously from 20 to 40 days. The O∶N ratios varied to different degrees, and minimized after 20 days. The low O∶N ratios implied that the protein was the main material for the metabolism of C. farreri.

Transcriptomic analysis of Saccharomyces cerevisiae upon honokiol treatment.

PubMed

Zhu, Xiaolong; Zou, Shenshen; Li, Youbin; Liang, Yongheng

2017-09-01

Honokiol (HNK), one of the main medicinal components in Magnolia officinalis, possesses antimicrobial activity against a variety of pathogenic bacteria and fungi. However, little is known of the molecular mechanisms underpinning the antimicrobial activity. To explore the molecular mechanism of its antifungal activity, we determined the effects of HNK on the mRNA expression profile of Saccharomyces cerevisiae using a DNA microarray approach. HNK markedly induced the expression of genes related to iron uptake and homeostasis. Conversely, genes associated with respiratory electron transport were downregulated, mirroring the effects of iron starvation. Meanwhile, HNK-induced growth deficiency was partly rescued by iron supplementation and HNK reacted with iron, producing iron complexes that depleted iron. These results suggest that HNK treatment induced iron starvation. Additionally, HNK treatment resulted in the upregulation of genes involved in protein synthesis and drug resistance networks. Furthermore, the deletion of PDR5, a gene encoding the plasma membrane ATP binding cassette (ABC) transporter, conferred sensitivity to HNK. Overexpression of PDR5 enhanced resistance of WT and pdr5Δ strains to HNK. Taken together, these findings suggest that HNK, which can be excluded by overexpression of Pdr5, functions in multiple cellular processes in S. cerevisiae, particularly in inducing iron starvation to inhibit cell growth. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

The role of transporters in supplying energy to plant plastids.

PubMed

Flügge, Ulf-Ingo; Häusler, Rainer E; Ludewig, Frank; Gierth, Markus

2011-04-01

The energy status of plant cells strongly depends on the energy metabolism in chloroplasts and mitochondria, which are capable of generating ATP either by photosynthetic or oxidative phosphorylation, respectively. Another energy-rich metabolite inside plastids is the glycolytic intermediate phosphoenolpyruvate (PEP). However, chloroplasts and most non-green plastids lack the ability to generate PEP via a complete glycolytic pathway. Hence, PEP import mediated by the plastidic PEP/phosphate translocator or PEP provided by the plastidic enolase are vital for plant growth and development. In contrast to chloroplasts, metabolism in non-green plastids (amyloplasts) of starch-storing tissues strongly depends on both the import of ATP mediated by the plastidic nucleotide transporter NTT and of carbon (glucose 6-phosphate, Glc6P) mediated by the plastidic Glc6P/phosphate translocator (GPT). Both transporters have been shown to co-limit starch biosynthesis in potato plants. In addition, non-photosynthetic plastids as well as chloroplasts during the night rely on the import of energy in the form of ATP via the NTT. During energy starvation such as prolonged darkness, chloroplasts strongly depend on the supply of ATP which can be provided by lipid respiration, a process involving chloroplasts, peroxisomes, and mitochondria and the transport of intermediates, i.e. fatty acids, ATP, citrate, and oxaloacetate across their membranes. The role of transporters involved in the provision of energy-rich metabolites and in pathways supplying plastids with metabolic energy is summarized here.

Different variations of tissue B-group vitamin concentrations in short- and long-term starved rats.

PubMed

Moriya, Aya; Fukuwatari, Tsutomu; Sano, Mitsue; Shibata, Katsumi

2012-01-01

Prolonged starvation changes energy metabolism; therefore, the metabolic response to starvation is divided into three phases according to changes in glucose, lipid and protein utilisation. B-group vitamins are involved in energy metabolism via metabolism of carbohydrates, fatty acids and amino acids. To determine how changes in energy metabolism alter B-group vitamin concentrations during starvation, we measured the concentration of eight kinds of B-group vitamins daily in rat blood, urine and in nine tissues including cerebrum, heart, lung, stomach, kidney, liver, spleen, testis and skeletal muscle during 8 d of starvation. Vitamin B1, vitamin B6, pantothenic acid, folate and biotin concentrations in the blood reduced after 6 or 8 d of starvation, and other vitamins did not change. Urinary excretion was decreased during starvation for all B-group vitamins except pantothenic acid and biotin. Less variation in B-group vitamin concentrations was found in the cerebrum and spleen. Concentrations of vitamin B1, vitamin B6, nicotinamide and pantothenic acid increased in the liver. The skeletal muscle and stomach showed reduced concentrations of five vitamins including vitamin B1, vitamin B2, vitamin B6, pantothenic acid and folate. Concentrations of two or three vitamins decreased in the kidney, testis and heart, and these changes showed different patterns in each tissue and for each vitamin. The concentration of pantothenic acid rapidly decreased in the heart, stomach, kidney and testis, whereas concentrations of nicotinamide were stable in all tissues except the liver. Different variations in B-group vitamin concentrations in the tissues of starved rats were found. The present findings will lead to a suitable supplementation of vitamins for the prevention of the re-feeding syndrome.

Nitrogen and photosynthetic function of hermatypic corals. Oxygen exchange of Stylophora pistillata coral under artificial feeding.

PubMed

Leletkin, V A

2005-01-01

The change of Stylophora pistillata coral photosynthetic function (oxygen exchange and biomass of symbionts) under starvation and food enrichment was studied to understand the role of heterotrophy in nitrogen supplements of zooxanthellae. The starvation caused the decrease of frequency of zooxanthellae cells division in 7-10 times. The number of degraded algae cells increased in same proportion and, as a result, the density of zooxanthellae in corals decreased about two times during one-two weeks. Under starvation corals kept their photosynthetic capacity at the level of corals in situ by means of enhancing the zooxanthellae gross photosynthesis. The respiration rate of coral had tendency to increase and the dry mass of polyp tissue to decrease. Under artificial feeding which was following starvation the zooxanthellae density increased in 1.5-2 times, and particular food caused more intensive accumulation of zooxanthellae comparing to dissolved inorganic ammonium. The feeding regime did not affect dry mass of polyp tissue and chlorophyll content as well as respiration and gross productivity of the corals. The conclusion about high effectiveness of particular feeding for supplying symbiotic algae with nitrogen was made and trophic status of zooxanthellae in hospite was determined as unlimited by nitrogen.

Phenformin-induced Hypoglycaemia in Normal Subjects*

PubMed Central

Lyngsøe, J.; Trap-Jensen, J.

1969-01-01

Study of the effect of phenformin on the blood glucose level in normal subjects before and during 70 hours of starvation showed a statistically significant hypoglycaemic effect after 40 hours of starvation. This effect was not due to increased glucose utilization. Another finding in this study was a statistically significant decrease in total urinary nitrogen excretion during starvation in subjects given phenformin. These findings show that the hypoglycaemic effect of phenformin in starved normal subjects is due to inhibition of gluconeogenesis. PMID:5780431

Post carbon removal nitrifying MBBR operation at high loading and exposure to starvation conditions.

PubMed

Young, Bradley; Delatolla, Robert; Kennedy, Kevin; LaFlamme, Edith; Stintzi, Alain

2017-09-01

This study investigates the performance of MBBR nitrifying biofilm post carbon removal at high loading and starvation conditions. The nitrifying MBBR, treating carbon removal lagoon effluent, achieved a maximum SARR of 2.13gN/m 2 d with complete conversion of ammonia to nitrate. The results also show the MBBR technology is capable of maintaining a stable biofilm under starvation conditions in systems that nitrify intermittently. The biomass exhibited a higher live fraction of total cells in the high loaded reactors (73-100%) as compared to the reactors operated in starvation condition (26-82%). For both the high loaded and starvation condition, the microbial communities significantly changed with time of operation. The nitrifying community, however, remained steady with the family Nitrosomonadacea as the primary AOBs and Nitrospira as the primary NOB. During starvation conditions, the relative abundance of AOBs decreased and Nitrospira increased corresponding to an NOB/AOB ratio of 5.2-12.1. Copyright © 2017 Elsevier Ltd. All rights reserved.

Analysis of starvation effects on hydrodynamic lubrication in nonconforming contacts

NASA Technical Reports Server (NTRS)

Brewe, D. E.; Hamrock, B. J.

1981-01-01

The effects of lubricant starvation on minimum film thickness, under conditions of a hydrodynamic point contact, are determined by numerical methods where: (1) starvation is effected by varying the fluid inlet level; (2) the Reynolds boundary conditions are applied at the cavitation boundary; and (3) zero pressure is stipulated at the meniscus or inlet boundary. Seventy-four cases were used to numerically determine a minimum-film-thickness equation, as a function of the ratio of dimensionless load to dimensionless speed for varying degrees of starvation. A film reduction factor was in turn determined as a function of the fluid inlet level, and a starved, fully-flooded boundary was defined along with an expression determining the onset of starvation. It is found that as the degree of starvation increases, the minimum film thickness decreases gradually until the fluid inlet becomes critical. The changes in the inlet pressure buildup due to changing the available lubricant supply are presented in the form of three-dimensional isometric plots.

Changes in the midgut cells in the European cave spider, Meta menardi, during starvation in spring and autumn.

PubMed

Lipovšek, Saška; Leitinger, Gerd; Novak, Tone; Janžekovič, Franc; Gorgoń, Szymon; Kamińska, Karolina; Rost-Roszkowska, Magdalena

2018-03-01

During the growth period, in surface habitats, spiders catch enough prey to feed normally. In contrast, in the cave entrance zone, prey may be relatively scarce. Meta menardi inhabits this cave section, resulting in temporary starvation. We studied structural changes in the midgut epithelial cells of M. menardi during a short-term and a medium-term controlled starvation to mimic the occasional starvation in caves, during spring and autumn. Digestive cells, secretory cells and adipocytes were examined before the experimental starvation, in the middle and at the end of starvation. We used light microscopy, transmission electron microscopy and specific histochemical methods for the detection of lipids, polysaccharides and proteins. Detection of lysosomes, autolysosomes and apoptosis was also carried out. The general structures of the cells did not change during the experimental starvation in either season, while some specific differences in the ultrastructure were observed. In both sexes, in both seasons, the amounts of lipids, glycogen and proteins decreased during starvation. Larger amounts of lipids were found in autumn, while there were no significant differences in the amounts of glycogen and proteins. In both sexes, in both seasons, autophagy and apoptosis intensified with starvation in progress, but more intensively in females. Thus, autumn individuals, in contrast to spring ones, compile energy-supplying stores to confront the subsequent winter deficiency of prey in caves, while the cellular ultrastructures undergo the same starvation-dependant changes at any time during the growth period.

p62-mediated autophagy affects nutrition-dependent insulin receptor substrate-1 dynamics in 3T3-L1 preadipocytes.

PubMed

Igawa, Hirobumi; Kikuchi, Akihiro; Misu, Hirofumi; Ishii, Kiyo-Aki; Kaneko, Shuichi; Takamura, Toshinari

2018-05-22

Previous studies have shown that the organism's nutritional status changes the protein levels of insulin receptor substrate 1 (IRS-1) in a tissue-specific manner. Although the mechanisms underlying the regulation of IRS-1 in the nutrient-rich conditions associated with diabetes and insulin resistance have been well studied, those under nutrient-poor conditions remain unknown. The aim of this study was to investigate how IRS-1 protein levels change depending on the nutritional status of 3T3-L1 preadipocytes. 3T3-L1 preadipocytes were treated with glucose-, amino acid- and serum-free medium for starvation. IRS-1 protein levels were detected by western blot. Autophagy activity was observed by western blot and fluorescence microscopy. The effect of autophagy and p62, an adaptor for selective autophagy, on IRS-1 protein levels under starvation conditions was examined by western blot and immunocytochemistry. We showed that that the levels of IRS-1, but not those of insulin receptor and Akt, decreased when starvation activated autophagy. The inhibition of autophagy by chloroquine or autophagy-related 7 (Atg7) RNA interference counteracted the starvation-induced decrease of IRS-1. Additionally, Atg7 knockdown increased insulin-stimulated phosphorylation of Akt under starvation conditions. Furthermore, p62 co-localized with IRS-1 under starvation conditions, and p62 knockdown counteracted the starvation-induced degradation of IRS-1. Autophagy through p62 plays an important role in regulating IRS-1 protein levels in response to nutritional deficiency. Our findings suggest that autophagy may function as energy depletion-sensing machinery that finely tunes insulin signal transduction. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

DIFFERENT PHYSIOLOGICAL RESPONSES OF FOUR MARINE SYNECHOCOCCUS STRAINS (CYANOPHYCEAE) TO NICKEL STARVATION UNDER IRON-REPLETE AND IRON-DEPLETE CONDITIONS(1).

PubMed

Qiu, Baosheng; Price, Neil M

2009-10-01

Synechococcus species are important primary producers in coastal and open-ocean ecosystems. When nitrate was provided as the sole nitrogen source, nickel starvation inhibited the growth of strains WH8102 and WH7803, while it had little effect on two euryhaline strains, WH5701 and PCC 7002. Nickel was required for the acclimation of Synechococcus WH7803 to low iron and high light. In WH8102 and WH7803, nickel starvation decreased the linear electron transport activity, slowed down QA reoxidation, but increased the connectivity factor between individual photosynthetic units. Under such conditions, the reduction of their intersystem electron transport chains was expected to increase, and their cyclic electron transport around PSI would be favored. Nickel starvation decreased the total superoxide dismutase (SOD) activity of WH8102 and WH7803 by 30% and 15% of the control, respectively. The protein-bound (63) Ni of the oceanic strain WH8102 comigrated with SOD activity on nondenaturing gels and thus provided additional evidence for the existence of active NiSOD in Synechococcus WH8102. In WH7803, it seems likely that nickel starvation affected other metabolic pathways and thus indirectly affected the total SOD activity. © 2009 Phycological Society of America.

Early Changes in the Ultrastructure of Streptococcus faecalis After Amino Acid Starvation

PubMed Central

Higgins, M. L.; Shockman, G. D.

1970-01-01

Thin sections of Streptococcus faecalis (ATCC 9790) starved of one essential amino acid (threonine or valine) initially show rapid increases in (i) cell wall thickness, (ii) the apparent size of the central nucleoid region, and (iii) mesosomal membranes. The most rapid increases in all three variables occurred during the first 1 to 2 hr of starvation. After this initial period, the rates progressively decreased over the 20-hr observation period. During threonine starvation, the mesosomal membrane that accumulated in the first hour was subsequently degraded and reached a level similar to that found in exponential-phase cells after 20 hr. With valine starvation, mesosomal membrane continued to slowly accumulate over the entire 20-hr observation period. The mesosomes of the starved cells retained the same “stalked-bag” morphology of those in exponential-phase cells. These cytological observations agree with previously published biochemical data on membrane lipid and wall content after starvation. Images PMID:4987306

Evidence for the adverse effect of starvation on bone quality: a review of the literature.

PubMed

Kueper, Janina; Beyth, Shaul; Liebergall, Meir; Kaplan, Leon; Schroeder, Josh E

2015-01-01

Malnutrition and starvation's possible adverse impacts on bone health and bone quality first came into the spotlight after the horrors of the Holocaust and the ghettos of World War II. Famine and food restrictions led to a mean caloric intake of 200-800 calories a day in the ghettos and concentration camps, resulting in catabolysis and starvation of the inhabitants and prisoners. Severely increased risks of fracture, poor bone mineral density, and decreased cortical strength were noted in several case series and descriptive reports addressing the medical issues of these individuals. A severe effect of severely diminished food intake and frequently concomitant calcium- and Vitamin D deficiencies was subsequently proven in both animal models and the most common cause of starvation in developed countries is anorexia nervosa. This review attempts to summarize the literature available on the impact of the metabolic response to Starvation on overall bone health and bone quality.

Temperature induces trade-offs between development and starvation resistance in Aedes aegypti (L.) larvae.

PubMed

Padmanabha, H; Lord, C C; Lounibos, L P

2011-12-01

Heightened temperature increases the development rate of mosquitoes. However, in Aedes aegypti (Diptera: Culicidae), the larvae of which commonly experience limited access to food in urban habitats, temperature effects on adult production may also be influenced by changes in the capacity of larvae to survive without food. We carried out experiments to investigate the effects of temperatures increasing at intervals of 2 °C from 20 °C to 30 °C on the growth, maturation rate and longevity of optimally fed larvae placed in starvation. Overall, both growth rate and starvation resistance were lower in the first three larval instars (L1-L3) compared with L4, in which growth of >75% occurred. Although increasing the temperature reduced the duration of each instar, it had a U-shaped impact in terms of the effect of initial growth on starvation resistance, which increased from L1 to L2 at 20 °C and 30 °C, remained constant at 22 °C and 28 °C, and decreased at 24 °C and 26 °C. Growth from L2 to L3 significantly increased starvation resistance only from 26 °C to 30 °C. Increased temperature (>22 °C) consistently reduced starvation resistance in L1. In L2-L4, increments of 2 °C decreased starvation resistance between 20 °C and 24 °C, but had weaker and instar-specific effects at >24 °C. These data show that starvation resistance in Ae. aegypti depends on both instar and temperature, indicating a trade-off between increased development rate and reduced starvation survival of early-instar larvae, particularly in the lower and middle temperatures of the dengue-endemic range of 20-30 °C. We suggest that anabolic and catabolic processes in larvae have distinct temperature dependencies, which may ultimately cause temperature to modify the density regulation of Ae. aegypti populations. © 2011 The Authors. Medical and Veterinary Entomology © 2011 The Royal Entomological Society.

Decreased ATP synthesis is phenotypically expressed during increased energy demand in fibroblasts containing mitochondrial tRNA mutations.

PubMed

James, A M; Sheard, P W; Wei, Y H; Murphy, M P

1999-01-01

Mutations in the tRNA genes of mitochondrial DNA (mtDNA) cause the debilitating MELAS (mitochondrial, myopathy, encephalopathy, lactic acidosis and stroke-like episodes) and MERRF (myoclonic epilepsy and ragged-red fibres) syndromes. These mtDNA mutations affect respiratory chain function, apparently without decreasing cellular ATP concentration [Moudy et al. (1995) PNAS, 92, 729-733]. To address this issue, we investigated the role of mitochondrial ATP synthesis in fibroblasts from MELAS and MERRF patients. The maximum rate of mitochondrial ATP synthesis was decreased by 60-88%, as a consequence of the decrease in the proton electrochemical potential gradient of MELAS and MERRF mitochondria. However, in quiescent fibroblasts neither ATP concentration or the ATP/ADP ratio was affected by the lowered rate of ATP synthesis. We hypothesized that the low ATP demand of quiescent fibroblasts masked the mitochondrial ATP synthesis defect and that this defect might become apparent during higher ATP use. To test this we simulated high energy demand by titrating cells with gramicidin, an ionophore that stimulates ATP hydrolysis by the plasma membrane Na+/K+-ATPase. We found a threshold gramicidin concentration in control cells at which both the ATP/ADP ratio and the plasma membrane potential decreased dramatically, due to ATP demand by the Na+/K+-ATPase outstripping mitochondrial ATP synthesis. In MELAS and MERRF fibroblasts the corresponding threshold concentrations of gramicidin were 2-20-fold lower than those for control cells. This is the first demonstration that cells containing mtDNA mutations are particularly sensitive to increased ATP demand and this has several implications for how mitochondrial dysfunction contributes to disease pathophysiology. In particular, the increased susceptibility to plasma membrane depolarization will render neurons with dysfunctional mitochondria susceptible to excitotoxic cell death.

TCDD decreases ATP levels and increases reactive oxygen production through changes in mitochondrial F F{sub 1}-ATP synthase and ubiquinone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shertzer, Howard G.; Genter, Mary Beth; Shen, Dongxiao

2006-12-15

Mitochondria generate ATP and participate in signal transduction and cellular pathology and/or cell death. TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) decreases hepatic ATP levels and generates mitochondrial oxidative DNA damage, which is exacerbated by increasing mitochondrial glutathione redox state and by inner membrane hyperpolarization. This study identifies mitochondrial targets of TCDD that initiate and sustain reactive oxygen production and decreased ATP levels. One week after treating mice with TCDD, liver ubiquinone (Q) levels were significantly decreased, while rates of succinoxidase and Q-cytochrome c oxidoreductase activities were increased. However, the expected increase in Q reduction state following TCDD treatment did not occur; instead, Q wasmore » more oxidized. These results could be explained by an ATP synthase defect, a premise supported by the unusual finding that TCDD lowers ATP/O ratios without concomitant changes in respiratory control ratios. Such results suggest either a futile cycle in ATP synthesis, or hydrolysis of newly synthesized ATP prior to release. The TCDD-mediated decrease in Q, concomitant with an increase in respiration, increases complex 3 redox cycling. This acts in concert with glutathione to increase membrane potential and reactive oxygen production. The proposed defect in ATP synthase explains both the greater respiratory rates and the lower tissue ATP levels.« less

Effect of tributyltin (TBT) on ATP levels in human natural killer (NK) cells: relationship to TBT-induced decreases in NK function.

PubMed

Dudimah, Fred D; Odman-Ghazi, Sabah O; Hatcher, Frank; Whalen, Margaret M

2007-01-01

The purpose of this study was to investigate the role that tributyltin (TBT)-induced decreases in ATP levels may play in TBT-induced decreases in the tumor lysing (lytic) function of natural killer (NK) cells. NK cells are a subset of lymphocytes that act as an initial immune defense against tumor cells and virally infected cells. TBT is an environmental contaminant that has been detected in human blood, which has been shown to interfere with ATP synthesis. Previous studies have shown that TBT is able to decrease very significantly the lytic function of NK cells. In this study NK cells were exposed to various concentrations of TBT and to two other compounds that interfere with ATP synthesis (rotenone a complex I inhibitor and oligomycin an ATP synthase inhibitor) for various lengths of time before determining the levels of ATP and lytic function. Exposures of NK cells to 10, 25, 50 and 100 nm TBT did not significantly reduce ATP levels after 24 h. However, these same exposures caused significant decreases in cytotoxic function. Studies of brief 1 h exposures to a range of TBT, rotenone and oligomycin concentrations followed by 24 h, 48 h and 6 day periods in compound-free media prior to assaying for ATP levels or cytotoxic function showed that each of the compounds caused persistent decreases in ATP levels and lytic function of NK cells. Exposures to 0.05-5 microm rotenone or oligomycin for 1 h reduced ATP levels by 20-25% but did not have any measurable effect on the ability of NK cells to lyse tumor cells. ATP levels were also decreased by about 20-25% after 24 h or 48 h exposures to rotenone or oligomycin (0.5 microm ), and the lytic function was decreased by about 50%. The results suggest that TBT-induced decreases in ATP levels were not responsible for the loss of cytotoxic function seen at 1 h and 24 h. However, TBT-induced decreases of NK-ATP levels may be at least in part responsible for losses of NK-cytotoxic function seen after 48 h and 6 day exposures. Copyright 2006 John Wiley & Sons, Ltd.

Genetic and nutrient modulation of acetyl-CoA levels in Synechocystis for n-butanol production.

PubMed

Anfelt, Josefine; Kaczmarzyk, Danuta; Shabestary, Kiyan; Renberg, Björn; Rockberg, Johan; Nielsen, Jens; Uhlén, Mathias; Hudson, Elton P

2015-10-16

There is a strong interest in using photosynthetic cyanobacteria as production hosts for biofuels and chemicals. Recent work has shown the benefit of pathway engineering, enzyme tolerance, and co-factor usage for improving yields of fermentation products. An n-butanol pathway was inserted into a Synechocystis mutant deficient in polyhydroxybutyrate synthesis. We found that nitrogen starvation increased specific butanol productivity up to threefold, but cessation of cell growth limited total n-butanol titers. Metabolite profiling showed that acetyl-CoA increased twofold during nitrogen starvation. Introduction of a phosphoketolase increased acetyl-CoA levels sixfold at nitrogen replete conditions and increased butanol titers from 22 to 37 mg/L at day 8. Flux balance analysis of photoautotrophic metabolism showed that a Calvin-Benson-Bassham-Phosphoketolase pathway had higher theoretical butanol productivity than CBB-Embden-Meyerhof-Parnas and a reduced butanol ATP demand. These results demonstrate that phosphoketolase overexpression and modulation of nitrogen levels are two attractive routes toward increased production of acetyl-CoA derived products in cyanobacteria and could be implemented with complementary metabolic engineering strategies.

pH regulation in glycosomes of procyclic form Trypanosoma brucei.

PubMed

Lin, Sheng; Voyton, Charles; Morris, Meredith T; Ackroyd, P Christine; Morris, James C; Christensen, Kenneth A

2017-05-12

Here we report the use of a fluorescein-tagged peroxisomal targeting sequence peptide (F-PTS1, acetyl-C{K(FITC)}GGAKL) for investigating pH regulation of glycosomes in live procyclic form Trypanosoma brucei When added to cells, this fluorescent peptide is internalized within vesicular structures, including glycosomes, and can be visualized after 30-60 min. Using F-PTS1 we are able to observe the pH conditions inside glycosomes in response to starvation conditions. Previous studies have shown that in the absence of glucose, the glycosome exhibits mild acidification from pH 7.4 ± 0.2 to 6.8 ± 0.2. Our results suggest that this response occurs under proline starvation as well. This pH regulation is found to be independent from cytosolic pH and requires a source of Na + ions. Glycosomes were also observed to be more resistant to external pH changes than the cytosol; placement of cells in acidic buffers (pH 5) reduced the pH of the cytosol by 0.8 ± 0.1 pH units, whereas glycosomal pH decreases by 0.5 ± 0.1 pH units. This observation suggests that regulation of glycosomal pH is different and independent from cytosolic pH regulation. Furthermore, pH regulation is likely to work by an active process, because cells depleted of ATP with 2-deoxyglucose and sodium azide were unable to properly regulate pH. Finally, inhibitor studies with bafilomycin and EIPA suggest that both V-ATPases and Na + /H + exchangers are required for glycosomal pH regulation. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Progesterone production requires activation of caspase-3 in preovulatory granulosa cells in a serum starvation model.

PubMed

An, Li-Sha; Yuan, Xiao-Hua; Hu, Ying; Shi, Zi-Yun; Liu, Xiao-Qin; Qin, Li; Wu, Gui-Qing; Han, Wei; Wang, Ya-Qin; Ma, Xu

2012-11-01

Granulosa cells proliferate, differentiate, and undergo apoptosis throughout follicular development. Previous studies have demonstrated that stimulation of progesterone production is accompanied by caspase-3 activation. Moreover, we previously reported that arsenic enhanced caspase-3 activity coupled with progesterone production. Inhibition of caspase-3 activity can significantly inhibit progesterone production induced by arsenic or follicle-stimulating hormone (FSH). Here, we report that serum starvation induces caspase-3 activation coupled with augmentation of progesterone production. Serum starvation also increased the levels of cytochrome P450 cholesterol side chain cleavage enzyme (P450scc) and steroidogenic acute regulatory (StAR) protein, both of which may contribute to progesterone synthesis in preovulatory granulosa cells. Inhibition of caspase-3 activity resulted in a decrease in progesterone production. Deactivation of caspase-3 activity by caspase-3 specific inhibitor also resulted in decreases in P450scc and StAR expression, which may partly contribute to the observed decrease in progesterone production. Our study demonstrates for the first time that progesterone production in preovulatory granulosa cells is required for caspase-3 activation in a serum starvation model. Inhibition of caspase-3 activity can result in decreased expression of the steroidogenic proteins P450scc and StAR. Our work provides further details on the relationship between caspase-3 activation and steroidogenesis and indicates that caspase-3 plays a critical role in progesterone production by granulosa cells. Copyright © 2012 Elsevier Inc. All rights reserved.

[Intensity of pentose phosphate metabolism of carbohydrates in various brain areas in normal and starved animals].

PubMed

Kerimov, B F

2002-01-01

The activities of key enzymes of pentose phosphate pathway, glucose-6-phosphate dehydrogenase (G-6 PD) and 6-phosphogluconate dehydrogenase (6-PGD), were studied in cytoplasmatic fractions of brain cortical (limbic, orbital, sensorimotor cortex) and subcortical (myelencefalon, mesencefalon, hypothalamus) structures of rats subjected to starvation for 1, 2, 3, 5 and 7 days. Short-term starvation (1-3 days) caused activation of 6-GPD and 6-PGD both in cortical and subcortical structures. Long-term starvation for 5-7 days caused a decrease of activities of the pentose phosphate pathway enzymes in all studied structures. It is suggested that enzymes of pentose phosphate pathway in nervous tissues are functionally and metabolically related to glutathione system and during starvation they indirectly participate in the regulation lipid peroxidation processes.

Effect of starvation and refeeding on digestive enzyme activities in sturgeon (Acipenser naccarii) and trout (Oncorhynchus mykiss).

PubMed

Furné, Miriam; García-Gallego, Manuel; Hidalgo, M Carmen; Morales, Amalia E; Domezain, Alberto; Domezain, Julio; Sanz, Ana

2008-04-01

The digestive enzyme activities were determined in Adriatic sturgeon and rainbow trout during starvation and refeeding period. Overall, the digestive enzyme activities are affected in the same sense in both species. The protease and lipase activities were decreased later than amylase activity. Even after 1 month of starvation, both species would be prepared to digest protein and lipids in an effective way. After 72 days of starvation, the digestive machinery of the sturgeon and of the trout shows an altered capacity to digest macronutrients. The capacity to digest proteins and lipids, after 60 days of refeeding, begins to become re-established in sturgeon and trout. In contrast, in this period, the capacity to digest carbohydrates remains depressed in both species.

MAPK14/p38α-dependent modulation of glucose metabolism affects ROS levels and autophagy during starvation.

PubMed

Desideri, Enrico; Vegliante, Rolando; Cardaci, Simone; Nepravishta, Ridvan; Paci, Maurizio; Ciriolo, Maria Rosa

2014-09-01

Increased glycolytic flux is a common feature of many cancer cells, which have adapted their metabolism to maximize glucose incorporation and catabolism to generate ATP and substrates for biosynthetic reactions. Indeed, glycolysis allows a rapid production of ATP and provides metabolic intermediates required for cancer cells growth. Moreover, it makes cancer cells less sensitive to fluctuations of oxygen tension, a condition usually occurring in a newly established tumor environment. Here, we provide evidence for a dual role of MAPK14 in driving a rearrangement of glucose metabolism that contributes to limiting reactive oxygen species (ROS) production and autophagy activation in condition of nutrient deprivation. We demonstrate that MAPK14 is phosphoactivated during nutrient deprivation and affects glucose metabolism at 2 different levels: on the one hand, it increases SLC2A3 mRNA and protein levels, resulting in a higher incorporation of glucose within the cell. This event involves the MAPK14-mediated enhancement of HIF1A protein stability. On the other hand, MAPK14 mediates a metabolic shift from glycolysis to the pentose phosphate pathway (PPP) through the modulation of PFKFB3 (6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase 3) degradation by the proteasome. This event requires the presence of 2 distinct degradation sequences, KEN box and DSG motif Ser273, which are recognized by 2 different E3 ligase complexes. The mutation of either motif increases PFKFB3 resistance to starvation-induced degradation. The MAPK14-driven metabolic reprogramming sustains the production of NADPH, an important cofactor for many reduction reactions and for the maintenance of the proper intracellular redox environment, resulting in reduced levels of ROS. The final effect is a reduced activation of autophagy and an increased resistance to nutrient deprivation.

Two Young MicroRNAs Originating from Target Duplication Mediate Nitrogen Starvation Adaptation via Regulation of Glucosinolate Synthesis in Arabidopsis thaliana1[W

PubMed Central

He, Hua; Liang, Gang; Li, Yang; Wang, Fang; Yu, Diqiu

2014-01-01

Nitrogen is an essential macronutrient required for plant growth and development. A number of genes respond to nitrogen starvation conditions. However, the functions of most of these nitrogen starvation-responsive genes are unclear. Our recent survey suggested that many microRNAs (miRNAs) are responsive to nitrogen starvation in Arabidopsis thaliana. Here, we identified a new miRNA (miR5090) from the complementary transcript of the MIR826 gene. Further investigation uncovered that both miRNA genes recently evolved from the inverse duplication of their common target gene, ALKENYL HYDROXALKYL PRODUCING2 (AOP2). Similar to miR826, miR5090 is induced by nitrogen starvation. By contrast, the AOP2 transcript level was negatively correlated with miR826 and miR5090 under nitrogen starvation. GUS-fused AOP2 expression suggested that AOP2 was posttranscriptionally suppressed by miR826 and miR5090. miRNA transgenic plants with significantly low AOP2 expression accumulated fewer Met-derived glucosinolates, phenocopying the aop2 mutants. Most glucosinolate synthesis-associated genes were repressed under nitrogen starvation conditions. Furthermore, miRNA transgenic plants with less glucosinolate displayed enhanced tolerance to nitrogen starvation, including high biomass, more lateral roots, increased chlorophyll, and decreased anthocyanin. Meanwhile, nitrogen starvation-responsive genes were up-regulated in transgenic plants, implying improved nitrogen uptake activity. Our study reveals a mechanism by which Arabidopsis thaliana regulates the synthesis of glucosinolates to adapt to environmental changes in nitrogen availability. PMID:24367020

Evidence for the Adverse Effect of Starvation on Bone Quality: A Review of the Literature

PubMed Central

Kueper, Janina; Beyth, Shaul; Liebergall, Meir; Kaplan, Leon; Schroeder, Josh E.

2015-01-01

Malnutrition and starvation's possible adverse impacts on bone health and bone quality first came into the spotlight after the horrors of the Holocaust and the ghettos of World War II. Famine and food restrictions led to a mean caloric intake of 200–800 calories a day in the ghettos and concentration camps, resulting in catabolysis and starvation of the inhabitants and prisoners. Severely increased risks of fracture, poor bone mineral density, and decreased cortical strength were noted in several case series and descriptive reports addressing the medical issues of these individuals. A severe effect of severely diminished food intake and frequently concomitant calcium- and Vitamin D deficiencies was subsequently proven in both animal models and the most common cause of starvation in developed countries is anorexia nervosa. This review attempts to summarize the literature available on the impact of the metabolic response to Starvation on overall bone health and bone quality. PMID:25810719

Effect of extended and daily short-term starvation/shut-down events on the performance of a biofilter treating toluene vapors.

PubMed

Jiménez, Lucero; Arriaga, Sonia; Muñoz, Raúl; Aizpuru, Aitor

2017-12-01

Industrial emissions of Volatile Organic Compounds are usually discontinuous. To assess the impact of interruptions in pollutant supply on the performance of biological treatment systems, two identical biofilters previously operated under continuous toluene loadings were subjected for 110 days to extended (12, 24, 36, 48, 60, 72, 84 and 96 h) and for a week to daily (8 h on, 16 h off) toluene starvation/shutdown events. One biofilter was operated under complete shutdowns (both air and toluene supply were interrupted), while the other maintained the air supply under toluene starvation. The biofilter operated under complete shutdowns was able to withstand both the extended and daily pollutant interruptions, while starvation periods >24 h severely impacted the performance of the other biofilter, with a removal efficiency decrease from 97.7 ± 0.1% to 45.4 ± 6.7% at the end of the extended starvation periods. This deterioration was likely due to a reduction in liquid lixiviation (from a total volume of 2380 mL to 1800 mL) mediated by the countercurrent airflow during the starvation periods. The presence of air under toluene starvation also favored the accumulation of inactive biomass, thus increasing the pressure drop from 337 to 700 mm H 2 O.m -1 , while decreasing the wash out of acidic by-products with a significantly higher pH of leachates (Student paired t-test <0.05). This study confirmed the need to prevent the accumulation of inhibitory compounds produced during process perturbation in order to increase biofiltration robustness. Process operation with sufficient drainage in the packing material and the absence of countercurrent airflow are highly recommended during toluene deprivation periods. Copyright © 2017. Published by Elsevier Ltd.

Effects of repeated cycles of starvation and refeeding on lungs of growing rats.

PubMed

Sahebjami, H; Domino, M

1992-12-01

Adult male rats were subjected to four cycles of mild starvation (2 wk) and refeeding (1 wk) and were compared with a fed group. Starvation was induced by giving rats one-third of their measured daily food consumption. During each starvation cycle, rats lost approximately 20% of their body weight. Despite catch-up growth and overall weight gain, starved rats had lower final body weight than fed rats. Lung dry weight and lung volumes were also reduced in the starved group. The mechanical properties of air- and saline-filled lungs did not change significantly with repeated cycles of starvation. Mean linear intercept was similar in the two groups, but alveolar surface area was reduced in the starved rats. Total content of crude connective tissue and concentration per lung dry weight of hydroxyproline and crude connective tissue were reduced in starved rats. We conclude that lung growth is retarded in growing rats subjected to repeated cycles of mild starvation and refeeding, as manifested by smaller lung volume and reduced alveolar surface area. Because alveolar size is unchanged, a reduced number of alveoli is most likely responsible for decreased lung volumes.

Proteomic analysis of rainbow trout (Oncorhynchus mykiss) intestinal epithelia: physiological acclimation to short-term starvation.

PubMed

Baumgarner, Bradley L; Bharadwaj, Anant S; Inerowicz, Dorota; Goodman, Angela S; Brown, Paul B

2013-03-01

The intestinal epithelia form the first line of defense against harmful agents in the gut lumen of most monogastric vertebrates, including teleost fishes. Previous investigations into the effect of starvation on the intestinal epithelia of teleost fishes have focused primarily on changes in morphological characteristics and targeted molecular analysis of specific enzymes. The goal of this study was to use a comprehensive approach to help reveal how the intestinal epithelia of carnivorous teleost fishes acclimate to short-term nutrient deprivation. We utilized two-dimensional gel electrophoresis (2-DE) to conduct the proteomic analysis of the mucosal and epithelial layer of the anterior gut intestinal tract (GIT) from satiation fed vs. 4 week starved rainbow trout (Oncorhynchus mykiss). A total of 40 proteins were determined to be differentially expressed and were subsequently picked for in-gel trypsin digestion. Peptide mass fingerprint analysis was conducted using matrix assisted laser desorption time-of-flight/time-of-flight. Nine of the 11 positively identified proteins were directly related to innate immunity. The expression of α-1 proteinase inhibitor decreased in starved vs. fed fish. Also, the concentration of one leukocyte elastase inhibitor (LEI) isomer decreased in starved fish, though the concentration of another LEI isomer increased in due to starvation. In addition, starvation promoted an increased concentration of the important xenobiotic-transporter p-glycoprotein. Finally, starvation resulted in a significant increase in type II keratin E2. Overall, our results indicate that starvation promoted a reduced capacity to inhibit enzymatic stress but increased xenobiotic resistance and paracellular permeability of epithelial cells in the anterior intestine of rainbow trout. Copyright © 2012 Elsevier Inc. All rights reserved.

Peptide trafficking and translocation across membranes in cellular signaling and self-defense strategies.

PubMed

Abele, Rupert; Tampé, Robert

2009-08-01

Cells are metastable per se and a fine-tuned balance of de novo protein synthesis and degradation shapes their proteome. The primary function of peptides is to supply amino acids for de novo protein synthesis or as an energy source during starvation. Peptides are intrinsically short-lived and steadily trimmed by an armada of intra and extracellular peptidases. However, peptides acquired additional, more sophisticated tasks already early in evolution. Here, we summarize current knowledge on intracellular peptide trafficking and translocation mediated by ATP-binding cassette (ABC) transport machineries with a focus on the functions of protein degradation products as important signaling molecules in self-defense mechanisms.

Long-term starvation and subsequent recovery of nitrifiers in aerated submerged fixed-bed biofilm reactors.

PubMed

Elawwad, Abdelsalam; Sandner, Hendrik; Kappelmeyer, Uwe; Koeser, Heinz

2013-01-01

The effectiveness of three operational strategies for maintaining nitrifiers in bench-scale, aerated, submerged fixed-bed biofilm reactors (SFBBRs) during long-term starvation at 20 degrees C were evaluated. The operational strategies were characterized by the resulting oxidation-reduction potential (ORP) in the SFBBRs. The activity rates of the nitrifiers were measured and the activity decay was expressed by half-life times. It was found that anoxic and alternating anoxic/aerobic conditions were the best ways to preserve ammonia-oxidizing bacteria (AOB) during long starvation periods and resulted in half-life times of up to 34 and 28 days, respectively. Extended anaerobic conditions caused the half-life for AOB to decrease to 21 days. In comparison, the activity decay of nitrite-oxidizing bacteria (NOB) tended to be slightly faster. The activity of AOB biofilms that were kept for 97 days under anoxic conditions could be completely recovered in less than one week, while over 4 weeks was needed for AOB kept under anaerobic conditions. NOB were more sensitive to starvation and required longer recovery periods than AOB. For complete recovery, NOB needed approximately 7 weeks, regardless of the starvation conditions applied. Using the fluorescence in situ hybridization (FISH) technique, Nitrospira was detected as the dominant NOB genus. Among the AOB, the terminal restriction fragment length polymorphism (TRFLP) technique showed that during starvation and recovery periods, the relative frequency of species shifted to Nitrosomonas europaea/eutropha, regardless of the starvation condition. The consequences of these findings for the operation of SFBBRs under low-load and starvation conditions are discussed.

Dormancy in Deinococcus sp. UDEC-P1 as a survival strategy to escape from deleterious effects of carbon starvation and temperature.

PubMed

Guerra, Matías; González, Karina; González, Carlos; Parra, Boris; Martínez, Miguel

2015-09-01

Dormancy is characterized by low metabolism and absence of protein synthesis and cellular division enabling bacterial cells to survive under stress. The aim was to determine if carbon starvation and low temperature are factors that modify the proportion of dormant/active cells in Deinococcus sp. UDEC-P1. By flow cytometry, RedoxSensor Green (RSG) was used to quantify metabolic activity and Propidium Iodide (PI) to evaluate membrane integrity in order to determine the percentage of dormant cells. Cell size and morphology were determined using scanning electronic microscopy. Under carbon starvation at 30°C, Deinococcus sp. UDEC-P1 increased its proportion of dormant cells from 0.1% to 20%, decreased the count of culturable cells and average cell volume decreased 7.1 times. At 4°C, however, the proportion of dormant cells increased only to 6%, without a change in the count of culturable cells and an average cellular volume decrease of 4.1 times and 3% of the dormant cells were able to be awakened. Results indicate a greater proportion of dormant Deinococcus sp. UDEC-P1 cells at 30ºC and it suggests that carbon starvation is more deleterious condition at 30ºC than 4ºC. For this reason Deinococcus sp. UDEC-P1 cells are more likely to enter into dormancy at higher temperature as a strategy to survive. Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.

The control of fatty acid metabolism in liver cells from fed and starved sheep.

PubMed Central

Lomax, M A; Donaldson, I A; Pogson, C I

1983-01-01

Isolated liver cells prepared from starved sheep converted palmitate into ketone bodies at twice the rate seen with cells from fed animals. Carnitine stimulated palmitate oxidation only in liver cells from fed sheep, and completely abolished the difference between fed and starved animals in palmitate oxidation. The rates of palmitate oxidation to CO2 and of octanoate oxidation to ketone bodies and CO2 were not affected by starvation or carnitine. Neither starvation nor carnitine altered the ratio of 3-hydroxybutyrate to acetoacetate or the rate of esterification of [1-14C]palmitate. Propionate, lactate, pyruvate and fructose inhibited ketogenesis from palmitate in cells from fed sheep. Starvation or the addition of carnitine decreased the antiketogenic effectiveness of gluconeogenic precursors. Propionate was the most potent inhibitor of ketogenesis, 0.8 mM producing 50% inhibition. Propionate, lactate, fructose and glycerol increased palmitate esterification under all conditions examined. Lactate, pyruvate and fructose stimulated oxidation of palmitate and octanoate to CO2. Starvation and the addition of gluconeogenic precursors stimulated apparent palmitate utilization by cells. Propionate, lactate and pyruvate decreased cellular long-chain acylcarnitine concentrations. Propionate decreased cell contents of CoA and acyl-CoA. It is suggested that propionate may control hepatic ketogenesis by acting at some point in the beta-oxidation sequence. The results are discussed in relation to the differences in the regulation of hepatic fatty acid metabolism between sheep and rats. PMID:6615480

The carbon starvation response of the ectomycorrhizal fungus Paxillus involutus

DOE PAGES

Ellstrom, Magnus; Shah, Firoz; Johansson, Tomas; ...

2015-03-16

The amounts of carbon allocated to the fungal partner in ectomycorrhizal associations can vary substantially depending on the plant growth and the soil nutrient conditions, and the fungus may frequently be confronted with limitations in carbon. We used chemical analysis and transcriptome profiling to examine the physiological response of the ectomycorrhizal fungus Paxillus involutus to carbon starvation during axenic cultivation. Carbon starvation induced a decrease in the biomass. Concomitantly, ammonium, cell wall material (chitin) and proteolytic enzymes were released into the medium, which suggest autolysis. Compared with the transcriptome of actively growing hyphae, about 45% of the transcripts analyzed weremore » differentially regulated during C-starvation. Induced during starvation were transcripts encoding extracellular enzymes such as peptidases, chitinases and laccases. In parallel, transcripts of N-transporters were upregulated, which suggest that some of the released nitrogen compounds were re-assimilated by the mycelium. The observed changes suggest that the carbon starvation response in P. involutus is associated with complex cellular changes that involves autolysis, recycling of intracellular compounds by autophagy and reabsorption of the extracellular released material. The study provides molecular markers that can be used to examine the role of autolysis for the turnover and survival of the ectomycorrhizal mycelium in soils.« less

The carbon starvation response of the ectomycorrhizal fungus Paxillus involutus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ellstrom, Magnus; Shah, Firoz; Johansson, Tomas

The amounts of carbon allocated to the fungal partner in ectomycorrhizal associations can vary substantially depending on the plant growth and the soil nutrient conditions, and the fungus may frequently be confronted with limitations in carbon. We used chemical analysis and transcriptome profiling to examine the physiological response of the ectomycorrhizal fungus Paxillus involutus to carbon starvation during axenic cultivation. Carbon starvation induced a decrease in the biomass. Concomitantly, ammonium, cell wall material (chitin) and proteolytic enzymes were released into the medium, which suggest autolysis. Compared with the transcriptome of actively growing hyphae, about 45% of the transcripts analyzed weremore » differentially regulated during C-starvation. Induced during starvation were transcripts encoding extracellular enzymes such as peptidases, chitinases and laccases. In parallel, transcripts of N-transporters were upregulated, which suggest that some of the released nitrogen compounds were re-assimilated by the mycelium. The observed changes suggest that the carbon starvation response in P. involutus is associated with complex cellular changes that involves autolysis, recycling of intracellular compounds by autophagy and reabsorption of the extracellular released material. The study provides molecular markers that can be used to examine the role of autolysis for the turnover and survival of the ectomycorrhizal mycelium in soils.« less

Glutamine starvation enhances PCV2 replication via the phosphorylation of p38 MAPK, as promoted by reducing glutathione levels.

PubMed

Chen, Xingxiang; Shi, Xiuli; Gan, Fang; Huang, Da; Huang, Kehe

2015-03-18

Glutamine has a positive effect on ameliorating reproductive failure caused by porcine circovirus type 2 (PCV2). However, the mechanism by which glutamine affects PCV2 replication remains unclear. This study was conducted to investigate the effects of glutamine on PCV2 replication and its underlying mechanisms in vitro. The results show that glutamine promoted PK-15 cell viability. Surprisingly, glutamine starvation significantly increased PCV2 replication. The promotion of PCV2 replication by glutamine starvation disappeared after fresh media with 4 mM glutamine was added. Likewise, promotion of PCV2 was observed after adding buthionine sulfoximine (BSO). Glutamine starvation or BSO treatment increased the level of p38 MAPK phosphorylation and PCV2 replication in PK-15 cells. Meanwhile, p38 MAPK phosphorylation and PCV2 replication significantly decreased in p38-knockdown PK-15 cells. Promotion of PCV2 replication caused by glutamine starvation could be blocked in p38-knockdown PK-15 cells. Therefore, glutamine starvation increased PCV2 replication by promoting p38 MAPK activation, which was associated with the down regulation of intracellular glutathione levels. Our findings may contribute toward interpreting the possible pathogenic mechanism of PCV2 and provide a theoretical reference for application of glutamine in controlling porcine circovirus-associated diseases.

[Offspring quality and its related factors of different Brachionus calyciflorus clones].

PubMed

Dong, Lili; Xi, Yilong; Zhang, Lei

2006-12-01

This paper studied the neonate starvation-endurance duration of four Brachionus calyciflorus clones (Clone A, B, C and D) with different biochemical-genetic characteristics at 15 degrees C, 20 degrees C, 25 degrees C and 30 degrees C, and the relationships of this duration with the temperature and the body- and egg volumes of B. calyciflorus. The results showed that at 15 degrees C, the neonates of Clone B had the shortest starvation-endurance duration (45.67 h); at 20 degrees C and 25 degrees C, the neonates' starvation-endurance duration of Clone C was the longest, being 61.33 h and 72.01 h, respectively; while at 30 degrees C, this duration of Clone A was the longest (40.11 h). The neonates' starvation-endurance duration of Clone A was the longest at 15 degrees C, those of Clone B and C were the shortest at 30 degrees C, while that of Clone D decreased with raising temperature. The neonates' starvation-endurance duration of all the four clones was negatively correlated with temperature. There was a negative correlation between this duration of Clone A and its egg volume, and the reverse was true for Clone C. The neonates' starvation-endurance duration of Clone B and D was positively correlated with the body volume of rotifer mother.

Integrated analysis of transcriptome and metabolites reveals an essential role of metabolic flux in starch accumulation under nitrogen starvation in duckweed.

PubMed

Yu, Changjiang; Zhao, Xiaowen; Qi, Guang; Bai, Zetao; Wang, Yu; Wang, Shumin; Ma, Yubin; Liu, Qian; Hu, Ruibo; Zhou, Gongke

2017-01-01

Duckweed is considered a promising source of energy due to its high starch content and rapid growth rate. Starch accumulation in duckweed involves complex processes that depend on the balanced expression of genes controlled by various environmental and endogenous factors. Previous studies showed that nitrogen starvation induces a global stress response and results in the accumulation of starch in duckweed. However, relatively little is known about the mechanisms underlying the regulation of starch accumulation under conditions of nitrogen starvation. In this study, we used next-generation sequencing technology to examine the transcriptome responses of Lemna aequinoctialis 6000 at three stages (0, 3, and 7 days) during nitrogen starvation in the presence of exogenously applied sucrose. Overall, 2522, 628, and 1832 differentially expressed unigenes (DEGs) were discovered for the treated and control samples. Clustering and enrichment analysis of DEGs revealed several biological processes occurring under nitrogen starvation. Genes involved in nitrogen metabolism showed the earliest responses to nitrogen starvation, whereas genes involved in carbohydrate biosynthesis were responded subsequently. The expression of genes encoding nitrate reductase, glutamine synthetase, and glutamate synthase was down-regulated under nitrogen starvation. The expression of unigenes encoding enzymes involved in gluconeogenesis was up-regulated, while the majority of unigenes involved in glycolysis were down-regulated. The metabolite results showed that more ADP-Glc was accumulated and lower levels of UDP-Glc were accumulated under nitrogen starvation, the activity of AGPase was significantly increased while the activity of UGPase was dramatically decreased. These changes in metabolite levels under nitrogen starvation are roughly consistent with the gene expression changes in the transcriptome. Based on these results, it can be concluded that the increase of ADP-glucose and starch contents under nitrogen starvation is a consequence of increased output from the gluconeogenesis and TCA pathways, accompanied with the reduction of lipids and pectin biosynthesis. The results provide novel insights into the underlying mechanisms of starch accumulation during nitrogen starvation, which provide a foundation for the improvement of advanced bioethanol production in duckweed.

Extracellular ATP decreases trophoblast invasion, spiral artery remodeling and immune cells in the mesometrial triangle in pregnant rats.

PubMed

Spaans, F; Melgert, B N; Chiang, C; Borghuis, T; Klok, P A; de Vos, P; van Goor, H; Bakker, W W; Faas, M M

2014-08-01

Preeclampsia is characterized by deficient trophoblast invasion and spiral artery remodeling, a process governed by inflammatory cells. High levels of the danger signal extracellular adenosine triphosphate (ATP) have been found in women with preeclampsia and infusion of ATP in pregnant rats induced preeclampsia-like symptoms such as albuminuria and placental ischemia. We hypothesized that ATP inhibits trophoblast invasion and spiral artery remodeling and affects macrophages and natural killer (NK) cells present in the rat mesometrial triangle. Pregnant rats were infused with ATP or saline (control) on day 14 of pregnancy. Rats were sacrificed on day 15, 17 or 20 of pregnancy and placentas with mesometrial triangle were collected. Sections were stained for trophoblast cells, α-smooth muscle actin (spiral artery remodeling), NK cells and various macrophage populations. Expression of various cytokines in the mesometrial triangle was analyzed using real-time RT-PCR. ATP infusion decreased interstitial trophoblast invasion on day 17 and spiral artery remodeling on day 17 and 20, increased activated tartrate resistant acid phosphatase (TRAP)-positive macrophages on day 15, decreased NK cells on day 17 and 20, and decreased inducible nitric oxide synthase (iNOS)-positive and CD206-positive macrophages and TNF-α and IL-33 expression at the end of pregnancy (day 20). Interstitial trophoblast invasion and spiral artery remodeling in the rat mesometrial triangle were decreased by infusion of ATP. These ATP-induced modifications were preceded by an increase in activated TRAP-positive macrophages and coincided with NK cell numbers, suggesting that they are involved. Trophoblast invasion and spiral artery remodeling may be inhibited by ATP-induced activated macrophages and decreased NK cells in the mesometrial triangle in rat pregnancy. Copyright © 2014 Elsevier Ltd. All rights reserved.

Studies on the Biochemistry and Fine Structure of Silica Shell Formation in Diatoms. Chemical Composition of Navicula pelliculosa during Silicon-Starvation Synchrony 1

PubMed Central

Coombs, J.; Darley, W. M.; Holm-Hansen, O.; Volcani, B. E.

1967-01-01

Changes are reported in total cellular organic carbon, nucleic acids, proteins, carbohydrates, lipids and chlorophylls during the course of silicon-starvation synchrony of Navicula pelliculosa. All constituents increased at the same rate, relative to cell number, for 30 hours of exponential growth during which silicon was depleted from the medium. Increase in cell number then stopped, but net synthesis of most components continued for a further 5 to 7 hours before ceasing. Deoxyribonucleic acids and lipids accumulated throughout the 14 hour silicon-starvation period. When silicon was resupplied, lipid synthesis ceased and organic carbon and carbohydrates decreased slightly. Net synthesis remained low during the 4 hour silicon uptake period but was resumed at higher rates as cell number began to rise. In cultures transferred to the dark 1 hour prior to readdition of silicon, total carbon, carbohydrates, and lipids decreased markedly during silicon uptake and cell separation. This was due in part to conversion of protein which maintained the protein level of the dark cells close to that of cells kept in the light. Mechanisms by which silicon starvation and reintroduction of silicon might affect rates of cellular synthesis are discussed. PMID:6080872

The changes in the biochemical compositions and enzymatic activities of rotifer (Brachionus plicatilis, Müller) and Artemia during the enrichment and starvation periods.

PubMed

Naz, Mehmet

2008-12-01

The changes in the biochemical compositions and enzymatic activities of rotifer (Brachionus plicatilis) and Artemia, enriched and stored at 4 degrees C temperature, were determined. The total starvation period was 16 h and samples were taken at the end of the 8th and 16th hours. In present study, the rotifer and nauplii catabolized a large proportion of the protein during the enrichment period. Lipid contents of both live preys increased during the enrichment period and decreased in nauplii and metanauplii throughout the starvation period but lipid content of the rotifer remained relatively constant during the starvation period. The changes observed in the amino acid compositions of Artemia and the rotifer were statistically significant (P < 0.05). The conspicuous decline the essential amino acid (EAA) and nonessential amino acid (NEAA) content of the rotifer was observed during the enrichment period. However, the essential amino acid (EAA) and nonessential amino acid (NEAA) contents of Artemia nauplii increased during the enrichment period. The unenriched and enriched rotifers contained more monounsaturated fatty acid (MUFAs) than polyunsaturated fatty acid (PUFAs) and saturated fatty acids (SFA). However, Artemia contained more PUFAs than MUFAs and SFA during the experimental period. A sharp increase in the amounts of docosahexaenoic acid (DHA) during the enrichment of the rotifer and Artemia nauplii was observed. However, the amount of DHA throughout the starvation period decreased in Artemia metanauplii but not in Artemia nauplii. Significant differences in tryptic, leucine aminopeptidase N (LAP), and alkaline phosphatase (AP) enzyme activities of Artemia and rotifer were observed during the enrichment and starvation period (P < 0.05). The digestive enzymes derived from live food to fish larvae provided the highest contribution at the end of the enrichment period. In conclusion, the results of the study provide important contributions to determine the most suitable live food offering time for marine fish larvae. Rotifer should be offered to fish larvae at the end of the enrichment period, Artemia nauplii just after hatching and before being stored at 4 degrees C, and Artemia metanauplii at the end of the enrichment and throughout the starvation period.

[Effect of starvation on blood protein levels in the population of Dobrinja (1992-1995)].

PubMed

Hasković, E

2000-01-01

In nutritional protein deficiency, numerous studies verified utilization of amino acids generated from tissue degradation in intensive protein synthesis. Unlike liver, muscle protein synthesis is extremely dependent on external supplies of essential amino acids. Prolonged nutritional protein deficiency results in decrease of body weight as well as total protein concentration, in particular in early days of starvation. In prolonged starvation during the war, significant decrease of body weight was registered in 70 subjects while their total protein concentration remained within the expected range and did not significantly differ the values recorded in the control group. Concentration of serum albumines in the control group was lower than the concentration recorded in the tested group, while the serum globulins concentration was higher in the control group. Although the difference in body weight between the tested and the control group was statistically significant, no significant difference in the concentration of total proteins, albumines and globulines was recorded.

Neuronal death induced by misfolded prion protein is due to NAD+ depletion and can be relieved in vitro and in vivo by NAD+ replenishment

PubMed Central

Zhou, Minghai; Ottenberg, Gregory; Sferrazza, Gian Franco; Hubbs, Christopher; Fallahi, Mohammad; Rumbaugh, Gavin; Brantley, Alicia F.

2015-01-01

The mechanisms of neuronal death in protein misfolding neurodegenerative diseases such as Alzheimer’s, Parkinson’s and prion diseases are poorly understood. We used a highly toxic misfolded prion protein (TPrP) model to understand neurotoxicity induced by prion protein misfolding. We show that abnormal autophagy activation and neuronal demise is due to severe, neuron-specific, nicotinamide adenine dinucleotide (NAD+) depletion. Toxic prion protein-exposed neuronal cells exhibit dramatic reductions of intracellular NAD+ followed by decreased ATP production, and are completely rescued by treatment with NAD+ or its precursor nicotinamide because of restoration of physiological NAD+ levels. Toxic prion protein-induced NAD+ depletion results from PARP1-independent excessive protein ADP-ribosylations. In vivo, toxic prion protein-induced degeneration of hippocampal neurons is prevented dose-dependently by intracerebral injection of NAD+. Intranasal NAD+ treatment of prion-infected sick mice significantly improves activity and delays motor impairment. Our study reveals NAD+ starvation as a novel mechanism of autophagy activation and neurodegeneration induced by a misfolded amyloidogenic protein. We propose the development of NAD+ replenishment strategies for neuroprotection in prion diseases and possibly other protein misfolding neurodegenerative diseases. PMID:25678560

Remote ischemic preconditioning with a specialized protocol activates the non-neuronal cardiac cholinergic system and increases ATP content in the heart.

PubMed

Oikawa, Shino; Mano, Asuka; Takahashi, Rina; Kakinuma, Yoshihiko

2015-11-01

Ischemic preconditioning (IPC) renders the targeted organ resistant to prolonged ischemic insults, leading to organoprotection. Among several means to achieve IPC, we reported that remote ischemic preconditioning (RIPC) activates the non-neuronal cardiac cholinergic system (NNCCS) to accelerate de novo ACh synthesis in cardiomyocytes. In the current study, we aimed to optimize a specific protocol to most efficiently activate NNCCS using RIPC. In this study, we elucidated that the protocol with 3 min of ischemia repeated three times increased cardiac ChAT expression (139.2 ± 0.4%; P < 0.05) as well as ACh (14.2 ± 2.0× 10(-8) M; P< 0.05) and ATP content (2.13 ± 0.19 μmol/g tissue; P < 0.05) in the heart. Moreover, in the specific protocol, several characteristic responses against energy starvation and for obtaining adequate energy were observed; therefore, it is suggested that RIPC evokes a robust response by the heart to activate NNCCS through the modification of energy metabolism. Copyright © 2015 Elsevier B.V. All rights reserved.

A Synthetic Analogue of Neopeltolide, 8,9-Dehydroneopeltolide, Is a Potent Anti-Austerity Agent against Starved Tumor Cells.

PubMed

Fuwa, Haruhiko; Sato, Mizuho

2017-10-20

Neopeltolide, an antiproliferative marine macrolide, is known to specifically inhibit complex III of the mitochondrial electron transport chain (mETC). However, details of the biological mode-of-action(s) remain largely unknown. This work demonstrates potent cytotoxic activity of synthetic neopeltolide analogue, 8,9-dehydroneopeltolide (8,9-DNP), against starved human pancreatic adenocarcinoma PANC-1 cells and human non-small cell lung adenocarcinoma A549 cells. 8,9-DNP induced rapid dissipation of the mitochondrial membrane potential and depletion of intracellular ATP level in nutrient-deprived medium. Meanwhile, in spite of mTOR inhibition under starvation conditions, impairment of cytoprotective autophagy was observed as the lipidation of LC3-I to form LC3-II and the degradation of p62 were suppressed. Consequently, cells were severely deprived of energy sources and underwent necrotic cell death. The autophagic flux inhibited by 8,9-DNP could be restored by glucose, and this eventually rescued cells from necrotic death. Thus, 8,9-DNP is a potent anti-austerity agent that impairs mitochondrial ATP synthesis and cytoprotective autophagy in starved tumor cells.

A Synthetic Analogue of Neopeltolide, 8,9-Dehydroneopeltolide, Is a Potent Anti-Austerity Agent against Starved Tumor Cells

PubMed Central

Sato, Mizuho

2017-01-01

Neopeltolide, an antiproliferative marine macrolide, is known to specifically inhibit complex III of the mitochondrial electron transport chain (mETC). However, details of the biological mode-of-action(s) remain largely unknown. This work demonstrates potent cytotoxic activity of synthetic neopeltolide analogue, 8,9-dehydroneopeltolide (8,9-DNP), against starved human pancreatic adenocarcinoma PANC-1 cells and human non-small cell lung adenocarcinoma A549 cells. 8,9-DNP induced rapid dissipation of the mitochondrial membrane potential and depletion of intracellular ATP level in nutrient-deprived medium. Meanwhile, in spite of mTOR inhibition under starvation conditions, impairment of cytoprotective autophagy was observed as the lipidation of LC3-I to form LC3-II and the degradation of p62 were suppressed. Consequently, cells were severely deprived of energy sources and underwent necrotic cell death. The autophagic flux inhibited by 8,9-DNP could be restored by glucose, and this eventually rescued cells from necrotic death. Thus, 8,9-DNP is a potent anti-austerity agent that impairs mitochondrial ATP synthesis and cytoprotective autophagy in starved tumor cells. PMID:29053565

Contractile activity of ATP and diadenosine tetraphosphate on urinary bladder in the rats: role of superoxide anion and urothelium.

PubMed

Khattab, M M; Al-Hrasen, M N

2006-04-01

Both ATP and diadenosine tetraphosphate (AP(4)A) produced a dose-dependent contraction of rat isolated urinary bladder rings. The AP(4)A dose-response curve was to the left of that of ATP, and the maximum response was greater than that produced by ATP. Mechanical removal of the urothelium increased the contractile response to ATP by between 53% and 71%, and that to AP(4)A by 42% (at highest AP(4)A concentration) to 68% at lower concentration. Inhibition of Cu/Zn superoxide dismutase with diethylthiocarbamate (DETCA, 5 mm) significantly reduced the ATP-evoked contraction by 31% (at high ATP concentration) to 40% at low ATP concentration. Similarly, the AP(4)A-induced contractions were significantly decreased by 27% at low AP(4)A level to 38% at higher concentrations. Induction of exogenous superoxide anion stress by the use of the superoxide anion generator, pyrogallol (0.5 mm), significantly decreased both ATP- and AP(4)A-induced contractions of the rat urinary bladder over the whole dose range. Contractile responses to ATP decreased by 36-40%, and those to AP(4)A by 44-49%. In conclusion, the urinary bladder urothelium exerts an inhibitory control over the purinergic contractility produced by adenine mononucleotides and dinucleotides. Superoxide anion stress, whether endogenous or exogenous, attenuates the ATP-induced as well as AP(4)A-induced contractility.

Increased mitochondrial energy efficiency in skeletal muscle after long-term fasting: its relevance to animal performance.

PubMed

Bourguignon, Aurore; Rameau, Anaïs; Toullec, Gaëlle; Romestaing, Caroline; Roussel, Damien

2017-07-01

In the final stage of fasting, skeletal muscle mass and protein content drastically decrease when the maintenance of efficient locomotor activity becomes crucial for animals to reactivate feeding behaviour and survive a very long period of starvation. As mitochondrial metabolism represents the main physiological link between the endogenous energy store and animal performance, the aim of this study was to determine how a very long, natural period of fasting affected skeletal muscle mitochondrial bioenergetics in king penguin ( Aptenodytes patagonicus ) chicks. Rates of mitochondrial oxidative phosphorylation were measured in pectoralis permeabilized fibres and isolated mitochondria. Mitochondrial ATP synthesis efficiency and the activities of respiratory chain complexes were measured in mitochondria isolated from pectoralis muscle. Results from long-term (4-5 months) naturally fasted chicks were compared with those from short-term (10 day) fasted birds. The respiratory activities of muscle fibres and isolated mitochondria were reduced by 60% and 45%, respectively, on average in long-term fasted chicks compared with short-term fasted birds. Oxidative capacity and mitochondrial content of pectoralis muscle were lowered by long-term fasting. Bioenergetic analysis of pectoralis muscle also revealed that mitochondria were, on average, 25% more energy efficient in the final stage of fasting (4-5 months) than after 10 days of fasting (short-term fasted birds). These results suggest that the strong reduction in respiratory capacity of pectoralis muscle was partly alleviated by increased mitochondrial ATP synthesis efficiency. Such oxidative phosphorylation optimization can impact animal performance, e.g. the metabolic cost of locomotion or the foraging efficiency. © 2017. Published by The Company of Biologists Ltd.

Starvation effects on nitrogen and carbon stable isotopes of animals: an insight from meta-analysis of fasting experiments

PubMed Central

Akamatsu, Fumikazu; González, Angélica L.

2017-01-01

Nitrogen and carbon stable isotopic compositions (δ15N and δ13C) of consumers have been used for physiological and food web studies. Previous studies have shown δ15N and δ13C values are affected by several biological and environmental factors during starvation, but the generality of the effect of starvation on δ15N and δ13C values has not yet been tested. Here, we performed a meta-analysis to evaluate the effects of starvation on δ15N and δ13C values of consumers, and the underlying factors that may explain the observed variation. The δ15N and δ13C values were calculated as the differences between the final δ15N and δ13C values of consumers (post-starvation) and the pre-starvation values on each experiment. Our meta-analysis showed a large variation in the δ15N and δ13C values of consumers (δ15N range: –0.82 to 4.30‰; mean: 0.47‰ and δ13C range: –1.92 to 2.62‰; mean: 0.01‰). The δ15N values of most consumers increased along the length of the starvation period and were influenced by nitrogen excretion and thermoregulation types, probably because differences in nitrogen metabolism and thermoregulation affect nitrogen processing and excretion rates. None of our predictor variables accounted for the variation in δ13C values, which showed both increases and decreases due to fasting. Our findings suggest that starvation results in changes in consumer δ15N values which are mainly explained by the length of the fasting period and by nitrogen and energy metabolism, but the underlying mechanisms of the starvation effects on δ13C values seem to be more complex than previously thought. PMID:28879005

Glycogen with short average chain length enhances bacterial durability

NASA Astrophysics Data System (ADS)

Wang, Liang; Wise, Michael J.

2011-09-01

Glycogen is conventionally viewed as an energy reserve that can be rapidly mobilized for ATP production in higher organisms. However, several studies have noted that glycogen with short average chain length in some bacteria is degraded very slowly. In addition, slow utilization of glycogen is correlated with bacterial viability, that is, the slower the glycogen breakdown rate, the longer the bacterial survival time in the external environment under starvation conditions. We call that a durable energy storage mechanism (DESM). In this review, evidence from microbiology, biochemistry, and molecular biology will be assembled to support the hypothesis of glycogen as a durable energy storage compound. One method for testing the DESM hypothesis is proposed.

A functional 12T-insertion polymorphism in the ATP1A1 promoter confers decreased susceptibility to hypertension in a male Sardinian population.

PubMed

Herrera, Victoria L; Pasion, Khristine A; Moran, Ann Marie; Zaninello, Roberta; Ortu, Maria Francesca; Fresu, Giovanni; Piras, Daniela Antonella; Argiolas, Giuseppe; Troffa, Chiara; Glorioso, Valeria; Masala, Wanda; Glorioso, Nicola; Ruiz-Opazo, Nelson

2015-01-01

Identification of susceptibility genes for essential hypertension in humans has been a challenge due to its multifactorial pathogenesis complicated by gene-gene and gene-environment interactions, developmental programing and sex specific differences. These concurrent features make identification of causal hypertension susceptibility genes with a single approach difficult, thus requiring multiple lines of evidence involving genetic, biochemical and biological experimentation to establish causal functional mutations. Here we report experimental evidence encompassing genetic, biochemical and in vivo modeling that altogether support ATP1A1 as a hypertension susceptibility gene in males in Sardinia, Italy. ATP1A1 encodes the α1Na,K-ATPase isoform, the sole sodium pump in vascular endothelial and renal tubular epithelial cells. DNA-sequencing detected a 12-nucleotide long thymidine (12T) insertion(ins)/deletion(del) polymorphism within a poly-T sequence (38T vs 26T) in the ATP1A1 5'-regulatory region associated with hypertension in a male Sardinian population. The 12T-insertion allele confers decreased susceptibility to hypertension (P = 0.035; OR = 0.50 [0.28-0.93]) accounting for 12.1 mmHg decrease in systolic BP (P = 0.02) and 6.6 mmHg in diastolic BP (P = 0.046). The ATP1A1 promoter containing the 12T-insertion exhibited decreased transcriptional activity in in vitro reporter-assay systems, indicating decreased α1Na,K-ATPase expression with the 12T-insertion, compared with the 12T-deletion ATP1A1 promoter. To test the effects of decreased α1Na,K-ATPase expression on blood pressure, we measured blood pressure by radiotelemetry in three month-old, highly inbred heterozygous knockout ATP1A1+/- male mice with resultant 58% reduction in ATP1A1 protein levels. Male ATP1A1+/- mice showed significantly lower blood pressure (P < 0.03) than age-matched male wild-type littermate controls. Concordantly, lower ATP1A1 expression is expected to lower Na-reabsorption in the kidney thereby decreasing sodium-associated risk for hypertension and sodium-induced endothelial stiffness and dysfunction. Altogether, data support ATP1A1 as a hypertension susceptibility gene in a male Sardinian population, and mandate further investigation of its involvement in hypertension in the general population.

A Functional 12T-Insertion Polymorphism in the ATP1A1 Promoter Confers Decreased Susceptibility to Hypertension in a Male Sardinian Population

PubMed Central

Herrera, Victoria L.; Pasion, Khristine A.; Moran, Ann Marie; Zaninello, Roberta; Ortu, Maria Francesca; Fresu, Giovanni; Piras, Daniela Antonella; Argiolas, Giuseppe; Troffa, Chiara; Glorioso, Valeria; Masala, Wanda; Glorioso, Nicola; Ruiz-Opazo, Nelson

2015-01-01

Identification of susceptibility genes for essential hypertension in humans has been a challenge due to its multifactorial pathogenesis complicated by gene-gene and gene-environment interactions, developmental programing and sex specific differences. These concurrent features make identification of causal hypertension susceptibility genes with a single approach difficult, thus requiring multiple lines of evidence involving genetic, biochemical and biological experimentation to establish causal functional mutations. Here we report experimental evidence encompassing genetic, biochemical and in vivo modeling that altogether support ATP1A1 as a hypertension susceptibility gene in males in Sardinia, Italy. ATP1A1 encodes the α1Na,K-ATPase isoform, the sole sodium pump in vascular endothelial and renal tubular epithelial cells. DNA-sequencing detected a 12-nucleotide long thymidine (12T) insertion(ins)/deletion(del) polymorphism within a poly-T sequence (38T vs 26T) in the ATP1A1 5’-regulatory region associated with hypertension in a male Sardinian population. The 12T-insertion allele confers decreased susceptibility to hypertension (P = 0.035; OR = 0.50 [0.28–0.93]) accounting for 12.1 mmHg decrease in systolic BP (P = 0.02) and 6.6 mmHg in diastolic BP (P = 0.046). The ATP1A1 promoter containing the 12T-insertion exhibited decreased transcriptional activity in in vitro reporter-assay systems, indicating decreased α1Na,K-ATPase expression with the 12T-insertion, compared with the 12T-deletion ATP1A1 promoter. To test the effects of decreased α1Na,K-ATPase expression on blood pressure, we measured blood pressure by radiotelemetry in three month-old, highly inbred heterozygous knockout ATP1A1+/− male mice with resultant 58% reduction in ATP1A1 protein levels. Male ATP1A1+/− mice showed significantly lower blood pressure (P < 0.03) than age-matched male wild-type littermate controls. Concordantly, lower ATP1A1 expression is expected to lower Na-reabsorption in the kidney thereby decreasing sodium-associated risk for hypertension and sodium-induced endothelial stiffness and dysfunction. Altogether, data support ATP1A1 as a hypertension susceptibility gene in a male Sardinian population, and mandate further investigation of its involvement in hypertension in the general population. PMID:25615575

Metabolic rate and rates of protein turnover in food-deprived cuttlefish, Sepia officinalis (Linnaeus 1758)

PubMed Central

MacCormack, Tyson J.; Sykes, Antonio V.; Hall, Jennifer R.; Speers-Roesch, Ben; Callaghan, Neal I.; Driedzic, William R.

2016-01-01

To determine the metabolic response to food deprivation, cuttlefish (Sepia officinalis) juveniles were either fed, fasted (3 to 5 days food deprivation), or starved (12 days food deprivation). Fasting resulted in a decrease in triglyceride levels in the digestive gland, and after 12 days, these lipid reserves were essentially depleted. Oxygen consumption was decreased to 53% and NH4 excretion to 36% of the fed group following 3–5 days of food deprivation. Oxygen consumption remained low in the starved group, but NH4 excretion returned to the level recorded for fed animals during starvation. The fractional rate of protein synthesis of fasting animals decreased to 25% in both mantle and gill compared with fed animals and remained low in the mantle with the onset of starvation. In gill, however, protein synthesis rate increased to a level that was 45% of the fed group during starvation. In mantle, starvation led to an increase in cathepsin A-, B-, H-, and L-like enzyme activity and a 2.3-fold increase in polyubiquitin mRNA that suggested an increase in ubiquitin-proteasome activity. In gill, there was a transient increase in the polyubiquitin transcript levels in the transition from fed through fasted to the starved state and cathepsin A-, B-, H-, and L-like activity was lower in starved compared with fed animals. The response in gill appears more complex, as they better maintain rates of protein synthesis and show no evidence of enhanced protein breakdown through recognized catabolic processes. PMID:27053650

Comparison of sister species identifies factors underpinning plastid compatibility in green sea slugs

PubMed Central

de Vries, Jan; Woehle, Christian; Christa, Gregor; Wägele, Heike; Tielens, Aloysius G. M.; Jahns, Peter; Gould, Sven B.

2015-01-01

The only animal cells known that can maintain functional plastids (kleptoplasts) in their cytosol occur in the digestive gland epithelia of sacoglossan slugs. Only a few species of the many hundred known can profit from kleptoplasty during starvation long-term, but why is not understood. The two sister taxa Elysia cornigera and Elysia timida sequester plastids from the same algal species, but with a very different outcome: while E. cornigera usually dies within the first two weeks when deprived of food, E. timida can survive for many months to come. Here we compare the responses of the two slugs to starvation, blocked photosynthesis and light stress. The two species respond differently, but in both starvation is the main denominator that alters global gene expression profiles. The kleptoplasts' ability to fix CO2 decreases at a similar rate in both slugs during starvation, but only E. cornigera individuals die in the presence of functional kleptoplasts, concomitant with the accumulation of reactive oxygen species (ROS) in the digestive tract. We show that profiting from the acquisition of robust plastids, and key to E. timida's longer survival, is determined by an increased starvation tolerance that keeps ROS levels at bay. PMID:25652835

Effects of starvation on intermolt development in Calanus finmarchicus copepodites: a comparison between theoretical models and field studies1

NASA Astrophysics Data System (ADS)

Crain, Jennifer A.; Miller, Charles B.

Campbell et al . (Deep Sea Research II, 48 (2001) 531) have shown that there was a localized starvation event affecting Calanus finmarchicus on the southern flank of Georges Bank in April 1997. Growth and molting rates of this dominant copepod were reduced. We have used the morphology of tooth development in field-collected samples to show that this starvation affected animals living continuously in the field, as well as those in Campbell et al .'s experimental tanks. Assuming a point of reserve saturation (PRS) response of Calanus to food limitation, and correspondence between PRS and advance from the postmolt jaw facies, the proportion of individuals with postmolt jaws should increase in all copepodite stages under starvation. Individuals that have developed past PRS should molt to the next stage, acquiring postmolt facies. Thus, the fraction of postmolt jaws should increase, while the fraction of jaws in later phases should decrease. This was observed for a drifter-marked station over five days. Numerical simulations of jaw phase distributions expected under full nutrition, and both total and patchy starvation were generated from individual-based models of development. Proportions of copepodites in postmolt phase do not increase with full nutrition. A simulation of a total starvation event showed a marked increase in postmolts during food limitation, but the increase was more extreme than the field data. A modification of the starvation simulation, representing patchy feeding conditions, matched the level of increase of postmolt individuals in all stages that was observed in the field samples.

Isothermal elastohydrodynamic lubrication of point contacts. 4: Starvation results

NASA Technical Reports Server (NTRS)

Hamrock, B. J.; Dowson, D.

1976-01-01

The influence of lubricant starvation on minimum film thickness was investigated by moving the inlet boundary closer to the contact center. The following expression was derived for the dimensionless inlet distance at the boundary between the fully flooded and starved conditions: m* = 1 + 3.06 ((R/b)(R/b)H) to the power 0.58, where R is the effective radius of curvature, b is the semiminor axis of the contact ellipse, and H is the central film thickness for fully flooded conditions. A corresponding expression was also given based on the minimum film thickness for fully flooded conditions. Therefore, for m m*, starvation occurs and, for m m*, a fully flooded condition exists. Two other expressions were also derived for the central and minimum film thicknesses for a starved condition. Contour plots of the pressure and the film thickness in and around the contact are shown for the fully flooded and starved lubricating conditions, from which the film thickness was observed to decrease substantially as starvation increases.

Antihyperlipidemic activity of adenosine triphosphate in rabbits fed a high-fat diet and hyperlipidemic patients.

PubMed

Zhang, Lianshan; Liang, Libin; Tong, Tong; Qin, Yuguo; Xu, Yanping; Tong, Xinglong

2016-10-01

Context Recently, adenosine triphosphate (ATP) was occasionally found to decrease the triglyceride (TG) levels in several hyperlipidemic patients in our clinical practice. Objective The study investigates the anti-hyperlipidemic effects of ATP in a high-fat fed rabbit model and hyperlipidemic patients. Materials and methods Twenty-four rabbits were randomly divided into three groups of eight animals each as follows: normal diet, high-fat diet and high-fat diet + ATP group. ATP supplementation (40 mg/day) was started at the 20th day and lasted for 10 days. Serum concentrations of total cholesterol (TC), TG, LDL-C, HDL-C were measured on the 20th day and 30th day. Heart, liver and aorta were subjected histopathological examination. Twenty outpatients diagnosed primary hyperlipidemia took ATP at a dose of 60 mg twice a day for 1 week. Results Feeding rabbits with a high-fat diet resulted in a significant elevation of lipid parameters including TC, TG, LDL-C, VLDL-C compared to the normal diet group (p < 0.01). ATP treatment significantly decreased serum TG level (p < 0.01), whilst other parameters remained statistically unaltered. Meanwhile, ATP significantly reduced the thickness of fat layer in cardiac epicardium (p < 0.05) and pathological gradation of ballooning degeneration in hepatocytes (p < 0.05). After taking ATP for 1 week, hyperlipidemia patients exhibited a significant decrease of TG (p < 0.01), but other lipid parameters had no significant change. Discussion and conclusion The study indicates that ATP selectively decreases serum TG levels in high-fat diet rabbits and hyperlipidemic patients. Therefore, ATP supplementation may provide an effective approach to control TG level.

A Network of Hydrophobic Residues Impeding Helix αC Rotation Maintains Latency of Kinase Gcn2, Which Phosphorylates the α Subunit of Translation Initiation Factor 2▿

PubMed Central

Gárriz, Andrés; Qiu, Hongfang; Dey, Madhusudan; Seo, Eun-Joo; Dever, Thomas E.; Hinnebusch, Alan G.

2009-01-01

Kinase Gcn2 is activated by amino acid starvation and downregulates translation initiation by phosphorylating the α subunit of translation initiation factor 2 (eIF2α). The Gcn2 kinase domain (KD) is inert and must be activated by tRNA binding to the adjacent regulatory domain. Previous work indicated that Saccharomyces cerevisiae Gcn2 latency results from inflexibility of the hinge connecting the N and C lobes and a partially obstructed ATP-binding site in the KD. Here, we provide strong evidence that a network of hydrophobic interactions centered on Leu-856 also promotes latency by constraining helix αC rotation in the KD in a manner relieved during amino acid starvation by tRNA binding and autophosphorylation of Thr-882 in the activation loop. Thus, we show that mutationally disrupting the hydrophobic network in various ways constitutively activates eIF2α phosphorylation in vivo and bypasses the requirement for a key tRNA binding motif (m2) and Thr-882 in Gcn2. In particular, replacing Leu-856 with any nonhydrophobic residue activates Gcn2, while substitutions with various hydrophobic residues maintain kinase latency. We further provide strong evidence that parallel, back-to-back dimerization of the KD is a step on the Gcn2 activation pathway promoted by tRNA binding and autophosphorylation. Remarkably, mutations that disrupt the L856 hydrophobic network or enhance hinge flexibility eliminate the need for the conserved salt bridge at the parallel dimer interface, implying that KD dimerization facilitates the reorientation of αC and remodeling of the active site for enhanced ATP binding and catalysis. We propose that hinge remodeling, parallel dimerization, and reorientation of αC are mutually reinforcing conformational transitions stimulated by tRNA binding and secured by the ensuing autophosphorylation of T882 for stable kinase activation. PMID:19114556

A network of hydrophobic residues impeding helix alphaC rotation maintains latency of kinase Gcn2, which phosphorylates the alpha subunit of translation initiation factor 2.

PubMed

Gárriz, Andrés; Qiu, Hongfang; Dey, Madhusudan; Seo, Eun-Joo; Dever, Thomas E; Hinnebusch, Alan G

2009-03-01

Kinase Gcn2 is activated by amino acid starvation and downregulates translation initiation by phosphorylating the alpha subunit of translation initiation factor 2 (eIF2alpha). The Gcn2 kinase domain (KD) is inert and must be activated by tRNA binding to the adjacent regulatory domain. Previous work indicated that Saccharomyces cerevisiae Gcn2 latency results from inflexibility of the hinge connecting the N and C lobes and a partially obstructed ATP-binding site in the KD. Here, we provide strong evidence that a network of hydrophobic interactions centered on Leu-856 also promotes latency by constraining helix alphaC rotation in the KD in a manner relieved during amino acid starvation by tRNA binding and autophosphorylation of Thr-882 in the activation loop. Thus, we show that mutationally disrupting the hydrophobic network in various ways constitutively activates eIF2alpha phosphorylation in vivo and bypasses the requirement for a key tRNA binding motif (m2) and Thr-882 in Gcn2. In particular, replacing Leu-856 with any nonhydrophobic residue activates Gcn2, while substitutions with various hydrophobic residues maintain kinase latency. We further provide strong evidence that parallel, back-to-back dimerization of the KD is a step on the Gcn2 activation pathway promoted by tRNA binding and autophosphorylation. Remarkably, mutations that disrupt the L856 hydrophobic network or enhance hinge flexibility eliminate the need for the conserved salt bridge at the parallel dimer interface, implying that KD dimerization facilitates the reorientation of alphaC and remodeling of the active site for enhanced ATP binding and catalysis. We propose that hinge remodeling, parallel dimerization, and reorientation of alphaC are mutually reinforcing conformational transitions stimulated by tRNA binding and secured by the ensuing autophosphorylation of T882 for stable kinase activation.

Increase in ubiquitin-protein conjugates concomitant with the increase in proteolysis in rat skeletal muscle during starvation and atrophy denervation

NASA Technical Reports Server (NTRS)

Wing, S. S.; Haas, A. L.; Goldberg, A. L.

1995-01-01

The rapid loss of skeletal-muscle protein during starvation and after denervation occurs primarily through increased rates of protein breakdown and activation of a non-lysosomal ATP-dependent proteolytic process. To investigate whether protein flux through the ubiquitin (Ub)-proteasome pathway is enhanced, as was suggested by related studies, we measured, using specific polyclonal antibodies, the levels of Ub-conjugated proteins in normal and atrophying muscles. The content of these critical intermediates had increased 50-250% after food deprivation in the extensor digitorum longus and soleus muscles 2 days after denervation. Like rates of proteolysis, the amount of Ub-protein conjugates and the fraction of Ub conjugated to proteins increased progressively during food deprivation and returned to normal within 1 day of refeeding. During starvation, muscles of adrenalectomized rats failed to increase protein breakdown, and they showed 50% lower levels of Ub-protein conjugates than those of starved control animals. The changes in the pools of Ub-conjugated proteins (the substrates for the 26S proteasome) thus coincided with and can account for the alterations in overall proteolysis. In this pathway, large multiubiquitinated proteins are preferentially degraded, and the Ub-protein conjugates that accumulated in atrophying muscles were of high molecular mass (> 100 kDa). When innervated and denervated gastrocnemius muscles were fractionated, a significant increase in ubiquitinated proteins was found in the myofibrillar fraction, the proteins of which are preferentially degraded on denervation, but not in the soluble fraction. Thus activation of this proteolytic pathway in atrophying muscles probably occurs initially by increasing Ub conjugation to cell proteins. The resulting accumulation of Ub-protein conjugates suggests that their degradation by the 26S proteasome complex subsequently becomes rate-limiting in these catabolic states.

General anesthetics cause mitochondrial dysfunction and reduction of intracellular ATP levels

PubMed Central

Kishikawa, Jun-ichi; Inoue, Yuki; Fujikawa, Makoto; Nishimura, Kenji; Nakanishi, Atsuko; Tanabe, Tsutomu; Imamura, Hiromi

2018-01-01

General anesthetics are indispensable for effective clinical care. Although, the mechanism of action of general anesthetics remains controversial, lipid bilayers and proteins have been discussed as their targets. In this study, we focused on the relationship between cellular ATP levels and general anesthetics. The ATP levels of nematodes and cultured mammalian cells were decreased by exposure to three general anesthetics: isoflurane, pentobarbital, and 1-phenoxy-2-propanol. Furthermore, these general anesthetics abolished mitochondrial membrane potential, resulting in the inhibition of mitochondrial ATP synthesis. These results suggest that the observed decrease of cellular ATP level is a common phenomenon of general anesthetics. PMID:29298324

Respiratory ATP cost and benefit of arbuscular mycorrhizal symbiosis with Nicotiana tabacum at different growth stages and under salinity.

PubMed

Del-Saz, Néstor Fernández; Romero-Munar, Antonia; Alonso, David; Aroca, Ricardo; Baraza, Elena; Flexas, Jaume; Ribas-Carbo, Miquel

2017-11-01

Growth and maintenance partly depend on both respiration and ATP production during oxidative phosphorylation in leaves. Under stress, ATP is needed to maintain the accumulated biomass. ATP production mostly proceeds from the cytochrome oxidase pathway (COP), while respiration via the alternative oxidase pathway (AOP) may decrease the production of ATP per oxygen consumed, especially under phosphorus (P) limitation and salinity conditions. Symbiosis with arbuscular mycorrhizal (AM) fungi is reputed by their positive effect on plant growth under stress at mature stages of colonization; however, fungal colonization may decrease plant growth at early stages. Thus, the present research is based on the hypothesis that AM fungus colonization will increase both foliar respiration and ATP production at mature stages of plant growth while decreasing them both at early stages. We used the oxygen-isotope-fractionation technique to study the in vivo respiratory activities and ATP production of the COP and AOP in AM and non-AM (NM) tobacco plants grown under P-limiting and saline conditions in sand at different growth stages (14, 28 and 49days). Our results suggest that AM symbiosis represents an ATP cost detrimental for shoot growth at early stages, whilst it represents a benefit on ATP allowing for faster rates of growth at mature stages, even under salinity conditions. Copyright © 2017 Elsevier GmbH. All rights reserved.

Monitoring protein turnover during phosphate starvation-dependent autophagic degradation using a photoconvertible fluorescent protein aggregate in tobacco BY-2 cells

PubMed Central

Tasaki, Maiko; Asatsuma, Satoru; Matsuoka, Ken

2014-01-01

We have developed a system for quantitative monitoring of autophagic degradation in transformed tobacco BY-2 cells using an aggregate-prone protein comprised of cytochrome b5 (Cyt b5) and a tetrameric red fluorescent protein (RFP). Unfortunately, this system is of limited use for monitoring the kinetics of autophagic degradation because the proteins synthesized before and after induction of autophagy cannot be distinguished. To overcome this problem, we developed a system using kikume green-red (KikGR), a photoconvertible and tetrameric fluorescent protein that changes its fluorescence from green to red upon irradiation with purple light. Using the fusion protein of Cyt b5 and KikGR together with a method for the bulk conversion of KikGR, which we had previously used to convert the Golgi-localized monomeric KikGR fusion protein, we were able to monitor both the growth and de novo formation of aggregates. Using this system, we found that tobacco cells do not cease protein synthesis under conditions of phosphate (Pi)-starvation. Induction of autophagy under Pi-starvation, but not under sugar- or nitrogen-starvation, was specifically inhibited by phosphite, which is an analog of Pi with a different oxidation number. Therefore, the mechanism by which BY-2 cells can sense Pi-starvation and induce autophagy does not involve sensing a general decrease in energy supply and a specific Pi sensor might be involved in the induction of autophagy under Pi-starvation. PMID:24817874

Monitoring protein turnover during phosphate starvation-dependent autophagic degradation using a photoconvertible fluorescent protein aggregate in tobacco BY-2 cells.

PubMed

Tasaki, Maiko; Asatsuma, Satoru; Matsuoka, Ken

2014-01-01

We have developed a system for quantitative monitoring of autophagic degradation in transformed tobacco BY-2 cells using an aggregate-prone protein comprised of cytochrome b5 (Cyt b5) and a tetrameric red fluorescent protein (RFP). Unfortunately, this system is of limited use for monitoring the kinetics of autophagic degradation because the proteins synthesized before and after induction of autophagy cannot be distinguished. To overcome this problem, we developed a system using kikume green-red (KikGR), a photoconvertible and tetrameric fluorescent protein that changes its fluorescence from green to red upon irradiation with purple light. Using the fusion protein of Cyt b5 and KikGR together with a method for the bulk conversion of KikGR, which we had previously used to convert the Golgi-localized monomeric KikGR fusion protein, we were able to monitor both the growth and de novo formation of aggregates. Using this system, we found that tobacco cells do not cease protein synthesis under conditions of phosphate (Pi)-starvation. Induction of autophagy under Pi-starvation, but not under sugar- or nitrogen-starvation, was specifically inhibited by phosphite, which is an analog of Pi with a different oxidation number. Therefore, the mechanism by which BY-2 cells can sense Pi-starvation and induce autophagy does not involve sensing a general decrease in energy supply and a specific Pi sensor might be involved in the induction of autophagy under Pi-starvation.

Behavior and stability of adenosine triphosphate (ATP) during chlorine disinfection.

PubMed

Nescerecka, Alina; Juhna, Talis; Hammes, Frederik

2016-09-15

Adenosine triphosphate (ATP) analysis is a cultivation-independent alternative method for the determination of bacterial viability in both chlorinated and non-chlorinated water. Here we investigated the behavior and stability of ATP during chlorination in detail. Different sodium hypochlorite doses (0-22.4 mg-Cl2 L(-1); 5 min exposure) were applied to an Escherichia coli pure culture suspended in filtered river water. We observed decreasing intracellular ATP with increasing chlorine concentrations, but extracellular ATP concentrations only increased when the chlorine dose exceeded 0.35 mg L(-1). The release of ATP from chlorine-damaged bacteria coincided with severe membrane damage detected with flow cytometry (FCM). The stability of extracellular ATP was subsequently studied in different water matrixes, and we found that extracellular ATP was stable in sterile deionized water and also in chlorinated water until extremely high chlorine doses (≤11.2 mg-Cl2 L(-1); 5 min exposure). In contrast, ATP decreased relatively slowly (k = 0.145 h(-1)) in 0.1 μm filtered river water, presumably due to degradation by either extracellular enzymes or the fraction of bacteria that were able to pass through the filter. Extracellular ATP decreased considerably faster (k = 0.368 h(-1)) during batch growth of a river water bacterial community. A series of growth potential tests showed that extracellular ATP molecules were utilized as a phosphorus source during bacteria proliferation. From the combined data we conclude that ATP released from bacteria at high chlorine doses could promote bacteria regrowth, contributing to biological instability in drinking water distribution systems. Copyright © 2016 Elsevier Ltd. All rights reserved.

Brain-midgut short neuropeptide F mechanism that inhibits digestive activity of the American cockroach, Periplaneta americana upon starvation.

PubMed

Mikani, Azam; Wang, Qiu-Shi; Takeda, Makio

2012-03-01

Immunohistochemical reactivity against short neuropeptide F (sNPF) was observed in the brain-corpus cardiacum and midgut paraneurons of the American cockroach, Periplaneta americana. Four weeks of starvation increased the number of sNPF-ir cells in the midgut epithelium but the refeeding decreased the number in 3h. Dramatic rises in sNPF contents in the midgut epithelium and hemolymph of roaches starved for 4 weeks were confirmed by ELISA. Starvation for 4 weeks reduced α-amylase, protease and lipase activities in the midgut of P. americana but refeeding restored these to high levels. Co-incubation of dissected midgut with sNPF at physiological concentrations inhibited α-amylase, protease and lipase activities. sNPF injection into the hemocoel led to a decrease in α-amylase, protease and lipase activities, whereas PBS injection had no effects. The injection of d-(+)-trehalose and l-proline into the hemocoel of decapitated adult male cockroaches that had been starved for 4 weeks had no effect on these digestive enzymes. However, injection into the hemocoel of head-intact starved cockroaches stimulated digestive activity. Injection of d-(+)-trehalose and l-proline into the lumen of decapitated cockroaches that had been starved for 4 weeks increased enzymes activities and suppressed sNPF in the midgut. Our data indicate that sNPF from the midgut paraneurons suppresses α-amylase, protease and lipase activities during starvation. Injection of d-(+)-trehalose/l-proline into the hemocoel of head-intact starved cockroach decreased the hemolymph sNPF content, which suggests that sNPF could be one of the brain factors, demonstrating brain-midgut interplay in the regulation of digestive activities and possibly nutrition-associated behavioral modifications. Copyright © 2011 Elsevier Inc. All rights reserved.

Growth, metabolism and physiological response of the sea cucumber, Apostichopus japonicus Selenka during periods of inactivity

NASA Astrophysics Data System (ADS)

Du, Rongbin; Zang, Yuanqi; Tian, Xiangli; Dong, Shuanglin

2013-03-01

The growth, metabolism and physiological response of the sea cucumber, Apostichopus japonicus, were investigated during periods of inactivity. The body weight, oxygen consumption rate (OCR), activities of acidic phosphatase (ACP), alkaline phosphatase (AKP), catalase (CAT) and superoxide dismutase (SOD), and content of heat shock protein 70 (Hsp70) in the body wall and coelomic fluid of A. japonicus were measured during starvation, experimental aestivation and aestivation. The results showed that the body weight of sea cucumber in the three treatments decreased significantly during the experimental period ( P < 0.05). The OCR of sea cucumber reduced in starvation and experimental aestivation treatments, but increased gradually in natural aestivation treatment. The activities of ACP and AKP of sea cucumber decreased gradually in all treatments, whereas those of SOD and CAT as well as Hsp70 content decreased in the starvation and experimental aestivation treatments and increased in natural aestivation treatment. The sea cucumber entered a state of aestivation at 24°C. To some extent, the animals in experimental aestivation were different from those in natural aestivation in metabolism and physiological response. These findings suggested that the aestivation mechanism of A. japonicus is complex and may not be attributed to the elevated temperature only.

The Greening after Extended Darkness1 Is an N-End Rule Pathway Mutant with High Tolerance to Submergence and Starvation1[OPEN

PubMed Central

Riber, Willi; Müller, Jana T.; Visser, Eric J.W.; Sasidharan, Rashmi; Voesenek, Laurentius A.C.J.; Mustroph, Angelika

2015-01-01

Plants respond to reductions in internal oxygen concentrations with adaptive mechanisms (for example, modifications of metabolism to cope with reduced supply of ATP). These responses are, at the transcriptional level, mediated by the group VII Ethylene Response Factor transcription factors, which have stability that is regulated by the N-end rule pathway of protein degradation. N-end rule pathway mutants are characterized by a constitutive expression of hypoxia response genes and abscisic acid hypersensitivity. Here, we identify a novel proteolysis6 (prt6) mutant allele, named greening after extended darkness1 (ged1), which was previously discovered in a screen for genomes uncoupled-like mutants and shows the ability to withstand long periods of darkness at the seedling stage. Interestingly, this ethyl methanesulfonate-derived mutant shows unusual chromosomal rearrangement instead of a point mutation. Furthermore, the sensitivity of N-end rule pathway mutants ged1 and prt6-1 to submergence was studied in more detail to understand previously contradicting experiments on this topic. Finally, it was shown that mutants for the N-end rule pathway are generally more tolerant to starvation conditions, such as prolonged darkness or submergence, which was partially associated with carbohydrate conservation. PMID:25667318

Effect of starvation and exercise on actual and total activity of the branched-chain 2-oxo acid dehydrogenase complex in rat tissues.

PubMed Central

Wagenmakers, A J; Schepens, J T; Veerkamp, J H

1984-01-01

Starvation does not change the actual activity per g of tissue of the branched-chain 2-oxo acid dehydrogenase in skeletal muscles, but affects the total activity to a different extent, depending on the muscle type. The activity state (proportion of the enzyme present in the active state) does not change in diaphragm and decreases in quadriceps muscle. Liver and kidney show an increase of both activities, without a change of the activity state. In heart and brain no changes were observed. Related to organ wet weights, the actual activity present in the whole-body muscle mass decreases on starvation, whereas the activities present in liver and kidney do not change, or increase slightly. Exercise (treadmill-running) of untrained rats for 15 and 60 min causes a small increase of the actual activity and the activity state of the branched-chain 2-oxo acid dehydrogenase complex in heart and skeletal muscle. Exercise for 1 h, furthermore, increased the actual and the total activity in liver and kidney, without a change of the activity state. In brain no changes were observed. The actual activity per g of tissue in skeletal muscle was less than 2% of that in liver and kidney, both before and after exercise and starvation. Our data indicate that the degradation of branched-chain 2-oxo acids predominantly occurs in liver and to a smaller extent in kidney and skeletal muscle in fed, starved and exercised rats. PMID:6508743

A novel approach to maintain gut mucosal integrity using an oral enzyme supplement.

PubMed

Hamarneh, Sulaiman R; Mohamed, Mussa M Rafat; Economopoulos, Konstantinos P; Morrison, Sara A; Phupitakphol, Tanit; Tantillo, Tyler J; Gul, Sarah S; Gharedaghi, Mohammad Hadi; Tao, Qingsong; Kaliannan, Kanakaraju; Narisawa, Sonoko; Millán, José L; van der Wilden, Gwendolyn M; Fagenholz, Peter J; Malo, Madhu S; Hodin, Richard A

2014-10-01

To determine the role of intestinal alkaline phosphatase (IAP) in enteral starvation-induced gut barrier dysfunction and to study its therapeutic effect as a supplement to prevent gut-derived sepsis. Critically ill patients are at increased risk for systemic sepsis and, in some cases, multiorgan failure leading to death. Years ago, the gut was identified as a major source for this systemic sepsis syndrome. Previously, we have shown that IAP detoxifies bacterial toxins, prevents endotoxemia, and preserves intestinal microbiotal homeostasis. WT and IAP-KO mice were used to examine gut barrier function and tight junction protein levels during 48-hour starvation and fed states. Human ileal fluid samples were collected from 20 patients postileostomy and IAP levels were compared between fasted and fed states. To study the effect of IAP supplementation on starvation-induced gut barrier dysfunction, WT mice were fasted for 48 hours +/- IAP supplementation in the drinking water. The loss of IAP expression is associated with decreased expression of intestinal junctional proteins and impaired barrier function. For the first time, we demonstrate that IAP expression is also decreased in humans who are deprived of enteral feeding. Finally, our data demonstrate that IAP supplementation reverses the gut barrier dysfunction and tight junction protein losses due to a lack of enteral feeding. IAP is a major regulator of gut mucosal permeability and is able to ameliorate starvation-induced gut barrier dysfunction. Enteral IAP supplementation may represent a novel approach to maintain bowel integrity in critically ill patients.

Effects of single amino acid deficiency on mRNA translation are markedly different for methionine versus leucine.

PubMed

Mazor, Kevin M; Dong, Leiming; Mao, Yuanhui; Swanda, Robert V; Qian, Shu-Bing; Stipanuk, Martha H

2018-05-24

Although amino acids are known regulators of translation, the unique contributions of specific amino acids are not well understood. We compared effects of culturing HEK293T cells in medium lacking either leucine, methionine, histidine, or arginine on eIF2 and 4EBP1 phosphorylation and measures of mRNA translation. Methionine starvation caused the most drastic decrease in translation as assessed by polysome formation, ribosome profiling, and a measure of protein synthesis (puromycin-labeled polypeptides) but had no significant effect on eIF2 phosphorylation, 4EBP1 hyperphosphorylation or 4EBP1 binding to eIF4E. Leucine starvation suppressed polysome formation and was the only tested condition that caused a significant decrease in 4EBP1 phosphorylation or increase in 4EBP1 binding to eIF4E, but effects of leucine starvation were not replicated by overexpressing nonphosphorylatable 4EBP1. This suggests the binding of 4EBP1 to eIF4E may not by itself explain the suppression of mRNA translation under conditions of leucine starvation. Ribosome profiling suggested that leucine deprivation may primarily inhibit ribosome loading, whereas methionine deprivation may primarily impair start site recognition. These data underscore our lack of a full understanding of how mRNA translation is regulated and point to a unique regulatory role of methionine status on translation initiation that is not dependent upon eIF2 phosphorylation.

Effect of Different Starvation Levels on Cognitive Ability in Mice

NASA Astrophysics Data System (ADS)

Li, Xiaobing; Zhi, Guoguo; Yu, Yi; Cai, Lingyu; Li, Peng; Zhang, Danhua; Bao, Shuting; Hu, Wenlong; Shen, Haiyan; Song, Fujuan

2018-01-01

Objective: To study the effect of different starvation levels on cognitive ability in mice. Method: Mice were randomly divided into four groups: normal group, dieting group A, dieting group B, dieting group C. The mice of normal group were given normal feeding amount, the rest of groups were given 3/4 of normal feeding amount, 2/4 of normal feeding amount and 1/4 of normal feeding amount. After feeding mice four days, the weight was observed and T-maze experiment, Morris water maze test, open field test and Serum Catalase activity were detected. Result: Compared with the normal group, the correct rate of the intervention group in the T-maze experiment was decreased and dieting group A> dieting group B> dieting group C. In the Morris water maze test, Compared with the normal group, the correct rate of the intervention group was increased. Among these three intervention groups, dieting group A had the highest correct rate and the difference of dieting group B and dieting group C were similar. In the open field test, Compared with the normal group, the exploration rate of the surrounding environment in the intervention group was increased. In the Serum Catalase test, Compared with the normal group, the activities of serum peroxidase in the intervention groups were decreased and dieting group A> dieting group B> dieting group C. Conclusion: A certain level of starvation could affect the cognitive ability of mice. In a certain range, the level of starvation is inversely proportional to cognitive ability in mice.

Effects of Long-Term Starvation on a Host Bivalve (Codakia orbicularis, Lucinidae) and Its Symbiont Population▿

PubMed Central

Caro, Audrey; Got, Patrice; Bouvy, Marc; Troussellier, Marc; Gros, Olivier

2009-01-01

The bivalve Codakia orbicularis, hosting sulfur-oxidizing gill endosymbionts, was starved (in artificial seawater filtered through a 0.22-μm-pore-size membrane) for a long-term experiment (4 months). The effects of starvation were observed using transmission electron microscopy, fluorescence in situ hybridization and catalyzed reporter deposition (CARD-FISH), and flow cytometry to monitor the anatomical and physiological modifications in the gill organization of the host and in the symbiotic population housed in bacteriocytes. The abundance of the symbiotic population decreased through starvation, with a loss of one-third of the bacterial population each month, as shown by CARD-FISH. At the same time, flow cytometry revealed significant changes in the physiology of symbiotic cells, with a decrease in cell size and modifications to the nucleic acid content, while most of the symbionts maintained a high respiratory activity (measured using the 5-cyano-2,3-ditolyl tetrazolium chloride method). Progressively, the number of symbiont subpopulations was reduced, and the subsequent multigenomic state, characteristic of this symbiont in freshly collected clams, turned into one and five equivalent genome copies for the two remaining subpopulations after 3 months. Concomitant structural modifications appeared in the gill organization. Lysosymes became visible in the bacteriocytes, while large symbionts disappeared, and bacteriocytes were gradually replaced by granule cells throughout the entire lateral zone. Those data suggested that host survival under these starvation conditions was linked to symbiont digestion as the main nutritional source. PMID:19346359

Potential use of carbon-11 labeled alpha-aminoisobutyric acid (AIB) as an in vivo tracer of amino acid uptake in differing metabolic states

DOE Office of Scientific and Technical Information (OSTI.GOV)

Conti, P.S.; Starnes, H.F.; Brennan, M.F.

1986-05-01

AIB has been used as a model amino acid for the evaluation of alanine-preferring amino acid transport. Hormonal factors and starvation alter the tissue distribution of amino acids, particularly in liver and muscle. With positron emission tomography and labeling of biochemical tracers with C-11, (t1/2=20.4 min), it is now possible to study amino acid kinetics in vivo using external imaging. In order to investigate the utility of C-11 AIB as an in vivo tracer of altered tissue metabolism, C-14 AIB was studied in groups of rats with either streptozotocin-induced diabetes, insulin-induced hypoglycemia or starvation. The data suggest an increased aminomore » acid uptake in liver in starvation, an increased uptake in muscle in response to insulin and associated hypoglycemia and decreased transport in muscle in starvation, as seen by other investigators. These results suggest that C-11 AIB may be useful as an in vivo monitor of metabolic changes in body tissues.« less

ATPase activity and light scattering of acto-heavy meromyosin: dependence on ATP concentration and on ionic strength.

PubMed

Dancker, P

1975-01-01

1. The dependence on ATP concentration of ATPase activity and light scattering decrease of acto-HMM could be described at very low ionic strength by one hyperbolic adsorption isotherm with a dissociation constant of 3 X 10(-6)M. Hence the increase of ATP ase activity was paralleled by a decrease in light scattering. At higher values of ionic strength ATPase activity stopped rising before HMM was completely saturated with ATP. Higher ionic strength prevented ATPase activity from further increasing when the rigor links (links between actin and nucleotide-free myosin), which have formerly protected the ATPase against the suppressing action of higher ionic strength have fallen below a certain amount. This protecting influence of rigor links did not require tropomyosin-troponin. 2. For complete activation of ATPase activity by actin less actin was needed when HMM was incompletely saturated with ATP than when it was completely saturated with ATP. 3. The apparent affinity of ATP to regulated acto-HMM (which contained tropomyosin-troponin) was lower than to unregulated acto-HMM (which was devoid of tropomyosin-troponin). In the presence of rigor complexes (indicated by an incomplete decrease of light scattering) the ATPase activity of regulated acto-HMM was higher than that of unregulated acto-HMM. At increasing ATP concentrations the ATPase activity of regulated acto-HMM stopped rising at a similar degree of saturation with ATP as the ATPase activity of unregulated acto-HMM at the same ionic strength.

Regulation of CDP-diacylglycerol synthesis and utilization by inositol and choline in Schizosaccharomyces pombe.

PubMed Central

Gaynor, P M; Greenberg, M L

1992-01-01

CDP-diacylglycerol (CDP-DG) is an important branchpoint intermediate in eucaryotic phospholipid biosynthesis and could be a key regulatory site in phospholipid metabolism. Therefore, we examined the effects of growth phase, phospholipid precursors, and the disruption of phosphatidylcholine (PC) synthesis on the membrane-associated phospholipid biosynthetic enzymes CDP-DG synthase, phosphatidylglycerolphosphate (PGP) synthase, phosphatidylinositol (PI) synthase, and phosphatidylserine (PS) synthase in cell extracts of the fission yeast Schizosaccharomyces pombe. In complete synthetic medium containing inositol, maximal expression of CDP-DG synthase, PGP synthase, PI synthase, and PS synthase in wild-type cells occurred in the exponential phase of growth and decreased two- to fourfold in the stationary phase of growth. In cells starved for inositol, this decrease in PGP synthase, PI synthase, and PS synthase expression was not observed. Starvation for inositol resulted in a twofold derepression of PGP synthase and PS synthase expression, while PI synthase expression decreased initially and then remained constant. Upon the addition of inositol to inositol-starved cells, there was a rapid and continued increase in PI synthase expression. We examined expression of these enzymes in cho2 and cho1 mutants, which are blocked in the methylation pathway for synthesis of PC. Choline starvation resulted in a decrease in PS synthase and CDP-DG synthase expression in cho1 but not cho2 cells. Expression of PGP synthase and PI synthase was not affected by choline starvation. Inositol starvation resulted in a 1.7-fold derepression of PGP synthase expression in cho2 but not cho1 cells when PC was synthesized. PS synthase expression was not depressed, while CDP-DG synthase and PI synthase expression decreased in cho2 and cho1 cells in the absence of inositol. These results demonstrate that (i) CDP-DG synthase, PGP synthase, PI synthase, and PS synthase are similarly regulated by growth phase; (ii) inositol affects the expression of PGP synthase, PI synthase, and PS synthase; (iii) disruption of the methylation pathway results in aberrant patterns of regulation of growth phase and phospholipid precursors. Important differences between S. pombe and Saccharomyces cerevisiae with regard to regulation of these enzymes are discussed. PMID:1324908

Combined effects of starvation and butyrate on autophagy-dependent gingival epithelial cell death.

PubMed

Evans, M; Murofushi, T; Tsuda, H; Mikami, Y; Zhao, N; Ochiai, K; Kurita-Ochiai, T; Yamamoto, M; Otsuka, K; Suzuki, N

2017-06-01

Bacteria in the dental biofilm surrounding marginal gingival grooves cause periodontal diseases. Numerous bacteria within the biofilm consume nutrients from the gingival crevicular fluid. Furthermore, some gram-negative bacteria in mature dental biofilms produce butyrate. Thus, gingival epithelial cells in close proximity to mature dental biofilms are at risk of both starvation and exposure to butyrate. In the present study, we determined the combined effects of starvation and butyrate exposure on gingival epithelial cell death and the underlying mechanisms. The Ca9-22 cell line was used as an in vitro counterpart of gingival epithelial cells. Cell death was measured as the amount of total DNA in the dead cells using SYTOX Green dye, which penetrates through membranes of dead cells and emits fluorescence when it intercalates into double-stranded DNA. AMP-activated protein kinase (AMPK) activity, the amount of autophagy, and acetylation of histone H3 were determined using western blot. Gene expression levels of microtubule-associated protein 1 light chain 3b (lc3b) were determined using quantitative reverse transcription-polymerase chain reaction. Butyrate-induced cell death occurred in a dose-dependent manner whether cells were starved or fed. However, the induction of cell death was two to four times higher when cells were placed under starvation conditions compared to when they were fed. Moreover, both starvation and butyrate exposure induced AMPK activity and autophagy. While AMPK inactivation resulted in decreased autophagy and butyrate-induced cell death under conditions of starvation, AMPK activation resulted in butyrate-induced cell death when cells were fed. Combined with the results of our previous report, which demonstrated butyrate-induced autophagy-dependent cell death, the results of this study suggest that the combination of starvation and butyrate exposure activates AMPK inducing autophagy and subsequent cell death. Notably, this combination markedly induced LC3B production and the induction was attenuated by AMPK inhibition. LC3B knockdown, in turn, significantly decreased butyrate-induced cell death. Therefore, AMPK-dependent LC3B induction apparently plays an important role in butyrate-induced cell death. There was a lack of correspondence between the levels of AMPK activation and LC3B induction; this may reflect the histone deacetylase-inhibitory capacity of butyrate on histone proteins. Taken together, starvation and butyrate exposure promote autophagy via AMPK signaling, while the histone deacetylase-inhibitory effects of butyrate alter chromatin to transcriptionally active state, resulting in strong LC3B induction and subsequent cell death. These findings may help improve the understanding of the cellular processes underlying periodontal disease initiation. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The effects of gamma radiation, UV and visible light on ATP levels in yeast cells depend on cellular melanization.

PubMed

Bryan, Ruth; Jiang, Zewei; Friedman, Matthew; Dadachova, Ekaterina

2011-10-01

Previously we have shown that growth of melanized fungi is stimulated by low levels of gamma radiation. The goal of this study was to examine the effects of visible light, UV light, and gamma radiation on the energy level (ATP concentration) in melanized Cryptococcus neoformans cells. Melanized C. neoformans cells as well as non-melanized controls were subjected to visible, UV or gamma radiation, and ATP was quantified by measuring the amount of light emitted by the ATP-dependent reaction of luciferase with luciferin. We found that all three forms of radiation led to a reduction in the ATP levels in melanized C. neoformans cells. This points to a universal melanin-related mechanism underlying observation of ATP decrease in irradiated melanized cells. In contrast, in non-melanized cells visible light led to increase in ATP levels; gamma radiation did not cause any changes while UV exposure resulted in some ATP decrease, however, much less pronounced than in melanized cells. Copyright © 2011 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

Heterologous desensitization of muscarinic receptors by P2Z purinoceptors in rat parotid acinar cells.

PubMed

Fukushi, Y

1999-01-01

We studied the heterologous desensitization of muscarinic receptors by ATP in fura-2-loaded rat parotid acinar cells. Exposure to ATP or 3'-o-(4-benzoyl) benzoyl-ATP shortened the duration and decreased the magnitude of acetylcholine-induced Ca2+ release from intracellular Ca2+ stores in a dose-dependent manner. The shortening was observed only in an early stage of desensitization (within 20 s), whereas the decrease in the magnitude of the response was dependent upon the time the cells were exposed to the nucleotides. Atropine induced a profound shortening during the progressive decrease in the magnitude of acetylcholine-induced Ca2+ release. 3'-o-(4-Benzoyl) benzoyl-ATP did not induce an increase in the cytosolic Ca2+ concentration when the cells were incubated in the Ca2+- and Na+-free medium, but it did induce a strong desensitization of muscarinic receptors. The specific protein kinase C inhibitor bisindoylmaleimide resensitized the 3'-o-(4-benzoyl) benzoyl-ATP-treated muscarinic receptors. Phorbol 12-myristate 13-acetate potentiated the desensitization of muscarinic receptors. Ceramides that prevent the activation of phospholipase D resensitized the 3'-o-(4-benzoyl) benzoyl-ATP-treated muscarinic receptors. These results suggest that ATP, acting through P2Z purinoceptor-mediated phospholipase D, may produce a Ca2+-independent protein kinase C. Heterologous desensitization of muscarinic receptors by protein kinase C may shorten the duration and decrease the magnitude of acetylcholine-induced Ca2+ release.

Intracellular and extracellular adenosine triphosphate in regulation of insulin secretion from pancreatic β cells (β).

PubMed

Wang, Chunjiong; Geng, Bin; Cui, Qinghua; Guan, Youfei; Yang, Jichun

2014-03-01

Adenosine triphosphate (ATP) synthesis and release in mitochondria play critical roles in regulating insulin secretion in pancreatic β cells. Mitochondrial dysfunction is mainly characterized by a decrease in ATP production, which is a central event in the progression of pancreatic β cell dysfunction and diabetes. ATP has been demonstrated to regulate insulin secretion via several pathways: (i) Intracellular ATP directly closes ATP-sensitive potassium channel to open L-type calcium channel, leading to an increase in free cytosolic calcium levels and exocytosis of insulin granules; (ii) A decrease in ATP production is always associated with an increase in production of reactive oxygen species, which exerts deleterious effects on pancreatic β cell survival and insulin secretion; and (iii) ATP can be co-secreted with insulin from pancreatic β cells, and the released ATP functions as an autocrine signal to modulate insulin secretory process via P2 receptors on the cell membrane. In this review, the recent findings regarding the role and mechanism of ATP synthesis and release in regulation of insulin secretion from pancreatic β cells will be summarized and discussed. © 2013 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Wiley Publishing Asia Pty Ltd.

[ATP content in cryopreserved sperm of Siberian white cranes Grus leucogeranus (Aves: Gruiformes)].

PubMed

Maksudov, G Iu; Erokhin, A S; Nesterenko, O N; Panchenko, V G

2002-01-01

ATP contents were studied in the native and cryoconserved sperm of Siberian white cranes Grus leucogeranus using bioluminescence analysis. The ATP content in freshly obtained spermatozoa was 12.7 nmol/10(8) cells. No ATP was found in the seminal plasma. In the process of freezing-thawing, the ATP concentration in the spermatozoa decreased by 30%. The differences in the dynamics of ATP content during cryoconservation of sperm of white cranes and other birds and mammals are discussed.

SIRT3 Deacetylates ATP Synthase F1 Complex Proteins in Response to Nutrient- and Exercise-Induced Stress

PubMed Central

Vassilopoulos, Athanassios; Pennington, J. Daniel; Andresson, Thorkell; Rees, David M.; Bosley, Allen D.; Fearnley, Ian M.; Ham, Amy; Flynn, Charles Robb; Hill, Salisha; Rose, Kristie Lindsey; Kim, Hyun-Seok; Walker, John E.

2014-01-01

Abstract Aims: Adenosine triphosphate (ATP) synthase uses chemiosmotic energy across the inner mitochondrial membrane to convert adenosine diphosphate and orthophosphate into ATP, whereas genetic deletion of Sirt3 decreases mitochondrial ATP levels. Here, we investigate the mechanistic connection between SIRT3 and energy homeostasis. Results: By using both in vitro and in vivo experiments, we demonstrate that ATP synthase F1 proteins alpha, beta, gamma, and Oligomycin sensitivity-conferring protein (OSCP) contain SIRT3-specific reversible acetyl-lysines that are evolutionarily conserved and bind to SIRT3. OSCP was further investigated and lysine 139 is a nutrient-sensitive SIRT3-dependent deacetylation target. Site directed mutants demonstrate that OSCPK139 directs, at least in part, mitochondrial ATP production and mice lacking Sirt3 exhibit decreased ATP muscle levels, increased ATP synthase protein acetylation, and an exercise-induced stress-deficient phenotype. Innovation: This work connects the aging and nutrient response, via SIRT3 direction of the mitochondrial acetylome, to the regulation of mitochondrial energy homeostasis under nutrient-stress conditions by deacetylating ATP synthase proteins. Conclusion: Our data suggest that acetylome signaling contributes to mitochondrial energy homeostasis by SIRT3-mediated deacetylation of ATP synthase proteins. Antioxid. Redox Signal. 21, 551–564. PMID:24252090

Ribose in the heart.

PubMed

Herrick, James; St Cyr, John

2008-01-01

Every cell needs energy, i.e., adenosine triphosphate (ATP), to carry out its function. Decreased oxygen levels, decreased blood flow, and other stressful conditions can drastically effect the intracellular concentrations of these energy compounds. Skeletal muscle, unlike the heart, can address this drop in ATP by employing the myokinase reaction, ultimately producing ATP with a subsequent elevation in adenosine monophosphate (AMP). Ribose, a naturally occurring 5-carbon monosaccharide, is a key component of RNA, DNA (which has deoxyribose), acetyl coenzyme A, and ATP. Each cell produces its own ribose, involved in the pentose phosphate pathway (PPP), to aid in ATP production. States of ischemia and/or hypoxia can severely lower levels of cellular energy compounds in the heart, with an associated compromise in cellular processes, ultimately reflected in altered function. Ribose appears to provide a solution to the problem in replenishing the depressed ATP levels and improving functional status of patients afflicted with cardiovascular diseases.

Study on ATP concentration changes in cytosol of individual cultured neurons during glutamate-induced deregulation of calcium homeostasis.

PubMed

Surin, A M; Gorbacheva, L R; Savinkova, I G; Sharipov, R R; Khodorov, B I; Pinelis, V G

2014-02-01

For the first time, simultaneous monitoring of changes in the concentration of cytosolic ATP ([ATP]c), pH (pHc), and intracellular free Ca2+ concentration ([Ca2+]i) of the individual neurons challenged with toxic glutamate (Glu) concentrations was performed. To this end, the ATP-sensor AT1.03, which binds to ATP and therefore enhances the efficiency of resonance energy transfer between blue fluorescent protein (energy donor) and yellow-green fluorescent protein (energy acceptor), was expressed in cultured hippocampal neurons isolated from 1-2-day-old rat pups. Excitation of fluorescence in the acceptor protein allowed monitoring changes in pHc. Cells were loaded with fluorescent low-affinity Ca2+ indicators Fura-FF or X-rhod-FF to register [Ca2+]i. It was shown that Glu (20 µM, glycine 10 µM, Mg2+-free) produced a rapid acidification of the cytosol and decrease in [ATP]c. An approximately linear relationship (r(2) = 0.56) between the rate of [ATP]c decline and latency of glutamate-induced delayed calcium deregulation (DCD) was observed: higher rate of [ATP]c decrease corresponded to shorter DCD latency period. DCD began with a decrease in [ATP]c of as much as 15.9%. In the phase of high [Ca2+]i, the plateau of [ATP]c dropped to 10.4% compared to [ATP]c in resting neurons (100%). In the presence of the Na+/K+-ATPase inhibitor ouabain (0.5 mM), glutamate-induced reduction in [ATP]c in the phase of the high [Ca2+]i plateau was only 36.6%. Changes in [ATP]c, [Ca2+]i, mitochondrial potential, and pHc in calcium-free or sodium-free buffers, as well as in the presence of the inhibitor of Na+/K+-ATPase ouabain (0.5 mM), led us to suggest that in addition to increase in proton conductivity and decline in [ATP]c, one of the triggering factors of DCD might be a reversion of the neuronal plasma membrane Na+/Ca2+ exchange.

Effect of K+ATP channel and adenosine receptor blockade during rest and exercise in congestive heart failure.

PubMed

Traverse, Jay H; Chen, YingJie; Hou, MingXiao; Li, Yunfang; Bache, Robert J

2007-06-08

K(+)(ATP) channels are important metabolic regulators of coronary blood flow (CBF) that are activated in the setting of reduced levels of ATP or perfusion pressure. In the normal heart, blockade of K(+)(ATP) channels results in a approximately 20% reduction in resting CBF but does not impair the increase in CBF that occurs during exercise. In contrast, adenosine receptor blockade fails to alter CBF or myocardial oxygen consumption (MVO(2)) in the normal heart but contributes to the increase in CBF during exercise when vascular K(+)(ATP) channels are blocked. Congestive heart failure (CHF) is associated with a decrease in CBF that is matched to a decrease in MVO(2) suggesting downregulation of myocardial energy utilization. Because myocardial ATP levels and coronary perfusion pressure are reduced in CHF, this study was undertaken to examine the role of K(+)(ATP) channels and adenosine in dogs with pacing-induced CHF. Myocardial blood flow (MBF) and MVO(2) were measured during rest and treadmill exercise before and after K(+)(ATP) channel blockade with glibenclamide (50 microg/kg/min ic) or adenosine receptor blockade with 8-phenyltheophylline (8-PT; 5 mg/kg iv). Inhibition of K(+)(ATP) channels resulted in a decrease in CBF and MVO(2) at rest and during exercise without a change in the relationship between CBF and MVO(2). In contrast, adenosine receptor blockade caused a significant increase in CBF that occurred secondary to an increase of MVO(2). These findings demonstrate that coronary K(+)(ATP) channel activity contribute to the regulation of resting MBF in CHF, and that endogenous adenosine may act to inhibit MVO(2) in the failing heart.

Role of hemolysis in red cell adenosine triphosphate release in simulated exercise conditions in vitro.

PubMed

Mairbäurl, Heimo; Ruppe, Florian A; Bärtsch, Peter

2013-10-01

Specific adenosine triphosphate (ATP) release from red blood cells has been discussed as a possible mediator controlling microcirculation in states of decreased tissue oxygen. Because intravascular hemolysis might also contribute to plasma ATP, we tested in vitro which portion of ATP release is due to hemolysis in typical exercise-induced strains to the red blood cells (shear stress, deoxygenation, and lactic acidosis). Human erythrocytes were suspended in dextran-containing media (hematocrit 10%) and were exposed to shear stress in a rotating Couette viscometer at 37°C. Desaturation (oxygen saturation of hemoglobin ∼20%) was achieved by tonometry with N2 before shear stress exposure. Cells not exposed to shear stress were used as controls. Na lactate (15 mM), lactic acid (15 mM, pH 7.0), and HCl (pH 7.0) were added to simulate exercise-induced lactic acidosis. After incubation, extracellular hemoglobin was measured to quantify hemolysis. ATP was measured with the luciferase assay. Shear stress increased extracellular ATP in a stress-related and time-dependent manner. Hypoxia induced a ∼10-fold increase in extracellular ATP in nonsheared cells and shear stress-exposed cells. Lactic acid had no significant effect on ATP release and hemolysis. In normoxic cells, approximately 20%-50% of extracellular ATP was due to hemolysis. This proportion decreased to less than 10% in hypoxic cells. Our results indicate that when exposing red blood cells to typical strains they encounter when passing through capillaries of exercising skeletal muscle, ATP release from red blood cells is caused mainly by deoxygenation and shear stress, whereas lactic acidosis had only a minor effect. Hemolysis effects were decreased when hemoglobin was deoxygenated. Together, by specific release and hemolysis, extracellular ATP reaches values that have been shown to cause local vasodilatation.

Defects in mitochondrial localization and ATP synthesis in the mdx mouse model of Duchenne muscular dystrophy are not alleviated by PDE5 inhibition

PubMed Central

Percival, Justin M.; Siegel, Michael P.; Knowels, Gary; Marcinek, David J.

2013-01-01

Given the crucial roles for mitochondria in ATP energy supply, Ca2+ handling and cell death, mitochondrial dysfunction has long been suspected to be an important pathogenic feature in Duchenne muscular dystrophy (DMD). Despite this foresight, mitochondrial function in dystrophin-deficient muscles has remained poorly defined and unknown in vivo. Here, we used the mdx mouse model of DMD and non-invasive spectroscopy to determine the impact of dystrophin-deficiency on skeletal muscle mitochondrial localization and oxidative phosphorylation function in vivo. Mdx mitochondria exhibited significant uncoupling of oxidative phosphorylation (reduced P/O) and a reduction in maximal ATP synthesis capacity that together decreased intramuscular ATP levels. Uncoupling was not driven by increased UCP3 or ANT1 expression. Dystrophin was required to maintain subsarcolemmal mitochondria (SSM) pool density, implicating it in the spatial control of mitochondrial localization. Given that nitric oxide-cGMP pathways regulate mitochondria and that sildenafil-mediated phosphodiesterase 5 inhibition ameliorates dystrophic pathology, we tested whether sildenafil's benefits result from decreased mitochondrial dysfunction in mdx mice. Unexpectedly, sildenafil treatment did not affect mitochondrial content or oxidative phosphorylation defects in mdx mice. Rather, PDE5 inhibition decreased resting levels of ATP, phosphocreatine and myoglobin, suggesting that sildenafil improves dystrophic pathology through other mechanisms. Overall, these data indicate that dystrophin-deficiency disrupts SSM localization, promotes mitochondrial inefficiency and restricts maximal mitochondrial ATP-generating capacity. Together these defects decrease intramuscular ATP and the ability of mdx muscle mitochondria to meet ATP demand. These findings further understanding of how mitochondrial bioenergetic dysfunction contributes to disease pathogenesis in dystrophin-deficient skeletal muscle in vivo. PMID:23049075

Molt cycle related changes and effect of short term starvation on the biochemical constituents of the blue swimmer crab Portunus pelagicus

PubMed Central

Sugumar, V.; Vijayalakshmi, G.; Saranya, K.

2012-01-01

Synthesis and hardening of a new exoskeleton are essential to the arthropod molting process. The present study emphasizes the variations in the levels of hemolymph total free sugars, hepatopancreas glycogen and cuticular proteins during the molting stages of Portunus pelagicus. It also reports the effect of short-term starvation conditions on the biochemical constituents of the hemolymph. Intermolt crabs were subjected to 6 days of starvation and hemolymph samples were taken. Standard biochemical procedures were followed toward the quantification of total proteins, total free sugars and total lipids. The total free sugar level in the hemolymph of P. pelagicus was observed to increase during early premolt D0 (3.108 ± 0.032 g/ml) and a gradual decrease till late postmolt B stage (0.552 ± 0.124 g/ml), suggesting the need for total free sugars to provide energy for the apolysis process. Increase in the levels of hepatopancreas glycogen was observed from 1225 ± 0.04 μg/mg in early premolt D0 to 1700 ± 0.3 μg/mg in late premolt D2–3. This is in correlation with the decreased levels of free sugars during premolt stages, suggesting an increase in the storage of glycogen reserves in the hepatopancreas. Cuticular proteins increased during stage B (2.702 ± 0.093 g/ml) and stage C (3.065 ± 0.012 g/ml), indicating exoskeleton hardening and mineralization. Results of the starvation studies clearly showed a steady decline in the level of total free sugars till day 6 (0.099 ± 0.00 g/ml) when compared to the control (8.646 ± 0.08 g/ml). Gradual decrease of total lipids was also observed from the first day of the experiment (6.088 ± 2.44 g/ml) to the last day of the study (0.401 ± 0.20 g/ml) which was 85% lesser than the control (8.450 ± 0.49 g/ml)suggesting the efficient usage of total sugars to consolidate the loss of energy reserves during starvation. The knowledge of Molt-cycle events can be used as a tool for the evaluation of the developmental state providing a morphological reference system for physiological and biochemical studies related to crab aquaculture. Starvation studies enlightens that increasing carbohydrate levels in crab feed together with good protein content could alleviate the natural effects of starvation, improve farm productivity and reduce the deleterious impact of nitrogen pollution generated by rich-protein feeds used in crab farming. PMID:23961226

[Effect of stress actions on some hematologic and biochemical parameters of rat blood and on energetic intermolecular interactions in lipid extract under effect of light radiation].

PubMed

Zabelinskiĭ, S A; Chebotareva, M A; Tavrovskaia, T V; Skverchinskaia, E A; Shukoliukova, E P; Maslov, M N; Krivchenko, A I

2012-01-01

Comparative study has been carried of effect of the three-day long starvation, running, and their combination on morphological parameters of rat blood, lipid metabolism, and activity of blood Na,K-ATPase. Different effect has been shown of these stress factors on the blood erythrocyte composition. Starvation is accompanied by the most pronounced release of stored erythrocyte into blood, which results in a significant decrease both of the total amount of reticulocytes and the complete absence of reticulocytes of the I stage of maturity (the youngest). The running on treadmill led to a significant increase of the total amount of blood reticulocytes and to multiple increase of immature reticulocytes (RC-I and RC-II), which can indicate some stress of the bone marrow erythroid stem line. The curve of acid resistance of blood reticulocytes has shown the animal to experience the greatest stress at a combination of starvation and running. Starvation and running produced different effects on blood lipid characteristics. The content of triacylglycerides (TAG) in blood rose by 40% at starvation and decreased by 30% at running, a similar tendency being found for index of atherogeneity. The fatty acid composition of blood phospholipids at running and its combination with starvation practically did not differ from control. A change of Na,K-ATPase, which is so characteristic of reaction to various kinds of stress, sharply fell at starvation (by 22%), but increased at running (by 13%) and decreased markedly at combination of these actions. Absorption spectra of lipid extracts of the whole blood of the rats submitted to various stress actions showed that extracted from blood (at different amount depending on the kind of action) is an organic substance with coupled bonds, which absorbs light in the diapason of 360-620 nm. The absorption of light in the diapason of 400-410 nm has been found to belong to the Soret band of ferroheme and ferriheme. The shift of the Soret band indicates electron transitions in the iron cation. By the change and disappearance of the Soret band, it is possible to judge about the processes occurring in the lipid extract. The disappearance of the Soret band in the lipid extract indicates formation in it of steady radicals as a result of the ferriheme disintegration due to accumulation of energy in porphyrin, which does not seem to occur in the blood cell membranes. The iron atom in the ferriheme molecule is known to accept electron and yields a part of energy probably to porphyrin. Then ferriheme yields electron and becomes ferriheme with excess of energy in porphyrin. Hence, at admission of the next electron to the iron atom the porphyrin molecule is to get rid of the energy obtained earlier to prevent its disintegration. The heme is possible to be an accumulator and distributor of energy in tissue.

Effect of periodical starvation on the life history of Brachionus plicatilis O.F. Müller (Rotifera): a possible strategy for population stability.

PubMed

Yoshinaga; Hagiwara; Tsukamoto

2000-10-25

To estimate the changes in the life history of the rotifer Brachionus plicatilis O.F. Müller under starvation, we carried out an individual culture and determined the effects of periodical food deprivation on its asexual reproductive characteristics such as lifespan, reproductive period, age at first egg and offspring production, and lifetime fecundity (total number of offspring produced in her lifetime). Rotifers were fed for 1-3 h daily, and were then starved until the next day. Control animals were fed throughout their lifespan. Starved rotifers matured and produced their first offspring at an older age than the control animals. The periodical starvation resulted in a decrease in the lifetime fecundity to less than half that of the non-starved control. The reproductive period and lifespan were 2-3 times longer in the starved animals than in the control animals. The negative relationship between lifespan and lifetime fecundity is interpreted as a trade-off in an alternative life-history strategy of rotifers under starved conditions. The great decrease in fecundity and extension of lifespan enables rotifers to compensate to keep the population in equilibrium.

31P-NMR measurements of ATP, ADP, 2,3-diphosphoglycerate and Mg2+ in human erythrocytes.

PubMed

Petersen, A; Kristensen, S R; Jacobsen, J P; Hørder, M

1990-08-17

Absolute 31P-NMR measurements of ATP, ADP and 2,3-diphosphoglycerate (2,3-DPG) in oxygenated and partly deoxygenated human erythrocytes, compared to measurements by standard assays after acid extraction, show that ATP is only 65% NMR visible, ADP measured by NMR is unexpectedly 400% higher than the enzymatic measurement and 2,3-DPG is fully NMR visible, regardless of the degree of oxygenation. These results show that binding to hemoglobin is unlikely to cause the decreased visibility of ATP in human erythrocytes as deoxyhemoglobin binds the phosphorylated metabolites more tightly than oxyhemoglobin. The high ADP visibility is unexplained. The levels of free Mg2+ [( Mg2+]free) in human erythrocytes are 225 mumol/l at an oxygen saturation of 98.6% and instead of the expected increase, the level decreased to 196 mumol/l at an oxygen saturation of 38.1% based on the separation between the alpha- and beta-ATP peaks. [Mg2+]free in the erythrocytes decreased to 104 mumol/l at a high 2,3-DPG concentration of 25.4 mmol/l red blood cells (RBC) and a normal ATP concentration of 2.05 mmol/l RBC. By increasing the ATP concentration to 3.57 mmol/l RBC, and with a high 2,3-DPG concentration of 24.7 mmol/l RBC, the 31P-NMR measured [Mg2+]free decreased to 61 mumol/l. These results indicate, that the 31P-NMR determined [Mg2+]free in human erythrocytes, based solely on the separation of the alpha- and beta-ATP peaks, does not give a true measure of intracellular free Mg2+ changes with different oxygen saturation levels. Furthermore the measurement is influenced by the concentration of the Mg2+ binding metabolites ATP and 2,3-DPG. Failure to take these factors into account when interpreting 31P-NMR data from human erythrocytes may explain some discrepancies in the literature regarding [Mg2+]free.

High-Throughput Profiling of Caenorhabditis elegans Starvation-Responsive microRNAs

PubMed Central

Garcia-Segura, Laura; Abreu-Goodger, Cei; Hernandez-Mendoza, Armando; Dimitrova Dinkova, Tzvetanka D.; Padilla-Noriega, Luis; Perez-Andrade, Martha Elva; Miranda-Rios, Juan

2015-01-01

MicroRNAs (miRNAs) are non-coding RNAs of ~22 nucleotides in length that regulate gene expression by interfering with the stability and translation of mRNAs. Their expression is regulated during development, under a wide variety of stress conditions and in several pathological processes. In nature, animals often face feast or famine conditions. We observed that subjecting early L4 larvae from Caenorhabditis elegans to a 12-hr starvation period produced worms that are thinner and shorter than well-fed animals, with a decreased lipid accumulation, diminished progeny, reduced gonad size, and an increased lifespan. Our objective was to identify which of the 302 known miRNAs of C. elegans changed their expression under starvation conditions as compared to well-fed worms by means of deep sequencing in early L4 larvae. Our results indicate that 13 miRNAs (miR-34-3p, the family of miR-35-3p to miR-41-3p, miR-39-5p, miR-41-5p, miR-240-5p, miR-246-3p and miR-4813-5p) were upregulated, while 2 miRNAs (let-7-3p and miR-85-5p) were downregulated in 12-hr starved vs. well-fed early L4 larvae. Some of the predicted targets of the miRNAs that changed their expression in starvation conditions are involved in metabolic or developmental process. In particular, miRNAs of the miR-35 family were upregulated 6–20 fold upon starvation. Additionally, we showed that the expression of gld-1, important in oogenesis, a validated target of miR-35-3p, was downregulated when the expression of miR-35-3p was upregulated. The expression of another reported target, the cell cycle regulator lin-23, was unchanged during starvation. This study represents a starting point for a more comprehensive understanding of the role of miRNAs during starvation in C. elegans. PMID:26554708

High-Throughput Profiling of Caenorhabditis elegans Starvation-Responsive microRNAs.

PubMed

Garcia-Segura, Laura; Abreu-Goodger, Cei; Hernandez-Mendoza, Armando; Dimitrova Dinkova, Tzvetanka D; Padilla-Noriega, Luis; Perez-Andrade, Martha Elva; Miranda-Rios, Juan

2015-01-01

MicroRNAs (miRNAs) are non-coding RNAs of ~22 nucleotides in length that regulate gene expression by interfering with the stability and translation of mRNAs. Their expression is regulated during development, under a wide variety of stress conditions and in several pathological processes. In nature, animals often face feast or famine conditions. We observed that subjecting early L4 larvae from Caenorhabditis elegans to a 12-hr starvation period produced worms that are thinner and shorter than well-fed animals, with a decreased lipid accumulation, diminished progeny, reduced gonad size, and an increased lifespan. Our objective was to identify which of the 302 known miRNAs of C. elegans changed their expression under starvation conditions as compared to well-fed worms by means of deep sequencing in early L4 larvae. Our results indicate that 13 miRNAs (miR-34-3p, the family of miR-35-3p to miR-41-3p, miR-39-5p, miR-41-5p, miR-240-5p, miR-246-3p and miR-4813-5p) were upregulated, while 2 miRNAs (let-7-3p and miR-85-5p) were downregulated in 12-hr starved vs. well-fed early L4 larvae. Some of the predicted targets of the miRNAs that changed their expression in starvation conditions are involved in metabolic or developmental process. In particular, miRNAs of the miR-35 family were upregulated 6-20 fold upon starvation. Additionally, we showed that the expression of gld-1, important in oogenesis, a validated target of miR-35-3p, was downregulated when the expression of miR-35-3p was upregulated. The expression of another reported target, the cell cycle regulator lin-23, was unchanged during starvation. This study represents a starting point for a more comprehensive understanding of the role of miRNAs during starvation in C. elegans.

Anethole dithiolethione, a putative neuroprotectant, increases intracellular and extracellular glutathione levels during starvation of cultured astroglial cells.

PubMed

Dringen, R; Hamprecht, B; Drukarch, B

1998-12-01

Astroglial cells protect neurons against oxidative damage. The antioxidant glutathione plays a pivotal role in the neuroprotective action of astroglial cells which is impaired following loss of glutathione. Anethole dithiolethione (ADT), a sulfur-containing compound which is used in humans as a secretagogue, increases glutathione levels in cultured astroglial cells under "physiological" conditions and is thought thereby to protect against oxidative damage. Presently, we report the effect of ADT (3-100 microM) on glutathione content of and efflux from rat primary astroglia-rich cultures under "pathological" conditions, i.e., extended deprivation of glucose and amino acids. Although cellular viability was not affected significantly, starvation of these cultures for 24 h in a bicarbonate buffer lacking glucose and amino acids led to a decrease in glutathione and protein content of approximately 43% and 40%, respectively. Although no effect on the protein loss occurred, the presence of ADT during starvation counteracted the starvation-induced loss of intracellular glutathione in a concentration-dependent way. At a concentration of 100 microM ADT even a significant increase in astroglial glutathione content was noted after 24 h of starvation. Alike intracellular glutathione levels, the amount of glutathione found in the buffer was elevated substantially if ADT was present during starvation. This ADT-mediated, apparent increase in glutathione efflux was additive to the stimulatory effect on extracellular glutathione levels of acivicin (100 microM), an inhibitor of extracellular enzymatic glutathione breakdown. However, the ADT-induced elevation of both intra- and extracellular glutathione content during starvation was prevented completely by coincubation with buthionine sulfoximine (10 microM), an inhibitor of glutathione synthesis. These results demonstrate that, most likely through stimulation of glutathione synthesis, ADT enables astroglial cells to maintain higher intra- and extracellular levels of glutathione under adverse conditions. Considering the lowered glutathione levels in neurodegenerative syndromes, we conclude that further evaluation of the therapeutic potential of the compound is warranted.

Tolbutamide attenuates diazoxide-induced aggravation of hypoxic cell injury.

PubMed

Pissarek, M; Reichelt, C; Krauss, G J; Illes, P

1998-11-23

ATP-dependent potassium (KATP) channels of neurons are closed in the presence of physiological levels of intracellular ATP and open when ATP is depleted during hypoxia or metabolic damage. The present study investigates hypoxic alterations of purine and pyrimidine nucleotide levels supposed to intracellularly modulate KATP channels. In addition, the effects of the KATP channel activator diazoxide and its antagonist tolbutamide were investigated on ATP, GTP, CTP and UTP levels in slices of the parietal cortex. Hypoxia was evoked by saturation of the medium with 95% N2-5% CO2 instead of 95% O2-5% CO2 for 5 min. Nucleotide contents were measured by anion-exchange HPLC in neutralized perchloric acid extracts obtained from slices frozen immediately at the end of incubation. Hypoxia per se decreased purine and pyrimidine nucleoside triphosphate contents. Thus, ATP and GTP contents were reduced to 69.9 and 77.6% of the respective normoxic levels. UTP and CTP contents were even more decreased (to 60.9 and 41.6%),, probably because the salvage pathway of these pyrimidine nucleotides is less effective than that of the purine nucleotides ATP and GTP. While tolbutamide (30 microM) had no effect on the hypoxia-induced decrease of nucleotides, diazoxide at 300, but not 30 microM aggravated the decline of ATP, UTP and CTP to 51.8, 37.5 and 28.5% of the contents observed at normoxia; GTP levels also showed a tendency to decrease after diazoxide application. Tolbutamide (300 microM) antagonized the effects of diazoxide (300 but not 30 microM aggravated the decline of ATP, UTP and CTP to 51.8, 37.5 and 28.5% of the contents observed at normoxia; GTP levels also showed a tendency to decrease after diazoxide application. Tolbutamide (300 microM) antagonized the effects of diazoxide (300 MicroM). Nucleoside diphosphate (ADP, GDP and UDP) levels were uniformly increased by hypoxia. There was no hypoxia-induced increase of ADP contents in the presence of tolbutamide (300 microM). The ATP/ADP, GTP/GDP and UTP/UDP ratios uniformly declined at a low pO2. However, only the ATP/ADP ratio was decreased further by diazoxide (300 microM). The observed alterations in nucleotide contents may be of importance for long- and short-term processes related to acute cerebral hypoxia. Thus, hypoxia-induced alterations of purine and pyrimidine nucleotide levels may influence the open state of KATP-channels during the period of reversible hypoxic cerebral injury. Furthermore, alterations during the irreversible period of cerebral injury may also arise, as a consequence of decreased pyrimidine nucleotide contents affecting cell survival viaprotein and DNA synthesis.

Analysis of the mechanism by which calcium negatively regulates the tyrosine phosphorylation cascade associated with sperm capacitation.

PubMed

Baker, Mark A; Hetherington, Louise; Ecroyd, Heath; Roman, Shaun D; Aitken, R John

2004-01-15

The capacitation of mammalian spermatozoa involves the activation of a cAMP-mediated signal transduction pathway that drives tyrosine phosphorylation via mechanisms that are unique to this cell type. Controversy surrounds the impact of extracellular calcium on this process, with positive and negative effects being recorded in independent publications. We clearly demonstrate that the presence of calcium in the external medium decreases tyrosine phosphorylation in both human and mouse spermatozoa. Under these conditions, a rise in intracellular pH was recorded, however, this event was not responsible for the observed changes in phosphotyrosine expression. Rather, the impact of calcium on tyrosine phosphorylation in these cells was associated with an unexpected change in the intracellular availability of ATP. Thus, the ATP content of both human and mouse spermatozoa fell significantly when these cells were incubated in the presence of external calcium. Furthermore, the removal of glucose, or addition of 2-deoxyglucose, decreased ATP levels within human spermatozoon populations and induced a corresponding decline in phosphotyrosine expression. In contrast, the mitochondrial inhibitor rotenone had no effect on either ATP levels or tyrosine phosphorylation. Addition of the affinity-labeling probe 8-N3 ATP confirmed our prediction that spermatozoa have many calcium-dependent ATPases. Moreover, addition of the ATPase inhibitor thapsigargin, increased intracellular calcium levels, decreased ATP and suppressed tyrosine phosphorylation. Based on these findings, the present study indicates that extracellular calcium suppresses tyrosine phosphorylation by decreasing the availability of intracellular ATP, and not by activating tyrosine phosphatases or inhibiting tyrosine kinases as has been previously suggested.

A Novel GLP1 Receptor Interacting Protein ATP6ap2 Regulates Insulin Secretion in Pancreatic Beta Cells*

PubMed Central

Dai, Feihan F.; Bhattacharjee, Alpana; Liu, Ying; Batchuluun, Battsetseg; Zhang, Ming; Wang, Xinye Serena; Huang, Xinyi; Luu, Lemieux; Zhu, Dan; Gaisano, Herbert; Wheeler, Michael B.

2015-01-01

GLP1 activates its receptor, GLP1R, to enhance insulin secretion. The activation and transduction of GLP1R requires complex interactions with a host of accessory proteins, most of which remain largely unknown. In this study, we used membrane-based split ubiquitin yeast two-hybrid assays to identify novel GLP1R interactors in both mouse and human islets. Among these, ATP6ap2 (ATPase H+-transporting lysosomal accessory protein 2) was identified in both mouse and human islet screens. ATP6ap2 was shown to be abundant in islets including both alpha and beta cells. When GLP1R and ATP6ap2 were co-expressed in beta cells, GLP1R was shown to directly interact with ATP6ap2, as assessed by co-immunoprecipitation. In INS-1 cells, overexpression of ATP6ap2 did not affect insulin secretion; however, siRNA knockdown decreased both glucose-stimulated and GLP1-induced insulin secretion. Decreases in GLP1-induced insulin secretion were accompanied by attenuated GLP1 stimulated cAMP accumulation. Because ATP6ap2 is a subunit required for V-ATPase assembly of insulin granules, it has been reported to be involved in granule acidification. In accordance with this, we observed impaired insulin granule acidification upon ATP6ap2 knockdown but paradoxically increased proinsulin secretion. Importantly, as a GLP1R interactor, ATP6ap2 was required for GLP1-induced Ca2+ influx, in part explaining decreased insulin secretion in ATP6ap2 knockdown cells. Taken together, our findings identify a group of proteins that interact with the GLP1R. We further show that one interactor, ATP6ap2, plays a novel dual role in beta cells, modulating both GLP1R signaling and insulin processing to affect insulin secretion. PMID:26272612

Influence of some mononucleotides and their corresponding nucleosides on the metabolism of carbohydrates in the isolated rat diaphragm muscle

PubMed Central

Beloff-Chain, Anne; Betto, P.; Bleszynski, W.; Catanzaro, Raffaella; Chain, E. B.; Dmitrovskii, A. A.; Longinotti, L.; Pocchiari, F.

1965-01-01

1. The influence of ATP on glucose metabolism was studied in the isolated rat diaphragm; it was shown that ATP increases the oxidation of glucose and the aerobic conversion of glucose into lactate, whereas it decreases glycogen synthesis. There was no influence of ATP on the anaerobic formation of lactate from glucose. 2. A maximum effect of ATP on the oxidation of glucose (about 160% increase) was obtained in the presence of 10mm-ATP; in the presence of 2mm-ATP the effect was about 65%, and was approximately constant from 10 to 90min. incubation period. 3. In a phosphate-free tris-buffered medium the oxidation of glucose was considerably decreased, but the percentage stimulation by ATP was about the same as in a phosphate-buffered medium. 4. ATP was shown to increase the oxidation of fructose, glucose 6-phosphate, glucose 1-phosphate, fructose 1,6-diphosphate and, to a much smaller extent, pyruvate. 5. ADP stimulated the oxidation of glucose to the same extent as ATP at a concentration of 2mm and the effect with AMP was only slightly less; IMP and adenosine had only a small stimulatory effect at this concentration, whereas inosine had no effect. PMID:16749165

Hepatic Complications of Anorexia Nervosa.

PubMed

Rosen, Elissa; Bakshi, Neeru; Watters, Ashlie; Rosen, Hugo R; Mehler, Philip S

2017-11-01

Anorexia nervosa (AN) has the highest mortality rate of all psychiatric illnesses due to the widespread organ dysfunction caused by the underlying severe malnutrition. Starvation causes hepatocyte injury and death leading to a rise in aminotransferases. Malnutrition-induced hepatitis is common among individuals with AN especially as body mass index decreases. Acute liver failure associated with coagulopathy and encephalopathy can rarely occur. Liver enzymes may also less commonly increase as part of the refeeding process due to hepatic steatosis and can be distinguished from starvation hepatitis by the finding of a fatty liver on ultrasonography. Individuals with AN and starvation-induced hepatitis are at increased risk of hypoglycemia due to depleted glycogen stores and impaired gluconeogenesis. Gastroenterology and hepatology consultations are often requested when patients with AN and signs of hepatitis are hospitalized. It should be noted that additional laboratory testing, imaging, or liver biopsy all have low diagnostic yield, are costly, and potentially invasive, therefore, not generally recommended for diagnostic purposes. While the hepatitis of AN can reach severe levels, a supervised increase in caloric intake and a return to a healthy body weight often quickly lead to normalization of elevated aminotransferases caused by starvation.

Chi-square analysis of the reduction of ATP levels in L-02 hepatocytes by hexavalent chromium.

PubMed

Yuan, Yang; Peng, Li; Gong-Hua, Hu; Lu, Dai; Xia-Li, Zhong; Yu, Zhou; Cai-Gao, Zhong

2012-06-01

This study explored the reduction of adenosine triphosphate (ATP) levels in L-02 hepatocytes by hexavalent chromium (Cr(VI)) using chi-square analysis. Cells were treated with 2, 4, 8, 16, or 32 μM Cr(VI) for 12, 24, or 36 h. Methyl thiazolyl tetrazolium (MTT) experiments and measurements of intracellular ATP levels were performed by spectrophotometry or bioluminescence assays following Cr(VI) treatment. The chi-square test was used to determine the difference between cell survival rate and ATP levels. For the chi-square analysis, the results of the MTT or ATP experiments were transformed into a relative ratio with respect to the control (%). The relative ATP levels increased at 12 h, decreased at 24 h, and increased slightly again at 36 h following 4, 8, 16, 32 μM Cr(VI) treatment, corresponding to a "V-shaped" curve. Furthermore, the results of the chi-square analysis demonstrated a significant difference of the ATP level in the 32-μM Cr(VI) group (P < 0.05). The results suggest that the chi-square test can be applied to analyze the interference effects of Cr(VI) on ATP levels in L-02 hepatocytes. The decreased ATP levels at 24 h indicated disruption of mitochondrial energy metabolism and the slight increase of ATP levels at 36 h indicated partial recovery of mitochondrial function or activated glycolysis in L-02 hepatocytes.

Effect of exogenous ATP on the postharvest properties and pectin degradation of mung bean sprouts (Vigna radiata).

PubMed

Chen, Lin; Zhou, Yige; He, Zhenyun; Liu, Qin; Lai, Shaojuan; Yang, Hongshun

2018-06-15

The effects of exogenous ATP on the postharvest quality, browning and softening of mung bean (Vigna radiata) sprouts were evaluated. ATP treatment significantly alleviated the quality loss and browning events during the storage of 3 days. It also reduced the oxidant damage by inducing high activities of peroxidase (9.3-13.9%) and superoxide dismutase (8.8-10.3%) which scavenged the reactive oxygen species (ROS) effectively. Transcriptional results indicated that ATP treatment decreased VrPL1, VrPME and VrPG1 gene expression levels more than 2 folds at some time points. Furthermore, the atomic force microscope (AFM) images revealed that the pectin degradation was notably slowed by ATP treatment and the width and height of pectin backbone were better maintained (47.1% and 45.6% higher than control without ATP treatment). The cooperative effects of ROS scavenging and decreased expressions of pectin-related genes might contribute to the deferred pectin deterioration and firmness loss by ATP treatment. Copyright © 2018 Elsevier Ltd. All rights reserved.

Effects and mechanism of acid rain on plant chloroplast ATP synthase.

PubMed

Sun, Jingwen; Hu, Huiqing; Li, Yueli; Wang, Lihong; Zhou, Qing; Huang, Xiaohua

2016-09-01

Acid rain can directly or indirectly affect plant physiological functions, especially photosynthesis. The enzyme ATP synthase is the key in photosynthetic energy conversion, and thus, it affects plant photosynthesis. To clarify the mechanism by which acid rain affects photosynthesis, we studied the effects of acid rain on plant growth, photosynthesis, chloroplast ATP synthase activity and gene expression, chloroplast ultrastructure, intracellular H(+) level, and water content of rice seedlings. Acid rain at pH 4.5 remained the chloroplast structure unchanged but increased the expression of six chloroplast ATP synthase subunits, promoted chloroplast ATP synthase activity, and increased photosynthesis and plant growth. Acid rain at pH 4.0 or less decreased leaf water content, destroyed chloroplast structure, inhibited the expression of six chloroplast ATP synthase subunits, decreased chloroplast ATP synthase activity, and reduced photosynthesis and plant growth. In conclusion, acid rain affected the chloroplast ultrastructure, chloroplast ATPase transcription and activity, and P n by changing the acidity in the cells, and thus influencing the plant growth and development. Finally, the effects of simulated acid rain on the test indices were found to be dose-dependent.

Consequences of the pathogenic T9176C mutation of human mitochondrial DNA on yeast mitochondrial ATP synthase

PubMed Central

Kucharczyk, Roza; Ezkurdia, Nahia; Couplan, Elodie; Procaccio, Vincent; Ackerman, Sharon H.; Blondel, Marc; di Rago, Jean-Paul

2010-01-01

Summary Several human neurological disorders have been associated with various mutations affecting mitochondrial enzymes involved in cellular ATP production. One of these mutations, T9176C in the mitochondrial DNA (mtDNA), changes a highly conserved leucine residue into proline at position 217 of the mitochondrially encoded Atp6p (or a) subunit of the F1FO-ATP synthase. The consequences of this mutation on the mitochondrial ATP synthase are still poorly defined. To gain insight into the primary pathogenic mechanisms induced by T9176C, we have investigated the consequences of this mutation on the ATP synthase of yeast where Atp6p is also encoded by the mtDNA. In vitro, yeast atp6-T9176C mitochondria showed a 30% decrease in the rate of ATP synthesis. When forcing the F1FO complex to work in the reverse mode, i.e. F1-catalyzed hydrolysis of ATP coupled to proton transport out of the mitochondrial matrix, the mutant showed a normal proton-pumping activity and this activity was fully sensitive to oligomycin, an inhibitor of the ATP synthase proton channel. However, under conditions of maximal ATP hydrolytic activity, using non-osmotically protected mitochondria, the mutant ATPase activity was less efficiently inhibited by oligomycin (60% inhibition versus 85% for the wild type control). BN-PAGE analyses revealed that atp6-T9176C yeast accumulated rather good levels of fully assembled ATP synthase complexes. However, a number of subcomplexes (F1, Atp9p-ring, unassembled α-F1 subunits) could be detected as well, presumably because of a decreased stability of Atp6p within the ATP synthase. Although the oxidative phosphorylation capacity was reduced in atp6-T9176C yeast, the number of ATP molecules synthesized per electron transferred to oxygen was similar compared with wild type yeast. It can therefore be inferred that the coupling efficiency within the ATP synthase was mostly unaffected and that the T9176C mutation did not increase the proton permeability of the mitochondrial inner membrane. PMID:20056103

Aldehyde dehydrogenase inhibition combined with phenformin treatment reversed NSCLC through ATP depletion

PubMed Central

Lee, Jae-Seon; Nam, Boas; Seong, Tae Wha; Son, Jaekyoung; Jang, Hyonchol; Hong, Kyeong Man; Lee, Cheolju; Kim, Soo-Youl

2016-01-01

Among ALDH isoforms, ALDH1L1 in the folate pathway showed highly increased expression in non-small-cell lung cancer cells (NSCLC). Based on the basic mechanism of ALDH converting aldehyde to carboxylic acid with by-product NADH, we suggested that ALDH1L1 may contribute to ATP production using NADH through oxidative phosphorylation. ALDH1L1 knockdown reduced ATP production by up to 60% concomitantly with decrease of NADH in NSCLC. ALDH inhibitor, gossypol, also reduced ATP production in a dose dependent manner together with decrease of NADH level in NSCLC. A combination treatment of gossypol with phenformin, mitochondrial complex I inhibitor, synergized ATP depletion, which efficiently induced cell death. Pre-clinical xenograft model using human NSCLC demonstrated a remarkable therapeutic response to the combined treatment of gossypol and phenformin. PMID:27384481

Aldehyde dehydrogenase inhibition combined with phenformin treatment reversed NSCLC through ATP depletion.

PubMed

Kang, Joon Hee; Lee, Seon-Hyeong; Lee, Jae-Seon; Nam, Boas; Seong, Tae Wha; Son, Jaekyoung; Jang, Hyonchol; Hong, Kyeong Man; Lee, Cheolju; Kim, Soo-Youl

2016-08-02

Among ALDH isoforms, ALDH1L1 in the folate pathway showed highly increased expression in non-small-cell lung cancer cells (NSCLC). Based on the basic mechanism of ALDH converting aldehyde to carboxylic acid with by-product NADH, we suggested that ALDH1L1 may contribute to ATP production using NADH through oxidative phosphorylation. ALDH1L1 knockdown reduced ATP production by up to 60% concomitantly with decrease of NADH in NSCLC. ALDH inhibitor, gossypol, also reduced ATP production in a dose dependent manner together with decrease of NADH level in NSCLC. A combination treatment of gossypol with phenformin, mitochondrial complex I inhibitor, synergized ATP depletion, which efficiently induced cell death. Pre-clinical xenograft model using human NSCLC demonstrated a remarkable therapeutic response to the combined treatment of gossypol and phenformin.

Long-term sulphur starvation of Arabidopsis thaliana modifies mitochondrial ultrastructure and activity and changes tissue energy and redox status.

PubMed

Ostaszewska, Monika; Juszczuk, Izabela M; Kołodziejek, Izabella; Rychter, Anna M

2014-04-15

Sulphur, as a constituent of amino acids (cysteine and methionine), iron-sulphur clusters, proteins, membrane sulpholipids, glutathione, glucosinolates, coenzymes, and auxin precursors, is essential for plant growth and development. Absence or low sulphur concentration in the soil results in severe growth retardation. Arabidopsis thaliana plants grown hydroponically for nine weeks on Knop nutrient medium without sulphur showed morphological symptoms of sulphur deficiency. The purpose of our study was to investigate changes that mitochondria undergo and the role of the highly branched respiratory chain in survival during sulphur deficiency stress. Ultrastructure analysis of leaf mesophyll cells of sulphur-deficient Arabidopsis showed heterogeneity of mitochondria; some of them were not altered, but the majority had swollen morphology. Dilated mitochondria displayed a lower matrix density and fewer cristae compared to control mitochondria. Disintegration of the inner and outer membranes of some mitochondria from the leaves of sulphur-deficient plants was observed. On the contrary, chloroplast ultrastructure was not affected. Sulphur deficiency changed the respiratory activity of tissues and isolated mitochondria; Complex I and IV capacities and phosphorylation rates were lower, but external NAD(P)H dehydrogenase activity increased. Higher external NAD(P)H dehydrogenase activity corresponded to increased cell redox level with doubled NADH/NAD ratio in the leaf and root tissues. Sulphur deficiency modified energy status in the tissues of Arabidopsis plants. The total concentration of adenylates (expressed as ATP+ADP), measured in the light, was lower in the leaves and roots of sulphur-deficient plants than in the controls, which was mainly due to the severely decreased ATP levels. We show that the changes in mitochondrial ultrastructure are compensated by the modifications in respiratory chain activity. Although mitochondria of Arabidopsis tissues are affected by sulphur deficiency, their metabolic and structural features, which readily reach new homeostasis, make these organelles crucial for adaptation of plants to survive sulphur deficiency. Copyright © 2013 Elsevier GmbH. All rights reserved.

Sensitivity of small myosin II ensembles from different isoforms to mechanical load and ATP concentration.

PubMed

Erdmann, Thorsten; Bartelheimer, Kathrin; Schwarz, Ulrich S

2016-11-01

Based on a detailed crossbridge model for individual myosin II motors, we systematically study the influence of mechanical load and adenosine triphosphate (ATP) concentration on small myosin II ensembles made from different isoforms. For skeletal and smooth muscle myosin II, which are often used in actomyosin gels that reconstitute cell contractility, fast forward movement is restricted to a small region of phase space with low mechanical load and high ATP concentration, which is also characterized by frequent ensemble detachment. At high load, these ensembles are stalled or move backwards, but forward motion can be restored by decreasing ATP concentration. In contrast, small ensembles of nonmuscle myosin II isoforms, which are found in the cytoskeleton of nonmuscle cells, are hardly affected by ATP concentration due to the slow kinetics of the bound states. For all isoforms, the thermodynamic efficiency of ensemble movement increases with decreasing ATP concentration, but this effect is weaker for the nonmuscle myosin II isoforms.

K+ depolarization evokes ATP, adenosine and glutamate release from glia in rat hippocampus: a microelectrode biosensor study

PubMed Central

Heinrich, A; Andó, RD; Túri, G; Rózsa, B; Sperlágh, B

2012-01-01

BACKGROUND AND PURPOSE This study was undertaken to characterize the ATP, adenosine and glutamate outflow evoked by depolarization with high K+ concentrations, in slices of rat hippocampus. EXPERIMENTAL APPROACH We utilized the microelectrode biosensor technique and extracellular electrophysiological recording for the real-time monitoring of the efflux of ATP, adenosine and glutamate. KEY RESULTS ATP, adenosine and glutamate sensors exhibited transient and reversible current during depolarization with 25 mM K+, with distinct kinetics. The ecto-ATPase inhibitor ARL67156 enhanced the extracellular level of ATP and inhibited the prolonged adenosine efflux, suggesting that generation of adenosine may derive from the extracellular breakdown of ATP. Stimulation-evoked ATP, adenosine and glutamate efflux was inhibited by tetrodotoxin, while exposure to Ca2+-free medium abolished ATP and adenosine efflux from hippocampal slices. Extracellular elevation of ATP and adenosine were decreased in the presence of NMDA receptor antagonists, D-AP-5 and ifenprodil, whereas non-NMDA receptor blockade by CNQX inhibited glutamate but not ATP and adenosine efflux. The gliotoxin fluoroacetate and P2X7 receptor antagonists inhibited the K+-evoked ATP, adenosine and glutamate efflux, while carbenoxolone in low concentration and probenecid decreased only the adenosine efflux. CONCLUSIONS AND IMPLICATIONS Our results demonstrated activity-dependent gliotransmitter release in the hippocampus in response to ongoing neuronal activity. ATP and glutamate were released by P2X7 receptor activation into extracellular space. Although the increased extracellular levels of adenosine did derive from released ATP, adenosine might also be released directly via pannexin hemichannels. LINKED ARTICLE This article is commented on by Sershen, pp. 1000–1002 of this issue. To view this commentary visit http://dx.doi.org/10.1111/j.1476-5381.2012.02072.x PMID:22394324

Influence of glucagon or 5-(tetradecyloxy)-2-furoic acid on binding to mitochondria and phosphorylation of ATP-citrate lyase.

PubMed

Janski, A M; Cornell, N W

1982-02-01

To study the binding to mitochondria and the phosphorylation of ATP-citrate lyase (EC 4.1.3.8), isolated rat hepatocytes were fractionated by exposure to digitonin. After incubation of hepatocytes with the hypolipidemic agent 5-(tetradecyloxy)-2-furoic acid, which decreases the cellular CoA, the amount of bound ATP-citrate lyase was increased, but the content of acid-stable phosphate in the enzyme was diminished. Glucagon, in contrast, decreased the amount of bound enzyme but increased phosphorylation. This inverse relationship might indicate either that the bound ATP-citrate lyase is less readily phosphorylated or that the phosphorylated enzyme binds less readily to mitochondria.

ATP promotes cell survival via regulation of cytosolic [Ca2+] and Bcl-2/Bax ratio in lung cancer cells

PubMed Central

Song, Shanshan; Jacobson, Krista N.; McDermott, Kimberly M.; Reddy, Sekhar P.; Cress, Anne E.; Tang, Haiyang; Dudek, Steven M.; Black, Stephen M.; Garcia, Joe G. N.; Makino, Ayako

2015-01-01

Adenosine triphosphate (ATP) is a ubiquitous extracellular messenger elevated in the tumor microenvironment. ATP regulates cell functions by acting on purinergic receptors (P2X and P2Y) and activating a series of intracellular signaling pathways. We examined ATP-induced Ca2+ signaling and its effects on antiapoptotic (Bcl-2) and proapoptotic (Bax) proteins in normal human airway epithelial cells and lung cancer cells. Lung cancer cells exhibited two phases (transient and plateau phases) of increase in cytosolic [Ca2+] ([Ca2+]cyt) caused by ATP, while only the transient phase was observed in normal cells. Removal of extracellular Ca2+ eliminated the plateau phase increase of [Ca2+]cyt in lung cancer cells, indicating that the plateau phase of [Ca2+]cyt increase is due to Ca2+ influx. The distribution of P2X (P2X1-7) and P2Y (P2Y1, P2Y2, P2Y4, P2Y6, P2Y11) receptors was different between lung cancer cells and normal cells. Proapoptotic P2X7 was nearly undetectable in lung cancer cells, which may explain why lung cancer cells showed decreased cytotoxicity when treated with high concentration of ATP. The Bcl-2/Bax ratio was increased in lung cancer cells following treatment with ATP; however, the antiapoptotic protein Bcl-2 demonstrated more sensitivity to ATP than proapoptotic protein Bax. Decreasing extracellular Ca2+ or chelating intracellular Ca2+ with BAPTA-AM significantly inhibited ATP-induced increase in Bcl-2/Bax ratio, indicating that a rise in [Ca2+]cyt through Ca2+ influx is the critical mediator for ATP-mediated increase in Bcl-2/Bax ratio. Therefore, despite high ATP levels in the tumor microenvironment, which would induce cell apoptosis in normal cells, the decreased P2X7 and elevated Bcl-2/Bax ratio in lung cancer cells may enable tumor cells to survive. Increasing the Bcl-2/Bax ratio by exposure to high extracellular ATP may, therefore, be an important selective pressure promoting transformation and cancer progression. PMID:26491047

Starvation stress during larval development facilitates an adaptive response in adult worker honey bees (Apis mellifera L.).

PubMed

Wang, Ying; Kaftanoglu, Osman; Brent, Colin S; Page, Robert E; Amdam, Gro V

2016-04-01

Most organisms are constantly faced with environmental changes and stressors. In diverse organisms, there is an anticipatory mechanism during development that can program adult phenotypes. The adult phenotype would be adapted to the predicted environment that occurred during organism maturation. However, whether this anticipatory mechanism is present in eusocial species is questionable because eusocial organisms are largely shielded from exogenous conditions by their stable nest environment. In this study, we tested whether food deprivation during development of the honey bee (Apis mellifera), a eusocial insect model, can shift adult phenotypes to better cope with nutritional stress. After subjecting fifth instar worker larvae to short-term starvation, we measured nutrition-related morphology, starvation resistance, physiology, endocrinology and behavior in the adults. We found that the larval starvation caused adult honey bees to become more resilient toward starvation. Moreover, the adult bees were characterized by reduced ovary size, elevated glycogen stores and juvenile hormone (JH) titers, and decreased sugar sensitivity. These changes, in general, can help adult insects survive and reproduce in food-poor environments. Overall, we found for the first time support for an anticipatory mechanism in a eusocial species, the honey bee. Our results suggest that this mechanism may play a role in honey bee queen-worker differentiation and worker division of labor, both of which are related to the responses to nutritional stress. © 2016. Published by The Company of Biologists Ltd.

Regulation of PRPP and nucleoside tri and tetraphosphate pools in Escherichia coli under conditions of nitrogen starvation.

PubMed Central

Villadsen, I S; Michelsen, O

1977-01-01

The ribonucleoside triphosphate, deoxyribonucleoside triphosphate, 3' -diphosphate guanosine 5' -diphosphate (ppGpp), and 5-phosphoribosyl 1-pyrophosphate (PRPP) pools in Escherichia coli B were determined by thin-layer chromatography during changing conditions to ammonium starvation. The intracellular concentrations of all nucleotides were found to change in a well-defined order several minutes before andy observed change in the optical density of the culture. The levels of purine nucleoside triphosphates (adenosine 5' -triphosphate [CTP], dCTP) and uridine nucleotides (uridine 5' -triphosphate, deoxythymidine 5'-triphosphate). The deoxyribonucleotides thus behaved as the ribonucleotides. The levels of ppGpp increased 11-fold after the decrease in uridine nucleotides, when the accumulation of stable ribonucleic acid (RNA) stopped. The level of the nucleotide pool did not stabilize until 30 min after the change in optical density. The pool of dGTP dropped concomitantly with the pool of CTP. The nucleotide precursor PRPP exhibited a transient increase, wtih maximum value of four times the exponential levels at the onset of starvation. Apparently the cell adjusts early to starvation by reducing either the phosphorylating activity or the nucleotide biosynthetic activity. As in other downshift systems, the accumulation of stable RNA stopped before the break in optical density and before the stop in protein accumulation. Cell divisions were quite insensitive to the control mechanisms operating on RNA and protein accumulation under ammonium starvation, since the cells continued to divide for 21 min without any net accumulation of RNA. Images PMID:323222

Starvation stress affects the interplay among shrimp gut microbiota, digestion and immune activities.

PubMed

Dai, Wen-Fang; Zhang, Jin-Jie; Qiu, Qiong-Fen; Chen, Jiong; Yang, Wen; Ni, Sui; Xiong, Jin-Bo

2018-05-24

Aquatic animals are frequently suffered from starvation due to restricted food availability or deprivation. It is currently known that gut microbiota assists host in nutrient acquisition. Thus, exploring the gut microbiota responses would improve our understanding on physiological adaptation to starvation. To achieve this, we investigated how the gut microbiota and shrimp digestion and immune activities were affected under starvation stress. The results showed that the measured digestion activities in starved shrimp were significantly lower than in normal cohorts; while the measured immune activities exhibited an opposite trend. A structural equation modeling (SEM) revealed that changes in the gut bacterial community were directly related to digestive and immune enzyme activities, which in turn markedly affected shrimp growth traits. Notably, several gut bacterial indicators that characterized the shrimp nutrient status were identified, with more abundant opportunistic pathogens in starved shrimp, although there were no statistical differences in the overall diversity and the structures of gut bacterial communities between starved and normal shrimp. Starved shrimp exhibited less connected and cooperative interspecies interaction as compared with normal cohorts. Additionally, the functional pathways involved in carbohydrate and protein digestion, glycan biosynthesis, lipid and enzyme metabolism remarkably decreased in starved shrimp. These attenuations could increase the susceptibility of starved shrimp to pathogens infection. In summary, this study provides novel insights into the interplay among shrimp digestion, immune activities and gut microbiota in response to starvation stress. Copyright © 2018 Elsevier Ltd. All rights reserved.

Targeted mitochondrial therapy using MitoQ shows equivalent renoprotection to angiotensin converting enzyme inhibition but no combined synergy in diabetes.

PubMed

Ward, Micheal S; Flemming, Nicole B; Gallo, Linda A; Fotheringham, Amelia K; McCarthy, Domenica A; Zhuang, Aowen; Tang, Peter H; Borg, Danielle J; Shaw, Hannah; Harvie, Benjamin; Briskey, David R; Roberts, Llion A; Plan, Manuel R; Murphy, Michael P; Hodson, Mark P; Forbes, Josephine M

2017-11-09

Mitochondrial dysfunction is a pathological mediator of diabetic kidney disease (DKD). Our objective was to test the mitochondrially targeted agent, MitoQ, alone and in combination with first line therapy for DKD. Intervention therapies (i) vehicle (D); (ii) MitoQ (DMitoQ;0.6 mg/kg/day); (iii) Ramipril (DRam;3 mg/kg/day) or (iv) combination (DCoAd) were administered to male diabetic db/db mice for 12 weeks (n = 11-13/group). Non-diabetic (C) db/m mice were followed concurrently. No therapy altered glycaemic control or body weight. By the study end, both monotherapies improved renal function, decreasing glomerular hyperfiltration and albuminuria. All therapies prevented tubulointerstitial collagen deposition, but glomerular mesangial expansion was unaffected. Renal cortical concentrations of ATP, ADP, AMP, cAMP, creatinine phosphate and ATP:AMP ratio were increased by diabetes and mostly decreased with therapy. A higher creatine phosphate:ATP ratio in diabetic kidney cortices, suggested a decrease in ATP consumption. Diabetes elevated glucose 6-phosphate, fructose 6-phosphate and oxidised (NAD+ and NADP+) and reduced (NADH) nicotinamide dinucleotides, which therapy decreased generally. Diabetes increased mitochondrial oxygen consumption (OCR) at complex II-IV. MitoQ further increased OCR but decreased ATP, suggesting mitochondrial uncoupling as its mechanism of action. MitoQ showed renoprotection equivalent to ramipril but no synergistic benefits of combining these agents were shown.

Contribution of proton leak to oxygen consumption in skeletal muscle during intense exercise is very low despite large contribution at rest

PubMed Central

2017-01-01

A computer model was used to simulate the dependence of protonmotive force (Δp), proton leak and phenomenological (involving proton leak) ATP/O2 ratio on work intensity in skeletal muscle. Δp, NADH and proton leak decreased with work intensity. The contribution of proton leak to oxygen consumption (V˙O2) decreased from about 60% at rest to about 3 and 1% at moderate and heavy/severe exercise, respectively, while the ATP/O2 ratio increased from 2.1 to 5.5 and 5.7. A two-fold increase in proton leak activity or its decrease to zero decreased/increased the ATP/O2 ratio by only about 3 and 1% during moderate and heavy/severe exercise, respectively. The low contribution of proton leak to V˙O2 in intensively working skeletal muscle was mostly caused by a huge increase in ATP usage intensity during rest-to-work transition, while OXPHOS, and thus oxidative ATP supply and V˙O2 related to it, was mostly stimulated by high each-step activation (ESA) of OXPHOS complexes. The contribution of proton leak to V˙O2 and ATP/O2 ratio in isolated mitochondria should not be directly extrapolated to working muscle, as mitochondria lack ESA, at least in the absence of Ca2+, and therefore V˙O2 cannot be elevated as much as in intact muscle. PMID:29045413

Contribution of proton leak to oxygen consumption in skeletal muscle during intense exercise is very low despite large contribution at rest.

PubMed

Korzeniewski, Bernard

2017-01-01

A computer model was used to simulate the dependence of protonmotive force (Δp), proton leak and phenomenological (involving proton leak) ATP/O2 ratio on work intensity in skeletal muscle. Δp, NADH and proton leak decreased with work intensity. The contribution of proton leak to oxygen consumption ([Formula: see text]) decreased from about 60% at rest to about 3 and 1% at moderate and heavy/severe exercise, respectively, while the ATP/O2 ratio increased from 2.1 to 5.5 and 5.7. A two-fold increase in proton leak activity or its decrease to zero decreased/increased the ATP/O2 ratio by only about 3 and 1% during moderate and heavy/severe exercise, respectively. The low contribution of proton leak to [Formula: see text] in intensively working skeletal muscle was mostly caused by a huge increase in ATP usage intensity during rest-to-work transition, while OXPHOS, and thus oxidative ATP supply and [Formula: see text] related to it, was mostly stimulated by high each-step activation (ESA) of OXPHOS complexes. The contribution of proton leak to [Formula: see text] and ATP/O2 ratio in isolated mitochondria should not be directly extrapolated to working muscle, as mitochondria lack ESA, at least in the absence of Ca2+, and therefore [Formula: see text] cannot be elevated as much as in intact muscle.

Ethanol Dose- and Time-dependently Increases α and β Subunits of Mitochondrial ATP Synthase of Cultured Neonatal Rat Cardiomyocytes.

PubMed

Mashimo, Keiko; Arthur, Peter G; Ohno, Youkichi

2015-01-01

Mitochondria are target subcellular organelles of ethanol. In this study, the effects of ethanol on protein composition was examined with 2-dimensional electrophoresis of protein extracts from cultured neonatal rat cardiomyocytes exposed to 100 mM ethanol for 24 hours. A putative β subunit of mitochondrial ATP synthase was increased, which was confirmed by Western blot. The cellular protein abundances in the α and β subunits of ATP synthase increased in dose (0, 10, 50, and 100 mM) - and time (0.5 hour and 24 hours) -dependent manners. The DNA microarray analysis of total RNA extract demonstrated that gene expression of the corresponding messenger RNAs of these subunit proteins did not significantly alter due to 24-hour ethanol exposure. Therefore, protein expression of these nuclear-encoded mitochondrial proteins may be regulated at the translational, rather than the transcriptional, level. Alternatively, degradation of these subunit proteins might be decreased. Additionally, cellular ATP content of cardiomyocytes scarcely decreased following 24-hour exposure to any examined concentrations of ethanol. Previous studies, together with this study, have demonstrated that protein abundance of the α subunit or β subunit or both subunits of ATP synthase after ethanol exposure or dysfunctional conditions might differ according to tissue: significant increases in heart but decreases in liver and brain. Thus, it is suggested that the abundance of subunit proteins of mitochondrial ATP synthase in the ethanol-exposed heart, being different from that in the liver and brain, should increase dose-dependently through either translational upregulation or decreased degradation or both to maintain ATP production, as the heart requires much more energy than other tissues for continuing sustained contractions.

Fur-dependent detoxification of organic acids by rpoS mutants during prolonged incubation under aerobic, phosphate starvation conditions.

PubMed

Guillemet, Mélanie L; Moreau, Patrice L

2008-08-01

The activity of amino acid-dependent acid resistance systems allows Escherichia coli to survive during prolonged incubation under phosphate (P(i)) starvation conditions. We show in this work that rpoS-null mutants incubated in the absence of any amino acid survived during prolonged incubation under aerobic, P(i) starvation conditions. Whereas rpoS(+) cells incubated with glutamate excreted high levels of acetate, rpoS mutants grew on acetic acid. The characteristic metabolism of rpoS mutants required the activity of Fur (ferric uptake regulator) in order to decrease the synthesis of the small RNA RyhB that might otherwise inhibit the synthesis of iron-rich proteins. We propose that RpoS (sigma(S)) and the small RNA RyhB contribute to decrease the synthesis of iron-rich proteins required for the activity of the tricarboxylic acid (TCA) cycle, which redirects the metabolic flux toward the production of acetic acid at the onset of stationary phase in rpoS(+) cells. In contrast, Fur activity, which represses ryhB, and the lack of RpoS activity allow a substantial activity of the TCA cycle to continue in stationary phase in rpoS mutants, which decreases the production of acetic acid and, eventually, allows growth on acetic acid and P(i) excreted into the medium. These data may help explain the fact that a high frequency of E. coli rpoS mutants is found in nature.

H2O2 treatment or serum deprivation induces autophagy and apoptosis in naked mole-rat skin fibroblasts by inhibiting the PI3K/Akt signaling pathway.

PubMed

Zhao, Shanmin; Li, Li; Wang, Shiyong; Yu, Chenlin; Xiao, Bang; Lin, Lifang; Cong, Wei; Cheng, Jishuai; Yang, Wenjing; Sun, Wei; Cui, Shufang

2016-12-20

Naked mole-rats (NMR; Heterocephalus glaber) display extreme longevity and resistance to cancer. Here, we examined whether autophagy contributes to the longevity of NMRs by assessing the effects of the PI3K/Akt pathway inhibitor LY294002 and the autophagy inhibitor chloroquine (CQ) on autophagy and apoptosis in NMR skin fibroblasts. Serum starvation, H2O2 treatment, and LY294002 treatment all increased the LC3-II/LC3-I ratio and numbers of double-membraned autophagosomes and autophagic vacuoles, and decreased levels of p70S6K, p-AktSer473, and p-AktThr308. By contrast, CQ treatment decreased p70S6K, AktSer473, and AktThr308 levels. The Bax/Bcl-2 ratio increased after 12 h of exposure to LY294002 or CQ. These data show that inhibiting the Akt pathway promotes autophagy and apoptosis in NMR skin fibroblasts. Furthermore, LY294002 or CQ treatment decreased caspase-3, p53, and HIF1-α levels, suggesting that serum starvation or H2O2 treatment increase autophagy and apoptosis in NMR skin fibroblasts by inhibiting the PI3K/Akt pathway. CQ-induced inhibition of late autophagy stages also prevented Akt activation and induced apoptosis. Finally, the HIF-1α and p53 pathways were involved in serum starvation- or H2O2-induced autophagy in NMR skin fibroblasts.

Comparative hemodynamic effects of hypotension induced by diadenosine tetraphosphate (AP4A) and ATP in dogs.

PubMed

Takeda, Shohei; Inada, Yutaka; Fukui, Noriyuki; Tomaru, Teruaki

1997-03-01

ATP and diadenosine tetraphosphate (AP 4 A) have been shown to produce vasodilation mediated by P 1 - and P 2 -purinoceptor, respectively. The differing mechanisms involved in this vasodilating activity may induce different systemic hemodynamic changes. We compared the hemodynamic effects of AP 4 A-induced hypotension with those induced by ATP. Fourteen mongrel dogs were anesthetized with 0.87% halothane in oxygen (1 MAC). After the baseline period, mean arterial pressure was reduced to 60 mmHg for 60 min by the infusion of AP 4 A or ATP. The ATP- and AP 4 A-induced hypotension resulted in a maximum reduction in systemic vascular resistance of 43% and 46%, respectively (P<0.01), associated with a significant increase in stroke volume index. With ATP, a 20% of maximum increase (P<0.05) in cardiac index (CI) was observed during the induced hypotension. In contrast, AP 4 A-induced hypotension did not result in any changes in CI throughout the observation period. The varying results concerning CI during the ATP- and AP 4 A-induced hypotension were probably due to differences in ventricular filling pressure, since AP 4 A-induced hypotension was associated with decreases (P<0.01) in both right atrial and pulmonary capillary wedge pressures, whereas neither of these variables significantly changed with ATP. The hypotension induced by either ATP or AP 4 A was associated with a significant decrease in heart rate (HR). However, both the magnitude and duration of decreases in HR due to ATP-induced hypotension were more pronounced than those seen with AP 4 A. In conclusion, while both drugs were equally capable of inducing hypotension, our results suggest that AP 4 A was more suitable for induced hypotension because of its potent vasodilatory action with venodilation and less negative chronotropic action.

N-Acetylcysteine-induced vasodilatation is modulated by KATP channels, Na+/K+-ATPase activity and intracellular calcium concentration: An in vitro study.

PubMed

Vezir, Özden; Çömelekoğlu, Ülkü; Sucu, Nehir; Yalın, Ali Erdinç; Yılmaz, Şakir Necat; Yalın, Serap; Söğüt, Fatma; Yaman, Selma; Kibar, Kezban; Akkapulu, Merih; Koç, Meryem İlkay; Seçer, Didem

2017-08-01

In this study, we aimed to investigate the role of ATP-sensitive potassium (K ATP ) channel, Na + /K + -ATPase activity, and intracellular calcium levels on the vasodilatory effect of N-acetylcysteine (NAC) in thoracic aorta by using electrophysiological and molecular techniques. Rat thoracic aorta ring preparations and cultured thoracic aorta cells were divided into four groups as control, 2mM NAC, 5mM NAC, and 10mM NAC. Thoracic aorta rings were isolated from rats for measurements of relaxation responses and Na + /K + -ATPase activity. In the cultured thoracic aorta cells, we measured the currents of K ATP channel, the concentration of intracellular calcium and mRNA expression level of K ATP channel subunits (KCNJ8, KCNJ11, ABCC8 and ABCC9). The relaxation rate significantly increased in all NAC groups compared to control. Similarly, Na + /K + - ATPase activity also significantly decreased in NAC groups. Outward K ATP channel current significantly increased in all NAC groups compared to the control group. Intracellular calcium concentration decreased significantly in all groups with compared control. mRNA expression level of ABCC8 subunit significantly increased in all NAC groups compared to the control group. Pearson correlation analysis showed that relaxation rate was significantly associated with K ATP current, intracellular calcium concentration, Na + /K + -ATPase activity and mRNA expression level of ABCC8 subunit. Our findings suggest that NAC relaxes vascular smooth muscle cells through a direct effect on K ATP channels, by increasing outward K+ flux, partly by increasing mRNA expression of K ATP subunit ABCC8, by decreasing in intracellular calcium and by decreasing in Na + /K + -ATPase activity. Copyright © 2017 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

Copper Transporter ATP7A Protects Against Endothelial Dysfunction in Type 1 Diabetic Mice by Regulating Extracellular Superoxide Dismutase

PubMed Central

Sudhahar, Varadarajan; Urao, Norifumi; Oshikawa, Jin; McKinney, Ronald D.; Llanos, Roxana M.; Mercer, Julian F.B.; Ushio-Fukai, Masuko; Fukai, Tohru

2013-01-01

Oxidative stress and endothelial dysfunction contribute to vascular complication in diabetes. Extracellular superoxide dismutase (SOD3) is one of the key antioxidant enzymes that obtains copper via copper transporter ATP7A. SOD3 is secreted from vascular smooth muscles cells (VSMCs) and anchors at the endothelial surface. The role of SOD3 and ATP7A in endothelial dysfunction in type 1 diabetes mellitus (T1DM) is entirely unknown. Here we show that the specific activity of SOD3, but not SOD1, is decreased, which is associated with increased O2•− production in aortas of streptozotocin-induced and genetically induced Ins2Akita T1DM mice. Exogenous copper partially rescued SOD3 activity in isolated T1DM vessels. Functionally, acetylcholine-induced, endothelium-dependent relaxation is impaired in T1DM mesenteric arteries, which is rescued by SOD mimetic tempol or gene transfer of SOD3. Mechanistically, ATP7A expression in T1DM vessels is dramatically decreased whereas other copper transport proteins are not altered. T1DM-induced endothelial dysfunction and decrease of SOD3 activity are rescued in transgenic mice overexpressing ATP7A. Furthermore, SOD3-deficient T1DM mice or ATP7A mutant T1DM mice augment endothelial dysfunction and vascular O2•− production versus T1DM mice. These effects are in part due to hypoinsulinemia in T1DM mice, since insulin treatment, but not high glucose, increases ATP7A expression in VSMCs and restores SOD3 activity in the organoid culture of T1DM vessels. In summary, a decrease in ATP7A protein expression contributes to impaired SOD3 activity, resulting in O2•− overproduction and endothelial dysfunction in blood vessels of T1DM. Thus, restoring copper transporter function is an essential therapeutic approach for oxidant stress–dependent vascular and metabolic diseases. PMID:23884884

Treatment of heterotopic ossification through remote ATP hydrolysis.

PubMed

Peterson, Jonathan R; De La Rosa, Sara; Eboda, Oluwatobi; Cilwa, Katherine E; Agarwal, Shailesh; Buchman, Steven R; Cederna, Paul S; Xi, Chuanwu; Morris, Michael D; Herndon, David N; Xiao, Wenzhong; Tompkins, Ronald G; Krebsbach, Paul H; Wang, Stewart C; Levi, Benjamin

2014-09-24

Heterotopic ossification (HO) is the pathologic development of ectopic bone in soft tissues because of a local or systemic inflammatory insult, such as burn injury or trauma. In HO, mesenchymal stem cells (MSCs) are inappropriately activated to undergo osteogenic differentiation. Through the correlation of in vitro assays and in vivo studies (dorsal scald burn with Achilles tenotomy), we have shown that burn injury enhances the osteogenic potential of MSCs and causes ectopic endochondral heterotopic bone formation and functional contractures through bone morphogenetic protein-mediated canonical SMAD signaling. We further demonstrated a prevention strategy for HO through adenosine triphosphate (ATP) hydrolysis at the burn site using apyrase. Burn site apyrase treatment decreased ATP, increased adenosine 3',5'-monophosphate, and decreased phosphorylation of SMAD1/5/8 in MSCs in vitro. This ATP hydrolysis also decreased HO formation and mitigated functional impairment in vivo. Similarly, selective inhibition of SMAD1/5/8 phosphorylation with LDN-193189 decreased HO formation and increased range of motion at the injury site in our burn model in vivo. Our results suggest that burn injury-exacerbated HO formation can be treated through therapeutics that target burn site ATP hydrolysis and modulation of SMAD1/5/8 phosphorylation. Copyright © 2014, American Association for the Advancement of Science.

A novel aptasensor for the ultra-sensitive detection of adenosine triphosphate via aptamer/quantum dot based resonance energy transfer.

PubMed

Li, Zheng; Wang, Yijing; Liu, Ying; Zeng, Yongyi; Huang, Aimin; Peng, Niancai; Liu, Xiaolong; Liu, Jingfeng

2013-09-07

We designed a novel aptamer based biosensor (aptasensor) for ultrasensitive detection of adenosine triphosphate (ATP) through resonance energy transfer (RET). The ATP aptamer was modified with Cy3 at the 3' end, and a green quantum dot (525) was attached to the 5' end of its complementary sequence respectively. The ATP aptamer and its complementary sequence could assemble into a duplex structure in the absence of target ATP, and then decrease the distance between the quantum dot and Cy3 which could produce significant RET signal. Upon ATP binding, the ATP aptamer could dissociate with its complementary sequence and then increase the distance between the quantum dot and Cy3 which would significantly decrease the RET signal. Therefore, the ATP detection could be easily achieved through detection of the fluorescence intensity ratio between 525 nm and 560 nm. The results show that the emission fluorescence intensity ratio of 525/560 is linearly related to the logarithmic concentration of ATP. The linear range of this aptasensor is from 0.1 nM to 1 μM, and the detection limit is lower down to 0.01 nM. Excellent selectivity of this aptasensor for ATP has been demonstrated through the detection of thymidine triphosphate (TTP), cytidine triphosphate (CTP), guanosine triphosphate (GTP) and adenosine diphosphate (ADP) respectively as control. The method we described here could easily detect ATP with excellent selectivity, linearity and sensitivity down to the nanomolar range, as well as avoid photobleaching.

The active transport of histidine and its role in ATP production in Trypanosoma cruzi.

PubMed

Barisón, M J; Damasceno, F S; Mantilla, B S; Silber, A M

2016-08-01

Trypanosoma cruzi, the aetiological agent of Chagas's disease, metabolizes glucose, and after its exhaustion, degrades amino acids as energy source. Here, we investigate histidine uptake and its participation in energy metabolism. No putative genes for the histidine biosynthetic pathway have been identified in genome databases of T. cruzi, suggesting that its uptake from extracellular medium is a requirement for the viability of the parasite. From this assumption, we characterized the uptake of histidine in T. cruzi, showing that this amino acid is incorporated through a single and saturable active system. We also show that histidine can be completely oxidised to CO2. This finding, together with the fact that genes encoding the putative enzymes for the histidine - glutamate degradation pathway were annotated, led us to infer its participation in the energy metabolism of the parasite. Here, we show that His is capable of restoring cell viability after long-term starvation. We confirm that as an energy source, His provides electrons to the electron transport chain, maintaining mitochondrial inner membrane potential and O2 consumption in a very efficient manner. Additionally, ATP biosynthesis from oxidative phosphorylation was found when His was the only oxidisable metabolite present, showing that this amino acid is involved in bioenergetics and parasite persistence within its invertebrate host.

Activation of an ATP-dependent K(+) conductance in Xenopus oocytes by expression of adenylate kinase cloned from renal proximal tubules.

PubMed

Brochiero, E; Coady, M J; Klein, H; Laprade, R; Lapointe, J Y

2001-02-09

In rabbit proximal convoluted tubules, an ATP-sensitive K(+) (K(ATP)) channel has been shown to be involved in membrane cross-talk, i.e. the coupling (most likely mediated through intracellular ATP) between transepithelial Na(+) transport and basolateral K(+) conductance. This K(+) conductance is inhibited by taurine. We sought to isolate this K(+) channel by expression cloning in Xenopus oocytes. Injection of renal cortex mRNA into oocytes induced a K(+) conductance, largely inhibited by extracellular Ba(2+) and intracellular taurine. Using this functional test, we isolated from our proximal tubule cDNA library a unique clone, which induced a large K(+) current which was Ba(2+)-, taurine- and glibenclamide-sensitive. Surprisingly, this clone is not a K(+) channel but an adenylate kinase protein (AK3), known to convert NTP+AMP into NDP+ADP (N could be G, I or A). AK3 expression resulted in a large ATP decrease and activation of the whole-cell currents including a previously unknown, endogenous K(+) current. To verify whether ATP decrease was responsible for the current activation, we demonstrated that inhibition of glycolysis greatly reduces oocyte ATP levels and increases an inwardly rectifying K(+) current. The possible involvement of AK in the K(ATP) channel's regulation provides a means of explaining their observed activity in cytosolic environments characterized by high ATP concentrations.

Food deprivation modulates gamma-aminobutyric acid receptors and peripheral benzodiazepine binding sites in rats.

PubMed

Weizman, A; Bidder, M; Fares, F; Gavish, M

1990-12-03

The effect of 5 days of food deprivation followed by 5 days of refeeding on gamma-aminobutyric acid (GABA) receptors, central benzodiazepine receptors (CBR), and peripheral benzodiazepine binding sites (PBzS) was studied in female Sprague-Dawley rats. Starvation induced a decrease in the density of PBzS in peripheral organs: adrenal (35%; P less than 0.001), kidney (33%; P less than 0.01), and heart (34%; P less than 0.001). Restoration of [3H]PK 11195 binding to normal values was observed in all three organs after 5 days of refeeding. The density of PBzS in the ovary, pituitary, and hypothalamus was not affected by starvation. Food deprivation resulted in a 35% decrease in cerebellar GABA receptors (P less than 0.01), while CBR in the hypothalamus and cerebral cortex remained unaltered. The changes in PBzS observed in the heart and kidney may be related to the long-term metabolic stress associated with starvation and to the functional changes occurring in these organs. The down-regulation of the adrenal PBzS is attributable to the suppressive effect of hypercortisolemia on pituitary ACTH release. The reduction in cerebellar GABA receptors may be an adaptive response to food deprivation stress and may be relevant to the proaggressive effect of hunger.

Modeling regulation of cardiac KATP and L-type Ca2+ currents by ATP, ADP, and Mg2+.

PubMed

Michailova, Anushka; Saucerman, Jeffrey; Belik, Mary Ellen; McCulloch, Andrew D

2005-03-01

Changes in cytosolic free Mg(2+) and adenosine nucleotide phosphates affect cardiac excitability and contractility. To investigate how modulation by Mg(2+), ATP, and ADP of K(ATP) and L-type Ca(2+) channels influences excitation-contraction coupling, we incorporated equations for intracellular ATP and MgADP regulation of the K(ATP) current and MgATP regulation of the L-type Ca(2+) current in an ionic-metabolic model of the canine ventricular myocyte. The new model: 1), quantitatively reproduces a dose-response relationship for the effects of changes in ATP on K(ATP) current, 2), simulates effects of ADP in modulating ATP sensitivity of K(ATP) channel, 3), predicts activation of Ca(2+) current during rapid increase in MgATP, and 4), demonstrates that decreased ATP/ADP ratio with normal total Mg(2+) or increased free Mg(2+) with normal ATP and ADP activate K(ATP) current, shorten action potential, and alter ionic currents and intracellular Ca(2+) signals. The model predictions are in agreement with experimental data measured under normal and a variety of pathological conditions.

Modeling regulation of cardiac KATP and L-type Ca2+ currents by ATP, ADP, and Mg2+

NASA Technical Reports Server (NTRS)

Michailova, Anushka; Saucerman, Jeffrey; Belik, Mary Ellen; McCulloch, Andrew D.

2005-01-01

Changes in cytosolic free Mg(2+) and adenosine nucleotide phosphates affect cardiac excitability and contractility. To investigate how modulation by Mg(2+), ATP, and ADP of K(ATP) and L-type Ca(2+) channels influences excitation-contraction coupling, we incorporated equations for intracellular ATP and MgADP regulation of the K(ATP) current and MgATP regulation of the L-type Ca(2+) current in an ionic-metabolic model of the canine ventricular myocyte. The new model: 1), quantitatively reproduces a dose-response relationship for the effects of changes in ATP on K(ATP) current, 2), simulates effects of ADP in modulating ATP sensitivity of K(ATP) channel, 3), predicts activation of Ca(2+) current during rapid increase in MgATP, and 4), demonstrates that decreased ATP/ADP ratio with normal total Mg(2+) or increased free Mg(2+) with normal ATP and ADP activate K(ATP) current, shorten action potential, and alter ionic currents and intracellular Ca(2+) signals. The model predictions are in agreement with experimental data measured under normal and a variety of pathological conditions.

Failure of ATP supply to match ATP demand: the mechanism of toxicity of the lampricide, 3-trifluoromethyl-4-nitrophenol (TFM), used to control sea lamprey (Petromyzon marinus) populations in the Great Lakes.

PubMed

Birceanu, Oana; McClelland, Grant B; Wang, Yuxiang S; Wilkie, Michael P

2009-10-04

Although the pesticide, 3-trifluoromethyl-4-nitrophenol (TFM), has been extensively used to control invasive sea lamprey (Petromyzon marinus) populations in the Great Lakes, it is surprising that its mechanism(s) of toxicity is unresolved. A better knowledge of the mode of toxicity of this pesticide is needed for predicting and improving the effectiveness of TFM treatments on lamprey, and for risk assessments regarding potential adverse effects on invertebrate and vertebrate non-target organisms. We investigated two hypotheses of TFM toxicity in larval sea lamprey. The first was that TFM interferes with oxidative ATP production by mitochondria, causing rapid depletion of energy stores in vital, metabolically active tissues such as the liver and brain. The second was that TFM toxicity resulted from disruption of gill-ion uptake, adversely affecting ion homeostasis. Exposure of larval sea lamprey to 4.6 m gl(-1) TFM (12-h LC50) caused glycogen concentrations in the brain to decrease by 80% after 12h, suggesting that the animals increased their reliance on glycolysis to generate ATP due to a shortfall in ATP supply. This conclusion was reinforced by a 9-fold increase in brain lactate concentration, a 30% decrease in brain ATP concentration, and an 80% decrease in phosphocreatine (PCr) concentration after 9 and 12h. A more pronounced trend was noted in the liver, where glycogen decreased by 85% and ATP was no longer detected after 9 and 12h. TFM led to marginal changes in whole body Na(+), Cl(-), Ca(2+) and K(+), as well as in plasma Na(+) and Cl(-), which were unlikely to have contributed to toxicity. TFM had no adverse effect on Na(+) uptake rates or gill Na(+)/K(+)-ATPase activity. We conclude that TFM toxicity in the sea lamprey is due to a mismatch between ATP consumption and ATP production rates, leading to a depletion of glycogen in the liver and brain, which ultimately leads to neural arrest and death.

The effect of severe starvation and captivity stress on plasma thyroxine and triiodothyronine concentrations in an antarctic bird (emperor penguin).

PubMed

Groscolas, R; Leloup, J

1989-01-01

The effect of confinement and severe starvation on the plasma thyroxine (T4) and triiodothyronine (T3) concentrations was determined in emperor penguins (Aptenodytes forsteri). During their annual cycle, emperor penguins fast freely for periods of up to 4 months and may thus represent a unique subject to study endocrine adaptations to fasting. Plasma T4 concentrations progressively decreased following capture and confinement of naturally fasting penguins, and within 15-20 days stabilized at levels three times lower than in free-living penguins. A transient fourfold increase in plasma T3 concentration developed within the day following confinement in parallel with a rise in daily body mass loss. Both plasma T3 concentration and mass loss subsided to normal levels within 15 days. The decrease in plasma T4 concentration is in accordance with the well-known inhibitory effect of stress on thyroid function in birds and mammals, whereas the transient increase in plasma T3 concentration seems related to enhancement of energy expenditure as a consequence of restlessness. Starvation severe enough to exhaust fat stores and to activate protein catabolism induced a 6- and 5 to 10-fold fall in plasma T4 and T3, respectively. This is in marked contrast with maintenance of plasma thyroid levels during long-term natural fasting associated with protein sparing (R. Groscolas and J. Leloup (1986) Gen. Comp. Endocrinol. 63, 264-274). Surprisingly, there was a final reincrease in plasma T4 concentration in very lean penguins. These results suggest that the effect of starvation on plasma thyroid hormones seems to depend on how much protein catabolism is activated and demonstrate the acute sensitivity of thyroid hormone balance to stress in penguins.

Short-term starvation and realimentation helps stave off Edwardsiella tarda infection in red sea bream (Pagrus major).

PubMed

Mohapatra, Sipra; Chakraborty, Tapas; Reza, Mohammad Ali Noman; Shimizu, Sonoko; Matsubara, Takahiro; Ohta, Kohei

2017-04-01

Dietary regime modifications have been an integral part of health and healing practices throughout the animal kingdom. Thus, to assess the effects of periodic starvation and refeeding schedule on the physiological and immunological perturbations in Edwardsiella tarda infected red sea bream, we conducted a 20day experiment using 4 treatment groups, namely, pre-fed placebo (PFP); pre-starved placebo (PSP); pre-fed infected (PFI); and pre-starved infected (PSI), wherein a 5h E. tarda infection was done on the 11th day. In the present investigation, the pre-starved groups showed significant (P<0.05) alterations in the liver Hexokinase and Glucose-6-phosphatase activity. The pre-starved fish also exhibited significant (P<0.05) increment in the hepatosomatic index, along with increased hepatic glycogen content, in a time dependent fashion. The PPAR (peroxisome proliferator activated receptors)α transcription in the pre-starved group decreased significantly (P<0.05) by 10dai, while the PPARγ showcased a reverse pattern. The transcription of Hepcidin1 and Transferrin (iron homeostasis related genes), and Cathepsin D and Ubiquitin (programmed cell death related genes) portrayed a time responsive decrease and increase in PSI and PFI groups, respectively. Additionally, in comparison to the PFI group, the PSI fish demonstrated substantially reduced oxidative stress level. Fluorescent Immunohistochemistry showed significant (P<0.05) increase in p63 positive cells in the 10dai PFI fish in relation to the PSI group. Therefore, these findings provide new insight into the beneficial role of alternating starvation and refeeding schedule, preferably short-term starvation prior to an infection, in order to obtain better capability to battle against E. tarda infection in red sea bream. Copyright © 2017 Elsevier Inc. All rights reserved.

Influence of Starvation on Potential Ammonia-Oxidizing Activity and amoA mRNA Levels of Nitrosospira briensis

PubMed Central

Bollmann, Annette; Schmidt, Ingo; Saunders, Aaron M.; Nicolaisen, Mette H.

2005-01-01

The effect of short-term ammonia starvation on Nitrosospira briensis was investigated. The ammonia-oxidizing activity was determined in a concentrated cell suspension with a NOx biosensor. The apparent half-saturation constant [Km(app)] value of the NH3 oxidation of N. briensis was 3 μM NH3 for cultures grown both in continuous and batch cultures as determined by a NOx biosensor. Cells grown on the wall of the vessel had a lower Km(app) value of 1.8 μM NH3. Nonstarving cultures of N. briensis showed potential ammonia-oxidizing activities of between 200 to 250 μM N h−1, and this activity decreased only slowly during starvation up to 10 days. Within 10 min after the addition of fresh NH4+, 100% activity was regained. Parallel with activity measurements, amoA mRNA and 16S rRNA were investigated. No changes were observed in the 16S rRNA, but a relative decrease of amoA mRNA was observed during the starvation period. During resuscitation, an increase in amoA mRNA expression was detected simultaneously. The patterns of the soluble protein fraction of a 2-week-starved culture of N. briensis showed only small differences in comparison to a nonstarved control. From these results we conclude that N. briensis cells remain in a state allowing fast recovery of ammonia-oxidizing activity after addition of NH4+ to a starved culture. Maintaining cells in this kind of active state could be the survival strategy of ammonia-oxidizing bacteria in nature under fluctuating NH4+ availability. PMID:15746329

Halophilic Archaea: Life with Desiccation, Radiation and Oligotrophy over Geological Times

PubMed Central

Stan-Lotter, Helga; Fendrihan, Sergiu

2015-01-01

Halophilic archaebacteria (Haloarchaea) can survive extreme desiccation, starvation and radiation, sometimes apparently for millions of years. Several of the strategies that are involved appear specific for Haloarchaea (for example, the formation of halomucin, survival in fluid inclusions of halite), and some are known from other prokaryotes (dwarfing of cells, reduction of ATP). Several newly-discovered haloarchaeal strategies that were inferred to possibly promote long-term survival—halomucin, polyploidy, usage of DNA as a phosphate storage polymer, production of spherical dormant stages—remain to be characterized in detail. More information on potential strategies is desirable, since evidence for the presence of halite on Mars and on several moons in the solar system increased interest in halophiles with respect to the search for extraterrestrial life. This review deals in particular with novel findings and hypotheses on haloarchaeal long-term survival. PMID:26226005

Halophilic Archaea: Life with Desiccation, Radiation and Oligotrophy over Geological Times.

PubMed

Stan-Lotter, Helga; Fendrihan, Sergiu

2015-07-28

Halophilic archaebacteria (Haloarchaea) can survive extreme desiccation, starvation and radiation, sometimes apparently for millions of years. Several of the strategies that are involved appear specific for Haloarchaea (for example, the formation of halomucin, survival in fluid inclusions of halite), and some are known from other prokaryotes (dwarfing of cells, reduction of ATP). Several newly-discovered haloarchaeal strategies that were inferred to possibly promote long-term survival-halomucin, polyploidy, usage of DNA as a phosphate storage polymer, production of spherical dormant stages-remain to be characterized in detail. More information on potential strategies is desirable, since evidence for the presence of halite on Mars and on several moons in the solar system increased interest in halophiles with respect to the search for extraterrestrial life. This review deals in particular with novel findings and hypotheses on haloarchaeal long-term survival.

Leptin does not mediate short-term fasting-induced changes in growth hormone pulsatility but increases IGF-I in leptin deficiency states.

PubMed

Chan, Jean L; Williams, Catherine J; Raciti, Patricia; Blakeman, Jennifer; Kelesidis, Theodore; Kelesidis, Iosif; Johnson, Michael L; Thorner, Michael O; Mantzoros, Christos S

2008-07-01

States of acute and chronic energy deficit are characterized by increased GH secretion and decreased IGF-I levels. The objective of the study was to determine whether changes in levels of leptin, a key mediator of the adaptation to starvation, regulate the GH-IGF system during energy deficit. We studied 14 healthy normal-weight men and women during three conditions: baseline fed and 72-h fasting (to induce hypoleptinemia) with administration of placebo or recombinant methionyl human leptin (r-metHuLeptin) (to reverse the fasting associated hypoleptinemia). We also studied eight normal-weight women with exercise-induced chronic energy deficit and hypothalamic amenorrhea at baseline and during 2-3 months of r-metHuLeptin treatment. GH pulsatility, IGF levels, IGF and GH binding protein (GHBP) levels were measured. During short-term energy deficit, measures of GH pulsatility and disorderliness and levels of IGF binding protein (IGFBP)-1 increased, whereas leptin, insulin, IGF-I (total and free), IGFBP-4, IGFBP-6, and GHBP decreased; r-metHuLeptin administration blunted the starvation-associated decrease of IGF-I. In chronic energy deficit, total and free IGF-I, IGFBP-6, and GHBP levels were lower, compared with euleptinemic controls; r-metHuLeptin administration had no major effect on GH pulsatility after 2 wk but increased total IGF-I levels and tended to increase free IGF-I and IGFBP-3 after 1 month. The GH/IGF system changes associated with energy deficit are largely independent of leptin deficiency. During acute energy deficit, r-metHuLeptin administration in replacement doses blunts the starvation-induced decrease of IGF-I, but during chronic energy deficit, r-metHuLeptin administration increases IGF-I and tends to increase free IGF-I and IGFBP-3.

ATP depletion during mitotic arrest induces mitotic slippage and APC/CCdh1-dependent cyclin B1 degradation.

PubMed

Park, Yun Yeon; Ahn, Ju-Hyun; Cho, Min-Guk; Lee, Jae-Ho

2018-04-27

ATP depletion inhibits cell cycle progression, especially during the G1 phase and the G2 to M transition. However, the effect of ATP depletion on mitotic progression remains unclear. We observed that the reduction of ATP after prometaphase by simultaneous treatment with 2-deoxyglucose and NaN 3 did not arrest mitotic progression. Interestingly, ATP depletion during nocodazole-induced prometaphase arrest resulted in mitotic slippage, as indicated by a reduction in mitotic cells, APC/C-dependent degradation of cyclin B1, increased cell attachment, and increased nuclear membrane reassembly. Additionally, cells successfully progressed through the cell cycle after mitotic slippage, as indicated by EdU incorporation and time-lapse imaging. Although degradation of cyclin B during normal mitotic progression is primarily regulated by APC/C Cdc20 , we observed an unexpected decrease in Cdc20 prior to degradation of cyclin B during mitotic slippage. This decrease in Cdc20 was followed by a change in the binding partner preference of APC/C from Cdc20 to Cdh1; consequently, APC/C Cdh1 , but not APC/C Cdc20 , facilitated cyclin B degradation following ATP depletion. Pulse-chase analysis revealed that ATP depletion significantly abrogated global translation, including the translation of Cdc20 and Cdh1. Additionally, the half-life of Cdh1 was much longer than that of Cdc20. These data suggest that ATP depletion during mitotic arrest induces mitotic slippage facilitated by APC/C Cdh1 -dependent cyclin B degradation, which follows a decrease in Cdc20 resulting from reduced global translation and the differences in the half-lives of the Cdc20 and Cdh1 proteins.

An ATP-gated cation channel with some P2Z-like characteristics in gastric smooth muscle cells of toad.

PubMed Central

Ugur, M; Drummond, R M; Zou, H; Sheng, P; Singer, J J; Walsh, J V

1997-01-01

1. Whole-cell and single-channel currents elicited by extracellular ATP were studied in freshly dissociated smooth muscle cells from the stomach of the toad Bufo marinus using standard patch clamp and microfluorimetric techniques. 2. This ATP-gated cation channel shares a number of pharmacological and functional properties with native rat myometrium receptors, certain native P2Z purinoceptors and the recently cloned P2X7 purinoceptor. But, unlike the last two, the ATP-gated channel does not mediate the formation of large non-specific pores. Thus, it may represent a novel member of the P2X or P2Z class. 3. Extracellular application of ATP (> or = 150 microM) elicited an inward whole-cell current at negative holding potentials that was inwardly rectifying and showed no sign of desensitization. Na+, Cs+ and, to a lesser degree, the organic cation choline served as charge carriers, but Cl- did not. Ratiometric fura-2 measurements indicated that the current is carried in part by Ca2+. The EC50 for ATP was 700 microM in solutions with a low divalent cation concentration. 4. ATP (> or = 100 microM) at the extracellular surface of cell-attached or excised patches elicited inwardly rectifying single-channel currents with a 22 pS conductance. Cl- did not serve as a charge carrier but both Na+ and Cs+ did, as did choline to a lesser extent. The mean open time of the channel was quite long, with a range in hundreds of milliseconds at a holding potential of -70 mV. 5. Mg2+ and Ca2+ decreased the magnitude of the ATP-induced whole-cell currents. Mg2+ decreased both the amplitude and the activity of ATP-activated single-channel currents. 6. ADP, UTP, P1, P5-di-adenosine pentaphosphate (AP5A), adenosine and alpha, beta-methylene ATP (alpha, beta-Me-ATP) did not induce significant whole-cell current. ATP-gamma-S and 2-methylthio ATP (2-Me-S-ATP) were significantly less effective than ATP in inducing whole-cell currents, whereas benzoylbenzoyl ATP (BzATP) was more effective. BzATP, alpha, beta-Me-ATP, ATP-gamma-S and 2-Me-S-ATP induced single-channel currents, but a higher concentration of alpha, beta-Me-ATP was required. 7. BzATP did not induce the formation of large non-specific pores, as assayed using mag-fura-2 as a high molecular mass probe. PMID:9032690

Controlled rotation of the F1-ATPase reveals differential and continuous binding changes for ATP synthesis

PubMed Central

Adachi, Kengo; Oiwa, Kazuhiro; Yoshida, Masasuke; Nishizaka, Takayuki; Kinosita, Kazuhiko

2012-01-01

F1-ATPase is an ATP-driven rotary molecular motor that synthesizes ATP when rotated in reverse. To elucidate the mechanism of ATP synthesis, we imaged binding and release of fluorescently labelled ADP and ATP while rotating the motor in either direction by magnets. Here we report the binding and release rates for each of the three catalytic sites for 360° of the rotary angle. We show that the rates do not significantly depend on the rotary direction, indicating ATP synthesis by direct reversal of the hydrolysis-driven rotation. ADP and ATP are discriminated in angle-dependent binding, but not in release. Phosphate blocks ATP binding at angles where ADP binding is essential for ATP synthesis. In synthesis rotation, the affinity for ADP increases by >104, followed by a shift to high ATP affinity, and finally the affinity for ATP decreases by >104. All these angular changes are gradual, implicating tight coupling between the rotor angle and site affinities. PMID:22929779

The Effects of Oxygen Level and Glucose Concentration on the Metabolism of Porcine TMJ Disc Cells

PubMed Central

Cisewski, Sarah E.; Zhang, Lixia; Kuo, Jonathan; Wright, Gregory J.; Wu, Yongren; Kern, Michael J.; Yao, Hai

2015-01-01

Objective To determine the combined effect of oxygen level and glucose concentration on cell viability, ATP production, and matrix synthesis of temporomandibular joint (TMJ) disc cells. Design TMJ disc cells were isolated from pigs aged 6-8 months and cultured in a monolayer. Cell cultures were preconditioned for 48 hours with 0, 1.5, 5, or 25mM glucose DMEM under 1%, 5%, 10%, or 21% O2 level, respectively. The cell viability was measured using the WST-1 assay. ATP production was determined using the Luciferin-Luciferase assay. Collagen and proteoglycan synthesis were determined by measuring the incorporation of [2, 3-3H]proline and [35S]sulfate into the cells, respectively. Results TMJ disc cell viability significantly decreased (P<0.0001) without glucose. With glucose present, decreased oxygen levels significantly increased viability (P<0.0001), while a decrease in glucose concentration significantly decreased viability (P<0.0001). With glucose present, decreasing oxygen levels significantly reduced ATP production (P<0.0001) and matrix synthesis (P<0.0001). A decreased glucose concentration significantly decreased collagen synthesis (P<0.0001). The interaction between glucose and oxygen was significant in regards to cell viability (P<0.0001), ATP production (P=0.00015), and collagen (P=0.0002) and proteoglycan synthesis (P<0.0001). Conclusions Although both glucose and oxygen are important, glucose is the limiting nutrient for TMJ disc cell survival. At low oxygen levels, the production of ATP, collagen, and proteoglycan are severely inhibited. These results suggest that steeper nutrient gradients may exist in the TMJ disc and it may be vulnerable to pathological events that impede nutrient supply. PMID:26033165

The effects of oxygen level and glucose concentration on the metabolism of porcine TMJ disc cells.

PubMed

Cisewski, S E; Zhang, L; Kuo, J; Wright, G J; Wu, Y; Kern, M J; Yao, H

2015-10-01

To determine the combined effect of oxygen level and glucose concentration on cell viability, ATP production, and matrix synthesis of temporomandibular joint (TMJ) disc cells. TMJ disc cells were isolated from pigs aged 6-8 months and cultured in a monolayer. Cell cultures were preconditioned for 48 h with 0, 1.5, 5, or 25 mM glucose DMEM under 1%, 5%, 10%, or 21% O2 level, respectively. The cell viability was measured using the WST-1 assay. ATP production was determined using the Luciferin-Luciferase assay. Collagen and proteoglycan synthesis were determined by measuring the incorporation of [2, 3-(3)H] proline and [(35)S] sulfate into the cells, respectively. TMJ disc cell viability significantly decreased (P < 0.0001) without glucose. With glucose present, decreased oxygen levels significantly increased viability (P < 0.0001), while a decrease in glucose concentration significantly decreased viability (P < 0.0001). With glucose present, decreasing oxygen levels significantly reduced ATP production (P < 0.0001) and matrix synthesis (P < 0.0001). A decreased glucose concentration significantly decreased collagen synthesis (P < 0.0001). The interaction between glucose and oxygen was significant in regards to cell viability (P < 0.0001), ATP production (P = 0.00015), and collagen (P = 0.0002) and proteoglycan synthesis (P < 0.0001). Although both glucose and oxygen are important, glucose is the limiting nutrient for TMJ disc cell survival. At low oxygen levels, the production of ATP, collagen, and proteoglycan are severely inhibited. These results suggest that steeper nutrient gradients may exist in the TMJ disc and it may be vulnerable to pathological events that impede nutrient supply. Copyright © 2015 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Gene dosage imbalance during DNA replication controls bacterial cell-fate decision

NASA Astrophysics Data System (ADS)

Igoshin, Oleg

Genes encoding proteins in a common regulatory network are frequently located close to one another on the chromosome to facilitate co-regulation or couple gene expression to growth rate. Contrasting with these observations, here we demonstrate a functional role for the arrangement of Bacillus subtilis sporulation network genes on opposite sides of the chromosome. We show that the arrangement of two sporulation network genes, one located close to the origin, the other close to the terminus leads to a transient gene dosage imbalance during chromosome replication. This imbalance is detected by the sporulation network to produce cell-cycle coordinated pulses of the sporulation master regulator Spo0A~P. This pulsed response allows cells to decide between sporulation and continued vegetative growth during each cell-cycle spent in starvation. Furthermore, changes in DNA replication and cell-cycle parameters with decreased growth rate in starvation conditions enable cells to indirectly detect starvation without the need for evaluating specific metabolites. The simplicity of the uncovered coordination mechanism and starvation sensing suggests that it may be widely applicable in a variety of gene regulatory and stress-response settings. This work is supported by National Science Foundation Grants MCB-1244135, EAGER-1450867, MCB-1244423, NIH NIGMS Grant R01 GM088428 and HHMI International Student Fellowship.

Modeling turbidity type and intensity effects on the growth and starvation mortality of age-0 yellow perch

USGS Publications Warehouse

Manning, Nathan M; Bossenbroek, Jonathan M.; Mayer, Christine M.; Bunnell, David B.; Tyson, Jeff T.; Rudstam, Lars G.; Jackson, James R.

2014-01-01

We sought to quantify the possible population-level influence of sediment plumes and algal blooms on yellow perch (Perca flavescens), a visual predator found in systems with dynamic water clarity. We used an individual-based model (IBM), which allowed us to include variance in water clarity and the distribution of individual sizes. Our IBM was built with laboratory data showing that larval yellow perch feeding rates increased slightly as sediment turbidity level increased, but that both larval and juvenile yellow perch feeding rates decreased as phytoplankton level increased. Our IBM explained a majority of the variance in yellow perch length in data from the western and central basins of Lake Erie and Oneida Lake, with R2 values ranging from 0.611 to 0.742. Starvation mortality was size dependent, as the greatest daily mortality rates in each simulation occurred within days of each other. Our model showed that turbidity-dependent consumption rates and temperature are key components in determining growth and starvation mortality of age-0 yellow perch, linking fish production to land-based processes that influence water clarity. These results suggest the timing and persistence of sediment plumes and algal blooms can drastically alter the growth potential and starvation mortality of a yellow perch cohort.

The significance of translation regulation in the stress response

PubMed Central

2013-01-01

Background The stress response in bacteria involves the multistage control of gene expression but is not entirely understood. To identify the translational response of bacteria in stress conditions and assess its contribution to the regulation of gene expression, the translational states of all mRNAs were compared under optimal growth condition and during nutrient (isoleucine) starvation. Results A genome-scale study of the translational response to nutritional limitation was performed in the model bacterium Lactococcus lactis. Two measures were used to assess the translational status of each individual mRNA: the fraction engaged in translation (ribosome occupancy) and ribosome density (number of ribosomes per 100 nucleotides). Under isoleucine starvation, half of the mRNAs considered were translationally down-regulated mainly due to decreased ribosome density. This pattern concerned genes involved in growth-related functions such as translation, transcription, and the metabolism of fatty acids, phospholipids and bases, contributing to the slowdown of growth. Only 4% of the mRNAs were translationally up-regulated, mostly related to prophagic expression in response to stress. The remaining genes exhibited antagonistic regulations of the two markers of translation. Ribosome occupancy increased significantly for all the genes involved in the biosynthesis of isoleucine, although their ribosome density had decreased. The results revealed complex translational regulation of this pathway, essential to cope with isoleucine starvation. To elucidate the regulation of global gene expression more generally, translational regulation was compared to transcriptional regulation under isoleucine starvation and to other post-transcriptional regulations related to mRNA degradation and mRNA dilution by growth. Translational regulation appeared to accentuate the effects of transcriptional changes for down-regulated growth-related functions under isoleucine starvation although mRNA stabilization and lower dilution by growth counterbalanced this effect. Conclusions We show that the contribution of translational regulation to the control of gene expression is significant in the stress response. Post-transcriptional regulation is complex and not systematically co-directional with transcription regulation. Post-transcriptional regulation is important to the understanding of gene expression control. PMID:23985063

Lipid droplets accumulation and other biochemical changes induced in the fungal pathogen Ustilago maydis under nitrogen-starvation.

PubMed

Aguilar, Lucero Romero; Pardo, Juan Pablo; Lomelí, Mónica Montero; Bocardo, Oscar Ivan Luqueño; Juárez Oropeza, Marco A; Guerra Sánchez, Guadalupe

2017-10-01

In many organisms, the growth under nitrogen-deprivation or a poor nitrogen source impacts on the carbon flow distribution and causes accumulation of neutral lipids, which are stored as lipid droplets (LDs). Efforts are in progress to find the mechanism of LDs synthesis and degradation, and new organisms capable of accumulating large amounts of lipids for biotechnological applications. In this context, when Ustilago maydis was cultured in the absence of a nitrogen source, there was a large accumulation of lipid bodies containing mainly triacylglycerols. The most abundant fatty acids in lipid bodies at the stationary phase were palmitic, linoleic, and oleic acids, and they were synthesized de novo by the fatty-acid synthase. In regard to the production of NADPH for the synthesis of fatty acids, the cytosolic NADP + -dependent isocitrate dehydrogenase and the glucose-6-phosphate and 6-phosphogluconate dehydrogenases couple showed the highest specific activities, with a lower activity of the malic enzyme. The ATP-citrate lyase activity was not detected in any of the culture conditions, which points to a different mechanism for the transfer of acetyl-CoA into the cytosol. Protein and RNA contents decreased when U. maydis was grown without a nitrogen source. Due to the significant accumulation of triacylglycerols and the particular composition of fatty acids, U. maydis can be considered an alternative model for biotechnological applications.

Bioanalytical Applications of Real-Time ATP Imaging Via Bioluminescence

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gruenhagen, Jason Alan

The research discussed within involves the development of novel applications of real-time imaging of adenosine 5'-triphosphate (ATP). ATP was detected via bioluminescence and the firefly luciferase-catalyzed reaction of ATP and luciferin. The use of a microscope and an imaging detector allowed for spatially resolved quantitation of ATP release. Employing this method, applications in both biological and chemical systems were developed. First, the mechanism by which the compound 48/80 induces release of ATP from human umbilical vein endothelial cells (HUVECs) was investigated. Numerous enzyme activators and inhibitors were utilized to probe the second messenger systems involved in release. Compound 48/80 activatedmore » a G{sub q}-type protein to initiate ATP release from HUVECs. Ca 2+ imaging along with ATP imaging revealed that activation of phospholipase C and induction of intracellular Ca 2+ signaling were necessary for release of ATP. Furthermore, activation of protein kinase C inhibited the activity of phospholipase C and thus decreased the magnitude of ATP release. This novel release mechanism was compared to the existing theories of extracellular release of ATP. Bioluminescence imaging was also employed to examine the role of ATP in the field of neuroscience. The central nervous system (CNS) was dissected from the freshwater snail Lymnaea stagnalis. Electrophysiological experiments demonstrated that the neurons of the Lymnaea were not damaged by any of the components of the imaging solution. ATP was continuously released by the ganglia of the CNS for over eight hours and varied from ganglion to ganglion and within individual ganglia. Addition of the neurotransmitters K + and serotonin increased release of ATP in certain regions of the Lymnaea CNS. Finally, the ATP imaging technique was investigated for the study of drug release systems. MCM-41-type mesoporous nanospheres were loaded with ATP and end-capped with mercaptoethanol functionalized CdS monocrystals. Aggregates of nanospheres were bathed in imaging solution, and ATP bioluminescence was monitored to investigated the release kinetics of the nanosphere drug delivery systems. Addition of disulfide bond-cleaving molecules induced uncapping of the nanospheres and subsequently, the release of ATP. Increasing the concentration of the uncapping molecule decreased the temporal maximum and increased the magnitude of release of encapsulated ATP from the nanospheres. Furthermore, the release kinetics from the nanospheres varied with the size of the particle aggregates.« less

Fungal mycelia show lag time before re-growth on endogenous carbon.

PubMed

Pollack, Judith K; Li, Zheng Jian; Marten, Mark R

2008-06-15

Nutrient starvation is a common occurrence for filamentous fungi. To better understand the effects of starvation, we used a parallel plate flow chamber to study individual fungal mycelia when subjected to a step change in glucose concentration. We report the presence of a finite "lag time" in starved mycelia during which they ceased to grow/extend while switching from growth on exogenous carbon to re-growth on endogenous carbon. This lag time precedes other morphological or physiological changes such as change in growth rate (50-70% reduction), vacuolation (up to 16%), and decreased hyphal diameter (almost 50% reduction). Data suggests that during lag time, vacuolar degradation produces sufficient endogenous carbon to support survival and restart hyphal extension. Lag time is inversely related to the size of the mycelium at the time of starvation, which suggests a critical flow of endogenous carbon to the apical tip. We present a mathematical model consistent with our experimental observations that relate lag time, area, and flow of endogenous carbon. (c) 2008 Wiley Periodicals, Inc.

Glyceroneogenesis in the hepatopancreas of the crab Neohelice granulata: Diet, starvation and season effects.

PubMed

Sarapio, E; Santos, J T; Model, J F A; De Fraga, L S; Vinagre, A S; Martins, T L; Da Silva, R S M; Trapp, M

2017-09-01

We determined the activity of glyceroneogenesis from [2- 14 C]-pyruvate, the phosphoenolpyruvate carboxykinase activity, [2- 14 C]-pyruvate oxidation and total lipid levels in the hepatopancreas of the crab Neohelice granulata fed with a carbohydrate-rich (HC) diet or a high-protein (HP) diet and then subjected to 5weeks of starvation, in summer and winter, to determine whether the seasonal adjustments of lipid metabolism to food scarcity are modulated by the composition of the diet previously given to the crabs. The results demonstrated that glyceroneogenesis is an active pathway in N. granulata hepatopancreas, and is regulated by seasonal variations, diet composition and starvation. This study showed that in summer the increase in the hepatopancreas glyceroneogenesis activity is among the strategies used by N. granulata fed an HP diet, to maintain the triglyceride/fatty acid cycle during starvation, a normal condition in the biological cycle of this crab. However, the administration of an HC diet reduced the glyceroneogenesis capacity in response to starvation in summer. In winter, the decrease in the glyceroneogenesis capacity in both fed (HP and HC diets) and starved crabs seems to be a strategy to reduce energy consumption and/or requirement. In contrast to the summer results, the incorporation of [2- 14 C]-pyruvate into 14 CO 2 was markedly higher in both diet (HC and HP) groups and in starved crabs during the winter. Four decades after the first study describing the glyceroneogenesis pathway in rat white adipose tissue, this pathway is evidenced for the first time in a crustacean. Copyright © 2017 Elsevier Inc. All rights reserved.

Global gene expression under nitrogen starvation in Xylella fastidiosa: contribution of the σ54 regulon

PubMed Central

2010-01-01

Background Xylella fastidiosa, a Gram-negative fastidious bacterium, grows in the xylem of several plants causing diseases such as citrus variegated chlorosis. As the xylem sap contains low concentrations of amino acids and other compounds, X. fastidiosa needs to cope with nitrogen limitation in its natural habitat. Results In this work, we performed a whole-genome microarray analysis of the X. fastidiosa nitrogen starvation response. A time course experiment (2, 8 and 12 hours) of cultures grown in defined medium under nitrogen starvation revealed many differentially expressed genes, such as those related to transport, nitrogen assimilation, amino acid biosynthesis, transcriptional regulation, and many genes encoding hypothetical proteins. In addition, a decrease in the expression levels of many genes involved in carbon metabolism and energy generation pathways was also observed. Comparison of gene expression profiles between the wild type strain and the rpoN null mutant allowed the identification of genes directly or indirectly induced by nitrogen starvation in a σ54-dependent manner. A more complete picture of the σ54 regulon was achieved by combining the transcriptome data with an in silico search for potential σ54-dependent promoters, using a position weight matrix approach. One of these σ54-predicted binding sites, located upstream of the glnA gene (encoding glutamine synthetase), was validated by primer extension assays, confirming that this gene has a σ54-dependent promoter. Conclusions Together, these results show that nitrogen starvation causes intense changes in the X. fastidiosa transcriptome and some of these differentially expressed genes belong to the σ54 regulon. PMID:20799976

Time-series resolution of gradual nitrogen starvation and its impact on photosynthesis in the cyanobacterium Synechocystis PCC 6803.

PubMed

Krasikov, Vladimir; Aguirre von Wobeser, Eneas; Dekker, Henk L; Huisman, Jef; Matthijs, Hans C P

2012-07-01

Sequential adaptation to nitrogen deprivation and ultimately to full starvation requires coordinated adjustment of cellular functions. We investigated changes in gene expression and cell physiology of the cyanobacterium Synechocystis PCC 6803 during 96 h of nitrogen starvation. During the first 6 h, the transcriptome showed activation of nitrogen uptake and assimilation systems and of the core nitrogen and carbon assimilation regulators. However, the nitrogen-deprived cells still grew at the same rate as the control and even showed transiently increased expression of phycobilisome genes. After 12 h, cell growth decreased and chlorosis started with degradation of the nitrogen-rich phycobilisomes. During this phase, the transcriptome showed suppression of genes for phycobilisomes, for carbon fixation and for de novo protein synthesis. Interestingly, photosynthetic activity of both photosystem I (PSI) and photosystem II was retained quite well. Excess electrons were quenched by the induction of terminal oxidase and hydrogenase genes, compensating for the diminished carbon fixation and nitrate reduction activity. After 48 h, the cells ceased most activities. A marked exception was the retained PSI gene transcription, possibly this supports the viability of Synechocystis cells and enables rapid recovery after relieving from nitrogen starvation. During early recovery, many genes changed expression, supporting the resumed cellular activity. In total, our results distinguished three phases during gradual nitrogen depletion: (1) an immediate response, (2) short-term acclimation and (3) long-term survival. This shows that cyanobacteria respond to nitrogen starvation by a cascade of physiological adaptations reflected by numerous changes in the transcriptome unfolding at different timescales. Copyright © Physiologia Plantarum 2012.

Adenosine triphosphate (ATP) reduces amyloid-β protein misfolding in vitro.

PubMed

Coskuner, Orkid; Murray, Ian V J

2014-01-01

Alzheimer's disease (AD) is a devastating disease of aging that initiates decades prior to clinical manifestation and represents an impending epidemic. Two early features of AD are metabolic dysfunction and changes in amyloid-β protein (Aβ) levels. Since levels of ATP decrease over the course of the disease and Aβ is an early biomarker of AD, we sought to uncover novel linkages between the two. First and remarkably, a GxxxG motif is common between both Aβ (oligomerization motif) and nucleotide binding proteins (Rossmann fold). Second, ATP was demonstrated to protect against Aβ mediated cytotoxicity. Last, there is structural similarity between ATP and amyloid binding/inhibitory compounds such as ThioT, melatonin, and indoles. Thus, we investigated whether ATP alters misfolding of the pathologically relevant Aβ42. To test this hypothesis, we performed computational and biochemical studies. Our computational studies demonstrate that ATP interacts strongly with Tyr10 and Ser26 of Aβ fibrils in solution. Experimentally, both ATP and ADP reduced Aβ misfolding at physiological intracellular concentrations, with thresholds at ~500 μM and 1 mM respectively. This inhibition of Aβ misfolding is specific; requiring Tyr10 of Aβ and is enhanced by magnesium. Last, cerebrospinal fluid ATP levels are in the nanomolar range and decreased with AD pathology. This initial and novel finding regarding the ATP interaction with Aβ and reduction of Aβ misfolding has potential significance to the AD field. It provides an underlying mechanism for published links between metabolic dysfunction and AD. It also suggests a potential role of ATP in AD pathology, as the occurrence of misfolded extracellular Aβ mirrors lowered extracellular ATP levels. Last, the findings suggest that Aβ conformation change may be a sensor of metabolic dysfunction.

Impaired cardiac energy metabolism in embryos lacking adrenergic stimulation

PubMed Central

Baker, Candice N.; Gidus, Sarah A.; Price, George F.; Peoples, Jessica N. R.

2014-01-01

As development proceeds from the embryonic to fetal stages, cardiac energy demands increase substantially, and oxidative phosphorylation of ADP to ATP in mitochondria becomes vital. Relatively little, however, is known about the signaling mechanisms regulating the transition from anaerobic to aerobic metabolism that occurs during the embryonic period. The main objective of this study was to test the hypothesis that adrenergic hormones provide critical stimulation of energy metabolism during embryonic/fetal development. We examined ATP and ADP concentrations in mouse embryos lacking adrenergic hormones due to targeted disruption of the essential dopamine β-hydroxylase (Dbh) gene. Embryonic ATP concentrations decreased dramatically, whereas ADP concentrations rose such that the ATP/ADP ratio in the adrenergic-deficient group was nearly 50-fold less than that found in littermate controls by embryonic day 11.5. We also found that cardiac extracellular acidification and oxygen consumption rates were significantly decreased, and mitochondria were significantly larger and more branched in adrenergic-deficient hearts. Notably, however, the mitochondria were intact with well-formed cristae, and there was no significant difference observed in mitochondrial membrane potential. Maternal administration of the adrenergic receptor agonists isoproterenol or l-phenylephrine significantly ameliorated the decreases in ATP observed in Dbh−/− embryos, suggesting that α- and β-adrenergic receptors were effective modulators of ATP concentrations in mouse embryos in vivo. These data demonstrate that adrenergic hormones stimulate cardiac energy metabolism during a critical period of embryonic development. PMID:25516547

Thyroid hormone action on intermediary metabolism. Part I: respiration, thermogenesis and carbohydrate metabolism.

PubMed

Müller, M J; Seitz, H J

1984-01-02

The effect of thyroid hormones on mitochondrial respiration are summarized: T3 directly stimulates mitochondrial respiration and the synthesis of adenosine 5'-triphosphate (ATP). Cytosolic ATP availability is increased by a thyroid hormone-induced increase in adenine nucleotide translocation across the mitochondrial membrane; the steady state ATP concentration and the cytosolic ATP/adenosine 5'-diphosphate (ADP) ratio is even decreased in hyperthyroid tissues because of the simultaneous stimulation of the synthesis and consumption of ATP. With regard to the thyroid hormone-induced energy wasting processes, heart work, intra- and interorgan futile cycling and Na+/K+-ATPase are involved to varying degrees. As a consequence of the thyroid hormone-induced hydrolysis of ATP, thermogenesis is increased in hyper- and decreased in hypothyroidism. Despite an increased rate of glucose utilization, clinical and experimental hyperthyroidism is often characterized by an abnormal oral glucose tolerance test. This finding is due to the thyroid hormone-induced increase in intestinal glucose absorption as well as the still enhanced endogenous glucose production in the liver. Hypothyroid patients show a reduced glucose tolerance test because of a decrease in intestinal glucose absorption and a sometimes reduced glucose turnover. The thyroid hormone-induced alterations in glucose metabolism are most probably not due to alterations in serum insulin levels and/or to a peripheral insulin resistance at the receptor level.

Mutations in the Atp1p and Atp3p subunits of yeast ATP synthase differentially affect respiration and fermentation in Saccharomyces cerevisiae.

PubMed

Francis, Brian R; White, Karen H; Thorsness, Peter E

2007-04-01

ATP1-111, a suppressor of the slow-growth phenotype of yme1Delta lacking mitochondrial DNA is due to the substitution of phenylalanine for valine at position 111 of the alpha-subunit of mitochondrial ATP synthase (Atp1p in yeast). The suppressing activity of ATP1-111 requires intact beta (Atp2p) and gamma (Atp3p) subunits of mitochondrial ATP synthase, but not the stator stalk subunits b (Atp4p) and OSCP (Atp5p). ATP1-111 and other similarly suppressing mutations in ATP1 and ATP3 increase the growth rate of wild-type strains lacking mitochondrial DNA. These suppressing mutations decrease the growth rate of yeast containing an intact mitochondrial chromosome on media requiring oxidative phosphorylation, but not when grown on fermentable media. Measurement of chronological aging of yeast in culture reveals that ATP1 and ATP3 suppressor alleles in strains that contain mitochondrial DNA are longer lived than the isogenic wild-type strain. In contrast, the chronological life span of yeast cells lacking mitochondrial DNA and containing these mutations is shorter than that of the isogenic wild-type strain. Spore viability of strains bearing ATP1-111 is reduced compared to wild type, although ATP1-111 enhances the survival of spores that lacked mitochondrial DNA.

Metabolic Flux Analysis of the Synechocystis sp. PCC 6803 ΔnrtABCD Mutant Reveals a Mechanism for Metabolic Adaptation to Nitrogen-Limited Conditions.

PubMed

Nakajima, Tsubasa; Yoshikawa, Katsunori; Toya, Yoshihiro; Matsuda, Fumio; Shimizu, Hiroshi

2017-03-01

Metabolic flux redirection during nitrogen-limited growth was investigated in the Synechocystis sp. PCC 6803 glucose-tolerant (GT) strain under photoautotrophic conditions by isotopically non-stationary metabolic flux analysis (INST-MFA). A ΔnrtABCD mutant of Synechocystis sp. PCC 6803 was constructed to reproduce phenotypes arising during nitrogen starvation. The ΔnrtABCD mutant and the wild-type GT strain were cultured under photoautotrophic conditions by a photobioreactor. Intracellular metabolites were labeled over a time course using NaH13CO3 as a carbon source. Based on these data, the metabolic flux distributions in the wild-type and ΔnrtABCD cells were estimated by INST-MFA. The wild-type GT and ΔnrtABCD strains displayed similar distribution patterns, although the absolute levels of metabolic flux were lower in ΔnrtABCD. Furthermore, the relative flux levels for glycogen metabolism, anaplerotic reactions and the oxidative pentose phosphate pathway were increased in ΔnrtABCD. This was probably due to the increased expression of enzyme genes that respond to nitrogen depletion. Additionally, we found that the ratio of ATP/NADPH demand increased slightly in the ΔnrtABCD mutant. These results indicated that futile ATP consumption increases under nitrogen-limited conditions because the Calvin-Benson cycle and the oxidative pentose phosphate pathway form a metabolic futile cycle that consumes ATP without CO2 fixation and NADPH regeneration. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

APSR1, a novel gene required for meristem maintenance, is negatively regulated by low phosphate availability.

PubMed

González-Mendoza, Víctor; Zurita-Silva, Andrés; Sánchez-Calderón, Lenin; Sánchez-Sandoval, María Eugenia; Oropeza-Aburto, Araceli; Gutiérrez-Alanís, Dolores; Alatorre-Cobos, Fulgencio; Herrera-Estrella, Luis

2013-05-01

Proper root growth is crucial for anchorage, exploration, and exploitation of the soil substrate. Root growth is highly sensitive to a variety of environmental cues, among them water and nutrient availability have a great impact on root development. Phosphorus (P) availability is one of the most limiting nutrients that affect plant growth and development under natural and agricultural environments. Root growth in the direction of the long axis proceeds from the root tip and requires the coordinated activities of cell proliferation, cell elongation and cell differentiation. Here we report a novel gene, APSR1 (Altered Phosphate Starvation Response1), involved in root meristem maintenance. The loss of function mutant apsr1-1 showed a reduction in primary root length and root apical meristem size, short differentiated epidermal cells and long root hairs. Expression of APSR1 gene decreases in response to phosphate starvation and apsr1-1 did not show the typical progressive decrease of undifferentiated cells at root tip when grown under P limiting conditions. Interestingly, APSR1 expression pattern overlaps with root zones of auxin accumulation. Furthermore, apsr1-1 showed a clear decrease in the level of the auxin transporter PIN7. These data suggest that APSR1 is required for the coordination of cell processes necessary for correct root growth in response to phosphate starvation conceivably by direct or indirect modulation of PIN7. We also propose, based on its nuclear localization and structure, that APSR1 may potentially be a member of a novel group of transcription factors. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Sensor kinase KinB and its pathway-associated key factors sense the signal of nutrition starvation in sporulation of Bacillus subtilis.

PubMed

Liu, Weipeng; He, Zeying; Gao, Feng; Yan, Jinyuan; Huang, Xiaowei

2018-01-03

Bacillus subtilis responds to environmental stress cues and develops endospores for survival. In the process of endospore formation, sporulation initiation is a vital stage and this stage is governed by autophosphorylation of the sensor histidine kinases. The second major sensor kinase KinB perceives the intracellular changes of GTP and ATP during sporulation. However, determination of the environmental signals as well as its related signaling pathway of KinB requires further elucidation. Our current study found that, contrary to the sporulation failure induced by ΔkinA in the nutrient-rich 2× SG medium, the sensor kinase KinB sensed the environmental cues in the nutrient-poor MM medium. Two other membrane proteins, KapB and KbaA, also responded similarly to the same external signal as KinB. Both KapB and KbaA acted upstream of KinB, but they exerted their regulation upon KinB independently. Furthermore, we demonstrated that both the SH3 domain and the α-helix structure in KapB are required for sensing or transducing the signal of sporulation initiation. Collectively, our work here supplied the direct evidences that KinB and its pathway sense the external signal of nutrient starvation in MM medium, and further analyzes the interrelationship among KinB, KbaA, and KapB. © 2018 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Effect of Starvation on the Turnover and Metabolic Response to Leucine

PubMed Central

Sherwin, Robert S.

1978-01-01

l-Leucine was administered as a primed continuous 3-4-h infusion in nonobese and obese subjects in the postabsorptive state and for 12 h in obese subjects after a 3-day and 4-wk fast. In nonobese and obese subjects studied in the post-absorptive state, the leucine infusion resulted in a 150-200% rise in plasma leucine above preinfusion levels, a small decrease in plasma glucose, and unchanged levels of plasma insulin and glucagon and blood ketones. Plasma isoleucine (60-70%) and valine (35-40%) declined to a greater extent than other amino acids (P < 0.001). After 3 days and 4 wk of fasting, equimolar infusions of leucine resulted in two- to threefold greater increments in plasma leucine as compared to post-absorptive subjects, a 30-40% decline in other plasma amino acids, and a 25-30% decrease in negative nitrogen balance. Urinary excretion of 3-methylhistidine was however, unchanged. Plasma glucose which declined in 3-day fasted subjects after leucine administration, surprisingly rose by 20 mg/100 ml after 4 wk of fasting. The rise in blood glucose occurred in the absence of changes in plasma glucagon and insulin and in the face of a 15% decline in endogenous glucose production (as measured by infusion of [3-3H]glucose). On the other hand, fractional glucose utilization fell by 30% (P < 0.001), thereby accounting for hyperglycemia. The estimated metabolic clearance rate of leucine fell by 48% after 3 days of fasting whereas the plasma delivery rate of leucine was unchanged, thereby accounting for a 40% rise in plasma leucine during early starvation. After a 4-wk fast, the estimated metabolic clearance rate of leucine declined further to 59% below base line. Plasma leucine nevertheless fell to postabsorptive levels as the plasma delivery rate of leucine decreased 65% below postabsorptive values. Conclusions: (a) Infusion of exogenous leucine in prolonged fasting results in a decline in plasma levels of other amino acids, improvement in nitrogen balance and unchanged excretion of 3-methylhistidine, thus suggesting stimulation of muscle protein synthesis, (b) leucine infusion also reduces glucose production and to an even greater extent, glucose consumption, thereby raising blood glucose concentration; and (c) the rise in plasma leucine in early starvation results primarily from a decrease in leucine clearance which drops progressively during starvation. PMID:659610

Profound bioenergetic abnormalities in peri-infarct myocardial regions.

PubMed

Hu, Qingsong; Wang, Xiaohong; Lee, Joseph; Mansoor, Abdul; Liu, Jingbo; Zeng, Lepeng; Swingen, Cory; Zhang, Ge; Feygin, Julia; Ochiai, Koichi; Bransford, Toni L; From, Arthur H L; Bache, Robert J; Zhang, Jianyi

2006-08-01

Regions of myocardial infarct (MI) are surrounded by a border zone (BZ) of normally perfused but dysfunctional myocardium. Although systolic dysfunction has been attributed to elevated wall stress in this region, there is evidence that intrinsic abnormalities of contractile performance exist in BZ myocardium. This study examined whether decreases of high-energy phosphates (HEP) and mitochondrial F(1)F(0)-ATPase (mtATPase) subunits typical of failing myocardium exist in BZ myocardium of compensated postinfarct remodeled hearts. Eight pigs were studied 6 wk after MI was produced by ligation of the left anterior descending coronary artery (LAD) distal to the second diagonal. Animals developed compensated LV remodeling with a decrease of ejection fraction from 54.6 +/- 5.4% to 31 +/- 2.1% (MRI) 5 wk after LAD occlusion. The remote zone (RZ) myocardium demonstrated modest decreases of ATP and mtATPase components. In contrast, BZ myocardium demonstrated profound abnormalities with ATP levels decreased to 42% of normal, and phosphocreatine-to-ATP ratio ((31)P-magnetic resonance spectroscopy) decreased from 2.06 +/- 0.19 in normal hearts to 1.07 +/- 0.10, with decreases in alpha-, beta-, OSCP, and IF(1) subunits of mtATPase, especially in the subendocardium. The reduction of myocardial creatine kinase isoform protein expression was also more severe in the BZ relative to the RZ myocardium. These abnormalities were independent of a change in mitochondrial content because the mitochondrial citrate synthase protein level was not different between the BZ and RZ. This regional heterogeneity of ATP content and expression of key enzymes in ATP production suggests that energetic insufficiency in the peri-infarct region may contribute to the transition from compensated LV remodeling to congestive heart failure.

Identification of a P2X7 receptor in GH(4)C(1) rat pituitary cells: a potential target for a bioactive substance produced by Pfiesteria piscicida.

PubMed Central

Kimm-Brinson, K L; Moeller, P D; Barbier, M; Glasgow, H; Burkholder, J M; Ramsdell, J S

2001-01-01

We examined the pharmacologic activity of a putative toxin (pPfTx) produced by Pfiesteria piscicida by characterizing the signaling pathways that induce the c-fos luciferase construct in GH(4)C(1) rat pituitary cells. Adenosine-5'-triphosphate (ATP) was determined to increase and, at higher concentrations, decrease luciferase activity in GH(4)C(1) rat pituitary cells that stably express c-fos luciferase. The inhibition of luciferase results from cytotoxicity, characteristic of the putative P. piscicida toxin (pPfTx). The actions of both pPfTx and ATP to induce c-fos luciferase were inhibited by the purinogenic receptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS). Further characterization of a P2X receptor on the GH(4)C(1) cell was determined by the analog selectivity of P2X agonists. The P2X1/P2X3 agonist alpha,beta-methylene ATP (alpha,beta-MeATP) failed to increase or decrease c-fos luciferase. However, the P2X7 agonist 2',3'-(4-benzoyl)benzoyl ATP (BzATP), which had a predominant cytotoxic effect, was more potent than ATP. Immunoblot analysis of GH(4)C(1) cell membranes confirmed the presence of a 70-kDa protein that was immunoreactive to an antibody directed against the carboxy-terminal domain unique to the P2X7 receptor. The P2X7 irreversible antagonist oxidized-ATP (oxATP) inhibited the action of ATP, BzATP, and pPfTx. These findings indicate that GH(4)C(1) cells express purinogenic receptors with selectivity consistent with the P2X7 subtype and that this receptor pathway mediates the induction of the c-fos luciferase reporter gene by ATP and the putative Pfiesteria toxin PMID:11401756

Electrical Stimuli Are Anti-Apoptotic in Skeletal Muscle via Extracellular ATP. Alteration of This Signal in Mdx Mice Is a Likely Cause of Dystrophy

PubMed Central

Valladares, Denisse; Almarza, Gonzalo; Contreras, Ariel; Pavez, Mario; Buvinic, Sonja; Jaimovich, Enrique; Casas, Mariana

2013-01-01

ATP signaling has been shown to regulate gene expression in skeletal muscle and to be altered in models of muscular dystrophy. We have previously shown that in normal muscle fibers, ATP released through Pannexin1 (Panx1) channels after electrical stimulation plays a role in activating some signaling pathways related to gene expression. We searched for a possible role of ATP signaling in the dystrophy phenotype. We used muscle fibers from flexor digitorum brevis isolated from normal and mdx mice. We demonstrated that low frequency electrical stimulation has an anti-apoptotic effect in normal muscle fibers repressing the expression of Bax, Bim and PUMA. Addition of exogenous ATP to the medium has a similar effect. In dystrophic fibers, the basal levels of extracellular ATP were higher compared to normal fibers, but unlike control fibers, they do not present any ATP release after low frequency electrical stimulation, suggesting an uncoupling between electrical stimulation and ATP release in this condition. Elevated levels of Panx1 and decreased levels of Cav1.1 (dihydropyridine receptors) were found in triads fractions prepared from mdx muscles. Moreover, decreased immunoprecipitation of Cav1.1 and Panx1, suggest uncoupling of the signaling machinery. Importantly, in dystrophic fibers, exogenous ATP was pro-apoptotic, inducing the transcription of Bax, Bim and PUMA and increasing the levels of activated Bax and cytosolic cytochrome c. These evidence points to an involvement of the ATP pathway in the activation of mechanisms related with cell death in muscular dystrophy, opening new perspectives towards possible targets for pharmacological therapies. PMID:24282497

Electrical stimuli are anti-apoptotic in skeletal muscle via extracellular ATP. Alteration of this signal in Mdx mice is a likely cause of dystrophy.

PubMed

Valladares, Denisse; Almarza, Gonzalo; Contreras, Ariel; Pavez, Mario; Buvinic, Sonja; Jaimovich, Enrique; Casas, Mariana

2013-01-01

ATP signaling has been shown to regulate gene expression in skeletal muscle and to be altered in models of muscular dystrophy. We have previously shown that in normal muscle fibers, ATP released through Pannexin1 (Panx1) channels after electrical stimulation plays a role in activating some signaling pathways related to gene expression. We searched for a possible role of ATP signaling in the dystrophy phenotype. We used muscle fibers from flexor digitorum brevis isolated from normal and mdx mice. We demonstrated that low frequency electrical stimulation has an anti-apoptotic effect in normal muscle fibers repressing the expression of Bax, Bim and PUMA. Addition of exogenous ATP to the medium has a similar effect. In dystrophic fibers, the basal levels of extracellular ATP were higher compared to normal fibers, but unlike control fibers, they do not present any ATP release after low frequency electrical stimulation, suggesting an uncoupling between electrical stimulation and ATP release in this condition. Elevated levels of Panx1 and decreased levels of Cav1.1 (dihydropyridine receptors) were found in triads fractions prepared from mdx muscles. Moreover, decreased immunoprecipitation of Cav1.1 and Panx1, suggest uncoupling of the signaling machinery. Importantly, in dystrophic fibers, exogenous ATP was pro-apoptotic, inducing the transcription of Bax, Bim and PUMA and increasing the levels of activated Bax and cytosolic cytochrome c. These evidence points to an involvement of the ATP pathway in the activation of mechanisms related with cell death in muscular dystrophy, opening new perspectives towards possible targets for pharmacological therapies.

Lipidomic and metabolic changes in the P4-type ATPase ATP10D deficient C57BL/6J wild type mice upon rescue of ATP10D function.

PubMed

Sigruener, Alexander; Wolfrum, Christian; Boettcher, Alfred; Kopf, Thomas; Liebisch, Gerhard; Orsó, Evelyn; Schmitz, Gerd

2017-01-01

Sequence variants near the human gene for P4-type ATPase, class V, type 10D (ATP10D) were shown to significantly associate with circulating hexosylceramide d18:1/16:0 and d18:1/24:1 levels, obesity, insulin resistance, plasma high density lipoprotein (HDL), coronary stenotic index and intracranial atherosclerotic index. In mice Atp10d is associated with HDL modulation and C57BL/6 mice expressing a truncated, non-functional form of ATP10D easily develop obesity and insulin resistance on high-fat diet. We analyzed metabolic differences of ATP10D deficient C57BL/6J wild type and ATP10D transgenic C57BL/6J BAC129 mice. ATP10D transgenic mice gain 25% less weight on high-fat diet concomitant with a reduced increase in fat cell mass but independent of adipocyte size change. ATP10D transgenic mice also had 26% lower triacylglycerol levels with approximately 76% bound to very low density lipoprotein while in ATP10D deficient wild type mice 57% are bound to low density lipoprotein. Furthermore increased oxygen consumption and CO2 production, 38% lower glucose and 69% lower insulin levels and better insulin sensitivity were observed in ATP10D transgenic mice. Besides decreased hexosylceramide species levels were detected. Part of these effects may be due to reduced hepatic stearoyl-CoA desaturase 1 (SCD1) expression in ATP10D transgenic mice, which was reflected by altered fatty acid and lipid species patterns. There was a significant decrease in the hepatic 18:1 to 18:0 free fatty acid ratio in transgenic mice. The ratio of 16:1 to 16:0 was not significantly different. Interestingly both ratios were significantly reduced in plasma total fatty acids. In summary we found that ATP10D reduces high-fat diet induced obesity and improves insulin sensitivity. ATP10D transgenic mice showed altered hepatic expression of lipid-metabolism associated genes, including Scd1, along with changes in hepatic and plasma lipid species and plasma lipoprotein pattern.

Human forearm metabolism during progressive starvation.

PubMed

Owen, O E; Reichard, G A

1971-07-01

Forearm muscle metabolism was studied in eight obese subjects after an overnight, 3 and 24 day fast. Arterio-deep-venous differences of oxygen, carbon dioxide, glucose, lactate, pyruvate, free fatty acids, acetoacetate, and beta-hydroxybutyrate with simultaneous forearm blood flow were measured. Rates of metabolite utilization and production were thus estimated. Oxygen consumption and lactate and pyruvate production remained relatively constant at each fasting period. Glucose, initially the major substrate consumed, showed decreased consumption after 3 and 24 days of fasting. Acetoacetate and beta-hydroxybutyrate consumption after an overnight fast was low. At 3 days of fasting with increased arterial concentrations of acetoactate and beta-hydroxybutyrate, consumption of these substrates rose dramatically. At 24 days of fasting, despite further elevation of arterial levels of acetoacetate and beta-hydroxybutyrate, the utilization of acetoacetate did not increase further and if anything decreased, while five out of eight subjects released beta-hydroxybutyrate across the forearm. Acetoacetate was preferentially extracted over beta-hydroxybutyrate. At 24 days of starvation, free fatty acids were the principal fuels extracted by forearm muscle; at this time there was a decreased glucose and also ketone-body consumption by skeletal muscle.

Nisin depletes ATP and proton motive force in mycobacteria.

PubMed

Chung, H J; Montville, T J; Chikindas, M L

2000-12-01

This study examined the inhibitory effect of nisin and its mode of action against Mycobacterium smegmatis, a non-pathogenic species of mycobacteria, and M. bovis-Bacill Carmette Guerin (BCG), a vaccine strain of pathogenic M. bovis. In agar diffusion assays, 2.5 mg ml(-1) nisin was required to inhibit M. bovis-BCG. Nisin caused a slow, gradual, time- and concentration-dependent decrease in internal ATP levels in M. bovis-BCG, but no ATP efflux was detected. In mycobacteria, nisin decreased both components of proton motive force (membrane potential, Delta Psi and Delta pH) in a time- and concentration-dependent manner. However, mycobacteria maintained their intracellular ATP levels during the initial time period of Delta Psi and Delta pH dissipation. These data suggest that the mechanism of nisin in mycobacteria is similar to that in food-borne pathogens.

Impaired cardiac energy metabolism in embryos lacking adrenergic stimulation.

PubMed

Baker, Candice N; Gidus, Sarah A; Price, George F; Peoples, Jessica N R; Ebert, Steven N

2015-03-01

As development proceeds from the embryonic to fetal stages, cardiac energy demands increase substantially, and oxidative phosphorylation of ADP to ATP in mitochondria becomes vital. Relatively little, however, is known about the signaling mechanisms regulating the transition from anaerobic to aerobic metabolism that occurs during the embryonic period. The main objective of this study was to test the hypothesis that adrenergic hormones provide critical stimulation of energy metabolism during embryonic/fetal development. We examined ATP and ADP concentrations in mouse embryos lacking adrenergic hormones due to targeted disruption of the essential dopamine β-hydroxylase (Dbh) gene. Embryonic ATP concentrations decreased dramatically, whereas ADP concentrations rose such that the ATP/ADP ratio in the adrenergic-deficient group was nearly 50-fold less than that found in littermate controls by embryonic day 11.5. We also found that cardiac extracellular acidification and oxygen consumption rates were significantly decreased, and mitochondria were significantly larger and more branched in adrenergic-deficient hearts. Notably, however, the mitochondria were intact with well-formed cristae, and there was no significant difference observed in mitochondrial membrane potential. Maternal administration of the adrenergic receptor agonists isoproterenol or l-phenylephrine significantly ameliorated the decreases in ATP observed in Dbh-/- embryos, suggesting that α- and β-adrenergic receptors were effective modulators of ATP concentrations in mouse embryos in vivo. These data demonstrate that adrenergic hormones stimulate cardiac energy metabolism during a critical period of embryonic development. Copyright © 2015 the American Physiological Society.

Measurement of adenosine triphosphate and 2,3-diphosphoglycerate in stored blood with 31P nuclear magnetic resonance spectroscopy.

PubMed

Ambruso, D R; Hawkins, B; Johnson, D L; Fritzberg, A R; Klingensmith, W C; McCabe, E R

1986-06-01

Conditions for blood storage are chosen to assure adequate levels of adenosine triphosphate (ATP) and 2,3-diphosphoglycerate (2,3-DPG). Because of the invasive nature of the techniques, biochemical assays are not routinely used to measure levels of these compounds in stored blood. However, 31P NMR spectroscopy measures phosphorylated intermediates in intact cells and could be used without disruption of the storage pack. We compared levels of ATP and 2,3-DPG measured by 31P spectroscopy and standard enzyme-linked biochemical assays in whole blood (WB) and packed red blood cells (PRBCs) at weekly intervals during a 35-day storage period. NMR demonstrated a marked decrease in 2,3-DPG and an increase in inorganic phosphate after the first week of storage. No significant differences in ATP concentrations were seen in WB during the storage period, but a significant decrease in ATP in PRBCs was documented. There was good agreement in levels of ATP and 2,3-DPG measured by NMR and biochemical techniques. 31P NMR spectroscopy is a noninvasive technique for measuring ATP and 2,3-DPG which has a potential use in quality assurance of stored blood.

The Molecular Mechanisms Affecting N-Acetylaspartate Homeostasis Following Experimental Graded Traumatic Brain Injury

PubMed Central

Di Pietro, Valentina; Amorini, Angela Maria; Tavazzi, Barbara; Vagnozzi, Roberto; Logan, Ann; Lazzarino, Giacomo; Signoretti, Stefano; Lazzarino, Giuseppe; Belli, Antonio

2014-01-01

To characterize the molecular mechanisms of N-acetylaspartate (NAA) metabolism following traumatic brain injury (TBI), we measured the NAA, adenosine triphosphate (ATP) and adenosine diphosphate (ADP) concentrations and calculated the ATP/ADP ratio at different times from impact, concomitantly evaluating the gene and protein expressions controlling NAA homeostasis (the NAA synthesizing and degrading enzymes N-acetyltransferase 8-like and aspartoacylase, respectively) in rats receiving either mild or severe TBI. The reversible changes in NAA induced by mild TBI were due to a combination of transient mitochondrial malfunctioning with energy crisis (decrease in ATP and in the ATP/ADP ratio) and modulation in the gene and protein levels of N-acetyltransferase 8-like and increase of aspartoacylase levels. The irreversible decrease in NAA following severe TBI, was instead characterized by profound mitochondrial malfunctioning (constant 65% decrease of the ATP/ADP indicating permanent impairment of the mitochondrial phosphorylating capacity), dramatic repression of the N-acetyltransferase 8-like gene and concomitant remarkable increase in the aspartoacylase gene and protein levels. The mechanisms underlying changes in NAA homeostasis following graded TBI might be of note for possible new therapeutic approaches and will help in understanding the effects of repeat concussions occurring during particular periods of the complex NAA recovery process, coincident with the so called window of brain vulnerability. PMID:24515258

[Stabilization of Cadmium Contaminated Soils by Ferric Ion Modified Attapulgite (Fe/ATP)--Characterizations and Stabilization Mechanism].

PubMed

Rong, Yang; Li, Rong-bo; Zhou, Yong-li; Chen, Jing; Wang, Lin-ling; Lu, Xiao-hua

2015-08-01

Ferric ion modified attapulgite (Fe/ATP) was prepared by impregnation and its structure and morphology were characterized. The toxicity characteristic leaching procedure (TCLP) was used to evaluate the effect of Cadmium( Cd) stabilization in soil with the addition of attapulgite (ATP) and Fe/ATP. The stabilization mechanism of Cd was further elucidated by comparing the morphologies and structure of ATP and Fe/ATP before and after Cd adsorption. Fe/ATP exhibited much better adsorption capacity than ATP, suggesting different adsorption mechanisms occurred between ATP and Fe/ATP. The leaching concentrations of Cd in soil decreased by 45% and 91% respectively, with the addition of wt. 20% ATP and Fe/ATP. The former was attributed to the interaction between Cd2 and --OH groups by chemical binding to form inner-sphere complexes in ATP and the attachment between Cd2+ and the defect sites in ATP framework. Whereas Cd stabilization with Fe/ATP was resulted from the fact that the active centers (--OH bonds or O- sites) on ATP could react with Fe3+ giving Fe--O--Cd-- bridges, which helped stabilize Cd in surface soil. What'more, the ferric oxides and metal hydroxides on the surface of ATP could interact with Cd, probably by the formation of cadmium ferrite. In conclusion, Fe/ATP, which can be easily prepared, holds promise as a potential low-cost and environmental friendly stabilizing agent for remediation of soil contaminated with heavy metals.

Light-dependent quenching of chlorophyll fluorescence in pea chloroplasts induced by adenosine 5'-triphosphate.

PubMed

Horton, P; Black, M T

1981-03-12

Addition of ATP to chloroplasts causes a reversible 25-30% decrease in chlorophyll fluorescence. This quenching is light-dependent, uncoupler insensitive but inhibited by DCMU and electron acceptors and has a half-time of 3 minutes. Electron donors to Photosystem I can not overcome the inhibitory effect of DCMU, suggesting that light activation depends on the reduced state of plastoquinone. Fluorescence emission spectra recorded at -196 degrees C indicate that ATP treatment increases the amount of excitation energy transferred to Photosystem I. Examination of fluorescence induction curves indicate that ATP treatment decreases both the initial (F0) and variable (Fv) fluorescence such that the ratio of Fv to the maximum (Fm) yield is unchanged. The initial sigmoidal phase of induction is slowed down by ATP treatment and is quenched 3-fold more than the exponential slow phase, the rate of which is unchanged. A plot of Fv against area above the induction curve was identical plus or minus ATP. Thus ATP treatment can alter quantal distribution between Photosystems II and I without altering Photosystem II-Photosystem II interaction. The effect of ATP strongly resembles in its properties the phosphorylation of the light-harvesting complex by a light activated, ATP-dependent protein kinase found in chloroplast membranes and could be the basis of physiological mechanisms which contribute to slow fluorescence quenching in vivo and regulate excitation energy distribution between Photosystem I and II. It is suggested that the sensor for this regulation is the redox state of plastoquinone.

Energy allocation in juvenile roach and burbot under different temperature and feeding regimes.

PubMed

Binner, Maaike; Kloas, Werner; Hardewig, Iris

2008-06-01

Cold-active burbot (Lota lota (L.)) display reduced food intake during the summer. The impact of temperature on their energy budget was investigated in starved fish in a laboratory setting, simulating summer (20 degrees C) and winter (4 degrees C) conditions, to elucidate the impact of high temperature on burbot metabolism. Metabolic effects in burbot were compared to roach (Rutilus rutilus (L.)), which typically fast in winter. During warm acclimation, starvation (four weeks) resulted in a metabolic depression of oxygen consumption in both species. In roach, metabolic rate decreased by 55% after two weeks of starvation. Burbot, in contrast, displayed an immediate depression of metabolic rate by 50%. In both species, no reductions were observed in the cold. The temperature-induced differences between the metabolic rates at 20 degrees C and 4 degrees C showed a lower thermal sensitivity in burbot (Q (10) = 1.9) compared to roach (Q (10) = 2.7). Notably, for each species, energy consumption during starvation was highest under experimental conditions simulating their natural active periods, respectively. Warm acclimated roach relied mainly on muscle reserves, whereas in cold acclimated burbot, liver metabolic stores made a major contribution to the energy turnover. In cold acclimated roach and warm acclimated burbot, however, starvation apparently reduced swimming activity, resulting in considerable savings of energy reserves. These lower energy expenditures in roach and burbot corresponded to their natural inactive periods. Thus, starvation in burbot caused a lower energy turnover when exposed to high temperatures. These season-dependent adaptations of metabolism represent an advantageous strategy in burbot to manage winter temperature and withstand metabolism-activating summer temperatures, whereas roach metabolism correlates with the seasonal temperature cycle.

Root developmental adaptation to phosphate starvation: better safe than sorry.

PubMed

Péret, Benjamin; Clément, Mathilde; Nussaume, Laurent; Desnos, Thierry

2011-08-01

Phosphorus is a crucial component of major organic molecules such as nucleic acids, ATP and membrane phospholipids. It is present in soils in the form of inorganic phosphate (Pi), which has low availability and poor mobility. To cope with Pi limitations, plants have evolved complex adaptive responses that include morphological and physiological modifications. This review describes how the model plant Arabidopsis thaliana adapts its root system architecture to phosphate deficiency through inhibition of primary root growth, increase in lateral root formation and growth and production of root hairs, which all promote topsoil foraging. A better understanding of plant adaptation to low phosphate will open the way to increased phosphorus use efficiency by crops. Such an improvement is needed in order to adjust how we manage limited phosphorus stocks and to reduce the disastrous environmental effects of phosphate fertilizers overuse. Copyright © 2011 Elsevier Ltd. All rights reserved.

Ecm33 is a novel factor involved in efficient glucose uptake for nutrition-responsive TORC1 signaling in yeast.

PubMed

Umekawa, Midori; Ujihara, Masato; Nakai, Daiki; Takematsu, Hiromu; Wakayama, Mamoru

2017-11-01

Glucose uptake is crucial for providing both an energy source and a signal that regulates cell proliferation. Therefore, it is important to clarify the mechanisms underlying glucose uptake and its transmission to intracellular signaling pathways. In this study, we searched for a novel regulatory factor involved in glucose-induced signaling by using Saccharomyces cerevisiae as a eukaryotic model. Requirement of the extracellular protein Ecm33 in efficient glucose uptake and full activation of the nutrient-responsive TOR kinase complex 1 (TORC1) signaling pathway is shown. Cells lacking Ecm33 elicit a series of starvation-induced pathways even in the presence of extracellular high glucose concentration. This results in delayed cell proliferation, reduced ATP, induction of autophagy, and dephosphorylation of the TORC1 substrates Atg13 and Sch9. © 2017 Federation of European Biochemical Societies.

FGF21 and the late adaptive response to starvation in humans.

PubMed

Fazeli, Pouneh K; Lun, Mingyue; Kim, Soo M; Bredella, Miriam A; Wright, Spenser; Zhang, Yang; Lee, Hang; Catana, Ciprian; Klibanski, Anne; Patwari, Parth; Steinhauser, Matthew L

2015-11-03

In mice, FGF21 is rapidly induced by fasting, mediates critical aspects of the adaptive starvation response, and displays a number of positive metabolic properties when administered pharmacologically. In humans, however, fasting does not consistently increase FGF21, suggesting a possible evolutionary divergence in FGF21 function. Moreover, many key aspects of FGF21 function in mice have been identified in the context of transgenic overexpression or administration of supraphysiologic doses, rather than in a physiologic setting. Here, we explored the dynamics and function of FGF21 in human volunteers during a 10-day fast. Unlike mice, which show an increase in circulating FGF21 after only 6 hours, human subjects did not have a notable surge in FGF21 until 7 to 10 days of fasting. Moreover, we determined that FGF21 induction was associated with decreased thermogenesis and adiponectin, an observation that directly contrasts with previous reports based on supraphysiologic dosing. Additionally, FGF21 levels increased after ketone induction, demonstrating that endogenous FGF21 does not drive starvation-mediated ketogenesis in humans. Instead, a longitudinal analysis of biologically relevant variables identified serum transaminases--markers of tissue breakdown--as predictors of FGF21. These data establish FGF21 as a fasting-induced hormone in humans and indicate that FGF21 contributes to the late stages of adaptive starvation, when it may regulate the utilization of fuel derived from tissue breakdown.

Fibroblast growth factor rescues brain endothelial cells lacking presenilin 1 from apoptotic cell death following serum starvation.

PubMed

Gama Sosa, Miguel A; De Gasperi, Rita; Hof, Patrick R; Elder, Gregory A

2016-07-22

Presenilin 1 (Psen1) is important for vascular brain development and is known to influence cellular stress responses. To understand the role of Psen1 in endothelial stress responses, we investigated the effects of serum withdrawal on wild type (wt) and Psen1-/- embryonic brain endothelial cells. Serum starvation induced apoptosis in Psen1-/- cells but did not affect wt cells. PI3K/AKT signaling was reduced in serum-starved Psen1-/- cells, and this was associated with elevated levels of phospho-p38 consistent with decreased pro-survival AKT signaling in the absence of Psen1. Fibroblast growth factor (FGF1 and FGF2), but not vascular endothelial growth factor (VEGF) rescued Psen1-/- cells from serum starvation induced apoptosis. Inhibition of FGF signaling induced apoptosis in wt cells under serum withdrawal, while blocking γ-secretase activity had no effect. In the absence of serum, FGF2 immunoreactivity was distributed diffusely in cytoplasmic and nuclear vesicles of wt and Psen1-/- cells, as levels of FGF2 in nuclear and cytosolic fractions were not significantly different. Thus, sensitivity of Psen1-/- cells to serum starvation is not due to lack of FGF synthesis but likely to effects of Psen1 on FGF release onto the cell surface and impaired activation of the PI3K/AKT survival pathway.

Binding of ATP by pertussis toxin and isolated toxin subunits

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hausman, S.Z.; Manclark, C.R.; Burns, D.L.

1990-07-03

The binding of ATP to pertussis toxin and its components, the A subunit and B oligomer, was investigated. Whereas, radiolabeled ATP bound to the B oligomer and pertussis toxin, no binding to the A subunit was observed. The binding of ({sup 3}H)ATP to pertussis toxin and the B oligomer was inhibited by nucleotides. The relative effectiveness of the nucleotides was shown to be ATP > GTP > CTP > TTP for pertussis toxin and ATP > GTP > TTP > CTP for the B oligomer. Phosphate ions inhibited the binding of ({sup 3}H)ATP to pertussis toxin in a competitive manner;more » however, the presence of phosphate ions was essential for binding of ATP to the B oligomer. The toxin substrate, NAD, did not affect the binding of ({sup 3}H)ATP to pertussis toxin, although the glycoprotein fetuin significantly decreased binding. These results suggest that the binding site for ATP is located on the B oligomer and is distinct from the enzymatically active site but may be located near the eukaryotic receptor binding site.« less

Electron transfer protein complexes in the thylakoid membranes of heterocysts from the cyanobacterium Nostoc punctiforme.

PubMed

Cardona, Tanai; Battchikova, Natalia; Zhang, Pengpeng; Stensjö, Karin; Aro, Eva-Mari; Lindblad, Peter; Magnuson, Ann

2009-04-01

Filamentous, heterocystous cyanobacteria are capable of nitrogen fixation and photoautotrophic growth. Nitrogen fixation takes place in heterocysts that differentiate as a result of nitrogen starvation. Heterocysts uphold a microoxic environment to avoid inactivation of nitrogenase, e.g. by downregulation of oxygenic photosynthesis. The ATP and reductant requirement for the nitrogenase reaction is considered to depend on Photosystem I, but little is known about the organization of energy converting membrane proteins in heterocysts. We have investigated the membrane proteome of heterocysts from nitrogen fixing filaments of Nostoc punctiforme sp. PCC 73102, by 2D gel electrophoresis and mass spectrometry. The membrane proteome was found to be dominated by the Photosystem I and ATP-synthase complexes. We could identify a significant amount of assembled Photosystem II complexes containing the D1, D2, CP43, CP47 and PsbO proteins from these complexes. We could also measure light-driven in vitro electron transfer from Photosystem II in heterocyst thylakoid membranes. We did not find any partially disassembled Photosystem II complexes lacking the CP43 protein. Several subunits of the NDH-1 complex were also identified. The relative amount of NDH-1M complexes was found to be higher than NDH-1L complexes, which might suggest a role for this complex in cyclic electron transfer in the heterocysts of Nostoc punctiforme.

The combination of energy-dependent internal adaptation mechanisms and external factors enables Listeria monocytogenes to express a strong starvation survival response during multiple-nutrient starvation.

PubMed

Lungu, Bwalya; Saldivar, Joshua C; Story, Robert; Ricke, Steven C; Johnson, Michael G

2010-05-01

The goal of this study was to characterize the starvation survival response (SSR) of a wild-type Listeria monocytogenes 10403S and an isogenic DeltasigB mutant strain during multiple-nutrient starvation conditions over 28 days. This study examined the effects of inhibitors of protein synthesis, the proton motive force, substrate level phosphorylation, and oxidative phosphorylation on the SSR of L. monocytogenes 10403S and a DeltasigB mutant during multiple-nutrient starvation. The effects of starvation buffer changes on viability were also examined. During multiple-nutrient starvation, both strains expressed a strong SSR, suggesting that L. monocytogenes possesses SigB-independent mechanism(s) for survival during multiple-nutrient starvation. Neither strain was able to express an SSR following starvation buffer changes, indicating that the nutrients/factors present in the starvation buffer could be a source of energy for cell maintenance and survival. Neither the wild-type nor the DeltasigB mutant strain was able to elicit an SSR when exposed to the protein synthesis inhibitor chloramphenicol within the first 4 h of starvation. However, both strains expressed an SSR when exposed to chloramphenicol after 6 h or more of starvation, suggesting that the majority of proteins required to elicit an effective SSR in L. monocytogenes are likely produced somewhere between 4 and 6 h of starvation. The varying SSRs of both strains to the different metabolic inhibitors under aerobic or anaerobic conditions suggested that (1) energy derived from the proton motive force is important for an effective SSR, (2) L. monocytogenes utilizes an anaerobic electron transport during multiple-nutrient starvation conditions, and (3) the glycolytic pathway is an important energy source during multiple-nutrient starvation when oxygen is available, and less important under anaerobic conditions. Collectively, the data suggest that the combination of energy-dependent internal adaptation mechanisms of cells and external nutrients/factors enables L. monocytogenes to express a strong SSR.

The active site of oxidative phosphorylation and the origin of hyperhomocysteinemia in aging and dementia.

PubMed

McCully, Kilmer S

2015-01-01

The active site of oxidative phosphorylation and adenosine triphosphate (ATP) synthesis in mitochondria is proposed to consist of two molecules of thioretinamide bound to cobalamin, forming thioretinaco, complexed with ozone, oxygen, nicotinamide adenine dinucleotide. and inorganic phosphate, TR2CoO3O2NAD(+)H2PO4(-). Reduction of the pyridinium nitrogen of the nicotinamide group by an electron from electron transport complexes initiates polymerization of phosphate with adenosine diphosphate, yielding nicotinamide riboside and ATP bound to thioretinaco ozonide oxygen. A second electron reduces oxygen to hydroperoxyl radical, releasing ATP from the active site. A proton gradient is created within F1F0 ATPase complexes of mitochondria by reaction of protons with reduced nicotinamide riboside and with hydroperoxyl radical, yielding reduced nicotinamide riboside and hydroperoxide. The hyperhomocysteinemia of aging and dementia is attributed to decreased synthesis of adenosyl methionine by thioretinaco ozonide and ATP, causing decreased allosteric activation of cystathionine synthase and decreased allosteric inhibition of methylenetetrahydrofolate reductase and resulting in dysregulation of methionine metabolism. © 2015 by the Association of Clinical Scientists, Inc.

Plasmodesmal-mediated cell-to-cell transport in wheat roots is modulated by anaerobic stress

NASA Technical Reports Server (NTRS)

Cleland, R. E.; Fujiwara, T.; Lucas, W. J.

1994-01-01

Cell-to-cell transport of small molecules and ions occurs in plants through plasmodesmata. Plant roots are frequently subjected to localized anaerobic stress, with a resultant decrease in ATP. In order to determine the effect of this stress on plasmodesmal transport, fluorescent dyes of increasing molecular weight (0.46 to 1OkDa) were injected into epidermal and cortical cells of 3-day-old wheat roots, and their movement into neighboring cells was determined by fluorescence microscopy. Anaerobiosis was generated by N2 gas or simulated by the presence of sodium azide, both of which reduced the ATP levels in the tissue by over 80%. In the absence of such stress, the upper limit for movement, or size exclusion limit (SEL), of cortical plasmodesmata was <1 kDa. The ATP analogue TNP-ADP (mw 681) moved across the plasmodesmata of unstressed roots, indicating that plasmodesmata may be conduits for nucleotide (ATP and ADP) exchange between cells. Upon imposition of stress, the SEL rose to between 5 and 10 kDa. This response of plasmodesmata to a decrease in the level of ATP suggests that they are constricted by an ATP-dependent process so as to maintain a restricted SEL. When roots are subjected to anaerobic stress, an increase in SEL may permit enhanced delivery of sugars to the affected cells of the root where anaerobic respiration could regenerate the needed ATP.

Effects of catecholamines on rat myocardial metabolism. I. Influence of catecholamines on energy-rich nucleotides and phosphorylated fraction contents.

PubMed

Merouze, P; Gaudemer, Y

1975-01-01

1. The influence of catecholamines (adrenaline and noradrenaline) on energy metabolism of the rat myocardium has been studied by incubating slices of this tissue with these hormones and by following the levels of the different phosphorylated fractions and adenylic nucleotides. 2. Similar effects are obtained with both hormones, adrenaline being more effective. 3. Catecholamines decrease significantly the total amount of phosphate while Pi content increases during the first 10 minutes of incubation; labile and residual phosphate contents increase at the beginning of incubation and decrease to the initial values afterwards. 4. ATP and ADP levels decrease significantly with both hormones; however, the effect of noradrenalin on the ATP level needs a longer time of incubation. The ATP/ADP ratios decrease after 5 minutes incubation and the total adenylic nucleotide content is severely decreased (35 per cent with adrenalin, after 20 minutes incubation). 5. Similar results have been obtained with other tissues; these results can explain the decrease of aerobic metabolism we observed under the same conditions.

Influence of starvation on the larval development of Hyas araneus (Decapoda, Majidae)

NASA Astrophysics Data System (ADS)

Anger, K.; Dawirs, R. R.

1981-09-01

The influence of starvation on larval development of the spider crab Hyas araneus (L.) was studied in laboratory experiments. No larval stage suffering from continual lack of food had sufficient energy reserves to reach the next instar. Maximal survival times were observed at four different constant temperatures (2°, 6°, 12° and 18 °C). In general, starvation resistance decreased as temperatures increased: from 72 to 12days in the zoea-1, from 48 to 18 days in the zoea-2, and from 48 to 15 days in the megalopa stage. The length of maximal survival is of the same order of magnitude as the duration of each instar at a given temperature. “Sublethal limits” of early starvation periods were investigated at 12 °C: Zoea larvae must feed right from the beginning of their stage (at high food concentration) and for more than one fifth, approximately, of that stage to have at least some chance of surviving to the next instar, independent of further prey availability. The minimum time in which enough reserves are accumulated for successfully completing the instar without food is called “point-of-reserve-saturation” (PRS). If only this minimum period of essential initial feeding precedes starvation, development in both zoeal stages is delayed and mortality is greater, when compared to the fed control. Starvation periods beginning right after hatching of the first zoea cause a prolongation of this instar and, surprisingly, a slight shortening of the second stage. The delay in the zoea-1 increases proportionally to the length of the initial fasting period. If more than approximately 70 % of the maximum possible survival time has elapsed without food supply, the larvae become unable to recover and to moult to the second stage even when re-fed (“point-of-no-return”, PNR). The conclusion, based on own observations and on literature data, is that initial feeding is of paramount importance in the early development of planktotrophic decapod larvae. Taking into account hormonal and other developmental processes during the first moult cycle, a general hypothesis is proposed to explain the key role of first food uptake as well as the response pattern of the zoea-1 stage to differential starvation periods.

The effect of experimental gastric dilatation-volvulus on adenosine triphosphate content and conductance of the canine gastric and jejunal mucosa.

PubMed

Peycke, Laura E; Hosgood, Giselle; Davidson, Jacqueline R; Tetens, Joanne; Taylor, H Wayne

2005-07-01

The objective of this study was to determine if experimental gastric dilatation volvulus (GDV) would decrease adenosine triphosphate (ATP) concentration and increase membrane conductance of the canine gastric and jejunal mucosa. Male dogs (n = 15) weighing between 20 and 30 kg were used. Dogs were randomly assigned to 1 of 3 equal groups: Group 1 was control, group 2 was GDV, and group 3 was ischemia. All dogs were anesthetized for 210 min. Group 1 had no manipulation. Group 2 had GDV experimentally induced for 120 min followed by decompression, derotation, and reperfusion for 90 min. Group 3 had GDV experimentally induced for 210 min. Gastric (fundus and pylorus) and jejunal tissue was taken at 0, 120, and 210 min from all of the dogs. Tissue was analyzed for ATP concentration, mucosal conductance, and microscopic changes. The ATP concentration in the fundus did not change significantly from baseline in group 2, but decreased significantly below baseline at 210 min in group 3. The ATP concentration in the jejunum decreased significantly below baseline in groups 2 and 3 at 120 min, remaining significantly decreased in group 3 but returning to baseline at 210 min in group 2. Mucosal conductance of the fundus did not change significantly in any dog. Mucosal conductance of the jejunum increased at 120 min in groups 2 and 3, and became significantly increased above baseline at 210 min. The jejunal mucosa showed more profound cellular changes than the gastric mucosa. The jejunum showed substantial decreases in ATP concentration with an increase in mucosal conductance, suggesting cell membrane dysfunction. Dogs sustaining a GDV are likely to have a change in the activity of mucosal cells in the jejunum, which may be important in the pathophysiology of GDV.

Keratinocytes mediate innocuous and noxious touch via ATP-P2X4 signaling

PubMed Central

Moehring, Francie; Cowie, Ashley M; Menzel, Anthony D; Weyer, Andy D; Grzybowski, Michael; Arzua, Thiago; Geurts, Aron M; Palygin, Oleg

2018-01-01

The first point of our body’s contact with tactile stimuli (innocuous and noxious) is the epidermis, the outermost layer of skin that is largely composed of keratinocytes. Here, we sought to define the role that keratinocytes play in touch sensation in vivo and ex vivo. We show that optogenetic inhibition of keratinocytes decreases behavioral and cellular mechanosensitivity. These processes are inherently mediated by ATP signaling, as demonstrated by complementary cutaneous ATP release and degradation experiments. Specific deletion of P2X4 receptors in sensory neurons markedly decreases behavioral and primary afferent mechanical sensitivity, thus positioning keratinocyte-released ATP to sensory neuron P2X4 signaling as a critical component of baseline mammalian tactile sensation. These experiments lay a vital foundation for subsequent studies into the dysfunctional signaling that occurs in cutaneous pain and itch disorders, and ultimately, the development of novel topical therapeutics for these conditions. PMID:29336303

Lipid markers of diet history and their retention during experimental starvation in the Bering Sea euphausiid Thysanoessa raschii

NASA Astrophysics Data System (ADS)

Pleuthner, Rachel L.; Shaw, C. Tracy; Schatz, Megan J.; Lessard, Evelyn J.; Harvey, H. Rodger

2016-12-01

Two extended pulsed feeding experiments, following the spring bloom period, investigated lipid retention in the prominent Bering Sea euphausiid (krill) Thysanoessa raschii. These experiments occurred during late spring and early summer of 2010. Concurrent taxonomic analysis of the natural algal community allowed prey type to be linked to lipid composition of the natural communities. In late spring, experimental periods of feeding followed by starvation showed an overall decrease in total lipid for T. raschii. In early summer, no consistent trend was observed for total lipid with the visible presence of storage lipid in some animals. Polar lipids, as phospholipids, were the dominant krill lipid class in both experiments constituting ≥88% of total lipid, and triacylglycerols reached a maximum of 5% of total lipid. The sterols cholesterol and brassicasterol+desmosterol comprised 98-99% of total sterol abundances in T. raschii throughout both experiments, even after feeding periods when alternative sterols (i.e. the algal sterol 24-methylenecholesterol) accounted for up to 39% of sterols in potential food particles. Cholesterol abundance and concentration increased during both incubations, likely due to the metabolism of dietary sterols. Major fatty acids observed in krill included C14:0n, C16:0n, C16:1(n-7), C18:1(n-7), C18:1(n-9), C20:5(n-3), and C22:6(n-3) with the diatom-attributed C16:1(n-7) decreasing in abundance and concentration during starvation. Low concentrations of the dinoflagellate-derived sterol and a novel C28:8 PUFA, typically found in dinoflagellates and prymnesiophytes, indicated predation on protozooplankton in early summer when diatom abundances were low. The stability of lipid distributions over periods of starvation and intermittent feeding suggest that fatty acid and sterol biomarkers present in this polar euphausiid principally reflect long-term diet history rather than short-term feeding episodes.

Diadenosine tetraphosphate-gating of recombinant pancreatic ATP-sensitive K(+) channels.

PubMed

Jovanovic, S; Jovanovic, A

2001-02-01

Diadenosine tetraphosphate (Ap4A) has been recently discovered in the pancreatic beta cells where targets ATP-sensitive K(+) (K(ATP)) channels, depolarizes the cell membrane and induces insulin secretion. However, whether Ap4A inhibit pancreatic K(ATP) channels by targeting protein channel complex itself was unknown. Therefore, we coexpressed pancreatic K(ATP) channel subunits, Kir6.2 and SUR1, in COS-7 cells and examined the effect of Ap4A on the single channel behavior using the inside-out configuration of the patch-clamp technique. Ap4A inhibited channel opening in a concentration-dependent manner. Analysis of single channels demonstrated that Ap4A did not change intraburst kinetic behavior of K(ATP) channels, but rather decreased burst duration and increased between-burst duration. It is concluded that Ap4A antagonizes K(ATP) channel opening by targeting channel subunits themselves and by keeping channels longer in closed interburst states.

Inhibition of ATP Synthase by Chlorinated Adenosine Analogue

PubMed Central

Chen, Lisa S.; Nowak, Billie J.; Ayres, Mary L.; Krett, Nancy L.; Rosen, Steven T.; Zhang, Shuxing; Gandhi, Varsha

2009-01-01

8-Chloroadenosine (8-Cl-Ado) is a ribonucleoside analogue that is currently in clinical trial for chronic lymphocytic leukemia. Based on the decline in cellular ATP pool following 8-Cl-Ado treatment, we hypothesized that 8-Cl-ADP and 8-Cl-ATP may interfere with ATP synthase, a key enzyme in ATP production. Mitochondrial ATP synthase is composed of two major parts; FO intermembrane base and F1 domain, containing α and β subunits. Crystal structures of both α and β subunits that bind to the substrate, ADP, are known in tight binding (αdpβdp) and loose binding (αtpβtp) states. Molecular docking demonstrated that 8-Cl-ADP/8-Cl-ATP occupied similar binding modes as ADP/ATP in the tight and loose binding sites of ATP synthase, respectively, suggesting that the chlorinated nucleotide metabolites may be functional substrates and inhibitors of the enzyme. The computational predictions were consistent with our whole cell biochemical results. Oligomycin, an established pharmacological inhibitor of ATP synthase, decreased both ATP and 8-Cl-ATP formation from exogenous substrates, however, did not affect pyrimidine nucleoside analogue triphosphate accumulation. Synthesis of ATP from ADP was inhibited in cells loaded with 8-Cl-ATP. These biochemical studies are in consent with the computational modeling; in the αtpβtp state 8-Cl-ATP occupies similar binding as ANP, a non-hydrolyzable ATP mimic that is a known inhibitor. Similarly, in the substrate binding site (αdpβdp) 8-Cl-ATP occupies a similar position as ATP mimic ADP-BeF3 −. Collectively, our current work suggests that 8-Cl-ADP may serve as a substrate and the 8-Cl-ATP may be an inhibitor of ATP synthase. PMID:19477165

Extracellular ATP inhibits Schwann cell dedifferentiation and proliferation in an ex vivo model of Wallerian degeneration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shin, Youn Ho; Lee, Seo Jin; Jung, Junyang, E-mail: jjung@khu.ac.kr

Highlights: Black-Right-Pointing-Pointer ATP-treated sciatic explants shows the decreased expression of p75NGFR. Black-Right-Pointing-Pointer Extracellular ATP inhibits the expression of phospho-ERK1/2. Black-Right-Pointing-Pointer Lysosomal exocytosis is involved in Schwann cell dedifferentiation. Black-Right-Pointing-Pointer Extracellular ATP blocks Schwann cell proliferation in sciatic explants. -- Abstract: After nerve injury, Schwann cells proliferate and revert to a phenotype that supports nerve regeneration. This phenotype-changing process can be viewed as Schwann cell dedifferentiation. Here, we investigated the role of extracellular ATP in Schwann cell dedifferentiation and proliferation during Wallerian degeneration. Using several markers of Schwann cell dedifferentiation and proliferation in sciatic explants, we found that extracellular ATP inhibitsmore » Schwann cell dedifferentiation and proliferation during Wallerian degeneration. Furthermore, the blockage of lysosomal exocytosis in ATP-treated sciatic explants is sufficient to induce Schwann cell dedifferentiation. Together, these findings suggest that ATP-induced lysosomal exocytosis may be involved in Schwann cell dedifferentiation.« less

Host and Pathogen Copper-Transporting P-Type ATPases Function Antagonistically during Salmonella Infection.

PubMed

Ladomersky, Erik; Khan, Aslam; Shanbhag, Vinit; Cavet, Jennifer S; Chan, Jefferson; Weisman, Gary A; Petris, Michael J

2017-09-01

Copper is an essential yet potentially toxic trace element that is required by all aerobic organisms. A key regulator of copper homeostasis in mammalian cells is the copper-transporting P-type ATPase ATP7A, which mediates copper transport from the cytoplasm into the secretory pathway, as well as copper export across the plasma membrane. Previous studies have shown that ATP7A-dependent copper transport is required for killing phagocytosed Escherichia coli in a cultured macrophage cell line. In this investigation, we expanded on these studies by generating Atp7a LysMcre mice, in which the Atp7a gene was specifically deleted in cells of the myeloid lineage, including macrophages. Primary macrophages isolated from Atp7a LysMcre mice exhibit decreased copper transport into phagosomal compartments and a reduced ability to kill Salmonella enterica serovar Typhimurium compared to that of macrophages isolated from wild-type mice. The Atp7a LysMcre mice were also more susceptible to systemic infection by S Typhimurium than wild-type mice. Deletion of the S Typhimurium copper exporters, CopA and GolT, was found to decrease infection in wild-type mice but not in the Atp7a LysMcre mice. These studies suggest that ATP7A-dependent copper transport into the phagosome mediates host defense against S Typhimurium, which is counteracted by copper export from the bacteria via CopA and GolT. These findings reveal unique and opposing functions for copper transporters of the host and pathogen during infection. Copyright © 2017 American Society for Microbiology.

Fluoride decreased the sperm ATP of mice through inhabiting mitochondrial respiration.

PubMed

Sun, Zilong; Zhang, Wen; Xue, Xingchen; Zhang, Yuliang; Niu, Ruiyan; Li, Xuying; Li, Baojun; Wang, Xiaowen; Wang, Jundong

2016-02-01

Fluoride-induced low sperm motility was observed in accumulated investigations. However, the effect of fluoride exposure on ATP generation which is essential to sperm motility remains to be elucidated. In this study, 120 healthy male mice were orally administrated with 0, 25, 50, and 100 mg L(-1) NaF for 90 d. Results showed that compared with controls, fluoride ingestion significantly reduced sperm count, survival, as well as mobility and total ATP level in sperm untreated with carbonyl cyanide m-chlorophenylhydrazone (CCCP) or pyruvate, which was used to establish glycolysis or mitochondrial respiration model, respectively. Data further revealed that sperm mobility and ATP level under mitochondrial respiration condition were significantly suppressed, while no statistical difference occurred in the model of glycolysis, indicating ATP derived from mitochondria was affected. Moreover, mRNA expressions of mitochondrial cytochrome b (mt-Cytb) and cytochrome c oxidase subunit 2 (mt-COX2), two important molecules in mitochondrial electron transport chain (ETC), were down-regulated in all fluoride treatment groups. Mitochondria in sperm of mice exposed to 100 mg L(-1) NaF appeared to be irregular and vacuolated. These findings suggested that decreased sperm motility induced by fluoride may result from low ATP generation due to the disturbed ETC in sperm mitochondrial. Copyright © 2015 Elsevier Ltd. All rights reserved.

The P2X7 receptor antagonist, oxidized adenosine triphosphate, ameliorates renal ischemia-reperfusion injury by expansion of regulatory T cells.

PubMed

Koo, Tai Yeon; Lee, Jae-Ghi; Yan, Ji-Jing; Jang, Joon Young; Ju, Kyung Don; Han, Miyeun; Oh, Kook-Hwan; Ahn, Curie; Yang, Jaeseok

2017-08-01

Extracellular adenosine triphosphate (ATP) binds to purinergic receptors and, as a danger molecule, promotes inflammatory responses. Here we tested whether periodate-oxidized ATP (oATP), a P2X7 receptor (P2X7R) antagonist can attenuate renal ischemia-reperfusion injury and clarify the related cellular mechanisms. Treatment with oATP prior to ischemia-reperfusion injury decreased blood urea nitrogen, serum creatinine, the tubular injury score, and tubular epithelial cell apoptosis after injury. The infiltration of dendritic cells, neutrophils, macrophages, CD69 + CD4 + , and CD44 + CD4 + T cells was attenuated, but renal Foxp3 + CD4 + Treg infiltration was increased by oATP. The levels of IL-6 and CCL2 were reduced in the oATP group. Additionally, oATP treatment following injury improved renal function, decreased the infiltration of innate and adaptive effector cells, and increased the renal infiltration of Foxp3 + CD4 + Tregs. Post-ischemia-reperfusion injury oATP treatment increased tubular cell proliferation and reduced renal fibrosis. oATP treatment attenuated renal functional deterioration after ischemia-reperfusion injury in RAG-1 knockout mice; however, Treg depletion using PC61 abrogated the beneficial effects of oATP in wild-type mice. Furthermore, oATP treatment after transfer of Tregs from wild-type mice improved the beneficial effects of Tregs on ischemia-reperfusion injury, but treatment after transfer of Tregs from P2X7R knockout mice did not. Renal ischemia-reperfusion injury was also attenuated in P2X7R knockout mice. Experiments using bone marrow chimeras established that P2X7R expression on hematopoietic cells rather than non-hematopoietic cells, such as tubular epithelial cells, plays a major role in ischemia-reperfusion injury. Thus, oATP attenuated acute renal damage and facilitated renal recovery in ischemia-reperfusion injury by expansion of Tregs. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Mechanisms for the control of local tissue blood flow during thermal interventions: influence of temperature‐dependent ATP release from human blood and endothelial cells

PubMed Central

Chiesa, Scott T.; Trangmar, Steven J.; Ali, Leena; Lotlikar, Makrand D.; González‐Alonso, José

2017-01-01

New Findings What is the central question of this study? Skin and muscle blood flow increases with heating and decreases with cooling, but the temperature‐sensitive mechanisms underlying these responses are not fully elucidated. What is the main finding and its importance? We found that local tissue hyperaemia was related to elevations in ATP release from erythrocytes. Increasing intravascular ATP augmented skin and tissue perfusion to levels equal or above thermal hyperaemia. ATP release from isolated erythrocytes was altered by heating and cooling. Our findings suggest that erythrocytes are involved in thermal regulation of blood flow via modulation of ATP release. Local tissue perfusion changes with alterations in temperature during heating and cooling, but the thermosensitivity of the vascular ATP signalling mechanisms for control of blood flow during thermal interventions remains unknown. Here, we tested the hypotheses that the release of the vasodilator mediator ATP from human erythrocytes, but not from endothelial cells or other blood constituents, is sensitive to both increases and reductions in temperature and that increasing intravascular ATP availability with ATP infusion would potentiate thermal hyperaemia in limb tissues. We first measured blood temperature, brachial artery blood flow and plasma [ATP] during passive arm heating and cooling in healthy men and found that they increased by 3.0 ± 1.2°C, 105 ± 25 ml min−1 °C−1 and twofold, respectively, (all P < 0.05) with heating, but decreased or remained unchanged with cooling. In additional men, infusion of ATP into the brachial artery increased skin and deep tissue perfusion to levels equal or above thermal hyperaemia. In isolated erythrocyte samples exposed to different temperatures, ATP release increased 1.9‐fold from 33 to 39°C (P < 0.05) and declined by ∼50% at 20°C (P < 0.05), but no changes were observed in cultured human endothelial cells, plasma or serum samples. In conclusion, increases in plasma [ATP] and skin and deep tissue perfusion with limb heating are associated with elevations in ATP release from erythrocytes, but not from endothelial cells or other blood constituents. Erythrocyte ATP release is also sensitive to temperature reductions, suggesting that erythrocytes may function as thermal sensors and ATP signalling generators for control of tissue perfusion during thermal interventions. PMID:27859767

Previous Repeated Exposure to Food Limitation Enables Rats to Spare Lipid Stores during Prolonged Starvation.

PubMed

McCue, Marshall D; Albach, Audrey; Salazar, Giovanni

The risk of food limitation and, ultimately, starvation dates back to the dawn of heterotrophy in animals, yet starvation remains a major factor in the regulation of modern animal populations. Researchers studying starvation more than a century ago suggested that animals subjected to sublethal periods of food limitation are somehow more tolerant of subsequent starvation events. This possibility has received little attention over the past decades, yet it is highly relevant to modern science for two reasons. First, animals in natural populations are likely to be exposed to bouts of food limitation once or more before they face prolonged starvation, during which the risk of mortality becomes imminent. Second, our current approach to studying starvation physiology in the laboratory focuses on nourished animals with no previous exposure to nutritional stress. We examined the relationship between previous exposure to food limitation and potentially adaptive physiological responses to starvation in adult rats and found several significant differences. On two occasions, rats were fasted until they lost 20% of their body mass maintained lower body temperatures, and had presumably lower energy requirements when subjected to prolonged starvation than their naive cohort that never experienced food limitation. These rats that were trained in starvation also had lower plasma glucose set -points and reduced their reliance on endogenous lipid oxidation. These findings underscore (1) the need for biologists to revisit the classic hypothesis that animals can become habituated to starvation, using a modern set of research tools; and (2) the need to design controlled experiments of starvation physiology that more closely resemble the dynamic nature of food availability.

Fatigue and changes of ATP, creatine phosphate, and lactate during the 400-m sprint.

PubMed

Hirvonen, J; Nummela, A; Rusko, H; Rehunen, S; Härkönen, M

1992-06-01

Fatigue during the 400-m sprint was studied by measuring muscle ATP, creatine phosphate (CP), lactate (M-La), and blood lactate (B-La) in six male runners before and after four experimental sprints (100, 200, 300, and 400 m). During the first 100 m, muscle CP decreased from 15.8 +/- 1.7 to 8.3 +/- 0.3 mmol/kg while M-La increased to 3.6 +/- 0.4 mmol/kg. After 200 m the CP had decreased to 6.5 +/- 0.5 mmol/kg and M-La had increased to 8.3 +/- 1.1 mmol/kg. At the end of the 400 meters, ATP and CP concentrations had decreased by 27% and 89%, respectively, and M-La had increased to 17.3 +/- 0.9 mmol/kg. It was concluded that after 200 m the speed of running decreased, although CP was not depleted and lactate concentration was not at maximum level. Complete fatigue occurred when CP stores were depleted and B-La and M-La attained an individual maximum.

Mechanism of erythrocyte death in human population exposed to arsenic through drinking water

DOE Office of Scientific and Technical Information (OSTI.GOV)

Biswas, Debabrata; Banerjee, Mayukh; Sen, Gargi

2008-07-01

Arsenic contamination in drinking water is one of the biggest natural calamities, which has become an imperative threat to human health throughout the world. Abbreviation of erythrocyte lifespan leading to the development of anemia is a common sequel in arsenic exposed population. This study was undertaken to explore the mechanism of cell death in human erythrocytes during chronic arsenic exposure. Results revealed transformation of smooth discoid red cells into evaginated echinocytic form in the exposed individuals. Further distortion converted reversible echinocytes to irreversible spheroechinocytes. Arsenic toxicity increased membrane microviscosity along with an elevation of cholesterol/phospholipid ratio, which hampered the flexibilitymore » of red cell membrane and made them less deformable. Significant increase in the binding of merocyanine 540 with erythrocyte membrane due to arsenic exposure indicated disruption of lipid packing in the outer leaflet of the cell membrane resulting from altered transbilayer phospholipid asymmetry. Arsenic induced eryptosis was characterized by cell shrinkage and exposure of phosphatidylserine at the cell surface. Furthermore, metabolic starvation with depletion of cellular ATP triggered apoptotic removal of erythrocytes from circulation. Significant decrease in reduced glutathione content indicating defective antioxidant capacity was coupled with enhancement of malondialdehyde and protein carbonyl levels, which pointed to oxidative damage to erythrocyte membrane. Arsenic toxicity intervened into red cell membrane integrity eventually leading to membrane destabilization and hemoglobin release. The study depicted the involvement of both erythrophagocytosis and hemolysis in the destruction of human erythrocytes during chronic arsenic exposure.« less

Clusterin and COMMD1 Independently Regulate Degradation of the Mammalian Copper ATPases ATP7A and ATP7B*

PubMed Central

Materia, Stephanie; Cater, Michael A.; Klomp, Leo W. J.; Mercer, Julian F. B.; La Fontaine, Sharon

2012-01-01

ATP7A and ATP7B are copper-transporting P1B-type ATPases (Cu-ATPases) that are critical for regulating intracellular copper homeostasis. Mutations in the genes encoding ATP7A and ATP7B lead to copper deficiency and copper toxicity disorders, Menkes and Wilson diseases, respectively. Clusterin and COMMD1 were previously identified as interacting partners of these Cu-ATPases. In this study, we confirmed that clusterin and COMMD1 interact to down-regulate both ATP7A and ATP7B. Overexpression and knockdown of clusterin/COMMD1 decreased and increased, respectively, endogenous levels of ATP7A and ATP7B, consistent with a role in facilitating Cu-ATPase degradation. We demonstrate that whereas the clusterin/ATP7B interaction was enhanced by oxidative stress or mutation of ATP7B, the COMMD1/ATP7B interaction did not change under oxidative stress conditions, and only increased with ATP7B mutations that led to its misfolding. Clusterin and COMMD1 facilitated the degradation of ATP7B containing the same Wilson disease-causing C-terminal mutations via different degradation pathways, clusterin via the lysosomal pathway and COMMD1 via the proteasomal pathway. Furthermore, endogenous ATP7B existed in a complex with clusterin and COMMD1, but these interactions were neither competitive nor cooperative and occurred independently of each other. Together these data indicate that clusterin and COMMD1 represent alternative and independent systems regulating Cu-ATPase quality control, and consequently contributing to the maintenance of copper homeostasis. PMID:22130675

Effects of adenosine triphosphate concentration on motor force regulation during skeletal muscle contraction

NASA Astrophysics Data System (ADS)

Wei, J.; Dong, C.; Chen, B.

2017-04-01

We employ a mechanical model of sarcomere to quantitatively investigate how adenosine triphosphate (ATP) concentration affects motor force regulation during skeletal muscle contraction. Our simulation indicates that there can be negative cross-bridges resisting contraction within the sarcomere and higher ATP concentration would decrease the resistance force from negative cross-bridges by promoting their timely detachment. It is revealed that the motor force is well regulated only when ATP concentration is above a certain level. These predictions may provide insights into the role of ATP in regulating coordination among multiple motors.

Nutrient sensing signaling integrates nutrient metabolism and intestinal immunity in grass carp, Ctenopharyngodon idellus after prolonged starvation.

PubMed

Li, Shan; Li, Jiabo; Zhao, Yongliang; Zhang, Qin; Wang, Qingchao

2017-12-01

Starvation has been shown to affect growth and nutrient metabolism in fish; however, little information about the nutrient sensing signaling and mucosal adaptive immunity in fish was known. In the present study, grass carp was starved for 8weeks to simulate the natural aquaculture practice in Hubei during winter. The histology of liver was significantly affected with decreased expression of tight junction proteins including claudin-3, claudin-b and ZO-1. Muscle gene expression was also affected, with decreased expression of muscle growth promoting factors such as Myogenin, MyoD, Myf5, and increased expression of muscle degradation factors, such as CathepsinD. In addition, mucosal adaptive immunity was also significantly affected, with decreased expression of antibodies including IgZ and IgM in gut. Along with these changes was the inhibition of several nutrient sensing signaling including MAPK and TOR signaling, which leads to the inhibition of the synthesis of protein including immunoglobulin. The increased phosphorylation of eIF2α not only inhibited the translation, but also resulted in the decreased expression of IkB and increased expression of NF-B, with the activation of pro-inflammatory genes including IL8 and TNF. Copyright © 2017 Elsevier Ltd. All rights reserved.

Awakening of a Dormant Cyanobacterium from Nitrogen Chlorosis Reveals a Genetically Determined Program.

PubMed

Klotz, Alexander; Georg, Jens; Bučinská, Lenka; Watanabe, Satoru; Reimann, Viktoria; Januszewski, Witold; Sobotka, Roman; Jendrossek, Dieter; Hess, Wolfgang R; Forchhammer, Karl

2016-11-07

The molecular and physiological mechanisms involved in the transition of microbial cells from a resting state to the active vegetative state are critically relevant for solving problems in fields ranging from microbial ecology to infection microbiology. Cyanobacteria that cannot fix nitrogen are able to survive prolonged periods of nitrogen starvation as chlorotic cells in a dormant state. When provided with a usable nitrogen source, these cells re-green within 48 hr and return to vegetative growth. Here we investigated the resuscitation of chlorotic Synechocystis sp. PCC 6803 cells at the physiological and molecular levels with the aim of understanding the awakening process of a dormant bacterium. Almost immediately upon nitrate addition, the cells initiated a highly organized resuscitation program. In the first phase, they suppressed any residual photosynthetic activity and activated respiration to gain energy from glycogen catabolism. Concomitantly, they restored the entire translational apparatus, ATP synthesis, and nitrate assimilation. After only 12-16 hr, the cells re-activated the synthesis of the photosynthetic apparatus and prepared for metabolic re-wiring toward photosynthesis. When the cells reached full photosynthetic capacity after ∼48 hr, they resumed cell division and entered the vegetative cell cycle. An analysis of the transcriptional dynamics during the resuscitation process revealed a perfect match to the observed physiological processes, and it suggested that non-coding RNAs play a major regulatory role during the lifestyle switch in awakening cells. This genetically encoded program ensures rapid colonization of habitats in which nitrogen starvation imposes a recurring growth limitation. Copyright © 2016 Elsevier Ltd. All rights reserved.

Extracellular ATP inhibits root gravitropism at concentrations that inhibit polar auxin transport

NASA Technical Reports Server (NTRS)

Tang, Wenqiang; Brady, Shari R.; Sun, Yu; Muday, Gloria K.; Roux, Stanley J.

2003-01-01

Raising the level of extracellular ATP to mM concentrations similar to those found inside cells can block gravitropism of Arabidopsis roots. When plants are grown in Murashige and Skoog medium supplied with 1 mM ATP, their roots grow horizontally instead of growing straight down. Medium with 2 mM ATP induces root curling, and 3 mM ATP stimulates lateral root growth. When plants are transferred to medium containing exogenous ATP, the gravity response is reduced or in some cases completely blocked by ATP. Equivalent concentrations of ADP or inorganic phosphate have slight but usually statistically insignificant effects, suggesting the specificity of ATP in these responses. The ATP effects may be attributable to the disturbance of auxin distribution in roots by exogenously applied ATP, because extracellular ATP can alter the pattern of auxin-induced gene expression in DR5-beta-glucuronidase transgenic plants and increase the response sensitivity of plant roots to exogenously added auxin. The presence of extracellular ATP also decreases basipetal auxin transport in a dose-dependent fashion in both maize (Zea mays) and Arabidopsis roots and increases the retention of [(3)H]indole-3-acetic acid in root tips of maize. Taken together, these results suggest that the inhibitory effects of extracellular ATP on auxin distribution may happen at the level of auxin export. The potential role of the trans-plasma membrane ATP gradient in auxin export and plant root gravitropism is discussed.

Prostatic relaxation induced by agmatine is decreased in spontaneously hypertensive rats.

PubMed

Lee, Liang-Ming; Tsai, Tsung-Chin; Chung, Hsien-Hui; Tong, Yat-Ching; Cheng, Juei-Tang

2012-09-01

What's known on the subject? and What does the study add? Neurotransmitters are known to control prostate contractility. Agmatine is one of them and induces relaxation through imidazoline receptors. The paper shows that the action of agmatine is reduced in hypertensive rats, and that this change is related to the decrease of ATP-sensitive potassium channels in the prostate. The findings can increase our understanding of the possible underlying mechanism for the development of clinical benign prostatic hyperplasia. To compare agmatine-induced prostatic relaxation in hypertensive and control rats. To investigate the responsible mechanism(s) and the role of the ATP-sensitive potassium channel. Prostate strips were isolated from male spontaneously hypertensive (SH) rats and normal Wistar-Kyoto (WKY) rats for measurement of isometric tension. The strips were precontracted with 1 µmol/L phenylephrine or 50 mmol/L KCl. Dose-dependent relaxation of the prostatic strips was studied by cumulative administration of agmatine, 1 to 100 µmol/L, into the organ bath. Effects of specific antagonists on agmatine-induced relaxation were studied. Western blotting analysis was used to measure the gene expression of the ATP-sensitive potassium channel in the rat prostate. Prostatic relaxation induced by agmatine was markedly reduced in SH rats compared with WKY rats. The relaxation caused by agmatine was abolished by BU224, a selective imidazoline I(2)-receptor antagonist, but was not modified by efaroxan at a dose sufficient to block imidazoline I(1)-receptors. The relaxation induced by diazoxide at a concentration sufficient to activate ATP-sensitive potassium channels was markedly reduced in the SH rat prostate. Expressions of ATP-sensitive potassium channel sulphonylurea receptor and inwardly rectifying potassium channel (Kir) 6.2 subunits were both decreased in the prostate of SH rats. The decrease of agmatine-induced prostatic relaxation in SH rats is related to the change in ATP-sensitive potassium channels. © 2012 THE AUTHORS. BJU INTERNATIONAL © 2012 BJU INTERNATIONAL.

The Response of Diatom Central Carbon Metabolism to Nitrogen Starvation Is Different from That of Green Algae and Higher Plants1[W

PubMed Central

Hockin, Nicola Louise; Mock, Thomas; Mulholland, Francis; Kopriva, Stanislav; Malin, Gill

2012-01-01

The availability of nitrogen varies greatly in the ocean and limits primary productivity over large areas. Diatoms, a group of phytoplankton that are responsible for about 20% of global carbon fixation, respond rapidly to influxes of nitrate and are highly successful in upwelling regions. Although recent diatom genome projects have highlighted clues to the success of this group, very little is known about their adaptive response to changing environmental conditions. Here, we compare the proteome of the marine diatom Thalassiosira pseudonana (CCMP 1335) at the onset of nitrogen starvation with that of nitrogen-replete cells using two-dimensional gel electrophoresis. In total, 3,310 protein spots were distinguishable, and we identified 42 proteins increasing and 23 decreasing in abundance (greater than 1.5-fold change; P < 0.005). Proteins involved in the metabolism of nitrogen, amino acids, proteins, and carbohydrates, photosynthesis, and chlorophyll biosynthesis were represented. Comparison of our proteomics data with the transcriptome response of this species under similar growth conditions showed good correlation and provided insight into different levels of response. The T. pseudonana response to nitrogen starvation was also compared with that of the higher plant Arabidopsis (Arabidopsis thaliana), the green alga Chlamydomonas reinhardtii, and the cyanobacterium Prochlorococcus marinus. We have found that the response of diatom carbon metabolism to nitrogen starvation is different from that of other photosynthetic eukaryotes and bears closer resemblance to the response of cyanobacteria. PMID:22065419

Macrophage migration inhibitory factor plays a permissive role in the maintenance of cardiac contractile function under starvation through regulation of autophagy.

PubMed

Xu, Xihui; Pacheco, Benjamin D; Leng, Lin; Bucala, Richard; Ren, Jun

2013-08-01

The cytokine macrophage migration inhibitory factor (MIF) protects the heart through AMPK activation. Autophagy, a conserved pathway for bulk degradation of intracellular proteins and organelles, helps preserve and recycle energy and nutrients for cells to survive under starvation. This study was designed to examine the role of MIF in cardiac homeostasis and autophagy regulation following an acute starvation challenge. Wild-type (WT) and MIF knockout mice were starved for 48 h. Echocardiographic data revealed little effect of starvation on cardiac geometry, contractile and intracellular Ca²⁺ properties. MIF deficiency unmasked an increase in left ventricular end-systolic diameter, a drop in fractional shortening associated with cardiomyocyte contractile and intracellular Ca²⁺ anomalies following starvation. Interestingly, the unfavourable effect of MIF deficiency was associated with interruption of starvation-induced autophagy. Furthermore, restoration of autophagy using rapamycin partially protected against starvation-induced cardiomyocyte contractile defects. In our in vitro model of starvation, neonatal mouse cardiomyocytes from WT and MIF-/- mice and H9C2 cells were treated with serum free-glucose free DMEM for 2 h. MIF depletion dramatically attenuated starvation-induced autophagic vacuole formation in neonatal mouse cardiomyocytes and exacerbated starvation-induced cell death in H9C2 cells. In summary, these results indicate that MIF plays a permissive role in the maintenance of cardiac contractile function under starvation by regulation of autophagy.

Phosphate starvation induced OsPHR4 mediates Pi-signaling and homeostasis in rice.

PubMed

Ruan, Wenyuan; Guo, Meina; Wu, Ping; Yi, Keke

2017-02-01

OsPHR4 mediates the regulation of Pi-starvation signaling and Pi-homeostasis in a PHR1-subfamily dependent manner in rice. Phosphate (Pi) starvation response is a sophisticated process for plant in the natural environment. In this process, PHOSPHATE STARVATION RESPONSE 1 (PHR1) subfamily genes play a central role in regulating Pi-starvation signaling and Pi-homeostasis. Besides the three PHR1 orthologs in Oryza sativa L. (Os) [(Os) PHR1, (Os) PHR2, and (Os) PHR3], which were reported to regulated Pi-starvation signaling and Pi-homeostasis redundantly, a close related PHR1 ortholog [designated as (Os) PHR4] is presented in rice genome with unknown function. In this study, we found that OsPHR4 is a Pi-starvation induced gene and mainly expresses in vascular tissues through all growth and development periods. The expression of OsPHR4 is positively regulated by OsPHR1, OsPHR2 and OsPHR3. The nuclear located OsPHR4 can respectively interact with other three PHR1 subfamily members to regulate downstream Pi-starvation induced genes. Consistent with the positive role of PHR4 in regulating Pi-starvation signaling, the OsPHR4 overexpressors display higher Pi accumulation in the shoot and elevated expression of Pi-starvation induced genes under Pi-sufficient condition. Besides, moderate growth retardation and repression of the Pi-starvation signaling in the OsPHR4 RNA interfering (RNAi) transgenic lines can be observed under Pi-deficient condition. Together, we propose that OsPHR4 mediates the regulation of Pi-starvation signaling and Pi-homeostasis in a PHR1-subfamily dependent manner in rice.

S-Sulfhydration of ATP synthase by hydrogen sulfide stimulates mitochondrial bioenergetics.

PubMed

Módis, Katalin; Ju, YoungJun; Ahmad, Akbar; Untereiner, Ashley A; Altaany, Zaid; Wu, Lingyun; Szabo, Csaba; Wang, Rui

2016-11-01

Mammalian cells can utilize hydrogen sulfide (H 2 S) to support mitochondrial respiration. The aim of our study was to explore the potential role of S-sulfhydration (a H 2 S-induced posttranslational modification, also known as S-persulfidation) of the mitochondrial inner membrane protein ATP synthase (F1F0 ATP synthase/Complex V) in the regulation of mitochondrial bioenergetics. Using a biotin switch assay, we have detected S-sulfhydration of the α subunit (ATP5A1) of ATP synthase in response to exposure to H 2 S in vitro. The H 2 S generator compound NaHS induced S-sulfhydration of ATP5A1 in HepG2 and HEK293 cell lysates in a concentration-dependent manner (50-300μM). The activity of immunocaptured mitochondrial ATP synthase enzyme isolated from HepG2 and HEK293 cells was stimulated by NaHS at low concentrations (10-100nM). Site-directed mutagenesis of ATP5A1 in HEK293 cells demonstrated that cysteine residues at positions 244 and 294 are subject to S-sulfhydration. The double mutant ATP synthase protein (C244S/C294S) showed a significantly reduced enzyme activity compared to control and the single-cysteine-mutated recombinant proteins (C244S or C294S). To determine whether endogenous H 2 S plays a role in the basal S-sulfhydration of ATP synthase in vivo, we compared liver tissues harvested from wild-type mice and mice deficient in cystathionine-gamma-lyase (CSE, one of the three principal mammalian H 2 S-producing enzymes). Significantly reduced S-sulfhydration of ATP5A1 was observed in liver homogenates of CSE -/- mice, compared to wild-type mice, suggesting a physiological role for CSE-derived endogenous H 2 S production in the S-sulfhydration of ATP synthase. Various forms of critical illness (including burn injury) upregulate H 2 S-producing enzymes and stimulate H 2 S biosynthesis. In liver tissues collected from mice subjected to burn injury, we detected an increased S-sulfhydration of ATP5A1 at the early time points post-burn. At later time points (when systemic H 2 S levels decrease) S-sulfhydration of ATP5A1 decreased as well. In conclusion, H 2 S induces S-sulfhydration of ATP5A1 at C244 and C294. This post-translational modification may be a physiological mechanism to maintain ATP synthase in a physiologically activated state, thereby supporting mitochondrial bioenergetics. The sulfhydration of ATP synthase may be a dynamic process, which may be regulated by endogenous H 2 S levels under various pathophysiological conditions. Copyright © 2016 Elsevier Ltd. All rights reserved.

Regulatory Response to Carbon Starvation in Caulobacter crescentus

PubMed Central

Britos, Leticia; Abeliuk, Eduardo; Taverner, Thomas; Lipton, Mary; McAdams, Harley; Shapiro, Lucy

2011-01-01

Bacteria adapt to shifts from rapid to slow growth, and have developed strategies for long-term survival during prolonged starvation and stress conditions. We report the regulatory response of C. crescentus to carbon starvation, based on combined high-throughput proteome and transcriptome analyses. Our results identify cell cycle changes in gene expression in response to carbon starvation that involve the prominent role of the FixK FNR/CAP family transcription factor and the CtrA cell cycle regulator. Notably, the SigT ECF sigma factor mediates the carbon starvation-induced degradation of CtrA, while activating a core set of general starvation-stress genes that respond to carbon starvation, osmotic stress, and exposure to heavy metals. Comparison of the response of swarmer cells and stalked cells to carbon starvation revealed four groups of genes that exhibit different expression profiles. Also, cell pole morphogenesis and initiation of chromosome replication normally occurring at the swarmer-to-stalked cell transition are uncoupled in carbon-starved cells. PMID:21494595

Systems biology approach to transplant tolerance: proof of concept experiments using RNA interference (RNAi) to knock down hub genes in Jurkat and HeLa cells in vitro.

PubMed

Lwin, Wint Wah; Park, Ken; Wauson, Matthew; Gao, Qin; Finn, Patricia W; Perkins, David; Khanna, Ajai

2012-07-01

Systems biology is gaining importance in studying complex systems such as the functional interconnections of human genes [1]. To investigate the molecular interactions involved in T cell immune responses, we used databases of physical gene-gene interactions to constructed molecular interaction networks (interconnections) with R language algorithms. This helped to identify highly interconnected "hub" genes AT(1)P5C1, IL6ST, PRKCZ, MYC, FOS, JUN, and MAPK1. We hypothesized that suppression of these hub genes in the gene network would result in significant phenotypic effects on T cells and examined this in vitro. The molecular interaction networks were then analyzed and visualized with Cytoscape. Jurkat and HeLa cells were transfected with siRNA for the selected hub genes. Cell proliferation was measured using ATP luminescence and BrdU labeling, which were measured 36, 72, and 96 h after activation. Following T cell stimulation, we found a significant decrease in ATP production (P < 0.05) when the hub genes ATP5C1 and PRKCZ were knocked down using siRNA transfection, whereas no difference in ATP production was observed in siRNA transfected HeLa cells. However, HeLa cells showed a significant (P < 0.05) decrease in cell proliferation when the genes MAPK1, IL6ST, ATP5C1, JUN, and FOS were knocked down. In both Jurkat and HeLa cells, targeted gene knockdown using siRNA showed decreased cell proliferation and ATP production in both Jurkat and HeLa cells. However, Jurkat T cells and HELA cells use different hub genes to regulate activation responses. This experiment provides proof of principle of applying siRNA knockdown of T cell hub genes to evaluate their proliferative capacity and ATP production. This novel concept outlines a systems biology approach to identify hub genes for targeted therapeutics. Published by Elsevier Inc.

Oral Adenosine-5'-triphosphate (ATP) Administration Increases Postexercise ATP Levels, Muscle Excitability, and Athletic Performance Following a Repeated Sprint Bout.

PubMed

Purpura, Martin; Rathmacher, John A; Sharp, Matthew H; Lowery, Ryan P; Shields, Kevin A; Partl, Jeremy M; Wilson, Jacob M; Jäger, Ralf

2017-01-01

Oral adenosine-5'-triphosphate (ATP) administration has failed to increase plasma ATP levels; however, chronic supplementation with ATP has shown to increase power, strength, lean body mass, and blood flow in trained athletes. The purpose of this study was to investigate the effects of ATP supplementation on postexercise ATP levels and on muscle activation and excitability and power following a repeated sprint bout. In a double-blind, placebo-controlled, randomized design, 42 healthy male individuals were given either 400 mg of ATP as disodium salt or placebo for 2 weeks prior to an exercise bout. During the exercise bout, muscle activation and excitability (ME, ratio of power output to muscle activation) and Wingate test peak power were measured during all sprints. ATP and metabolites were measured at baseline, after supplementation, and immediately following exercise. Oral ATP supplementation prevented a drop in ATP, adenosine-5'-diphosphate (ADP), and adenosine-5'-monophosphate (AMP) levels postexercise (p < 0.05). No group by time interaction was observed for muscle activation. Following the supplementation period, muscle excitability significantly decreased in later bouts 8, 9, and 10 in the placebo group (-30.5, -28.3, and -27.9%, respectively; p < 0.02), whereas ATP supplementation prevented the decline in later bouts. ATP significantly increased Wingate peak power in later bouts compared to baseline (bout 8: +18.3%, bout 10: +16.3%). Oral ATP administration prevents exercise-induced declines in ATP and its metabolite and enhances peak power and muscular excitability, which may be beneficial for sports requiring repeated high-intensity sprinting bouts.

Regulation of respiration in brain mitochondria and synaptosomes: restrictions of ADP diffusion in situ, roles of tubulin, and mitochondrial creatine kinase.

PubMed

Monge, Claire; Beraud, Nathalie; Kuznetsov, Andrey V; Rostovtseva, Tatiana; Sackett, Dan; Schlattner, Uwe; Vendelin, Marko; Saks, Valdur A

2008-11-01

The role of ubiquitous mitochondrial creatine kinase (uMtCK) reaction in regulation of mitochondrial respiration was studied in purified preparations of rat brain synaptosomes and mitochondria. In permeabilized synaptosomes, apparent Km for exogenous ADP, Km (ADP), in regulation of respiration in situ was rather high (110 +/- 11 microM) in comparison with isolated brain mitochondria (9 +/- 1 microM). This apparent Km for ADP observed in isolated mitochondria in vitro dramatically increased to 169 +/- 52 microM after their incubation with 1 muM of dimeric tubulin showing that in rat brain, particularly in synaptosomes, mitochondrial outer membrane permeability for ADP, and ATP may be restricted by tubulin binding to voltage dependent anion channel (VDAC). On the other hand, in synaptosomes apparent Km (ADP) decreased to 25 +/- 1 microM in the presence of 20 mM creatine. To fully understand this effect of creatine on kinetics of respiration regulation, complete kinetic analysis of uMtCK reaction in isolated brain mitochondria was carried out. This showed that oxidative phosphorylation specifically altered only the dissociation constants for MgATP, by decreasing that from ternary complex MtCK.Cr.MgATP (K (a)) from 0.13 +/- 0.02 to 0.018 +/- 0.007 mM and that from binary complex MtCK.MgATP (K (ia)) from 1.1 +/- 0.29 mM to 0.17 +/- 0.07 mM. Apparent decrease of dissociation constants for MgATP reflects effective cycling of ATP and ADP between uMtCK and adenine nucleotide translocase (ANT). These results emphasize important role and various pathophysiological implications of the phosphocreatine-creatine kinase system in energy transfer in brain cells, including synaptosomes.

Cross-talk between Phosphate Starvation and Other Environmental Stress Signaling Pathways in Plants

PubMed Central

Baek, Dongwon; Chun, Hyun Jin; Yun, Dae-Jin; Kim, Min Chul

2017-01-01

The maintenance of inorganic phosphate (Pi) homeostasis is essential for plant growth and yield. Plants have evolved strategies to cope with Pi starvation at the transcriptional, post-transcriptional, and post-translational levels, which maximizes its availability. Many transcription factors, miRNAs, and transporters participate in the Pi starvation signaling pathway where their activities are modulated by sugar and phytohormone signaling. Environmental stresses significantly affect the uptake and utilization of nutrients by plants, but their effects on the Pi starvation response remain unclear. Recently, we reported that Pi starvation signaling is affected by abiotic stresses such as salt, abscisic acid, and drought. In this review, we identified transcription factors, such as MYB, WRKY, and zinc finger transcription factors with functions in Pi starvation and other environmental stress signaling. In silico analysis of the promoter regions of Pi starvation-responsive genes, including phosphate transporters, microRNAs, and phosphate starvation–induced genes, suggest that their expression may be regulated by other environmental stresses, such as hormones, drought, cold, heat, and pathogens as well as by Pi starvation. Thus, we suggest the possibility of cross-talk between Pi starvation signaling and other environmental stress signaling pathways. PMID:29047263

A Transient Rise in Free Mg2+ Ions Released from ATP-Mg Hydrolysis Contributes to Mitotic Chromosome Condensation.

PubMed

Maeshima, Kazuhiro; Matsuda, Tomoki; Shindo, Yutaka; Imamura, Hiromi; Tamura, Sachiko; Imai, Ryosuke; Kawakami, Syoji; Nagashima, Ryosuke; Soga, Tomoyoshi; Noji, Hiroyuki; Oka, Kotaro; Nagai, Takeharu

2018-02-05

For cell division, negatively charged chromatin, in which nucleosome fibers (10 nm fibers) are irregularly folded [1-5], must be condensed into chromosomes and segregated. While condensin and other proteins are critical for organizing chromatin into the appropriate chromosome shape [6-17], free divalent cations such as Mg 2+ and Ca 2+ , which condense chromatin or chromosomes in vitro [18-28], have long been considered important, especially for local condensation, because the nucleosome fiber has a net negative charge and is by itself stretched like "beads on a string" by electrostatic repulsion. For further folding, other positively charged factors are required to decrease the charge and repulsion [29]. However, technical limitations to measure intracellular free divalent cations, but not total cations [30], especially Mg 2+ , have prevented us from elucidating their function. Here, we developed a Förster resonance energy transfer (FRET)-based Mg 2+ indicator that monitors free Mg 2+ dynamics throughout the cell cycle. By combining this indicator with Ca 2+ [31] and adenosine triphosphate (ATP) [32] indicators, we demonstrate that the levels of free Mg 2+ , but not Ca 2+ , increase during mitosis. The Mg 2+ increase is coupled with a decrease in ATP, which is normally bound to Mg 2+ in the cell [33]. ATP inhibited Mg 2+ -dependent chromatin condensation in vitro. Chelating Mg 2+ induced mitotic cell arrest and chromosome decondensation, while ATP reduction had the opposite effect. Our results suggest that ATP-bound Mg 2+ is released by ATP hydrolysis and contributes to mitotic chromosome condensation with increased rigidity, suggesting a novel regulatory mechanism for higher-order chromatin organization by the intracellular Mg 2+ -ATP balance. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Substrate recognition by the hetero-octameric ATP phosphoribosyltransferase from Lactococcus lactis†

PubMed Central

Champagne, Karen S.; Piscitelli, Elise; Francklyn, Christopher S.

2008-01-01

Two families of ATP phosphoribosyl transferases (ATP-PRT) join ATP and 5-phosphoribosyl-1 pyrophosphate (PRPP) in the first reaction of histidine biosynthesis. These consist of a homohexameric form found in all three kingdoms, and a hetero-octameric form largely restricted to bacteria. Hetero-octameric ATP-PRTs consist of four HisGS catalytic subunits related to periplasmic binding proteins, and four HisZ regulatory subunits that resemble histidyl-tRNA synthetases. To clarify the relationship between the two families of ATP-PRTs, and among phosphoribosyltransferases in general, we determined the steady state kinetics for the hetero-octameric form, and characterized the active site by mutagenesis. The Km PRPP (18.4 ± 3.5 μM) and kcat (2.7 ± 0.3 sec−1) values for the PRPP substrate are similar to those of hexameric ATP-PRTs, but the Km for ATP (2.7 ± 0.3 mM) is 4-fold higher, suggestive of tighter regulation by energy charge. Histidine and AMP were determined to be non-competitive (Ki = 81.1 μM) and competitive (Ki= 1.44 mM) inhibitors, respectively, with values that approximate their intracellular concentrations. Mutagenesis experiments investigating the side chains recognizing PRPP showed that 5′ phosphate contacts (T159A and T162A) had the largest (25- and 155-fold) decreases in kcat/Km, while smaller decreases were seen with mutants making cross subunit contacts (K50A and K8A) to the pyrophosphate moiety, or contacts to the 2′ OH. Despite their markedly different quaternary structures, hexameric and hetero-octameric ATRP-PRTs exhibit similar functional parameters, and employ mechanistic strategies reminiscent of the broader PRT superfamily. PMID:17154531

Extracellular adenosine triphosphate affects systemic and kidney immune cell populations in pregnant rats.

PubMed

Spaans, Floor; Melgert, Barbro N; Borghuis, Theo; Klok, Pieter A; de Vos, Paul; Bakker, Winston W; van Goor, Harry; Faas, Marijke M

2014-09-01

Changes in the systemic immune response are found in preeclampsia. This may be related to high extracellular adenosine triphosphate (ATP) levels. The question arose whether ATP could affect immune responses in pregnancy. Previously, we investigated whether ATP affected monocyte activation and subpopulations. Here, we investigated ATP-induced changes in other immune cell populations in pregnant rats, systemically and in the kidney, an affected organ in preeclampsia. Using flow cytometry or immunohistochemistry, blood and kidney leukocytes were studied in pregnant and non-pregnant rats at different intervals after ATP or saline infusion. Adenosine triphosphate (ATP) infusion induced increased peripheral blood non-classical monocytes and decreased T lymphocyte subsets in pregnant rats only, higher glomerular macrophage and T lymphocyte numbers in non-pregnant animals 1 day after infusion, and higher glomerular macrophage numbers in pregnant rats 6 days after infusion. Adenosine triphosphate (ATP) infusion in pregnant rats induced a pregnancy-specific inflammatory response. Increased ATP levels could potentially contribute to development of the inflammatory response of preeclampsia. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Exercise-Induced Changes in Metabolic Resposes to Infection in Trained Rats.

DTIC Science & Technology

1981-07-20

superimposed on fasting reduced it in sedentary rats (24). Swimming amplified fasting ketosis and diminished infection ketonemia and markedly decreased...Ketone body metabolism in the ketosis of starvation and alloxan diabetes. J. Biol. Chem. 245:4382-4390, 1970. 22. MOSES, L. E. Determination of oxygen

Novel VCP modulators mitigate major pathologies of rd10, a mouse model of retinitis pigmentosa

PubMed Central

Ikeda, Hanako Ohashi; Sasaoka, Norio; Koike, Masaaki; Nakano, Noriko; Muraoka, Yuki; Toda, Yoshinobu; Fuchigami, Tomohiro; Shudo, Toshiyuki; Iwata, Ayana; Hori, Seiji; Yoshimura, Nagahisa; Kakizuka, Akira

2014-01-01

Neuroprotection may prevent or forestall the progression of incurable eye diseases, such as retinitis pigmentosa, one of the major causes of adult blindness. Decreased cellular ATP levels may contribute to the pathology of this eye disease and other neurodegenerative diseases. Here we describe small compounds (Kyoto University Substances, KUSs) that were developed to inhibit the ATPase activity of VCP (valosin-containing protein), the most abundant soluble ATPase in the cell. Surprisingly, KUSs did not significantly impair reported cellular functions of VCP but nonetheless suppressed the VCP-dependent decrease of cellular ATP levels. Moreover, KUSs, as well as exogenous ATP or ATP-producing compounds, e.g. methylpyruvate, suppressed endoplasmic reticulum stress, and demonstrably protected various types of cultured cells from death, including several types of retinal neuronal cells. We then examined their in vivo efficacies in rd10, a mouse model of retinitis pigmentosa. KUSs prevented photoreceptor cell death and preserved visual function. These results reveal an unexpected, crucial role of ATP consumption by VCP in determining cell fate in this pathological context, and point to a promising new neuroprotective strategy for currently incurable retinitis pigmentosa. PMID:25096051

Why Do Fast-Growing Bacteria Enter Overflow Metabolism? Testing the Membrane Real Estate Hypothesis.

PubMed

Szenk, Mariola; Dill, Ken A; de Graff, Adam M R

2017-08-23

Bacteria and other cells show a puzzling behavior. At high growth rates, E. coli switch from respiration (which is ATP-efficient) to using fermentation for additional ATP (which is inefficient). This overflow metabolism results in a several-fold decrease in ATP produced per glucose molecule provided as food. By integrating diverse types of experimental data into a simple biophysical model, we give evidence that this onset is the result of the membrane real estate hypothesis: Fast growth drives cells to be bigger, reducing their surface-to-volume ratios. This decreases the membrane area available for respiratory proteins despite growing demand, causing increased crowding. Only when respiratory proteins reach their crowding limit does the cell activate fermentation, since fermentation allows faster ATP production per unit membrane area. Surface limitation thus creates a Pareto trade-off between membrane efficiency and ATP yield that links metabolic choice to the size and shape of a bacterial cell. By exploring the predictions that emerge from this trade-off, we show how consideration of molecular structures, energetics, rates, and equilibria can provide important insight into cellular behavior. Copyright © 2017 Elsevier Inc. All rights reserved.

Establishment of a chronic activity-based anorexia rat model.

PubMed

Frintrop, Linda; Trinh, Stefanie; Liesbrock, Johanna; Paulukat, Lisa; Kas, Martien J; Tolba, Rene; Konrad, Kerstin; Herpertz-Dahlmann, Beate; Beyer, Cordian; Seitz, Jochen

2018-01-01

Anorexia nervosa (AN) is often a chronic eating disorder characterised by body image disturbance and low body weight often associated with starvation-induced amenorrhoea and excessive exercise. Activity-based anorexia (ABA) is an animal model representing many somatic aspects of this psychiatric illness. We systematically manipulated the extent and length of starvation and animal age to find the optimal parameters to study chronic starvation. Wistar rats had 24h/day running wheel access and received 40% of their baseline food intake until a 20% or 25% weight reduction was reached (acute starvation). This body weight was then maintained for two weeks (chronic starvation). The rats of different ages of 4 or 8 weeks were used to represent early and late adolescent animals, respectively. The complete absence of a menstrual cycle was defined as the primary outcome parameter. Acute starvation caused a disruption of the oestrous cycle in 58% of the animals. During chronic starvation, a complete loss of the oestrous cycle could be found. Furthermore, 4-week-old rats exhibited higher levels of hyperactivity and amenorrhoea than 8-week-old animals. A 20% starvation level led to 90% loss of cycle, while a 25% starvation level triggered complete loss. Most current ABA models focus on acute starvation, while most patients are chronically ill. The optimal parameters to achieve complete amenorrhoea included early adolescence, chronic starvation and 25% weight loss. The new ABA model allows studying the effects of chronic AN on underlying behavioural, hormonal and brain pathobiology. Copyright © 2017 Elsevier B.V. All rights reserved.

The interactome of the copper transporter ATP7A belongs to a network of neurodevelopmental and neurodegeneration factors

PubMed Central

Comstra, Heather S; McArthy, Jacob; Rudin-Rush, Samantha; Hartwig, Cortnie; Gokhale, Avanti; Zlatic, Stephanie A; Blackburn, Jessica B; Werner, Erica; Petris, Michael; D’Souza, Priya; Panuwet, Parinya; Barr, Dana Boyd; Lupashin, Vladimir; Vrailas-Mortimer, Alysia; Faundez, Victor

2017-01-01

Genetic and environmental factors, such as metals, interact to determine neurological traits. We reasoned that interactomes of molecules handling metals in neurons should include novel metal homeostasis pathways. We focused on copper and its transporter ATP7A because ATP7A null mutations cause neurodegeneration. We performed ATP7A immunoaffinity chromatography and identified 541 proteins co-isolating with ATP7A. The ATP7A interactome concentrated gene products implicated in neurodegeneration and neurodevelopmental disorders, including subunits of the Golgi-localized conserved oligomeric Golgi (COG) complex. COG null cells possess altered content and subcellular localization of ATP7A and CTR1 (SLC31A1), the transporter required for copper uptake, as well as decreased total cellular copper, and impaired copper-dependent metabolic responses. Changes in the expression of ATP7A and COG subunits in Drosophila neurons altered synapse development in larvae and copper-induced mortality of adult flies. We conclude that the ATP7A interactome encompasses a novel COG-dependent mechanism to specify neuronal development and survival. DOI: http://dx.doi.org/10.7554/eLife.24722.001 PMID:28355134

A futile cycle, formed between two ATP-dependant gamma-glutamyl cycle enzymes, gamma-glutamyl cysteine synthetase and 5-oxoprolinase: the cause of cellular ATP depletion in nephrotic cystinosis?

PubMed

Kumar, Akhilesh; Bachhawat, Anand Kumar

2010-03-01

Cystinosis, an inherited disease caused by a defect in the lysosomal cystine transporter (CTNS), is characterized by renal proximal tubular dysfunction. Adenosine triphosphate (ATP) depletion appears to be a key event in the pathophysiology of the disease, even though the manner in which ATP depletion occurs is still a puzzle. We present a model that explains how a futile cycle that is generated between two ATP-utilizing enzymes of the gamma-glutamyl cycle leads to ATP depletion. The enzyme gamma-glutamyl cysteine synthetase (gamma-GCS), in the absence of cysteine, forms 5-oxoproline (instead of the normal substrate, gamma-glutamyl cysteine) and the 5-oxoproline is converted into glutamate by the ATP-dependant enzyme, 5-oxoprolinase. Thus, in cysteine-limiting conditions, glutamate is cycled back into glutamate via 5-oxoproline at the cost of two ATP molecules without production of glutathione and is the cause of the decreased levels of glutathione synthesis, as well as the ATP depletion observed in these cells. The model is also compatible with the differences seen in the human patients and the mouse model of cystinosis, where renal failure is not observed.

Starvation Promotes Odor/Feeding-Time Associations in Flies

ERIC Educational Resources Information Center

Chouhan, Nitin Singh; Wolf, Reinard; Heisenberg, Martin

2017-01-01

Starvation causes a motivational state that facilitates diverse behaviors such as feeding, walking, and search. Starved "Drosophila" can form odor/feeding-time associations but the role of starvation in encoding of "time" is poorly understood. Here we show that the extent of starvation is correlated with the fly's ability to…

Enhancement of succinate yield by manipulating NADH/NAD+ ratio and ATP generation.

PubMed

Li, Jiaojiao; Li, Yikui; Cui, Zhiyong; Liang, Quanfeng; Qi, Qingsheng

2017-04-01

We previously engineered Escherichia coli YL104 to efficiently produce succinate from glucose. In this study, we investigated the relationships between the NADH/NAD + ratio, ATP level, and overall yield of succinate production by using glucose as the carbon source in YL104. First, the use of sole NADH dehydrogenases increased the overall yield of succinate by 7% and substantially decreased the NADH/NAD + ratio. Second, the soluble fumarate reductase from Saccharomyces cerevisiae was overexpressed to manipulate the anaerobic NADH/NAD + ratio and ATP level. Third, another strategy for reducing the ATP level was applied by introducing ATP futile cycling for improving succinate production. Finally, a combination of these methods exerted a synergistic effect on improving the overall yield of succinate, which was 39% higher than that of the previously engineered strain YL104. The study results indicated that regulation of the NADH/NAD + ratio and ATP level is an efficient strategy for succinate production.

Magnetic field affects enzymatic ATP synthesis.

PubMed

Buchachenko, Anatoly L; Kuznetsov, Dmitry A

2008-10-01

The rate of ATP synthesis by creatine kinase extracted from V. xanthia venom was shown to depend on the magnetic field. The yield of ATP produced by enzymes with 24Mg2+ and 26Mg2+ ions in catalytic sites increases by 7-8% at 55 mT and then decreases at 80 mT. For enzyme with 25Mg2+ ion in a catalytic site, the ATP yield increases by 50% and 70% in the fields 55 and 80 mT, respectively. In the Earth field the rate of ATP synthesis by enzyme, in which Mg2+ ion has magnetic nucleus 25Mg, is 2.5 times higher than that by enzymes, in which Mg2+ ion has nonmagnetic, spinless nuclei 24Mg or 26Mg. Both magnetic field effect and magnetic isotope effect demonstrate that the ATP synthesis is an ion-radical process, affected by Zeeman interaction and hyperfine coupling in the intermediate ion-radical pair.

Accumulation of nonesterified fatty acids causes the sustained energetic deficit in kidney proximal tubules after hypoxia-reoxygenation.

PubMed

Feldkamp, Thorsten; Kribben, Andreas; Roeser, Nancy F; Senter, Ruth A; Weinberg, Joel M

2006-02-01

Kidney proximal tubules exhibit decreased ATP and reduced, but not absent, mitochondrial membrane potential (Deltapsi(m)) during reoxygenation after severe hypoxia. This energetic deficit, which plays a pivotal role in overall cellular recovery, cannot be explained by loss of mitochondrial membrane integrity, decreased electron transport, or compromised F1F0-ATPase and adenine nucleotide translocase activities. Addition of oleate to permeabilized tubules produced concentration-dependent decreases of Deltapsi(m) measured by safranin O uptake (threshold for oleate = 0.25 microM, 1.6 nmol/mg protein; maximal effect = 4 microM, 26 nmol/mg) that were reversed by delipidated BSA (dBSA). Cell nonesterified fatty acid (NEFA) levels increased from <1 to 17.4 nmol/mg protein during 60- min hypoxia and remained elevated at 7.6 nmol/mg after 60 min reoxygenation, at which time ATP had recovered to only 10% of control values. Safranin O uptake in reoxygenated tubules, which was decreased 85% after 60-min hypoxia, was normalized by dBSA, which improved ATP synthesis as well. dBSA also almost completely normalized Deltapsi(m) when the duration of hypoxia was increased to 120 min. In intact tubules, the protective substrate combination of alpha-ketoglutarate + malate (alpha-KG/MAL) increased ATP three- to fourfold, limited NEFA accumulation during hypoxia by 50%, and lowered NEFA during reoxygenation. Notably, dBSA also improved ATP recovery when added to intact tubules during reoxygenation and was additive to the effect of alpha-KG/MAL. We conclude that NEFA overload is the primary cause of energetic failure of reoxygenated proximal tubules and lowering NEFA substantially contributes to the benefit from supplementation with alpha-KG/MAL.

Electrochemiluminescence aptasensor for adenosine triphosphate detection using host-guest recognition between metallocyclodextrin complex and aptamer.

PubMed

Chen, Hong; Chen, Qiong; Zhao, Yingying; Zhang, Fan; Yang, Fan; Tang, Jie; He, Pingang

2014-04-01

A sensitive and label-free electrochemiluminescence (ECL) aptasensor for the detection of adenosine triphosphate (ATP) was successfully designed using host-guest recognition between a metallocyclodextrin complex, i.e., tris(bipyridine)ruthenium(II)-β-cyclodextrin [tris(bpyRu)-β-CD], and an ATP-binding aptamer. In the protocol, the NH2-terminated aptamer was immobilized on a glassy carbon electrode (GCE) by a coupling interaction. After host-guest recognition between tris(bpyRu)-β-CD and aptamer, the tris(bpyRu)-β-CD/aptamer/GCE produced a strong ECL signal as a result of the photoactive properties of tris(bpyRu)-β-CD. However, in the presence of ATP, the ATP/aptamer complex was formed preferentially, which restricted host-guest recognition, and therefore less tris(bpyRu)-β-CD was attached to the GCE surface, resulting in an obvious decrease in the ECL intensity. Under optimal determination conditions, an excellent logarithmic linear relationship between the ECL decrease and ATP concentration was obtained in the range 10.0-0.05 nM, with a detection limit of 0.01 nM at the S/N ratio of 3. The proposed ECL-based ATP aptasensor exhibited high sensitivity and selectivity, without time-consuming signal-labeling procedures, and is considered to be a promising model for detection of aptamer-specific targets. Copyright © 2014. Published by Elsevier B.V.

Increased Glycolytic ATP Synthesis Is Associated with Tafenoquine Resistance in Leishmania major▿

PubMed Central

Manzano, José Ignacio; Carvalho, Luis; Pérez-Victoria, José M.; Castanys, Santiago; Gamarro, Francisco

2011-01-01

Tafenoquine (TFQ), an 8-aminoquinoline used to treat and prevent Plasmodium infections, could represent an alternative therapy for leishmaniasis. Indeed, TFQ has shown significant leishmanicidal activity both in vitro and in vivo, where it targets Leishmania mitochondria and activates a final apoptosis-like process. In order not to jeopardize the life span of this potential antileishmania drug, it is important to determine the likelihood that Leishmania will develop resistance to TFQ and the mechanisms of resistance induced. To address this issue, a TFQ-resistant Leishmania major promastigote line (R4) was selected. This resistance, which is unstable in a drug-free medium (revertant line), was maintained in intramacrophage amastigote forms, and R4 promastigotes were found to be cross-resistant to other 8-aminoquinolines. A decreased TFQ uptake, which is probably associated with an alkalinization of the intracellular pH rather than drug efflux, was observed for both the R4 and revertant lines. TFQ induces a decrease in ATP synthesis in all Leishmania lines, although total ATP levels were maintained at higher values in R4 parasites. In contrast, ATP synthesis by glycolysis was significantly increased in R4 parasites, whereas mitochondrial ATP synthesis was similar to that in wild-type parasites. We therefore conclude that increased glycolytic ATP synthesis is the main mechanism underlying TFQ resistance in Leishmania. PMID:21199921

Increased glycolytic ATP synthesis is associated with tafenoquine resistance in Leishmania major.

PubMed

Manzano, José Ignacio; Carvalho, Luis; Pérez-Victoria, José M; Castanys, Santiago; Gamarro, Francisco

2011-03-01

Tafenoquine (TFQ), an 8-aminoquinoline used to treat and prevent Plasmodium infections, could represent an alternative therapy for leishmaniasis. Indeed, TFQ has shown significant leishmanicidal activity both in vitro and in vivo, where it targets Leishmania mitochondria and activates a final apoptosis-like process. In order not to jeopardize the life span of this potential antileishmania drug, it is important to determine the likelihood that Leishmania will develop resistance to TFQ and the mechanisms of resistance induced. To address this issue, a TFQ-resistant Leishmania major promastigote line (R4) was selected. This resistance, which is unstable in a drug-free medium (revertant line), was maintained in intramacrophage amastigote forms, and R4 promastigotes were found to be cross-resistant to other 8-aminoquinolines. A decreased TFQ uptake, which is probably associated with an alkalinization of the intracellular pH rather than drug efflux, was observed for both the R4 and revertant lines. TFQ induces a decrease in ATP synthesis in all Leishmania lines, although total ATP levels were maintained at higher values in R4 parasites. In contrast, ATP synthesis by glycolysis was significantly increased in R4 parasites, whereas mitochondrial ATP synthesis was similar to that in wild-type parasites. We therefore conclude that increased glycolytic ATP synthesis is the main mechanism underlying TFQ resistance in Leishmania.

2-Oxoglutarate levels control adenosine nucleotide binding by Herbaspirillum seropedicae PII proteins.

PubMed

Oliveira, Marco A S; Gerhardt, Edileusa C M; Huergo, Luciano F; Souza, Emanuel M; Pedrosa, Fábio O; Chubatsu, Leda S

2015-12-01

Nitrogen metabolism in Proteobacteria is controlled by the Ntr system, in which PII proteins play a pivotal role, controlling the activity of target proteins in response to the metabolic state of the cell. Characterization of the binding of molecular effectors to these proteins can provide information about their regulation. Here, the binding of ATP, ADP and 2-oxoglutarate (2-OG) to the Herbaspirillum seropedicae PII proteins, GlnB and GlnK, was characterized using isothermal titration calorimetry. Results show that these proteins can bind three molecules of ATP, ADP and 2-OG with homotropic negative cooperativity, and 2-OG binding stabilizes the binding of ATP. Results also show that the affinity of uridylylated forms of GlnB and GlnK for nucleotides is significantly lower than that of the nonuridylylated proteins. Furthermore, fluctuations in the intracellular concentration of 2-OG in response to nitrogen availability are shown. Results suggest that under nitrogen-limiting conditions, PII proteins tend to bind ATP and 2-OG. By contrast, after an ammonium shock, a decrease in the 2-OG concentration is observed causing a decrease in the affinity of PII proteins for ATP. This phenomenon may facilitate the exchange of ATP for ADP on the ligand-binding pocket of PII proteins, thus it is likely that under low ammonium, low 2-OG levels would favor the ADP-bound state. © 2015 FEBS.

Distinguishing starvation from cachexia.

PubMed

Thomas, David R

2002-11-01

The poor response to hypercaloric feeding in ill adults may be caused by failure to distinguish cachexia from starvation (Table 1). The chief difference between starvation and cachexia is that refeeding reverses starvation but is less effective for cachexia. The ineffectiveness of refeeding in treating cachexia may explain some of the poor results from direct nutritional interventions in clinical trials. Simple starvation should respond to voluntary or involuntary hypercaloric feedings. The failure to demonstrate a more positive response may be caused by underlying cachexic states.

Amino acid starvation in Escherichia coli K-12: characteristics of the translation process.

PubMed Central

Subrahmanyam, C S; Das, H K

1976-01-01

Some characteristics of the translation process during amino acid starvation in Escherichia coli have been examined. Once starvation has been established, premature termination of polypeptides is negligible and complete proteins are formed. There is some preference for the synthesis of shorter proteins. The number of ribosomes involved in protein synthesis appears to decline to about half during amino acid-starvation. The assembly time of proteins during amino acid starvation is increased to only about fourfold, though protein synthesis maintained by turnover is reduced to 10%. To explain these observations, a model has been proposed for the course of events that possibly take place from the onset of starvation. PMID:780337

Elastohydrodynamics of elliptical contacts for materials of low elastic modulus

NASA Technical Reports Server (NTRS)

Hamrock, B. J.; Dowson, D.

1983-01-01

The influence of the ellipticity parameter k and the dimensionless speed U, load W, and materials G parameters on minimum film thickness for materials of low elastic modulus was investigated. The ellipticity parameter was varied from 1 (a ball-on-plane configuration) to 12 (a configuration approaching a line contact); U and W were each varied by one order of magnitude. Seventeen cases were used to generate the minimum- and central-film-thickness relations. The influence of lubricant starvation on minimum film thickness in starved elliptical, elastohydrodynamic configurations was also investigated for materials of low elastic modulus. Lubricant starvation was studied simply by moving the inlet boundary closer to the center of the conjunction in the numerical solutions. Contour plots of pressure and film thickness in and around the contact were presented for both fully flooded and starved lubrication conditions. It is evident from these figures that the inlet pressure contours become less circular and closer to the edge of the Hertzian contact zone and that the film thickness decreases substantially as the serverity of starvation increases. The results presented reveal the essential features of both fully flooded and starved, elliptical, elastohydrodynamic conjunctions for materials of low elastic modulus.

Effect of lappaconitine on neuropathic pain mediated by P2X3 receptor in rat dorsal root ganglion.

PubMed

Ou, Shan; Zhao, Yan-Dong; Xiao, Zhi; Wen, Hui-Zhong; Cui, Jian; Ruan, Huai-Zhen

2011-04-01

ATP facilitates initiation and transmission of the neuropathic pain at the dorsal root ganglion (DRG) level via the P2X receptors, especially the subtype P2X(3). Lappaconitine (LA) is an active principle isolated from Chinese herbal medicine and possesses analgesic effect. The aim of this study was to investigate the effect of LA on chronic constriction injury (CCI)-induced neuropathic pain mediated by P2X(3) receptor in the DRG neurons. In the presence of CCI and/or LA, the mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were measured and P2X(3) receptor expression in the DRG neurons was evaluated by immunohistochemistry and Western blotting. Following intrathecal administration of P2X(3) receptor oligonucleotide, the effect of LA on pain thresholds was assessed. Furthermore, the effect of LA on the P2X(3) receptor agonists ATP- and α,β-meATP-induced inward currents (I(ATP) and I(α,β-meATP)) in the acutely dissociated rat DRG neurons was investigated by whole cell patch-clamp. The results included: (1) There showed reduction of pain thresholds, enhancement of I(ATP) and I(α,β-meATP) and up-regulation of P2X(3) receptor expression in rat DRG neurons when neuropathic pain occurred. (2) In the presence of LA, the decreased pain thresholds, the up-regulated P2X(3) receptor expression and the enhanced I(ATP) and I(α,β-meATP) were reversible in the CCI rats. (3) The down-regulated P2X(3) receptor expression with pretreatment of P2X(3) receptor antisense oligonucleotide significantly attenuated the analgesic effect of LA. These results indicate that the analgesic effect of LA involves decrease of expression and sensitization of the P2X(3) receptors of the rat DRG neurons following CCI. Copyright © 2011 Elsevier Ltd. All rights reserved.

Identification of Genes Associated with Resilience/Vulnerability to Sleep Deprivation and Starvation in Drosophila

PubMed Central

Thimgan, Matthew S.; Seugnet, Laurent; Turk, John; Shaw, Paul J.

2015-01-01

Background and Study Objectives: Flies mutant for the canonical clock protein cycle (cyc01) exhibit a sleep rebound that is ∼10 times larger than wild-type flies and die after only 10 h of sleep deprivation. Surprisingly, when starved, cyc01 mutants can remain awake for 28 h without demonstrating negative outcomes. Thus, we hypothesized that identifying transcripts that are differentially regulated between waking induced by sleep deprivation and waking induced by starvation would identify genes that underlie the deleterious effects of sleep deprivation and/or protect flies from the negative consequences of waking. Design: We used partial complementary DNA microarrays to identify transcripts that are differentially expressed between cyc01 mutants that had been sleep deprived or starved for 7 h. We then used genetics to determine whether disrupting genes involved in lipid metabolism would exhibit alterations in their response to sleep deprivation. Setting: Laboratory. Patients or Participants: Drosophila melanogaster. Interventions: Sleep deprivation and starvation. Measurements and Results: We identified 84 genes with transcript levels that were differentially modulated by 7 h of sleep deprivation and starvation in cyc01 mutants and were confirmed in independent samples using quantitative polymerase chain reaction. Several of these genes were predicted to be lipid metabolism genes, including bubblegum, cueball, and CG4500, which based on our data we have renamed heimdall (hll). Using lipidomics we confirmed that knockdown of hll using RNA interference significantly decreased lipid stores. Importantly, genetically modifying bubblegum, cueball, or hll resulted in sleep rebound alterations following sleep deprivation compared to genetic background controls. Conclusions: We have identified a set of genes that may confer resilience/vulnerability to sleep deprivation and demonstrate that genes involved in lipid metabolism modulate sleep homeostasis. Citation: Thimgan MS, Seugnet L, Turk J, Shaw PJ. Identification of genes associated with resilience/vulnerability to sleep deprivation and starvation in Drosophila. SLEEP 2015;38(5):801–814. PMID:25409104

Energy reserves mobilization: Strategies of three decapod species.

PubMed

Sacristán, Hernán Javier; Rodríguez, Yamila Eliana; De Los Angeles Pereira, Nair; López Greco, Laura Susana; Lovrich, Gustavo Alejandro; Fernández Gimenez, Analía Verónica

2017-01-01

In food deprivation assays, several different responses have been observed in crustaceans. However, studying energy reserves utilization among more than one species during the same starvation period has not yet been performed, particularly to discern whether the responses are due to intrinsic and/or environmental factors. We hypothesize that decapod species with similar feeding habits have the same strategies in the use of energetic reserves during starvation, even though they inhabit different environments. The aim of this study was to compare the energy reserves mobilization of three decapods species (Cherax quadricarinatus, Palaemon argentinus and Munida gregaria) with similar feeding habits, exposed to similar food deprivation conditions. The crayfish, shrimp and squat-lobster were experimentally kept at continuous feeding or continuous starvation throughout 15 days. Every 3rd day, the midgut gland index (MGI), and the glycogen, lipid and protein contents were measured in the midgut gland (MG) and pleon muscle. Palaemon argentinus mobilized more reserves during starvation, followed by C. quadricarinatus, and the last M. gregaria. The starved shrimps presented low MGI, whereas MG showed a reduction in glycogen (from day 6 to 15), lipid (from day 3 to 15), and protein levels (at day 9 and 15) while in their muscle, lipid reserves decreased at days 3 and 6. In C. quadricarinatus, the most affected parameters in the MG were MGI, glycogen (from day 6 to 15), and lipids (at day 12 and 15). In the MG of M. gregaria only the glycogen was reduced during fasting from 3 to 15 days. Even though the three studied species have similar feeding habitats, we found that their energetic profile utilization is different and it could be explained by the habitat, life span, temperature, organ/tissue, and metabolism of the species. Our results may be useful to understand the several different responses of crustaceans during starvation.

Energy reserves mobilization: Strategies of three decapod species

PubMed Central

Rodríguez, Yamila Eliana; De los Angeles Pereira, Nair; López Greco, Laura Susana; Lovrich, Gustavo Alejandro; Fernández Gimenez, Analía Verónica

2017-01-01

In food deprivation assays, several different responses have been observed in crustaceans. However, studying energy reserves utilization among more than one species during the same starvation period has not yet been performed, particularly to discern whether the responses are due to intrinsic and/or environmental factors. We hypothesize that decapod species with similar feeding habits have the same strategies in the use of energetic reserves during starvation, even though they inhabit different environments. The aim of this study was to compare the energy reserves mobilization of three decapods species (Cherax quadricarinatus, Palaemon argentinus and Munida gregaria) with similar feeding habits, exposed to similar food deprivation conditions. The crayfish, shrimp and squat-lobster were experimentally kept at continuous feeding or continuous starvation throughout 15 days. Every 3rd day, the midgut gland index (MGI), and the glycogen, lipid and protein contents were measured in the midgut gland (MG) and pleon muscle. Palaemon argentinus mobilized more reserves during starvation, followed by C. quadricarinatus, and the last M. gregaria. The starved shrimps presented low MGI, whereas MG showed a reduction in glycogen (from day 6 to 15), lipid (from day 3 to 15), and protein levels (at day 9 and 15) while in their muscle, lipid reserves decreased at days 3 and 6. In C. quadricarinatus, the most affected parameters in the MG were MGI, glycogen (from day 6 to 15), and lipids (at day 12 and 15). In the MG of M. gregaria only the glycogen was reduced during fasting from 3 to 15 days. Even though the three studied species have similar feeding habitats, we found that their energetic profile utilization is different and it could be explained by the habitat, life span, temperature, organ/tissue, and metabolism of the species. Our results may be useful to understand the several different responses of crustaceans during starvation. PMID:28886062

FTIR Study of ATP-Induced Changes in Na+/K+-ATPase from Duck Supraorbital Glands

PubMed Central

Pratap, Promod R.; Dediu, Oana; Nienhaus, G. Ulrich

2003-01-01

The Na+/K+-ATPase uses energy from the hydrolysis of ATP to pump Na+ ions out of and K+ ions into the cell. ATP-induced conformational changes in the protein have been examined in the Na+/K+-ATPase isolated from duck supraorbital salt glands using Fourier transform infrared spectroscopy. Both standard transmission and attenuated total internal reflection sample geometries have been employed. Under transmission conditions, enzyme at 75 mg/ml was incubated with dimethoxybenzoin-caged ATP. ATP was released by flashing with a UV laser pulse at 355 nm, which resulted in a large change in the amide I band. The absorbance at 1659 cm−1 decreased with a concomitant increase in the absorbance at 1620 cm−1. These changes are consistent with a partial conversion of protein secondary structure from α-helix to β-sheet. The changes were ∼8% of the total absorbance, much larger than those seen with other P-type ATPases. Using attenuated total internal reflection Fourier transform infrared spectroscopy, the decrease in absorbance at ∼1650 cm−1 was titrated with ATP, and the titration midpoint K0.5 was determined under different ionic conditions. In the presence of metal ions (Na+, Na+ and K+, or Mg2+), K0.5 was on the order of a few μM. In the absence of these ions, K0.5 was an order of magnitude lower (0.1 μM), indicating a higher apparent affinity. This effect suggests that the equilibrium for the ATP-induced conformational changes is dependent on the presence of metal ions. PMID:14645062

Adenosine Phosphates in Germinating Radish (Raphanus sativus L.) Seeds 1

PubMed Central

Moreland, Donald E.; Hussey, Griscelda G.; Shriner, Carole R.; Farmer, Fred S.

1974-01-01

Changes in concentrations of adenosine phosphates (AMP, ADP, and ATP), oxygen utilization, and fresh weights were measured during the first 48 hours after imbibition of water by quiescent radish seeds (Raphanus sativus L.) at 22.5 C. The changes in ATP concentrations, oxygen utilization, and fresh weights followed a triphasic time course, characterized by a rapid initial increase, which extended from 0 to approximately 1.5 hours, a lag phase from 1.5 to 16 hours, and a sharp linear increase from 16 to 48 hours. In unimbibed seeds, the concentrations of ATP, ADP, and AMP were <0.1, 0.9, and 2.2 nmoles/seed, respectively. After imbibition of water by the quiescent seeds, for 1 hour, the ATP concentration had increased to 2.5, and ADP and AMP concentrations had decreased to 0.3 and 0.1 nmole/seed, respectively. These early changes occurred also in seeds maintained under anaerobic conditions (argon), or when treated with either 5 mm fluoroacetate, or 5 mm iodoacetate. The concentrations of ADP and AMP did not change significantly from 1 to 48 hours. The termination of the lag phase at 16 hours correlated with radicle emergence. Cell division in the radicles was initiated at approximately 28 hours. ATP concentrations in seeds maintained under argon or treated with fluoroacetate remained relatively constant from approximately 2 to 48 hours. In contrast, the ATP concentration of iodoacetate-treated seeds decreased curvilinearly from 4 to 48 hours. Oxidative phosphorylation was estimated to have contributed 15, 20, and 65% of the pool ATP at 1.5, 16, and 48 hours, respectively. PMID:16658928

GAPDH: the missing link between glycolysis and mitochondrial oxidative phosphorylation?

PubMed

Ramzan, Rabia; Weber, Petra; Linne, Uwe; Vogt, Sebastian

2013-10-01

The main function of glycolysis and oxidative phosphorylation is to produce cellular energy in the form of ATP. In the present paper we propose a link between both of these energy-regulatory processes in the form of GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and CytOx (cytochrome c oxidase). GAPDH is the sixth enzyme of glycolysis, whereas CytOx is the fourth complex of the mitochondrial oxidative phosphorylation system. In MS analysis, GAPDH was found to be associated with a BN-PAGE (blue native PAGE)-isolated complex of CytOx from bovine heart tissue homogenates. Both GAPDH and CytOx are highly regulated under normal energy metabolic conditions, but both of these enzymes are highly deregulated in the presence of oxidative stress. The interaction of GAPDH with CytOx could be the point of interest as it has already been shown that GAPDH protein damage results in a marked decrease in cellular ATP levels. On the other hand, decreasing the ATP/ADP ratio may ultimately result in switching off the allosteric ATP inhibition of CytOx leading to increased ROS (reactive oxygen species), cytochrome c release and apoptosis. Moreover, we have previously reported that allosteric ATP inhibition of CytOx is responsible for keeping the membrane potential at low healthy values, thus avoiding the production of ROS and this allosteric ATP inhibition is switched on at a high ATP/ADP ratio. So, in the present paper, we propose a scheme that could prove to be a link between these two enzymes and their role in the prevalence of diseases.

The effect of experimental gastric dilatation-volvulus on adenosine triphosphate content and conductance of the canine gastric and jejunal mucosa

PubMed Central

2005-01-01

Abstract The objective of this study was to determine if experimental gastric dilatation volvulus (GDV) would decrease adenosine triphosphate (ATP) concentration and increase membrane conductance of the canine gastric and jejunal mucosa. Male dogs (n = 15) weighing between 20 and 30 kg were used. Dogs were randomly assigned to 1 of 3 equal groups: Group 1 was control, group 2 was GDV, and group 3 was ischemia. All dogs were anesthetized for 210 min. Group 1 had no manipulation. Group 2 had GDV experimentally induced for 120 min followed by decompression, derotation, and reperfusion for 90 min. Group 3 had GDV experimentally induced for 210 min. Gastric (fundus and pylorus) and jejunal tissue was taken at 0, 120, and 210 min from all of the dogs. Tissue was analyzed for ATP concentration, mucosal conductance, and microscopic changes. The ATP concentration in the fundus did not change significantly from baseline in group 2, but decreased significantly below baseline at 210 min in group 3. The ATP concentration in the jejunum decreased significantly below baseline in groups 2 and 3 at 120 min, remaining significantly decreased in group 3 but returning to baseline at 210 min in group 2. Mucosal conductance of the fundus did not change significantly in any dog. Mucosal conductance of the jejunum increased at 120 min in groups 2 and 3, and became significantly increased above baseline at 210 min. The jejunal mucosa showed more profound cellular changes than the gastric mucosa. The jejunum showed substantial decreases in ATP concentration with an increase in mucosal conductance, suggesting cell membrane dysfunction. Dogs sustaining a GDV are likely to have a change in the activity of mucosal cells in the jejunum, which may be important in the pathophysiology of GDV. PMID:16187546

Epigenetic regulation of starvation-induced autophagy in Drosophila by histone methyltransferase G9a.

PubMed

An, Phan Nguyen Thuy; Shimaji, Kouhei; Tanaka, Ryo; Yoshida, Hideki; Kimura, Hiroshi; Fukusaki, Eiichiro; Yamaguchi, Masamitsu

2017-08-04

Epigenetics is now emerging as a key regulation in response to various stresses. We herein identified the Drosophila histone methyltransferase G9a (dG9a) as a key factor to acquire tolerance to starvation stress. The depletion of dG9a led to high sensitivity to starvation stress in adult flies, while its overexpression induced starvation stress resistance. The catalytic domain of dG9a was not required for starvation stress resistance. dG9a plays no apparent role in tolerance to other stresses including heat and oxidative stresses. Metabolomic approaches were applied to investigate global changes in the metabolome due to the loss of dG9a during starvation stress. The results obtained indicated that dG9a plays an important role in maintaining energy reservoirs including amino acid, trehalose, glycogen, and triacylglycerol levels during starvation. Further investigations on the underlying mechanisms showed that the depletion of dG9a repressed starvation-induced autophagy by controlling the expression level of Atg8a, a critical gene for the progression of autophagy, in a different manner to that in cancer cells. These results indicate a positive role for dG9a in starvation-induced autophagy.

Modulation of nucleotide sensitivity of ATP-sensitive potassium channels by phosphatidylinositol-4-phosphate 5-kinase.

PubMed

Shyng, S L; Barbieri, A; Gumusboga, A; Cukras, C; Pike, L; Davis, J N; Stahl, P D; Nichols, C G

2000-01-18

ATP-sensitive potassium channels (K(ATP) channels) regulate cell excitability in response to metabolic changes. K(ATP) channels are formed as a complex of a sulfonylurea receptor (SURx), a member of the ATP-binding cassette protein family, and an inward rectifier K(+) channel subunit (Kir6.x). Membrane phospholipids, in particular phosphatidylinositol (PI) 4,5-bisphosphate (PIP(2)), activate K(ATP) channels and antagonize ATP inhibition of K(ATP) channels when applied to inside-out membrane patches. To examine the physiological relevance of this regulatory mechanism, we manipulated membrane PIP(2) levels by expressing either the wild-type or an inactive form of PI-4-phosphate 5-kinase (PIP5K) in COSm6 cells and examined the ATP sensitivity of coexpressed K(ATP) channels. Channels from cells expressing the wild-type PIP5K have a 6-fold lower ATP sensitivity (K(1/2), the half maximal inhibitory concentration, approximately 60 microM) than the sensitivities from control cells (K(1/2) approximately 10 microM). An inactive form of the PIP5K had little effect on the K(1/2) of wild-type channels but increased the ATP-sensitivity of a mutant K(ATP) channel that has an intrinsically lower ATP sensitivity (from K(1/2) approximately 450 microM to K(1/2) approximately 100 microM), suggesting a decrease in membrane PIP(2) levels as a consequence of a dominant-negative effect of the inactive PIP5K. These results show that PIP5K activity, which regulates PIP(2) and PI-3,4,5-P(3) levels, is a significant determinant of the physiological nucleotide sensitivity of K(ATP) channels.

Larval starvation improves metabolic response to adult starvation in honey bees (Apis mellifera L.).

PubMed

Wang, Ying; Campbell, Jacob B; Kaftanoglu, Osman; Page, Robert E; Amdam, Gro V; Harrison, Jon F

2016-04-01

Environmental changes during development have long-term effects on adult phenotypes in diverse organisms. Some of the effects play important roles in helping organisms adapt to different environments, such as insect polymorphism. Others, especially those resulting from an adverse developmental environment, have a negative effect on adult health and fitness. However, recent studies have shown that those phenotypes influenced by early environmental adversity have adaptive value under certain (anticipatory) conditions that are similar to the developmental environment, though evidence is mostly from morphological and behavioral observations and it is still rare at physiological and molecular levels. In the companion study, we applied a short-term starvation treatment to fifth instar honey bee larvae and measured changes in adult morphology, starvation resistance, hormonal and metabolic physiology and gene expression. Our results suggest that honey bees can adaptively respond to the predicted nutritional stress. In the present study, we further hypothesized that developmental starvation specifically improves the metabolic response of adult bees to starvation instead of globally affecting metabolism under well-fed conditions. Here, we produced adult honey bees that had experienced a short-term larval starvation, then we starved them for 12 h and monitored metabolic rate, blood sugar concentrations and metabolic reserves. We found that the bees that experienced larval starvation were able to shift to other fuels faster and better maintain stable blood sugar levels during starvation. However, developmental nutritional stress did not change metabolic rates or blood sugar levels in adult bees under normal conditions. Overall, our study provides further evidence that early larval starvation specifically improves the metabolic responses to adult starvation in honey bees. © 2016. Published by The Company of Biologists Ltd.

A Mechanism of Intracellular P2X Receptor Activation*

PubMed Central

Sivaramakrishnan, Venketesh; Fountain, Samuel J.

2012-01-01

P2X receptors (P2XRs) are ATP-activated calcium-permeable ligand-gated ion channels traditionally viewed as sensors of extracellular ATP during diverse physiological processes including pain, inflammation, and taste. However, in addition to a cell surface residency P2XRs also populate the membranes of intracellular compartments, including mammalian lysosomes, phagosomes, and the contractile vacuole (CV) of the amoeba Dictyostelium. The function of intracellular P2XRs is unclear and represents a major gap in our understanding of ATP signaling. Here, we exploit the genetic versatility of Dictyostelium to investigate the effects of physiological concentrations of ATP on calcium signaling in isolated CVs. Within the CV, an acidic calcium store, P2XRs are orientated to sense luminal ATP. Application of ATP to isolated vacuoles leads to luminal translocation of ATP and release of calcium. Mechanisms of luminal ATP translocation and ATP-evoked calcium release share common pharmacology, suggesting that they are linked processes. The ability of ATP to mobilize stored calcium is reduced in vacuoles isolated from P2XAR knock-out amoeba and ablated in cells devoid of P2XRs. Pharmacological inhibition of luminal ATP translocation or depletion of CV calcium attenuates CV function in vivo, manifesting as a loss of regulatory cell volume decrease following osmotic swelling. We propose that intracellular P2XRs regulate vacuole activity by acting as calcium release channels, activated by translocation of ATP into the vacuole lumen. PMID:22736763

Alanine mutation of the catalytic sites of Pantothenate Synthetase causes distinct conformational changes in the ATP binding region.

PubMed

Pandey, Bharati; Grover, Sonam; Goyal, Sukriti; Kumari, Anchala; Singh, Aditi; Jamal, Salma; Kaur, Jagdeep; Grover, Abhinav

2018-01-17

The enzyme Pantothenate synthetase (PS) represents a potential drug target in Mycobacterium tuberculosis. Its X-ray crystallographic structure has demonstrated the significance and importance of conserved active site residues including His44, His47, Asn69, Gln72, Lys160 and Gln164 in substrate binding and formation of pantoyl adenylate intermediate. In the current study, molecular mechanism of decreased affinity of the enzyme for ATP caused by alanine mutations was investigated using molecular dynamics (MD) simulations and free energy calculations. A total of seven systems including wild-type + ATP, H44A + ATP, H47A + ATP, N69A + ATP, Q72A + ATP, K160A + ATP and Q164A + ATP were subjected to 50 ns MD simulations. Docking score, MM-GBSA and interaction profile analysis showed weak interactions between ATP (substrate) and PS (enzyme) in H47A and H160A mutants as compared to wild-type, leading to reduced protein catalytic activity. However, principal component analysis (PCA) and free energy landscape (FEL) analysis revealed that ATP was strongly bound to the catalytic core of the wild-type, limiting its movement to form a stable complex as compared to mutants. The study will give insight about ATP binding to the PS at the atomic level and will facilitate in designing of non-reactive analogue of pantoyl adenylate which will act as a specific inhibitor for PS.

Acute starvation alters lipopolysaccharide-induced fever in leptin-dependent and -independent mechanisms in rats.

PubMed

Inoue, Wataru; Luheshi, Giamal N

2010-12-01

A decrease in leptin levels with the onset of starvation triggers a myriad of physiological responses including immunosuppression and hypometabolism/hypothermia, both of which can counteract the fever response to pathogens. Here we examined the role of leptin in LPS-induced fever in rats that were fasted for 48 h prior to inflammation with or without leptin replacement (12 μg/day). The preinflammation fasting alone caused a progressive hypothermia that was almost completely reversed by leptin replacement. The LPS (100 μg/kg)-induced elevation in core body temperature (T(core)) was attenuated in the fasted animals at 2-6 h after the injection, an effect that was not reversed by leptin replacement. Increasing the LPS dose to 1,000 μg/kg caused a long-lasting fever that remained unabated for up to 36 h after the injection in the fed rats. This sustained response was strongly attenuated in the fasted rats whose T(core) started to decrease by 18 h after the injection. Leptin replacement almost completely restored the prolonged fever. The attenuation of the prolonged fever in the fasted animals was accompanied by the diminution of proinflammatory PGE(2) in the cerebrospinal fluid and mRNA of proopiomelanocortin (POMC) in the hypothalamus. Leptin replacement prevented the fasting-induced reduction of POMC but not PGE(2). Moreover, the leptin-dependent fever maintenance correlated closely with hypothalamic POMC levels (r = 0.77, P < 0.001). These results suggest that reduced leptin levels during starvation attenuate the sustained fever response by lowering hypothalamic POMC tone but not PGE(2) synthesis.

The plasma membrane of Saccharomyces cerevisiae: structure, function, and biogenesis.

PubMed Central

van der Rest, M E; Kamminga, A H; Nakano, A; Anraku, Y; Poolman, B; Konings, W N

1995-01-01

The composition of phospholipids, sphingolipids, and sterols in the plasma membrane has a strong influence on the activity of the proteins associated or embedded in the lipid bilayer. Since most lipid-synthesizing enzymes in Saccharomyces cerevisiae are located in intracellular organelles, an extensive flux of lipids from these organelles to the plasma membrane is required. Although the pathway of protein traffic to the plasma membrane is similar to that of most of the lipids, the bulk flow of lipids is separate from vesicle-mediated protein transport. Recent advances in the analysis of membrane budding and membrane fusion indicate that the mechanisms of protein transport from the endoplasmic reticulum to the Golgi and from the Golgi to plasma membrane are similar. The majority of plasma membrane proteins transport solutes across the membrane. A number of ATP-dependent export systems have been detected that couple the hydrolysis of ATP to transport of molecules out of the cell. The hydrolysis of ATP by the plasma membrane H(+)-ATPase generates a proton motive force which is used to drive secondary transport processes. In S. cerevisiae, many substrates are transported by more than one system. Transport of monosaccharide is catalyzed by uniport systems, while transport of disaccharides, amino acids, and nucleosides is mediated by proton symport systems. Transport activity can be regulated at the level of transcription, e.g., induction and (catabolite) repression, but transport proteins can also be affected posttranslationally by a process termed catabolite inactivation. Catabolite inactivation is triggered by the addition of fermentable sugars, intracellular acidification, stress conditions, and/or nitrogen starvation. Phosphorylation and/or ubiquitination of the transport proteins has been proposed as an initial step in the controlled inactivation and degradation of the target enzyme. The use of artificial membranes, like secretory vesicles and plasma membranes fused with proteoliposomes, as model systems for studies on the mechanism and regulation of transport is evaluated. PMID:7603412

High fat diet-fed obese rats are highly sensitive to doxorubicin-induced cardiotoxicity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mitra, Mayurranjan S.; Donthamsetty, Shashikiran; White, Brent

Often, chemotherapy by doxorubicin (Adriamycin) is limited due to life threatening cardiotoxicity in patients during and posttherapy. Recently, we have shown that moderate diet restriction remarkably protects against doxorubicin-induced cardiotoxicity. This cardioprotection is accompanied by decreased cardiac oxidative stress and triglycerides and increased cardiac fatty-acid oxidation, ATP synthesis, and upregulated JAK/STAT3 pathway. In the current study, we investigated whether a physiological intervention by feeding 40% high fat diet (HFD), which induces obesity in male Sprague-Dawley rats (250-275 g), sensitizes to doxorubicin-induced cardiotoxicity. A LD{sub 10} dose (8 mg doxorubicin/kg, ip) administered on day 43 of the HFD feeding regimen ledmore » to higher cardiotoxicity, cardiac dysfunction, lipid peroxidation, and 80% mortality in the obese (OB) rats in the absence of any significant renal or hepatic toxicity. Doxorubicin toxicokinetics studies revealed no change in accumulation of doxorubicin and doxorubicinol (toxic metabolite) in the normal diet-fed (ND) and OB hearts. Mechanistic studies revealed that OB rats are sensitized due to: (1) higher oxyradical stress leading to upregulation of uncoupling proteins 2 and 3, (2) downregulation of cardiac peroxisome proliferators activated receptor-{alpha}, (3) decreased plasma adiponectin levels, (4) decreased cardiac fatty-acid oxidation (666.9 {+-} 14.0 nmol/min/g heart in ND versus 400.2 {+-} 11.8 nmol/min/g heart in OB), (5) decreased mitochondrial AMP-{alpha}2 protein kinase, and (6) 86% drop in cardiac ATP levels accompanied by decreased ATP/ADP ratio after doxorubicin administration. Decreased cardiac erythropoietin and increased SOCS3 further downregulated the cardioprotective JAK/STAT3 pathway. In conclusion, HFD-induced obese rats are highly sensitized to doxorubicin-induced cardiotoxicity by substantially downregulating cardiac mitochondrial ATP generation, increasing oxidative stress and downregulating the JAK/STAT3 pathway.« less

Metabolic studies with NMR spectroscopy of the alga Dunaliella salina trapped within agarose beads.

PubMed

Bental, M; Pick, U; Avron, M; Degani, H

1990-02-22

A technique for the entrapment of the unicellular algae Dunaliella salina in agarose beads and their perfusion during NMR measurements is presented. The trapped cells maintained their ability to proliferate under normal growth conditions, and remained viable and stable under steady-state conditions for long periods during NMR measurements. Following osmotic shock in the dark, prominent changes were observed in the intracellular level of ATP and polyphosphates, but little to no changes in the intracellular pH or orthoposphate content. When cells were subjected to hyperosmotic shock, the ATP level decreased. The content of NMR-visible polyphosphates decreased as well, presumably due to the production of longer, NMR-invisible structures. Following hypoosmotic shock, the ATP content increased and longer polyphosphates were broken down to shorter, more mobile polymers.

Circulating vaspin and visfatin are not affected by acute or chronic energy deficiency or leptin administration in humans.

PubMed

Kang, Eun Seok; Magkos, Faidon; Sienkiewicz, Elizabeth; Mantzoros, Christos S

2011-06-01

Animal and in vitro studies indicate that leptin alleviates starvation-induced reduction in circulating vaspin and stimulates the production of visfatin. We thus examined whether vaspin and visfatin are affected by short- and long-term energy deprivation and leptin administration in human subjects in vivo. We measured circulating levels of vaspin and visfatin i) before and after 72 h of starvation (leading to severe hypoleptinemia) with or without leptin administration in replacement doses in 13 normal-weight subjects, ii) before and after 72 h of starvation with leptin administration in pharmacological doses in 13 lean and obese subjects, iii) during chronic energy deficiency in eight women with hypothalamic amenorrhea on leptin replacement for 3 months, and iv) during chronic energy deficiency in 18 women with hypothalamic amenorrhea on leptin replacement or placebo for 3 months. Acute starvation decreased serum leptin to 21% of baseline values, (P=0.002) but had no significant effect on vaspin and visfatin concentrations (P>0.05). Nor did normalization of leptin levels affect the concentrations of these two adipokines (P>0.9). Leptin replacement in women with hypothalamic amenorrhea did not significantly alter vaspin and visfatin concentrations, whether relative to baseline or placebo administration (P>0.25). Pharmacological doses of leptin did not affect circulating vaspin and visfatin concentrations (P>0.9). Circulating vaspin and visfatin are not affected by acute or chronic energy deficiency leading to hypoleptinemia and are not regulated by leptin in human subjects, indicating that these adipocyte-secreted hormonal regulators of metabolism are independently regulated in humans.

Glucose Tolerance Testing and Anthropometric Comparisons Among Rural Residents of Kyiv Region: Investigating the Possible Effect of Childhood Starvation-A Community-Based Study.

PubMed

Khalangot, Mykola D; Kovtun, Volodymir A; Okhrimenko, Nadia V; Gurianov, Vitaly G; Kravchenko, Victor I

2017-01-01

A relationship between childhood starvation and type 2 diabetes mellitus (T2D) in adulthood was previously indicated. Ukraine suffered a series of artificial famines between 1921 and 1947. Famines of 1932 to 1933 and 1946 were most severe among them. Long-term health consequences of these famines remain insufficiently investigated. Type 2 diabetes mellitus screening was conducted between June 2013 and December 2014. A total of 198 rural residents of Kyiv region more than 44 years of age, not registered as patients with T2D, were randomly selected. In all, 159 persons answered the question about starvation of parental family, including 73 born before 1947. Among them, 62 persons answered positive. Anthropometric measurements and glucose tolerance tests were performed. A logistic regression model was used to evaluate results. Type 2 diabetes mellitus was detected in 7 of 62 persons (11.3%), who starved during childhood vs 6 of 11 (54.5%) who did not ( P  = .002), age-adjusted and sex-adjusted odds ratio (OR) (95% confidence interval): 0.063 (0.007-0.557). Analysis of the anthropometric data revealed a negative connection between adulthood height and neck circumference (cm, continued variables) and childhood starvation: age-adjusted and sex-adjusted ORs 0.86 (0.76-0.97) and 0.73 (0.54-0.97), respectively. Individuals who starved during famines of 1932 to 1933 and 1946 in Ukraine had a decreased T2D prevalence several decades after the famine episodes.

A Plant Bacterial Pathogen Manipulates Its Insect Vector's Energy Metabolism

PubMed Central

Hijaz, Faraj; Ebert, Timothy A.; Rogers, Michael E.

2016-01-01

ABSTRACT Insect-transmitted plant-pathogenic bacteria may alter their vectors' fitness, survival, behavior, and metabolism. Because these pathogens interact with their vectors on the cellular and organismal levels, potential changes at the biochemical level might occur. “Candidatus Liberibacter asiaticus” (CLas) is transmitted in a persistent, circulative, and propagative manner. The genome of CLas revealed the presence of an ATP translocase that mediates the uptake of ATP and other nucleotides from medium to achieve its biological processes, such as growth and multiplication. Here, we showed that the levels of ATP and many other nucleotides were significantly higher in CLas-infected than healthy psyllids. Gene expression analysis showed upregulation for ATP synthase subunits, while ATPase enzyme activity showed a decrease in ATPase activity. These results indicated that CLas stimulated Diaphorina citri to produce more ATP and many other energetic nucleotides, while it may inhibit their consumption by the insect. As a result of ATP accumulation, the adenylated energy charge (AEC) increased and the AMP/ATP and ADP/ATP ratios decreased in CLas-infected D. citri psyllids. Survival analysis confirmed a shorter life span for CLas-infected D. citri psyllids. In addition, electropenetrography showed a significant reduction in total nonprobing time, salivation time, and time from the last E2 (phloem ingestion) to the end of recording, indicating that CLas-infected psyllids were at a higher hunger level and they tended to forage more often. This increased feeding activity reflects the CLas-induced energetic stress. In conclusion, CLas alters the energy metabolism of its psyllid vector, D. citri, in order to secure its need for energetic nucleotides. IMPORTANCE Insect transmission of plant-pathogenic bacteria involves propagation and circulation of the bacteria within their vectors. The transmission process is complex and requires specific interactions at the molecular and biochemical levels. The growth of the plant-pathogenic bacteria in the hemolymph of their vectors indicated that the hemolymph contains all the necessary nutrients for their growth. In addition to nutrients, “Candidatus Liberibacter asiaticus” (CLas) can take up energetic nucleotides, such as ATP, from its vector, Diaphorina citri, using ATP translocase. In this study, we found that the CLas pathogen manipulates the energy metabolism of its insect vector. The accumulation of ATP in CLas-infected D. citri psyllids indicated that CLas induces ATP production to fulfill its need for this energetic compound. As a result of ATP accumulation, a shorter life span and altered feeding behavior were observed. These findings increase our knowledge of insect transmission of the persistent-circulative-propagative type of plant pathogens vectored by insects. PMID:28039132

The regulation of nucleotide metabolism of immune cells: papaverine induced nucleotide breakdown.

PubMed

Sheppard, H; Sass, S; Tsien, W H

1980-06-01

During a period of prelabeling of mouse thymus cells with any nucleoside at 4 degrees C, nucleoside phosphates accumulated, but no nucleic acid synthesis occurred. Elevating the temperature to 37 degrees C then led to incorporation into the respective nucleic acid reaching a maximum in 5--15 min. Papaverine inhibited this incorporation (IC50:50 muM) and caused an efflux of label into the medium as a nonphosphorylated product. The responses of the different nucleotide phosphate pools showed more dependency on the base then the sugar moeity. The effect of papaverine could not be altered or mimicked by deprivation of oxygen, glucose, or calcium. Mouse spleen cells responded like thymocytes to papaverine, but rat GH3 pituitary cell DNA syntesis was only transiently inhibited with no concomitant efflux of 3H into the medium. As expected, thymus cellular adenosine triphosphate (ATP), determined by the luciferin-luciferase reaction, decreased in the presence of papaverine; suprisingly, extracellular ATP fell as well. The results suggest that decreases in cellular ATP of mouse thymus cells leads to reductions of all nucleoside phosphates and the efflux of the resultant nucleosides. Papaverine may effect a decrease in the ATP levels by activating a phosphohydrolase rather than, or in addition to, the previously suggested inhibition of mitochondrial electron transport.

Stimulation of ceramide formation and suicidal erythrocyte death by vitamin K(3) (menadione).

PubMed

Qadri, Syed M; Eberhard, Matthias; Mahmud, Hasan; Föller, Michael; Lang, Florian

2009-11-25

Vitamin K(3) is an essential micronutrient required for the activation of coagulation factors and thus hemostasis. Administration of vitamin K(3) analogues may cause anemia, which at least in theory could be due to stimulation of suicidal erythrocyte death or eryptosis characterized by cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane leading to exposure of phosphatidylserine at the erythrocyte surface. Eryptosis is triggered by an increase in the cytosolic Ca(2+) activity, by ceramide and by energy depletion (decrease of cytosolic ATP). The present experiments explored, whether vitamin K(3) may influence eryptosis. Hemolysis was estimated from the supernatant hemoglobin concentration, phosphatidylserine-exposing erythrocytes from annexin V-binding in fluorescence-activated cell sorter (FACS) analysis, erythrocyte volume from forward scatter in FACS analysis, ceramide formation from binding of fluorescent antibodies, and erythrocyte ATP content from a luciferin-luciferase assay. As a result, vitamin K(3) (> or =1microM) caused lysis of an only small fraction of erythrocytes, but significantly increased ceramide formation, significantly increased the percentage of annexin V-binding erythrocytes, significantly decreased forward scatter and, at higher concentrations, significantly decreased the cellular ATP content. In conclusion, vitamin K(3) stimulates suicidal erythrocyte death, an effect at least partially due to ceramide formation and ATP depletion.

Beneficial effect of the bioflavonoid quercetin on cholecystokinin-induced mitochondrial dysfunction in isolated rat pancreatic acinar cells.

PubMed

Weber, Heike; Jonas, Ludwig; Wakileh, Michael; Krüger, Burkhard

2014-03-01

The pathogenesis of acute pancreatitis (AP) is still poorly understood. Thus, a reliable pharmacological therapy is currently lacking. In recent years, an impairment of the energy metabolism of pancreatic acinar cells, caused by Ca(2+)-mediated depolarization of the inner mitochondrial membrane and a decreased ATP supply, has been implicated as an important pathological event. In this study, we investigated whether quercetin exerts protection against mitochondrial dysfunction. Following treatment with or without quercetin, rat pancreatic acinar cells were stimulated with supramaximal cholecystokinin-8 (CCK). CCK caused a decrease in the mitochondrial membrane potential (MMP) and ATP concentration, whereas the mitochondrial dehydrogenase activity was significantly increased. Quercetin treatment before CCK application exerted no protection on MMP but increased ATP to a normal level, leading to a continuous decrease in the dehydrogenase activity. The protective effect of quercetin on mitochondrial function was accompanied by a reduction in CCK-induced changes to the cell membrane. Concerning the molecular mechanism underlying the protective effect of quercetin, an increased AMP/ATP ratio suggests that the AMP-activated protein kinase system may be activated. In addition, quercetin strongly inhibited CCK-induced trypsin activity. The results indicate that the use of quercetin may be a therapeutic strategy for reducing the severity of AP.

Potentiation of adenosine triphosphate-induced contractile responses of the guinea-pig isolated vas deferens by adenosine monophosphate and adenosine 5'-monophosphorothioate.

PubMed Central

Fedan, J. S.

1987-01-01

The effects of incubating the guinea-pig isolated vas deferens in the presence of adenine nucleotides (adenosine triphosphate, ATP; adenosine diphosphate, ADP; and adenosine monophosphate, AMP), or in the presence of their phosphorothioate analogues (adenosine 5'-O-(3-thiotriphosphate), ATP gamma S; adenosine 5'-O-(2-thiodiphosphate), ADP beta S; and adenosine 5'-monophosphorothioate, AMP alpha S), on contractile responses to ATP were compared. After challenge with a low (1 microM) or high (300 microM) concentration of ATP to obtain control responses, one vas deferens of a pair was incubated for 5 min with one of the adenine nucleotides, while the contralateral preparation was incubated with the corresponding phosphorothioate analogue. At the conclusion of the incubation the preparations were challenged again with ATP. Incubation with AMP or AMP alpha S resulted in a transient potentiation of responses to 1 microM and 300 microM ATP. The potentiation following incubation with AMP alpha S was larger than that produced by AMP. After incubation with ADP, ADP beta S, ATP and ATP gamma S, responses to 1 microM ATP were decreased, while those to 300 microM ATP were unaffected. Thus, incubation with AMP and AMP alpha S results in potentiation, rather than inhibition, of ATP-induced responses. On the other hand, 5'-diphosphate, 5'-triphosphate, 5'-O-(2-thiodiphosphate) and 5'-O-(3-thiotriphosphate) moieties on adenosine have no effect or cause autoinhibition. These results indicate that AMP exerts a potentiating effect on reactivity to exogenous ATP. AMP arising from the enzymatic degradation of ATP might modulate the level of response to ATP released endogenously as a cotransmitter. PMID:3038248

Vacuolar transporter Avt4 is involved in excretion of basic amino acids from the vacuoles of Saccharomyces cerevisiae.

PubMed

Sekito, Takayuki; Chardwiriyapreecha, Soracom; Sugimoto, Naoko; Ishimoto, Masaya; Kawano-Kawada, Miyuki; Kakinuma, Yoshimi

2014-01-01

Basic amino acids (lysine, histidine and arginine) accumulated in Saccharomyces cerevisiae vacuoles should be mobilized to cytosolic nitrogen metabolism under starvation. We found that the decrease of vacuolar basic amino acids in response to nitrogen starvation was impaired by the deletion of AVT4 gene encoding a vacuolar transporter. In addition, overexpression of AVT4 reduced the accumulation of basic amino acids in vacuoles under nutrient-rich condition. In contrast to AVT4, the deletion and overexpression of AVT3, which encodes the closest homologue of Avt4p, did not affect the contents of vacuolar basic amino acids. Consistent with these, arginine uptake into vacuolar membrane vesicles was decreased by Avt4p-, but not by Avt3p-overproduction, whereas various neutral amino acids were excreted from vacuolar membrane vesicles in a manner dependent on either Avt4p or Avt3p. These results suggest that Avt4p is a vacuolar amino acid exporter involving in the recycling of basic amino acids.

Tributyltin-induced apoptosis requires glycolytic adenosine trisphosphate production.

PubMed

Stridh, H; Fava, E; Single, B; Nicotera, P; Orrenius, S; Leist, M

1999-10-01

The toxicity of tributyltin chloride (TBT) involves Ca(2+) overload, cytoskeletal damage, and mitochondrial failure leading to cell death by apoptosis or necrosis. Here, we examined whether the intracellular ATP level modulates the mode of cell death after exposure to TBT. When Jurkat cells were energized by the mitochondrial substrate, pyruvate, low concentrations of TBT (1-2 microM) triggered an immediate depletion of intracellular ATP followed by necrotic death. When ATP levels were maintained by the addition of glucose, the mode of cell death was typically apoptotic. Glycolytic ATP production was required for apoptosis at two distinct steps. First, maintenance of adequate ATP levels accelerated the decrease of mitochondrial membrane potential, and the release of the intermembrane proteins adenylate kinase and cytochrome c from mitochondria. A possible role of the adenine nucleotide exchanger in this first ATP-dependent step is suggested by experiments performed with the specific inhibitor, bongkrekic acid. This substance delayed cytochrome c release in a manner similar to that caused by ATP depletion. Second, caspase activation following cytochrome c release was only observed in ATP-containing cells. Bcl-2 had only a minor effect on TBT-triggered caspase activation or cell death. We conclude that intracellular ATP concentrations control the mode of cell death in TBT-treated Jurkat cells at both the mitochondrial and caspase activation levels.

Monte Carlo modeling of single-molecule cytoplasmic dynein.

PubMed

Singh, Manoranjan P; Mallik, Roop; Gross, Steven P; Yu, Clare C

2005-08-23

Molecular motors are responsible for active transport and organization in the cell, underlying an enormous number of crucial biological processes. Dynein is more complicated in its structure and function than other motors. Recent experiments have found that, unlike other motors, dynein can take different size steps along microtubules depending on load and ATP concentration. We use Monte Carlo simulations to model the molecular motor function of cytoplasmic dynein at the single-molecule level. The theory relates dynein's enzymatic properties to its mechanical force production. Our simulations reproduce the main features of recent single-molecule experiments that found a discrete distribution of dynein step sizes, depending on load and ATP concentration. The model reproduces the large steps found experimentally under high ATP and no load by assuming that the ATP binding affinities at the secondary sites decrease as the number of ATP bound to these sites increases. Additionally, to capture the essential features of the step-size distribution at very low ATP concentration and no load, the ATP hydrolysis of the primary site must be dramatically reduced when none of the secondary sites have ATP bound to them. We make testable predictions that should guide future experiments related to dynein function.

Action of diclofenac on kidney mitochondria and cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ng, Lin Eng; Vincent, Annette S.; Halliwell, Barry

2006-09-22

The mitochondrial membrane potential measured in isolated rat kidney mitochondria and in digitonin-permeabilized MDCK type II cells pre-energized with succinate, glutamate, and/or malate was reduced by micromolar diclofenac dose-dependently. However, ATP biosynthesis from glutamate/malate was significantly more compromised compared to that from succinate. Inhibition of the malate-aspartate shuttle by diclofenac with a resultant decrease in the ability of mitochondria to generate NAD(P)H was demonstrated. Diclofenac however had no effect on the activities of NADH dehydrogenase, glutamate dehydrogenase, and malate dehydrogenase. In conclusion, decreased NAD(P)H production due to an inhibition of the entry of malate and glutamate via the malate-aspartate shuttlemore » explained the more pronounced decreased rate of ATP biosynthesis from glutamate and malate by diclofenac. This drug, therefore affects the bioavailability of two major respiratory complex I substrates which would normally contribute substantially to supplying the reducing equivalents for mitochondrial electron transport for generation of ATP in the renal cell.« less

Glucose Starvation Inhibits Autophagy via Vacuolar Hydrolysis and Induces Plasma Membrane Internalization by Down-regulating Recycling*

PubMed Central

Lang, Michael J.; Martinez-Marquez, Jorge Y.; Prosser, Derek C.; Ganser, Laura R.; Buelto, Destiney; Wendland, Beverly; Duncan, Mara C.

2014-01-01

Cellular energy influences all aspects of cellular function. Although cells can adapt to a gradual reduction in energy, acute energy depletion poses a unique challenge. Because acute depletion hampers the transport of new energy sources into the cell, the cell must use endogenous substrates to replenish energy after acute depletion. In the yeast Saccharomyces cerevisiae, glucose starvation causes an acute depletion of intracellular energy that recovers during continued glucose starvation. However, how the cell replenishes energy during the early phase of glucose starvation is unknown. In this study, we investigated the role of pathways that deliver proteins and lipids to the vacuole during glucose starvation. We report that in response to glucose starvation, plasma membrane proteins are directed to the vacuole through reduced recycling at the endosomes. Furthermore, we found that vacuolar hydrolysis inhibits macroautophagy in a target of rapamycin complex 1-dependent manner. Accordingly, we found that endocytosis and hydrolysis are required for survival in glucose starvation, whereas macroautophagy is dispensable. Together, these results suggest that hydrolysis of components delivered to the vacuole independent of autophagy is the cell survival mechanism used by S. cerevisiae in response to glucose starvation. PMID:24753258

Renal oxygenation and hemodynamics in acute kidney injury and chronic kidney disease

PubMed Central

Singh, Prabhleen; Ricksten, Sven-Erik; Bragadottir, Gudrun; Redfors, Bengt; Nordquist, Lina

2013-01-01

Summary 1. Acute kidney injury (AKI) puts a major burden on health systems that may arise from multiple initiating insults, including ischemia-reperfusion injury, cardiovascular surgery, radio-contrast administration as well as sepsis. Similarly, the incidence and prevalence of chronic kidney disease (CKD) continues to increase with significant morbidity and mortality. Moreover, an increasing number of AKI patients survive to develop CKD and end-stage kidney disease (ESRD). 2. Although the mechanisms for development of AKI and progression of CKD remain poorly understood, initial impairment of oxygen balance is likely to constitute a common pathway, causing renal tissue hypoxia and ATP starvation that will in turn induce extracellular matrix production, collagen deposition and fibrosis. Thus, possible future strategies for one or both conditions may involve dopamine, loop-diuretics, inducible nitric oxide synthase inhibitors and atrial natriuretic peptide, substances that target kidney oxygen consumption and regulators of renal oxygenation such as nitric oxide and heme oxygenase-1. PMID:23360244

Regulation of autophagy by cytoplasmic p53.

PubMed

Tasdemir, Ezgi; Maiuri, M Chiara; Galluzzi, Lorenzo; Vitale, Ilio; Djavaheri-Mergny, Mojgan; D'Amelio, Marcello; Criollo, Alfredo; Morselli, Eugenia; Zhu, Changlian; Harper, Francis; Nannmark, Ulf; Samara, Chrysanthi; Pinton, Paolo; Vicencio, José Miguel; Carnuccio, Rosa; Moll, Ute M; Madeo, Frank; Paterlini-Brechot, Patrizia; Rizzuto, Rosario; Szabadkai, Gyorgy; Pierron, Gérard; Blomgren, Klas; Tavernarakis, Nektarios; Codogno, Patrice; Cecconi, Francesco; Kroemer, Guido

2008-06-01

Multiple cellular stressors, including activation of the tumour suppressor p53, can stimulate autophagy. Here we show that deletion, depletion or inhibition of p53 can induce autophagy in human, mouse and nematode cells subjected to knockout, knockdown or pharmacological inhibition of p53. Enhanced autophagy improved the survival of p53-deficient cancer cells under conditions of hypoxia and nutrient depletion, allowing them to maintain high ATP levels. Inhibition of p53 led to autophagy in enucleated cells, and cytoplasmic, not nuclear, p53 was able to repress the enhanced autophagy of p53(-/-) cells. Many different inducers of autophagy (for example, starvation, rapamycin and toxins affecting the endoplasmic reticulum) stimulated proteasome-mediated degradation of p53 through a pathway relying on the E3 ubiquitin ligase HDM2. Inhibition of p53 degradation prevented the activation of autophagy in several cell lines, in response to several distinct stimuli. These results provide evidence of a key signalling pathway that links autophagy to the cancer-associated dysregulation of p53.

Regulation of autophagy by cytoplasmic p53

PubMed Central

Tasdemir, Ezgi; Maiuri, M. Chiara; Galluzzi, Lorenzo; Vitale, Ilio; Djavaheri-Mergny, Mojgan; D'Amelio, Marcello; Criollo, Alfredo; Morselli, Eugenia; Zhu, Changlian; Harper, Francis; Nannmark, Ulf; Samara, Chrysanthi; Pinton, Paolo; Vicencio, José Miguel; Carnuccio, Rosa; Moll, Ute M.; Madeo, Frank; Paterlini-Brechot, Patrizia; Rizzuto, Rosario; Szabadkai, Gyorgy; Pierron, Gérard; Blomgren, Klas; Tavernarakis, Nektarios; Codogno, Patrice; Cecconi, Francesco; Kroemer, Guido

2009-01-01

Multiple cellular stressors, including activation of the tumour suppressor p53, can stimulate autophagy. Here we show that knockout, knockdown or pharmacological inhibition of p53 can induce autophagy in human, mouse and nematode cells. Enhanced autophagy improved the survival of p53-deficient cancer cells under conditions of hypoxia and nutrient depletion, allowing them to maintain high ATP levels. Inhibition of p53 led to autophagy in enucleated cells, and cytoplasmic, not nuclear, p53 was able to repress the enhanced autophagy of p53-/- cells. Many different inducers of autophagy (for example, starvation, rapamycin and toxins affecting the endoplasmic reticulum) stimulated proteasome-mediated degradation of p53 through a pathway relying on the E3 ubiquitin ligase HDM2. Inhibition of p53 degradation prevented the activation of autophagy in several cell lines, in response to several distinct stimuli. These results provide evidence of a key signalling pathway that links autophagy to the cancer-associated dysregulation of p53. PMID:18454141

Regulatory assembly of the vacuolar proton pump VoV1-ATPase in yeast cells by FLIM-FRET

NASA Astrophysics Data System (ADS)

Ernst, Stefan; Batisse, Claire; Zarrabi, Nawid; Böttcher, Bettina; Börsch, Michael

2010-02-01

We investigate the reversible disassembly of VOV1-ATPase in life yeast cells by time resolved confocal FRET imaging. VOV1-ATPase in the vacuolar membrane pumps protons from the cytosol into the vacuole. VOV1-ATPase is a rotary biological nanomotor driven by ATP hydrolysis. The emerging proton gradient is used for secondary transport processes as well as for pH and Ca2+ homoeostasis in the cell. The activity of the VOV1-ATPase is regulated through assembly / disassembly processes. During starvation the two parts of VOV1-ATPase start to disassemble. This process is reversed after addition of glucose. The exact mechanisms are unknown. To follow the disassembly / reassembly in vivo we tagged two subunits C and E with different fluorescent proteins. Cellular distributions of C and E were monitored using a duty cycle-optimized alternating laser excitation scheme (DCO-ALEX) for time resolved confocal FRET-FLIM measurements.

ATP synthesis in the energy metabolism pathway: a new perspective for manipulating CdSe quantum dots biosynthesized in Saccharomyces cerevisiae.

PubMed

Zhang, Rong; Shao, Ming; Han, Xu; Wang, Chuan; Li, Yong; Hu, Bin; Pang, Daiwen; Xie, Zhixiong

2017-01-01

Due to a growing trend in their biomedical application, biosynthesized nanomaterials are of great interest to researchers nowadays with their biocompatible, low-energy consumption, economic, and tunable characteristics. It is important to understand the mechanism of biosynthesis in order to achieve more efficient applications. Since there are only rare studies on the influences of cellular energy levels on biosynthesis, the influence of energy is often overlooked. Through determination of the intracellular ATP concentrations during the biosynthesis process, significant changes were observed. In addition, ATP synthesis deficiency caused great decreases in quantum dots (QDs) biosynthesis in the Δ atp1 , Δ atp2 , Δ atp14 , and Δ atp17 strains. With inductively coupled plasma-atomic emission spectrometry and atomic absorption spectroscopy analyses, it was found that ATP affected the accumulation of the seleno-precursor and helped with the uptake of Cd and the formation of QDs. We successfully enhanced the fluorescence intensity 1.5 or 2 times through genetic modification to increase ATP or SeAM (the seleno analog of S -adenosylmethionine, the product that would accumulate when ATP is accrued). This work explains the mechanism for the correlation of the cellular energy level and QDs biosynthesis in living cells, demonstrates control of the biosynthesis using this mechanism, and thus provides a new manipulation strategy for the biosynthesis of other nanomaterials to widen their applications.

ATP synthesis in the energy metabolism pathway: a new perspective for manipulating CdSe quantum dots biosynthesized in Saccharomyces cerevisiae

PubMed Central

Zhang, Rong; Shao, Ming; Han, Xu; Wang, Chuan; Li, Yong; Hu, Bin; Pang, Daiwen; Xie, Zhixiong

2017-01-01

Due to a growing trend in their biomedical application, biosynthesized nanomaterials are of great interest to researchers nowadays with their biocompatible, low-energy consumption, economic, and tunable characteristics. It is important to understand the mechanism of biosynthesis in order to achieve more efficient applications. Since there are only rare studies on the influences of cellular energy levels on biosynthesis, the influence of energy is often overlooked. Through determination of the intracellular ATP concentrations during the biosynthesis process, significant changes were observed. In addition, ATP synthesis deficiency caused great decreases in quantum dots (QDs) biosynthesis in the Δatp1, Δatp2, Δatp14, and Δatp17 strains. With inductively coupled plasma-atomic emission spectrometry and atomic absorption spectroscopy analyses, it was found that ATP affected the accumulation of the seleno-precursor and helped with the uptake of Cd and the formation of QDs. We successfully enhanced the fluorescence intensity 1.5 or 2 times through genetic modification to increase ATP or SeAM (the seleno analog of S-adenosylmethionine, the product that would accumulate when ATP is accrued). This work explains the mechanism for the correlation of the cellular energy level and QDs biosynthesis in living cells, demonstrates control of the biosynthesis using this mechanism, and thus provides a new manipulation strategy for the biosynthesis of other nanomaterials to widen their applications. PMID:28579774

Mitochondrial Genome Integrity Mutations Uncouple the Yeast Saccharomyces cerevisiae ATP Synthase*║

PubMed Central

Wang, Yamin; Singh, Usha; Mueller, David M.

2013-01-01

The mitochondrial ATP synthase is a molecular motor, which couples the flow of rotons with phosphorylation of ADP. Rotation of the central stalk within the core of ATP synthase effects conformational changes in the active sites driving the synthesis of ATP. Mitochondrial genome integrity (mgi) mutations have been previously identified in the α-, β-, and γ-subunits of ATP synthase in yeast Kluyveromyces lactis and trypanosome Trypanosoma brucei. These mutations reverse the lethality of the loss of mitochondrial DNA in petite negative strains. Introduction of the homologous mutations in Saccharomyces cerevisiae results in yeast strains that lose mitochondrial DNA at a high rate and accompanied decreases in the coupling of the ATP synthase. The structure of yeast F1-ATPase reveals that the mgi residues cluster around the γ-subunit and selectively around the collar region of F1. These results indicate that residues within the mgi complementation group are necessary for efficient coupling of ATP synthase, possibly acting as a support to fix the axis of rotation of the central stalk. PMID:17244612

Reduced Warburg Effect in Cancer Cells Undergoing Autophagy: Steady- State 1H-MRS and Real-Time Hyperpolarized 13C-MRS Studies

PubMed Central

Wong Te Fong, Anne-Christine; Hill, Deborah K.; Orton, Matthew R.; Parkes, Harry G.; Koh, Dow-Mu; Robinson, Simon P.; Leach, Martin O.; Eykyn, Thomas R.; Chung, Yuen-Li

2014-01-01

Autophagy is a highly regulated, energy dependent cellular process where proteins, organelles and cytoplasm are sequestered in autophagosomes and digested to sustain cellular homeostasis. We hypothesized that during autophagy induced in cancer cells by i) starvation through serum and amino acid deprivation or ii) treatment with PI-103, a class I PI3K/mTOR inhibitor, glycolytic metabolism would be affected, reducing flux to lactate, and that this effect may be reversible. We probed metabolism during autophagy in colorectal HT29 and HCT116 Bax knock-out cells using hyperpolarized 13C-magnetic resonance spectroscopy (MRS) and steady-state 1H-MRS. 24 hr PI103-treatment or starvation caused significant reduction in the apparent forward rate constant (kPL) for pyruvate to lactate exchange compared with controls in HT29 (100 μM PI-103: 82%, p = 0.05) and HCT116 Bax-ko cells (10 μM PI-103: 53%, p = 0.05; 20 μM PI-103: 42%, p<0.0001; starvation: 52%, p<0.001), associated with reduced lactate excretion and intracellular lactate in all cases, and unchanged lactate dehydrogenase (LDH) activity and increased NAD+/NADH ratio following PI103 treatment or decreased LDH activity and unchanged NAD+/NADH ratio following starvation. After 48 hr recovery from PI103 treatment, kPL remained below control levels in HT29 cells (74%, p = 0.02), and increased above treated values, but remained below 24 hr vehicle-treated control levels in HCT116 Bax-ko cells (65%, p = 0.004) both were accompanied by sustained reduction in lactate excretion, recovery of NAD+/NADH ratio and intracellular lactate. Following recovery from starvation, kPL was significantly higher than 24 hr vehicle-treated controls (140%, p = 0.05), associated with increased LDH activity and total cellular NAD(H). Changes in kPL and cellular and excreted lactate provided measureable indicators of the major metabolic processes accompanying starvation- and drug-induced autophagy. The changes are reversible, returning towards and exceeding control values on cellular recovery, which potentially identifies resistance. kPL (hyperpolarized 13C-MRS) and lactate (1H-MRS) provide useful biomarkers for the autophagic process, enabling non-invasive monitoring of the Warburg effect. PMID:24667972

Regulation of Cl^- Channels in Normal and Cystic Fibrosis Airway Epithelial Cells by Extracellular ATP

NASA Astrophysics Data System (ADS)

Stutts, M. J.; Chinet, T. C.; Mason, S. J.; Fullton, J. M.; Clarke, L. L.; Boucher, R. C.

1992-03-01

The rate of Cl^- secretion by human airway epithelium is determined, in part, by apical cell membrane Cl^- conductance. In cystic fibrosis airway epithelia, defective regulation of Cl^- conductance decreases the capability to secrete Cl^-. Here we report that extracytosolic ATP in the luminal bath of cultured human airway epithelia increased transepithelial Cl^- secretion and apical membrane Cl^- permeability. Single-channel studies in excised membrane patches revealed that ATP increased the open probability of outward rectifying Cl^- channels. The latter effect occurs through a receptor mechanism that requires no identified soluble second messengers and is insensitive to probes of G protein function. These results demonstrate a mode of regulation of anion channels by binding ATP at the extracellular surface. Regulation of Cl^- conductance by external ATP is preserved in cystic fibrosis airway epithelia.

Loss of the clock protein PER2 shortens the erythrocyte life span in mice.

PubMed

Sun, Qi; Zhao, Yue; Yang, Yunxia; Yang, Xiao; Li, Minghui; Xu, Xi; Wen, Dan; Wang, Junsong; Zhang, Jianfa

2017-07-28

Cell proliferation and release from the bone marrow have been demonstrated to be controlled by circadian rhythms in both humans and mice. However, it is unclear whether local circadian clocks in the bone marrow influence physiological functions and life span of erythrocytes. Here, we report that loss of the clock gene Per2 significantly decreased erythrocyte life span. Mice deficient in Per2 were more susceptible to acute stresses in the erythrocytes, becoming severely anemic upon phenylhydrazine, osmotic, and H 2 O 2 challenges. 1 H NMR-based metabolomics analysis revealed that the Per2 depletion causes significant changes in metabolic profiles of erythrocytes, including increased lactate and decreased ATP levels compared with wild-type mice. The lower ATP levels were associated with hyperfunction of Na + /K + -ATPase activity in Per2 -null erythrocytes, and inhibition of Na + /K + -ATPase activity by ouabain efficiently rescued ATP levels. Per2 -null mice displayed increased levels of Na + /K + -ATPase α1 (ATP1A1) in the erythrocyte membrane, and transfection of Per2 cDNA into the erythroleukemic cell line TF-1 inhibited Atp1a1 expression. Furthermore, we observed that PER2 regulates Atp1a1 transcription through interacting with trans-acting transcription factor 1 (SP1). Our findings reveal that Per2 function in the bone marrow is required for the regulation of life span in circulating erythrocytes. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

A peripheral governor regulates muscle contraction.

PubMed

MacIntosh, Brian R; Shahi, M Reza S

2011-02-01

Active skeletal muscles are capable of keeping the global [adenosine triphosphate (ATP)] reasonably constant during exercise, whether it is mild exercise, activating a few motor units, or all-out exercise using a substantial mass of muscle. This could only be accomplished if there were regulatory processes in place not only to replenish ATP as quickly as possible, but also to modulate the rate of ATP use when that rate threatens to exceed the rate of ATP replenishment, a situation that could lead to metabolic catastrophe. This paper proposes that there is a regulatory process or "peripheral governor" that can modulate activation of muscle to avoid metabolic catastrophe. A peripheral governor, working at the cellular level, should be able to reduce the cellular rate of ATP hydrolysis associated with muscle contraction by attenuating activation. This would necessarily cause something we call peripheral fatigue (i.e., reduced contractile response to a given stimulation). There is no doubt that peripheral fatigue occurs. It has been demonstrated in isolated muscles, in muscles in situ with no central nervous system input, and in intact human subjects performing voluntary exercise with small muscle groups or doing whole-body exercise. The regulation of muscle activation is achieved in at least 3 ways (decreasing membrane excitability, inhibiting Ca2+ release through ryanodine receptors, and decreasing the availability of Ca2+ in the sarcoplasmic reticulum), making this a highly redundant control system. The peripheral governor attenuates cellular activation to reduce the metabolic demand, thereby preserving ATP and the integrity of the cell.

Management of starvation in a Role 1 setting.

PubMed

Jeffery, S M T; Freshwater, D A

2012-01-01

Historical reports from war and natural disasters first identified the dangers of reintroducing food after a period of starvation or malnutrition. The development of advanced nutritional support for hospitalised patients gave rise to the concept of refeeding syndrome, further highlighting the problems and leading to the development of guidelines and protocols for managing malnutrition. In this paper we present a case of starvation in the maritime setting and review the pathophysiology of starvation and refeeding. We discuss the problems associated with managing acute starvation in a Role 1 setting without access to higher medical care, and present guidance for its management.

Roles of Plasmalemma Aquaporin Gene StPIP1 in Enhancing Drought Tolerance in Potato

PubMed Central

Wang, Li; Liu, Yuhui; Feng, Shoujiang; Yang, Jiangwei; Li, Dan; Zhang, Junlian

2017-01-01

Survival and mortality of plants in response to severe drought may be related to carbon starvation, but little is known about how plasma membrane intrinsic proteins may help alleviate the drought-induced damage. Here, we determined the roles of plasmalemma aquaporin gene in improving plant water status, maintaining carbon accumulation, and thereby enhancing drought tolerance. Seven StPIP1 transformed potato (Solanum tuberosum L.) lines (namely T1, T2…T7) were compared with non-transgenic control plant at molecule and whole-plant levels. The relative expression of StPIP1 gene was found in leaves, stems and roots, with the most abundant expression being in the roots. The transgenic lines T6 and T7 had the highest StPIP1 expression, averaging 7.2 times that of the control and the greatest differences occurred 48 h after mannitol osmotic stress treatment. Using an evaluation index to quantifying the degree of drought tolerance, we found that the StPIP1 transgenic lines T6 and T7 had the highest drought tolerance, averaging 8.5 times that of the control. Measured at 30 days in drought stress treatment, the control plant decreased net photosynthetic rate by 33 and 56%, respectively, under moderate and severe stresses; also decreased stomatal conductance by 39 and 65%; and lowered transpiration rate by 49 and 69%, compared to the no-stress treatment, whereas the transgenic lines T6 and T7 maintained a relatively stable level with slight decreases in these properties. The constitutive overexpression of StPIP1 in potato improved plant water use efficiency and increased nonstructural carbohydrate concentration, which helped alleviate carbon starvation and minimized the loss of biomass and tuber yield due to drought stress. We conclude that the expression of StPIPs improves overall water relations in the plant and helps maintain photosynthesis and stomatal conductance; these help minimize carbon starvation and enhance the whole plant tolerance to drought stress. PMID:28487712

High flavonoid accompanied with high starch accumulation triggered by nutrient starvation in bioenergy crop duckweed (Landoltia punctata).

PubMed

Tao, Xiang; Fang, Yang; Huang, Meng-Jun; Xiao, Yao; Liu, Yang; Ma, Xin-Rong; Zhao, Hai

2017-02-15

As the fastest growing plant, duckweed can thrive on anthropogenic wastewater. The purple-backed duckweed, Landoltia punctata, is rich in starch and flavonoids. However, the molecular biological basis of high flavonoid and low lignin content remains largely unknown, as does the best method to combine nutrients removed from sewage and the utilization value improvement of duckweed biomass. A combined omics study was performed to investigate the biosynthesis of flavonoid and the metabolic flux changes in L. punctata grown in different culture medium. Phenylalanine metabolism related transcripts were identified and carefully analyzed. Expression quantification results showed that most of the flavonoid biosynthetic transcripts were relatively highly expressed, while most lignin-related transcripts were poorly expressed or failed to be detected by iTRAQ based proteomic analyses. This explains why duckweed has a much lower lignin percentage and higher flavonoid content than most other plants. Growing in distilled water, expression of most flavonoid-related transcripts were increased, while most were decreased in uniconazole treated L. punctata (1/6 × Hoagland + 800 mg•L -1 uniconazole). When L. punctata was cultivated in full nutrient medium (1/6 × Hoagland), more than half of these transcripts were increased, however others were suppressed. Metabolome results showed that a total of 20 flavonoid compounds were separated by HPLC in L. punctata grown in uniconazole and full nutrient medium. The quantities of all 20 compounds were decreased by uniconazole, while 11 were increased and 6 decreased when grown in full nutrient medium. Nutrient starvation resulted in an obvious purple accumulation on the underside of each frond. The high flavonoid and low lignin content of L. punctata appears to be predominantly caused by the flavonoid-directed metabolic flux. Nutrient starvation is the best option to obtain high starch and flavonoid accumulation simultaneously in a short time for biofuels fermentation and natural products isolation.

Amperometric biosensor system for simultaneous determination of adenosine-5'-triphosphate and glucose.

PubMed

Kucherenko, Ivan S; Didukh, Daria Yu; Soldatkin, Oleksandr O; Soldatkin, Alexei P

2014-06-03

The majority of biosensors for adenosine-5'-triphosphate (ATP) determination are based on cascades of enzymatic reactions; therefore, they are sensitive to glucose or glycerol (depending on the enzymatic system) as well as to ATP. The presence of unknown concentrations of these substances in the sample greatly complicates the determination of ATP. To overcome this disadvantage of known biosensors, we developed a biosensor system consisting of two biosensors: the first one is based on glucose oxidase and is intended for measuring glucose concentration, and the second one is based on glucose oxidase and hexokinase and is sensitive toward both glucose and ATP. Using glucose concentration measured by the first biosensor, we can analyze the total response to glucose and ATP obtained by the second biosensor. Platinum disc electrodes were used as amperometric transducers. The polyphenilenediamine membrane was deposited onto the surface of platinum electrodes to avoid the response to electroactive substances. The effect of glucose concentration on biosensor determination of ATP was studied. The reproducibility of biosensor responses to glucose and ATP during a day was tested (relative standard deviation, RSD, of responses to glucose was 3-6% and to ATP was 8-12%) as well as storage stability of the biosensors (no decrease of glucose responses and 43% drop of ATP responses during 50 days). The measurements of ATP and glucose in pharmaceutical vials (including mixtures of ATP and glucose) were carried out. It was shown that the developed biosensor system can be used for simultaneous analysis of glucose and ATP concentrations in water solutions.

Global gene profiling of aging lungs in Atp8b1 mutant mice.

PubMed

Soundararajan, Ramani; Stearns, Timothy M; Czachor, Alexander; Fukumoto, Jutaro; Turn, Christina; Westermann-Clark, Emma; Breitzig, Mason; Tan, Lee; Lockey, Richard F; King, Benjamin L; Kolliputi, Narasaiah

2016-09-29

Recent studies implicate cardiolipin oxidation in several age-related diseases. Atp8b1 encoding Type 4 P-type ATPases is a cardiolipin transporter. Mutation in Atp8b1 gene or inflammation of the lungs impairs the capacity of Atp8b1 to clear cardiolipin from lung fluid. However, the link between Atp8b1 mutation and age-related gene alteration is unknown. Therefore, we investigated how Atp8b1 mutation alters age-related genes. We performed Affymetrix gene profiling of lungs isolated from young (7-9 wks, n=6) and aged (14 months, 14 M, n=6) C57BL/6 and Atp8b1 mutant mice. In addition, Ingenuity Pathway Analysis (IPA) was performed. Differentially expressed genes were validated by quantitative real-time PCR (qRT-PCR). Global transcriptome analysis revealed 532 differentially expressed genes in Atp8b1 lungs, 157 differentially expressed genes in C57BL/6 lungs, and 37 overlapping genes. IPA of age-related genes in Atp8b1 lungs showed enrichment of Xenobiotic metabolism and Nrf2-mediated signaling pathways. The increase in Adamts2 and Mmp13 transcripts in aged Atp8b1 lungs was validated by qRT-PCR. Similarly, the decrease in Col1a1 and increase in Cxcr6 transcripts was confirmed in both Atp8b1 mutant and C57BL/6 lungs. Based on transcriptome profiling, our study indicates that Atp8b1 mutant mice may be susceptible to age-related lung diseases.

Hydrostatic pressure activates ATP-sensitive K+ channels in lung epithelium by ATP release through pannexin and connexin hemichannels.

PubMed

Richter, Katrin; Kiefer, Kevin P; Grzesik, Benno A; Clauss, Wolfgang G; Fronius, Martin

2014-01-01

Lungs of air-breathing vertebrates are constantly exposed to mechanical forces and therefore are suitable for investigation of mechanotransduction processes in nonexcitable cells and tissues. Freshly dissected Xenopus laevis lungs were used for transepithelial short-circuit current (ISC) recordings and were exposed to increased hydrostatic pressure (HP; 5 cm fluid column, modified Ussing chamber). I(SC) values obtained under HP (I(5cm)) were normalized to values before HP (I(0cm)) application (I(5cm)/I(0cm)). Under control conditions, HP decreased I(SC) (I(5cm)/I(0cm)=0.84; n=68; P<0.0001). This effect was reversible and repeatable ≥30 times. Preincubation with ATP-sensitive K(+) channel (K(ATP)) inhibitors (HMR1098 and glibenclamide) prevented the decrease in I(SC) (I(5cm)/I(0cm): HMR1098=1.19, P<0.0001; glibenclamide=1.11, P<0.0001). Similar effects were observed with hemichannel inhibitors (I(5cm)/I(0cm): meclofenamic acid=1.09, P<0.0001; probenecid=1.0, P<0.0001). The HP effect was accompanied by release of ATP (P<0.05), determined by luciferin-luciferase luminescence in perfusion solution from the luminal side of an Ussing chamber. ATP release was abrogated by both meclofenamic acid and probenecid. RT-PCR experiments revealed the expression of pannexin and connexin hemichannels and KATP subunit transcripts in X. laevis lung. These data show an activation of KATP in pulmonary epithelial cells in response to HP that is induced by ATP release through mechanosensitive pannexin and connexin hemichannels. These findings represent a novel mechanism of mechanotransduction in nonexcitable cells.

Capture and quality control mechanisms for ATP binding

PubMed Central

Li, Li; Martinis, Susan A.

2013-01-01

The catalytic events in members of the nucleotidylyl transferase superfamily are initiated by a millisecond binding of ATP in the active site. Through metadynamics simulations on a class I aminoacyl-tRNA synthetase (aaRSs), the largest group in the superfamily, we calculate the free energy landscape of ATP selection and binding. Mutagenesis studies and fluorescence spectroscopy validated the identification of the most populated intermediate states. The rapid first binding step involves formation of encounter complexes captured through a fly-casting mechanism that acts up on the triphosphate moiety of ATP. In the slower nucleoside binding step, a conserved histidine in the HxxH motif orients the incoming ATP through base-stacking interactions resulting in a deep minimum in the free energy surface. Mutation of this histidine significantly decreases the binding affinity measured experimentally and computationally. The metadynamics simulations further reveal an intermediate quality control state that the synthetases and most likely other members of the superfamily use to select ATP over other nucleoside triphosphates. PMID:23276298

A G-quadruplex-based Label-free Fluorometric Aptasensor for Adenosine Triphosphate Detection.

PubMed

Li, Li Juan; Tian, Xue; Kong, Xiang Juan; Chu, Xia

2015-01-01

A G-quadruplex-based, label-free fluorescence assay was demonstrated for the detection of adenosine triphosphate (ATP). A double-stranded DNA (dsDNA), hybridized by ATP-aptamer and its complementary sequence, was employed as a substrate for ATP binding. SYBR Green I (SG I) was a fluorescent probe and exonuclease III (Exo III) was a nuclease to digest the dsDNA. Consequently, in the absence of ATP, the dsDNA was inset with SG I and was digested by Exo III, resulting in a low background signal. In the presence of ATP, the aptamer in dsDNA folded into a G-quadruplex structure that resisted the digestion of Exo III. SG I was inserted into the structure, showing high fluorescence. Owing to a decrease of the background noise, a high signal-to-noise ratio could be obtained. This sensor can detect ATP with a concentration ranging from 50 μM to 5 mM, and possesses a capacity for the sensitive determination of other targets.

Amino Acid Availability Modulates Vacuolar H+-ATPase Assembly*

PubMed Central

Stransky, Laura A.; Forgac, Michael

2015-01-01

The vacuolar H+-ATPase (V-ATPase) is an ATP-dependent proton pump composed of a peripheral ATPase domain (V1) and a membrane-integral proton-translocating domain (V0) and is involved in many normal and disease processes. An important mechanism of regulating V-ATPase activity is reversible assembly of the V1 and V0 domains. Increased assembly in mammalian cells occurs under various conditions and has been shown to involve PI3K. The V-ATPase is necessary for amino acid-induced activation of mechanistic target of rapamycin complex 1 (mTORC1), which is important in controlling cell growth in response to nutrient availability and growth signals. The V-ATPase undergoes amino acid-dependent interactions with the Ragulator complex, which is involved in recruitment of mTORC1 to the lysosomal membrane during amino acid sensing. We hypothesized that changes in the V-ATPase/Ragulator interaction might involve amino acid-dependent changes in V-ATPase assembly. To test this, we measured V-ATPase assembly by cell fractionation in HEK293T cells treated with and without amino acids. V-ATPase assembly increases upon amino acid starvation, and this effect is reversed upon readdition of amino acids. Lysosomes from amino acid-starved cells possess greater V-ATPase-dependent proton transport, indicating that assembled pumps are catalytically active. Amino acid-dependent changes in both V-ATPase assembly and activity are independent of PI3K and mTORC1 activity, indicating the involvement of signaling pathways distinct from those implicated previously in controlling assembly. By contrast, lysosomal neutralization blocks the amino acid-dependent change in assembly and reactivation of mTORC1 after amino acid starvation. These results identify an important new stimulus for controlling V-ATPase assembly. PMID:26378229

Hypoxic HepG2 cell adaptation decreases ATP synthase dimers and ATP production in inflated cristae by mitofilin down-regulation concomitant to MICOS clustering.

PubMed

Plecitá-Hlavatá, Lydie; Engstová, Hana; Alán, Lukáš; Špaček, Tomáš; Dlasková, Andrea; Smolková, Katarína; Špačková, Jitka; Tauber, Jan; Strádalová, Vendula; Malínský, Jan; Lessard, Mark; Bewersdorf, Joerg; Ježek, Petr

2016-05-01

The relationship of the inner mitochondrial membrane (IMM) cristae structure and intracristal space (ICS) to oxidative phosphorylation (oxphos) is not well understood. Mitofilin (subunit Mic60) of the mitochondrial contact site and cristae organizing system (MICOS) IMM complex is attached to the outer membrane (OMM) via the sorting and assembly machinery/topogenesis of mitochondrial outer membrane β-barrel proteins (SAM/TOB) complex and controls the shape of the cristae. ATP synthase dimers determine sharp cristae edges, whereas trimeric OPA1 tightens ICS outlets. Metabolism is altered during hypoxia, and we therefore studied cristae morphology in HepG2 cells adapted to 5% oxygen for 72 h. Three dimensional (3D), super-resolution biplane fluorescence photoactivation localization microscopy with Eos-conjugated, ICS-located lactamase-β indicated hypoxic ICS expansion with an unchanged OMM (visualized by Eos-mitochondrial fission protein-1). 3D direct stochastic optical reconstruction microscopy immunocytochemistry revealed foci of clustered mitofilin (but not MICOS subunit Mic19) in contrast to its even normoxic distribution. Mitofilin mRNA and protein decreased by ∼20%. ATP synthase dimers vs monomers and state-3/state-4 respiration ratios were lower during hypoxia. Electron microscopy confirmed ICS expansion (maximum in glycolytic cells), which was absent in reduced or OMM-detached cristae of OPA1- and mitofilin-silenced cells, respectively. Hypoxic adaptation is reported as rounding sharp cristae edges and expanding cristae width (ICS) by partial mitofilin/Mic60 down-regulation. Mitofilin-depleted MICOS detaches from SAM while remaining MICOS with mitofilin redistributes toward higher interdistances. This phenomenon causes partial oxphos dormancy in glycolytic cells via disruption of ATP synthase dimers.-Plecitá-Hlavatá, L., Engstová, H., Alán, L., Špaček, T., Dlasková, A., Smolková, K., Špačková, J., Tauber, J., Strádalová, V., Malínský, J., Lessard, M., Bewersdorf, J., Ježek, P. Hypoxic HepG2 cell adaptation decreases ATP synthase dimers and ATP production in inflated cristae by mitofilin down-regulation concomitant to MICOS clustering. © FASEB.

Biallelic Mutations in ATP5F1D , which Encodes a Subunit of ATP Synthase, Cause a Metabolic Disorder

DOE Office of Scientific and Technical Information (OSTI.GOV)

Oláhová, Monika; Yoon, Wan Hee; Thompson, Kyle

ATP synthase, H + transporting, mitochondrial F1 complex, δ subunit (ATP5F1D; formerly ATP5D) is a subunit of mitochondrial ATP synthase and plays an important role in coupling proton translocation and ATP production. Here, we describe two individuals, each with homozygous missense variants in ATP5F1D, who presented with episodic lethargy, metabolic acidosis, 3-methylglutaconic aciduria, and hyperammonemia. Subject 1, homozygous for c.245C>T (p.Pro82Leu), presented with recurrent metabolic decompensation starting in the neonatal period, and subject 2, homozygous for c.317T>G (p.Val106Gly), presented with acute encephalopathy in childhood. Cultured skin fibroblasts from these individuals exhibited impaired assembly of F 1F O ATP synthase andmore » subsequent reduced complex V activity. Cells from subject 1 also exhibited a significant decrease in mitochondrial cristae. Knockdown of Drosophila ATPsynδ, the ATP5F1D homolog, in developing eyes and brains caused a near complete loss of the fly head, a phenotype that was fully rescued by wild-type human ATP5F1D. In contrast, expression of the ATP5F1D c.245C>T and c.317T>G variants rescued the head-size phenotype but recapitulated the eye and antennae defects seen in other genetic models of mitochondrial oxidative phosphorylation deficiency. Our data establish c.245C>T (p.Pro82Leu) and c.317T>G (p.Val106Gly) in ATP5F1D as pathogenic variants leading to a Mendelian mitochondrial disease featuring episodic metabolic decompensation.« less

Impaired ATP release from red blood cells promotes their adhesion to endothelial cells: A mechanism of hypoxemia after transfusion

PubMed Central

Zhu, Hongmei; Zennadi, Rahima; Xu, Bruce X.; Eu, Jerry P.; Torok, Jordan A.; Telen, Marilyn J.; McMahon, Timothy J.

2011-01-01

Objective Transfusion of red blood cells (RBCs) has been linked to disappointing clinical outcomes in the critically ill, but specific mechanisms of organ dysfunction after transfusion remain poorly understood. We tested the hypothesis that RBC storage impairs the ability of RBCs to release ATP and that impaired ATP-release was injurious in vivo, in part through increased RBC adhesion. Design Prospective, controlled, mechanistic study. Setting University research laboratory. Subjects Human and mouse blood donors; nude mouse transfusion recipients. Interventions Manipulation of ATP release, supplemental ATP, and antibodies to RBC and endothelial adhesion receptors were used in vitro and in vivo to probe the roles of released ATP and adhesion in responses to (transfused) RBCs. Measurements and main results The ability of stored RBCs to release ATP declined markedly within 14 days after collection, despite relatively stable levels of ATP within the RBCs. Inhibiting ATP release promoted the adhesion of stored RBCs to endothelial cells in vitro and RBC sequestration in the lungs of transfused mice in vivo. Unlike transfusion of fresh human RBCs, stored-RBC transfusion in mice decreased blood oxygenation and increased extravasation of RBCs into the lung’s alveolar airspaces. Similar findings were seen with transfusion of fresh RBCs treated with the ATP-release inhibitors glibenclamide and carbenoxolone. These findings were prevented by either co-infusion of an ATP analog or pre-transfusion incubation of the RBCs with an antibody against the erythrocyte adhesion receptor LW (Landsteiner-Wiener; ICAM-4). Conclusions The normal flow of RBCs in pulmonary microvessels depends in part on the release of anti-adhesive ATP from RBCs, and storage-induced deficiency in ATP release from transfused RBCs may promote or exacerbate microvascular pathophysiology in the lung, in part through increased RBC adhesion. PMID:21765360

The effect of salts on catecholamine fluxes and adenosine triphosphatase activity in storage vesicles from the adrenal medulla

PubMed Central

Taugner, G.

1971-01-01

1. Influx and efflux of catecholamine and adenosine triphosphatase activity in storage vesicles from the adrenal medulla were studied with dl-[14C]adrenaline in different media. 2. The lowest values for flux and adenosine triphosphatase activity were observed in sucrose media in which an ATP-dependent influx of catecholamine compensated for an efflux of the same magnitude. Efflux in the presence or absence of ATP was similar. 3. In media containing sodium succinate or glutarate adenosine triphosphatase activity was higher and the ATP-dependent influx of catecholamine was about twice that observed in iso-osmotic sucrose medium. In the presence of ATP influx and efflux of catecholamine were balanced; in its absence there was a net release of catecholamine, since efflux was more than twice the influx. Efflux in the presence or absence of ATP was similar. 4. In media containing sodium or potassium chloride and in the presence of ATP influx and adenosine triphosphatase activity were further enhanced, but in the absence of ATP there was no further increase in influx, since catecholamine was released with or without ATP at the same rate. Efflux was therefore twice as high in the presence of ATP as in its absence. 5. Sodium nitrate suppressed the ATP-dependent influx nearly completely, but caused a greatly enhanced efflux, which was twice as high in the presence of ATP as in its absence. 6. The extinction of vesicular suspensions remained unchanged in the presence of ATP under conditions where the catecholamine efflux was balanced by the influx. Under conditions where the efflux was not compensated by influx, the extinction of the suspensions decreased in the presence of ATP more than in its absence. PMID:4256794

Biallelic Mutations in ATP5F1D, which Encodes a Subunit of ATP Synthase, Cause a Metabolic Disorder.

PubMed

Oláhová, Monika; Yoon, Wan Hee; Thompson, Kyle; Jangam, Sharayu; Fernandez, Liliana; Davidson, Jean M; Kyle, Jennifer E; Grove, Megan E; Fisk, Dianna G; Kohler, Jennefer N; Holmes, Matthew; Dries, Annika M; Huang, Yong; Zhao, Chunli; Contrepois, Kévin; Zappala, Zachary; Frésard, Laure; Waggott, Daryl; Zink, Erika M; Kim, Young-Mo; Heyman, Heino M; Stratton, Kelly G; Webb-Robertson, Bobbie-Jo M; Snyder, Michael; Merker, Jason D; Montgomery, Stephen B; Fisher, Paul G; Feichtinger, René G; Mayr, Johannes A; Hall, Julie; Barbosa, Ines A; Simpson, Michael A; Deshpande, Charu; Waters, Katrina M; Koeller, David M; Metz, Thomas O; Morris, Andrew A; Schelley, Susan; Cowan, Tina; Friederich, Marisa W; McFarland, Robert; Van Hove, Johan L K; Enns, Gregory M; Yamamoto, Shinya; Ashley, Euan A; Wangler, Michael F; Taylor, Robert W; Bellen, Hugo J; Bernstein, Jonathan A; Wheeler, Matthew T

2018-03-01

ATP synthase, H + transporting, mitochondrial F1 complex, δ subunit (ATP5F1D; formerly ATP5D) is a subunit of mitochondrial ATP synthase and plays an important role in coupling proton translocation and ATP production. Here, we describe two individuals, each with homozygous missense variants in ATP5F1D, who presented with episodic lethargy, metabolic acidosis, 3-methylglutaconic aciduria, and hyperammonemia. Subject 1, homozygous for c.245C>T (p.Pro82Leu), presented with recurrent metabolic decompensation starting in the neonatal period, and subject 2, homozygous for c.317T>G (p.Val106Gly), presented with acute encephalopathy in childhood. Cultured skin fibroblasts from these individuals exhibited impaired assembly of F 1 F O ATP synthase and subsequent reduced complex V activity. Cells from subject 1 also exhibited a significant decrease in mitochondrial cristae. Knockdown of Drosophila ATPsynδ, the ATP5F1D homolog, in developing eyes and brains caused a near complete loss of the fly head, a phenotype that was fully rescued by wild-type human ATP5F1D. In contrast, expression of the ATP5F1D c.245C>T and c.317T>G variants rescued the head-size phenotype but recapitulated the eye and antennae defects seen in other genetic models of mitochondrial oxidative phosphorylation deficiency. Our data establish c.245C>T (p.Pro82Leu) and c.317T>G (p.Val106Gly) in ATP5F1D as pathogenic variants leading to a Mendelian mitochondrial disease featuring episodic metabolic decompensation. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Biallelic Mutations in ATP5F1D , which Encodes a Subunit of ATP Synthase, Cause a Metabolic Disorder

DOE PAGES

Oláhová, Monika; Yoon, Wan Hee; Thompson, Kyle; ...

2018-02-22

ATP synthase, H + transporting, mitochondrial F1 complex, δ subunit (ATP5F1D; formerly ATP5D) is a subunit of mitochondrial ATP synthase and plays an important role in coupling proton translocation and ATP production. Here, we describe two individuals, each with homozygous missense variants in ATP5F1D, who presented with episodic lethargy, metabolic acidosis, 3-methylglutaconic aciduria, and hyperammonemia. Subject 1, homozygous for c.245C>T (p.Pro82Leu), presented with recurrent metabolic decompensation starting in the neonatal period, and subject 2, homozygous for c.317T>G (p.Val106Gly), presented with acute encephalopathy in childhood. Cultured skin fibroblasts from these individuals exhibited impaired assembly of F 1F O ATP synthase andmore » subsequent reduced complex V activity. Cells from subject 1 also exhibited a significant decrease in mitochondrial cristae. Knockdown of Drosophila ATPsynδ, the ATP5F1D homolog, in developing eyes and brains caused a near complete loss of the fly head, a phenotype that was fully rescued by wild-type human ATP5F1D. In contrast, expression of the ATP5F1D c.245C>T and c.317T>G variants rescued the head-size phenotype but recapitulated the eye and antennae defects seen in other genetic models of mitochondrial oxidative phosphorylation deficiency. Our data establish c.245C>T (p.Pro82Leu) and c.317T>G (p.Val106Gly) in ATP5F1D as pathogenic variants leading to a Mendelian mitochondrial disease featuring episodic metabolic decompensation.« less

A common modality of action of simulated space stresses on the oxidative metabolism of ethylmorphine, aniline and p-nitroanisole by male rat liver.

NASA Technical Reports Server (NTRS)

Furner, R. L.; Neville, E. D.; Talarico, K. S.; Feller, D. D.

1972-01-01

High gravity, cold and starvation elicited similar responses in male Simonson rats. These responses included a decreased rate in body weight gain, increased metabolism of aniline and p-nitroanisole, and no consistent pattern of change in the metabolism of ethylmorphine. Cold and starvation increased the amount of hepatic cytochrome P-450, while hypobaric-hyperoxia caused no change in any of the parameters measured. When 1% acetone was given to the rats in their drinking water, the effects on drug metabolism were similar to those produced by food restriction in that the metabolism of aniline and p-nitroanisole was increased, and the metabolism of ethylmorphine unchanged. The type I binding spectrum of acetone suggests that it is either a substrate, inhibitor, or both for hepatitic oxidative enzymes.

Sarcolemmal cardiac K(ATP) channels as a target for the cardioprotective effects of the fluorine-containing pinacidil analogue, flocalin.

PubMed

Voitychuk, Oleg I; Strutynskyi, Ruslan B; Yagupolskii, Lev M; Tinker, Andrew; Moibenko, Olexiy O; Shuba, Yaroslav M

2011-02-01

A class of drugs known as K(ATP) -channel openers induce cardioprotection. This study examined the effects of the novel K(ATP) -channel opener, the fluorine-containing pinacidil derivative, flocalin, on cardiac-specific K(ATP) -channels, excitability of native cardiac myocytes and on the ischaemic heart. The action of flocalin was investigated on: (i) membrane currents through cardiac-specific K(ATP) -channels (I(KATP) ) formed by K(IR) 6.2/SUR2A heterologously expressed in HEK-293 cells (HEK-293(₆.₂/₂A) ); (ii) excitability and intracellular Ca²(+) ([Ca²(+) ](i) ) transients of cultured rat neonatal cardiac myocytes; and (iii) functional and ultrastructural characteristics of isolated guinea-pig hearts subjected to ischaemia-reperfusion. Flocalin concentration-dependently activated a glibenclamide-sensitive I(KATP) in HEK-293(₆.₂/₂A) cells with an EC₅₀= 8.1 ± 0.4 µM. In cardiac myocytes, flocalin (5 µM) hyperpolarized resting potential by 3-5 mV, markedly shortened action potential duration, reduced the amplitude of [Ca²(+) ](i) transients by 2-3-fold and suppressed contraction. The magnitude and extent of reversibility of these effects depended on the type of cardiac myocytes. In isolated hearts, perfusion with 5 µmol·L⁻¹ flocalin, before inducing ischaemia, facilitated restoration of contraction during reperfusion, decreased the number of extrasystoles, prevented the appearance of coronary vasoconstriction and reduced damage to the cardiac tissue at the ultrastructural level (state of myofibrils, membrane integrity, mitochondrial cristae structure). Flocalin induced potent cardioprotection by activating cardiac-type K(ATP) -channels with all the benefits of the presence of fluorine group in the drug structure: higher lipophilicity, decreased toxicity, resistance to oxidation and thermal degradation, decreased metabolism in the organism and prolonged therapeutic action. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

Attenuation of Phosphate Starvation Responses by Phosphite in Arabidopsis1

PubMed Central

Ticconi, Carla A.; Delatorre, Carla A.; Abel, Steffen

2001-01-01

When inorganic phosphate is limiting, Arabidopsis has the facultative ability to metabolize exogenous nucleic acid substrates, which we utilized previously to identify insensitive phosphate starvation response mutants in a conditional genetic screen. In this study, we examined the effect of the phosphate analog, phosphite (Phi), on molecular and morphological responses to phosphate starvation. Phi significantly inhibited plant growth on phosphate-sufficient (2 mm) and nucleic acid-containing (2 mm phosphorus) media at concentrations higher than 2.5 mm. However, with respect to suppressing typical responses to phosphate limitation, Phi effects were very similar to those of phosphate. Phosphate starvation responses, which we examined and found to be almost identically affected by both anions, included changes in: (a) the root-to-shoot ratio; (b) root hair formation; (c) anthocyanin accumulation; (d) the activities of phosphate starvation-inducible nucleolytic enzymes, including ribonuclease, phosphodiesterase, and acid phosphatase; and (e) steady-state mRNA levels of phosphate starvation-inducible genes. It is important that induction of primary auxin response genes by indole-3-acetic acid in the presence of growth-inhibitory Phi concentrations suggests that Phi selectively inhibits phosphate starvation responses. Thus, the use of Phi may allow further dissection of phosphate signaling by genetic selection for constitutive phosphate starvation response mutants on media containing organophosphates as the only source of phosphorus. PMID:11706178

Starvation induced cell death in autophagy-defective yeast mutants is caused by mitochondria dysfunction.

PubMed

Suzuki, Sho W; Onodera, Jun; Ohsumi, Yoshinori

2011-02-25

Autophagy is a highly-conserved cellular degradation and recycling system that is essential for cell survival during nutrient starvation. The loss of viability had been used as an initial screen to identify autophagy-defective (atg) mutants of the yeast Saccharomyces cerevisiae, but the mechanism of cell death in these mutants has remained unclear. When cells grown in a rich medium were transferred to a synthetic nitrogen starvation media, secreted metabolites lowered the extracellular pH below 3.0 and autophagy-defective mutants mostly died. We found that buffering of the starvation medium dramatically restored the viability of atg mutants. In response to starvation, wild-type (WT) cells were able to upregulate components of the respiratory pathway and ROS (reactive oxygen species) scavenging enzymes, but atg mutants lacked this synthetic capacity. Consequently, autophagy-defective mutants accumulated the high level of ROS, leading to deficient respiratory function, resulting in the loss of mitochondria DNA (mtDNA). We also showed that mtDNA deficient cells are subject to cell death under low pH starvation conditions. Taken together, under starvation conditions non-selective autophagy, rather than mitophagy, plays an essential role in preventing ROS accumulation, and thus in maintaining mitochondria function. The failure of response to starvation is the major cause of cell death in atg mutants.

Nutrient deprivation-related OXPHOS/glycolysis interconversion via HIF-1α/C-MYC pathway in U251 cells.

PubMed

Liu, Zhongjian; Sun, Yang; Tan, Shirui; Liu, Liang; Hu, Suqiong; Huo, Hongyu; Li, Meizhang; Cui, Qinghua; Yu, Min

2016-05-01

Although the Warburg effect is a dominant metabolic phenotype observed in cancers, the metabolic changes and adaptation occurring in tumors have been demonstrated to extend beyond the Warburg effect and thus considered a secondary effect to the transformation process of carcinogenesis, including nutritional deficiencies. However, the role of nutritional deficiencies in this metabolic reprogramming (e. g., oxidative phosphorylation (OXPHOS)/glycolysis interconversion) is not completely known yet. Here, we showed that under regular culture condition, the proliferation of U251 cells, but not other tumor cell lines, preferentially performed the Warburg effect and was remarkably inhibited by oxamic acid which can inhibit the activity of lactate dehydrogenase (LDH); whereas under serum starvation, glycolysis was depressed, tricarboxylic acid cycle (TCA) was enhanced, and the activity of OXPHOS was reinforced to maintain cellular ATP content in a high level, but interestingly, we observed a decreased expression of reactive oxygen species (ROS). Moreover, the upregulated activity of mitochondrial complex I was confirmed by Western blots and showed that the mitochondrial-related protein, NDUFA9, NDUFB8, ND1, and VDAC1 were remarkably increased after serum starved. Mechanistically, nutritional deficiencies could reduce hypoxia-inducible factor α (HIF-1α) protein expression to increase C-MYC protein level, which in turn increased nuclear respiratory factor 1 (NRF1) and mitochondrial transcription factor A (TFAM) transcription to enhance the activity of OXPHOS, suggesting that metabolic reprogramming by the changes of microenvironment during the carcinogenesis can provide some novel therapeutic clues to traditional cancer treatments.

Mechanisms of appearance of the Pasteur effect in Saccharomyces cerevisiae: inactivation of sugar transport systems.

PubMed

Lagunas, R; Dominguez, C; Busturia, A; Sáez, M J

1982-10-01

Saccharomyces cerevisiae does not show a noticeable Pasteur effect (activation of sugar catabolism by anaerobiosis) when growing with an excess of sugar and nitrogen source, but it does do so after exhaustion of the nitrogen source in the medium (resting state). We have found that this different behavior of growing and resting S. cerevisiae seems due to differences in the contribution of respiration to catabolism under both states. Growing S. cerevisiae respired only 3 to 20% of the catabolized sugar, depending on the sugar present; the remainder was fermented. In contrast, resting S. cerevisiae respired as much as 25 to 100% of the catabolized sugar. These results suggest that a shift to anaerobiosis would have much greater energetic consequences in resting than in growing S. cerevisiae. In resting S. cerevisiae anaerobiosis would strongly decrease the formation of ATP; as a consequence, various regulatory mechanisms would switch on, producing the observed increase of the rate of glycolysis. The greater significance that respiration reached in resting cells was not due to an increase of the respiratory capacity itself, but to a loss of fermentation which turned respiration into the main catabolic pathway. The main mechanism involved in the loss of fermentation observed during nitrogen starvation was a progressive inactivation of the sugar transport systems that reduced the rate of fermentation to less than 10% of the value observed in growing cells. Inactivation of the sugar transports seems a consequence of the turnover of the sugar carriers whose apparent half-lives were 2 to 7 h.

[The ability of cells to adjust to the low oxigen content associated with Na,K-ATPase glutationilation].

PubMed

Petrushanko, I Iu; Simonenko, O V; Burnysheva, K M; Klimanova, E A; Dergousova, E A; Mit'kevich, V A; Lopina, O D; Makarov, A A

2015-01-01

Decreasing the amount of oxygen in the tissues under hypoxic and ischemic conditions, observed at a number of pathologic processes, inevitably leads to their damage. One of the main causes of cell damage and death is a violation of the systems maintaining ionic balance. Na,K-ATPaseis a basic ion-transporting protein of animal cell plasma membrane and inhibition of the Na,K-ATPase activity at lower concentrations of oxygen is one of the earliest and most critical events for cell viability. Currently there is an active search for modulators of Na,K-ATPase activity. For this purpose traditionally used cardiac glycosides but the existence of serious adverse effects forced to look for alternative inhibitors of Na,K-ATPase. Previously we have found that the glutathionylation of Na,K-ATPase catalytic subunit leads to a complete-inhibition of the enzyme. In this paper it is shown that the agents which increase the level of Na,K-ATPase glutathionylation: ethyl glutathione (et-GSH), oxidized glutathione (GSSG) and N-acetyl cysteine (NAC), increase cell survival under oxygen deficiency conditions, prevent decline of ATP in the cells and normalize their redox status. Concentration range in which these substances have a maximum protective effect, and does not exhibit cytotoxic properties was defined: for et-GSH 0.2-0.5 mM, for GSSG 0.2-1 mM, for NAC 10 to 15 mM. The results show prospects for development of methods for tissues protection from damage caused by oxygen starvation by varying the degree of Na,K-ATPase glutathionylation.

Atp1a3-deficient heterozygous mice show lower rank in the hierarchy and altered social behavior.

PubMed

Sugimoto, H; Ikeda, K; Kawakami, K

2018-06-01

Atp1a3 is the Na-pump alpha3 subunit gene expressed mainly in neurons of the brain. Atp1a3-deficient heterozygous mice (Atp1a3 +/- ) show altered neurotransmission and deficits of motor function after stress loading. To understand the function of Atp1a3 in a social hierarchy, we evaluated social behaviors (social interaction, aggression, social approach and social dominance) of Atp1a3 +/- and compared the rank and hierarchy structure between Atp1a3 +/- and wild-type mice within a housing cage using the round-robin tube test and barbering observations. Formation of a hierarchy decreases social conflict and promote social stability within the group. The hierarchical rank is a reflection of social dominance within a cage, which is heritable and can be regulated by specific genes in mice. Here we report: (1) The degree of social interaction but not aggression was lower in Atp1a3 +/- than wild-type mice, and Atp1a3 +/- approached Atp1a3 +/- mice more frequently than wild type. (2) The frequency of barbering was lower in the Atp1a3 +/- group than in the wild-type group, while no difference was observed in the mixed-genotype housing condition. (3) Hierarchy formation was not different between Atp1a3 +/- and wild type. (4) Atp1a3 +/- showed a lower rank in the mixed-genotype housing condition than that in the wild type, indicating that Atp1a3 regulates social dominance. In sum, Atp1a3 +/- showed unique social behavior characteristics of lower social interaction and preference to approach the same genotype mice and a lower ranking in the hierarchy. © 2017 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

Ca2+-regulated-cAMP/PKA signaling in cardiac pacemaker cells links ATP supply to demand.

PubMed

Yaniv, Yael; Juhaszova, Magdalena; Lyashkov, Alexey E; Spurgeon, Harold A; Sollott, Steven J; Lakatta, Edward G

2011-11-01

In sinoatrial node cells (SANC), Ca(2+) activates adenylate cyclase (AC) to generate a high basal level of cAMP-mediated/protein kinase A (PKA)-dependent phosphorylation of Ca(2+) cycling proteins. These result in spontaneous sarcoplasmic-reticulum (SR) generated rhythmic Ca(2+) oscillations during diastolic depolarization, that not only trigger the surface membrane to generate rhythmic action potentials (APs), but, in a feed-forward manner, also activate AC/PKA signaling. ATP is consumed to pump Ca(2+) to the SR, to produce cAMP, to support contraction and to maintain cell ionic homeostasis. Since feedback mechanisms link ATP-demand to ATP production, we hypothesized that (1) both basal ATP supply and demand in SANC would be Ca(2+)-cAMP/PKA dependent; and (2) due to its feed-forward nature, a decrease in flux through the Ca(2+)-cAMP/PKA signaling axis will reduce the basal ATP production rate. O(2) consumption in spontaneous beating SANC was comparable to ventricular myocytes (VM) stimulated at 3 Hz. Graded reduction of basal Ca(2+)-cAMP/PKA signaling to reduce ATP demand in rabbit SANC produced graded ATP depletion (r(2)=0.96), and reduced O(2) consumption and flavoprotein fluorescence. Neither inhibition of glycolysis, selectively blocking contraction nor specific inhibition of mitochondrial Ca(2+) flux reduced the ATP level. Feed-forward basal Ca(2+)-cAMP/PKA signaling both consumes ATP to drive spontaneous APs in SANC and is tightly linked to mitochondrial ATP production. Interfering with Ca(2+)-cAMP/PKA signaling not only slows the firing rate and reduces ATP consumption, but also appears to reduce ATP production so that ATP levels fall. This distinctly differs from VM, which lack this feed-forward basal cAMP/PKA signaling, and in which ATP level remains constant when the demand changes. Published by Elsevier Ltd.

Variable effects of the mitoK(ATP) channel modulators diazoxide and 5-HD in ATP-depleted renal epithelial cells.

PubMed

Nilakantan, Vani; Liang, Huanling; Mortensen, Jordan; Taylor, Erin; Johnson, Christopher P

2010-02-01

The role of mitochondrial K(ATP) (mitoK(ATP)) channels in renal ischemia-reperfusion injury is controversial with studies showing both protective and deleterious effects. In this study, we compared the effects of the putative mitoK(ATP) opener, diazoxide, and the mitoK(ATP) blocker, 5-hydroxydecanoate (5-HD) on cytotoxicity and apoptosis in tubular epithelial cells derived from rat (NRK-52E) and pig (LLC-PK1) following in vitro ischemic injury. Following ATP depletion-recovery, there was a significant increase in cytotoxicity in both NRK cells and LLC-PK1 cells although NRK cells were more sensitive to the injury. Diazoxide treatment attenuated cytotoxicity in both cell types and 5-HD treatment-increased cytotoxicity in the sensitive NRK cells in a superoxide-dependant manner. The protective effect of diazoxide was also reversed in the presence of 5-HD in ATP-depleted NRK cells. The ATP depletion-mediated increase in superoxide was enhanced by both diazoxide and 5-HD with the effect being more pronounced in the cells undergoing 5-HD treatment. Further, ATP depletion-induced activation of caspase-3 was decreased by diazoxide in NRK cells. In order to determine the signaling pathways involved in apoptosis, we examined the activation of Erk and JNK in ATP-depleted NRK cells. Diazoxide-activated Erk in ATP-depleted cells, but did not have any effect on JNK activation. In contrast, 5-HD did not impact Erk levels but increased JNK activation even under controlled conditions. Further, the use of a JNK inhibitor with 5-HD reversed the deleterious effects of 5-HD. This study demonstrates that in cells that are sensitive to ATP depletion-recovery, mitoK(ATP) channels protect against ATP depletion-mediated cytotoxicity and apoptosis through Erk- and JNK-dependant mechanisms.

The regulation of ATP release from the urothelium by adenosine and transepithelial potential.

PubMed

Dunning-Davies, Bryony M; Fry, Christopher H; Mansour, Dina; Ferguson, Douglas R

2013-03-01

WHAT'S KNOWN ON THE SUBJECT? AND WHAT DOES THE STUDY ADD?: Stretch of the urothelium, as occurs during bladder filling, is associated with a release of ATP that is postulated to act as a sensory neurotransmitter. The regulation of ATP release is poorly understood and in particular if there is a feedback mechanism provided by ATP itself. Adenosine, a breakdown product of ATP, is a potent inhibitor of stretch-induced ATP release, acting through and A1 receptor; endogenous levels are about 0.6μM. Data are consistent with ATP release relying on the rise of intracellular Ca2+. Transepithelial potential also controls ATP release, also acting via an A1 receptor-dependent pathway. To test the hypothesis that distension-induced ATP release from the bladder urothelium is regulated by adenosine as well as changes to transurothelial potential (TEP). To examine the role of changes to intracellular [Ca(2+) ] in ATP release. Rabbit urothelium/suburothelium membranes were used in an Ussing chamber system. Distension was induced by fluid removal from the chamber bathing the serosal (basolateral) membrane face. The TEP and short-circuit current were measured. ATP was measured in samples aspirated from the serosal chamber by a luciferin-luciferase assay. Intracellular [Ca(2+) ] was measured in isolated urothelial cells using the fluorochrome Fura-2. All experiments were performed at 37°C. Distension-induced ATP release was decreased by adenosine (1-10 μm) and enhanced by adenosine deaminase and A1- (but not A2-) receptor antagonists. Distension-induced ATP release was reduced by 2-APB, nifedipine and capsazepine; capsaicin induced ATP release in the absence of distension. ATP and capsaicin, but not adenosine, generated intracellular Ca(2+) transients; adenosine did not affect the ATP-generated Ca(2+) transient. ATP release was dependent on a finite transepithelial potential. Changes to TEP, in the absence of distension, generated ATP release that was in turn reduced by adenosine. Adenosine exerts a powerful negative feedback control of ATP release from the urothelium via A1 receptor activation. Distension-induced ATP release may be mediated by a rise of the intracellular [Ca(2+) ]. Modulation of distension-induced ATP release by adenosine and TEP may have a common pathway. © 2012 BJU International.

High fat, high sucrose diet causes cardiac mitochondrial dysfunction due in part to oxidative post-translational modification of mitochondrial complex II

PubMed Central

Sverdlov, Aaron L.; Elezaby, Aly; Behring, Jessica B.; Bachschmid, Markus M.; Luptak, Ivan; Tu, Vivian H.; Siwik, Deborah A.; Miller, Edward J.; Liesa, Marc; Shirihai, Orian S; Pimentel, David R.; Cohen, Richard A.; Colucci, Wilson S.

2014-01-01

Background Diet-induced obesity leads to metabolic heart disease (MHD) characterized by increased oxidative stress that may cause oxidative post-translational modifications (OPTM) of cardiac mitochondrial proteins. The functional consequences of OPTM of cardiac mitochondrial proteins in MHD are unknown. Our objective was to determine whether cardiac mitochondrial dysfunction in MHD due to diet-induced obesity is associated with cysteine OPTM. Methods and results Male C57Bl/6J mice were fed either a high-fat, high-sucrose (HFHS) or control diet for 8 months. Cardiac mitochondria from HFHS-fed mice (vs. control diet) had an increased rate of H2O2 production, a decreased GSH/GSSG ratio, a decreased rate of complex II substrate-driven ATP synthesis and decreased complex II activity. Complex II substrate-driven ATP synthesis and complex II activity were partially restored ex-vivo by reducing conditions. A biotin switch assay showed that HFHS feeding increased cysteine OPTM in complex II subunits A (SDHA) and B (SDHB). Using iodo-TMT multiplex tags we found that HFHS feeding is associated with reversible oxidation of cysteines 89 and 231 in SDHA, and 100, 103 and 115 in SDHB. Conclusions MHD due to consumption of a HFHS “Western” diet causes increased H2O2 production and oxidative stress in cardiac mitochondria associated with decreased ATP synthesis and decreased complex II activity. Impaired complex II activity and ATP production are associated with reversible cysteine OPTM of complex II. Possible sites of reversible cysteine OPTM in SDHA and SDHB were identified by iodo-TMT tag labeling. Mitochondrial ROS may contribute to the pathophysiology of MHD by impairing the function of complex II. PMID:25109264

Cytoprotection by fructose and other ketohexoses during bile salt-induced apoptosis of hepatocytes.

PubMed

Zeid, I M; Bronk, S F; Fesmier, P J; Gores, G J

1997-01-01

Toxic bile salts cause hepatocyte necrosis at high concentrations and apoptosis at lower concentrations. Although fructose prevents bile salt-induced necrosis, the effect of fructose on bile salt-induced apoptosis is unclear. Our aim was to determine if fructose also protects against bile salt-induced apoptosis. Fructose inhibited glycochenodeoxycholate (GCDC)-induced apoptosis in a concentration-dependent manner with a maximum inhibition of 72% +/- 10% at 10 mmol/L. First, we determined if fructose inhibited apoptosis by decreasing adenosine triphosphate (ATP) and intracellular pH (pHi). Although fructose decreased ATP to <25% of basal values, oligomycin (an ATP synthase inhibitor) did not inhibit apoptosis despite decreasing ATP to similar values. Fructose (10 mmol/L) decreased intracellular pH (pHi) by 0.2 U. However, extracellular acidification (pH 6.8), which decreased hepatocyte pHi 0.35 U and is known to inhibit necrosis, actually potentiated apoptosis 1.6-fold. Fructose cytoprotection also could not be explained by induction of bcl-2 transcription or metal chelation. Because we could not attribute fructose cytoprotection to metabolic effects, alterations in the expression of bcl-2, or metal chelation, we next determined if the poorly metabolized ketohexoses, tagatose and sorbose, also inhibited apoptosis; unexpectedly, both ketohexoses inhibited apoptosis. Because bile salt-induced apoptosis and necrosis are inhibited by fructose, these data suggest that similar processes initiate bile salt-induced hepatocyte necrosis and apoptosis. In contrast, acidosis, which inhibits necrosis, potentiates apoptosis. Thus, ketohexose-sensitive pathways appear to initiate both bile salt-induced cell apoptosis and necrosis, whereas dissimilar, pH-sensitive, effector mechanisms execute these two different cell death processes.

Breast feeding increases vasoconstriction induced by electrical field stimulation in rat mesenteric artery. Role of neuronal nitric oxide and ATP.

PubMed

Blanco-Rivero, Javier; Sastre, Esther; Caracuel, Laura; Granado, Miriam; Balfagón, Gloria

2013-01-01

The aim of this study was to investigate in rat mesenteric artery whether breast feeding (BF) affects the vasomotor response induced by electrical field stimulation (EFS), participation by different innervations in the EFS-induced response and the mechanism/s underlying these possible modifications. Experiments were performed in female Sprague-Dawley rats (3 months old), divided into three groups: Control (in oestrous phase), mothers after 21 days of BF, and mothers that had recovered their oestral cycle (After BF, in oestrous phase). Vasomotor response to EFS, noradrenaline (NA) and nitric oxide (NO) donor DEA-NO were studied. Neuronal NO synthase (nNOS) and phosphorylated nNOS (P-nNOS) protein expression were analysed and NO, superoxide anion (O(2)(.-)), NA and ATP releases were also determined. EFS-induced contraction was higher in the BF group, and was recovered after BF. 1 µmol/L phentolamine decreased the response to EFS similarly in control and BF rats. NA vasoconstriction and release were similar in both experimental groups. ATP release was higher in segments from BF rats. 0.1 mmol/L L-NAME increased the response to EFS in both control and BF rats, but more so in control animals. BF decreased NO release and did not modify O(2)(.-) production. Vasodilator response to DEA-NO was similar in both groups, while nNOS and P-nNOS expressions were decreased in segments from BF animals. Breast feeding increases EFS-induced contraction in mesenteric arteries, mainly through the decrease of neuronal NO release mediated by decreased nNOS and P-nNOS expression. Sympathetic function is increased through the increased ATP release in BF rats.

Breast Feeding Increases Vasoconstriction Induced by Electrical Field Stimulation in Rat Mesenteric Artery. Role of Neuronal Nitric Oxide and ATP

PubMed Central

Caracuel, Laura; Granado, Miriam; Balfagón, Gloria

2013-01-01

Objectives The aim of this study was to investigate in rat mesenteric artery whether breast feeding (BF) affects the vasomotor response induced by electrical field stimulation (EFS), participation by different innervations in the EFS-induced response and the mechanism/s underlying these possible modifications. Methods Experiments were performed in female Sprague-Dawley rats (3 months old), divided into three groups: Control (in oestrous phase), mothers after 21 days of BF, and mothers that had recovered their oestral cycle (After BF, in oestrous phase). Vasomotor response to EFS, noradrenaline (NA) and nitric oxide (NO) donor DEA-NO were studied. Neuronal NO synthase (nNOS) and phosphorylated nNOS (P-nNOS) protein expression were analysed and NO, superoxide anion (O2 .–), NA and ATP releases were also determined. Results EFS-induced contraction was higher in the BF group, and was recovered after BF. 1 µmol/L phentolamine decreased the response to EFS similarly in control and BF rats. NA vasoconstriction and release were similar in both experimental groups. ATP release was higher in segments from BF rats. 0.1 mmol/L L-NAME increased the response to EFS in both control and BF rats, but more so in control animals. BF decreased NO release and did not modify O2 .– production. Vasodilator response to DEA-NO was similar in both groups, while nNOS and P-nNOS expressions were decreased in segments from BF animals. Conclusion Breast feeding increases EFS-induced contraction in mesenteric arteries, mainly through the decrease of neuronal NO release mediated by decreased nNOS and P-nNOS expression. Sympathetic function is increased through the increased ATP release in BF rats. PMID:23342008

HNP-3 enhanced the antimicrobial activity of CIP by promoting ATP efflux from P. aeruginosa cells.

PubMed

Wang, Hao; Dong, Birong; Lou, Li

2011-04-01

To establish a novel strategy of P. aeruginosa control, we acquired recombination HNP-3 by gene recombination. Then we examined HNP-3 bio-activities and the influences of antimicrobial peptide on the efflux of ATP. Consequently, we obtained target protein with a molecular mass of 3,000 D consistent with the Anticipation. FIC index of Ciprofloxacin added HNP-3 was less than 0.5 and HNP3 synergistically cooperated with CIP to suppress P. aeruginosa colony formation revealed that there was significant synergy. ATP efflux was however up-regulated by low concentrations of HNP-3, although CIP did not exert any influence on ATP efflux. Conclusively, recombination protein HNP-3 displayed antimicrobial and synergic effects. HNP-3 enhanced the antimicrobial activity of CIP by promoting ATP efflux from P. aeruginosa cells and decreasing efflux of the drugs, which could have useful clinical applications.

ATP11B mediates platinum resistance in ovarian cancer

PubMed Central

Moreno-Smith, Myrthala; Halder, J.B.; Meltzer, Paul S.; Gonda, Tamas A.; Mangala, Lingegowda S.; Rupaimoole, Rajesha; Lu, Chunhua; Nagaraja, Archana S.; Gharpure, Kshipra M.; Kang, Yu; Rodriguez-Aguayo, Cristian; Vivas-Mejia, Pablo E.; Zand, Behrouz; Schmandt, Rosemarie; Wang, Hua; Langley, Robert R.; Jennings, Nicholas B.; Ivan, Cristina; Coffin, Jeremy E.; Armaiz, Guillermo N.; Bottsford-Miller, Justin; Kim, Sang Bae; Halleck, Margaret S.; Hendrix, Mary J.C.; Bornman, William; Bar-Eli, Menashe; Lee, Ju-Seog; Siddik, Zahid H.; Lopez-Berestein, Gabriel; Sood, Anil K.

2013-01-01

Platinum compounds display clinical activity against a wide variety of solid tumors; however, resistance to these agents is a major limitation in cancer therapy. Reduced platinum uptake and increased platinum export are examples of resistance mechanisms that limit the extent of DNA damage. Here, we report the discovery and characterization of the role of ATP11B, a P-type ATPase membrane protein, in cisplatin resistance. We found that ATP11B expression was correlated with higher tumor grade in human ovarian cancer samples and with cisplatin resistance in human ovarian cancer cell lines. ATP11B gene silencing restored the sensitivity of ovarian cancer cell lines to cisplatin in vitro. Combined therapy of cisplatin and ATP11B-targeted siRNA significantly decreased cancer growth in mice bearing ovarian tumors derived from cisplatin-sensitive and -resistant cells. In vitro mechanistic studies on cellular platinum content and cisplatin efflux kinetics indicated that ATP11B enhances the export of cisplatin from cells. The colocalization of ATP11B with fluorescent cisplatin and with vesicular trafficking proteins, such as syntaxin-6 (STX6) and vesicular-associated membrane protein 4 (VAMP4), strongly suggests that ATP11B contributes to secretory vesicular transport of cisplatin from Golgi to plasma membrane. In conclusion, inhibition of ATP11B expression could serve as a therapeutic strategy to overcome cisplatin resistance. PMID:23585472

ATP13A2 variability in Parkinson disease

PubMed Central

Vilariño-Güell, Carles; Soto, Alexandra I.; Lincoln, Sarah J.; Yahmed, Samia Ben; Kefi, Mounir; Heckman, Michael G.; Hulihan, Mary M.; Chai, Hua; Diehl, Nancy N.; Amouri, Rim; Rajput, Alex; Mash, Deborah C.; Dickson, Dennis W.; Middleton, Lefkos T.; Gibson, Rachel A.; Hentati, Faycal; Farrer, Matthew J.

2008-01-01

Recessively inherited mutations in ATP13A2 result in Kufor-Rakeb syndrome, whereas genetic variability and elevated ATP13A2 expression have been implicated in Parkinson disease (PD). Given this background, ATP13A2 was comprehensively assessed to support or refute its contribution to PD. Sequencing of ATP13A2 exons and intron-exon boundaries was performed in 89 probands with familial parkinsonism from Tunisia. The segregation of mutations with parkinsonism was subsequently assessed within pedigrees. The frequency of genetic variants and evidence for association was also examined in 240 patients with non-familial PD and 372 healthy controls. ATP13A2 mRNA expression was also quantified in brain tissues from 38 patients with non-familial PD and 38 healthy subjects from the US. Sequencing analysis revealed 37 new variants; seven missense, six silent and 24 that were noncoding. However, no single ATP13A2 mutation segregated with familial parkinsonism in either a dominant or recessive manner. Four markers showed marginal association with non-familial PD, prior to correction for multiple testing. ATP13A2 mRNA expression was marginally decreased in PD brains compared with tissue from control subjects. In conclusion, neither ATP13A2 genetic variability nor quantitative gene expression in brain appears to contribute to familial parkinsonism or non-familial PD. PMID:19085912

Uridylylation of Herbaspirillum seropedicae GlnB and GlnK proteins is differentially affected by ATP, ADP and 2-oxoglutarate in vitro.

PubMed

Bonatto, Ana C; Souza, Emanuel M; Oliveira, Marco A S; Monteiro, Rose A; Chubatsu, Leda S; Huergo, Luciano F; Pedrosa, Fábio O

2012-08-01

PII are signal-transducing proteins that integrate metabolic signals and transmit this information to a large number of proteins. In proteobacteria, PII are modified by GlnD (uridylyltransferase/uridylyl-removing enzyme) in response to the nitrogen status. The uridylylation/deuridylylation cycle of PII is also regulated by carbon and energy signals such as ATP, ADP and 2-oxoglutarate (2-OG). These molecules bind to PII proteins and alter their tridimensional structure/conformation and activity. In this work, we determined the effects of ATP, ADP and 2-OG levels on the in vitro uridylylation of Herbaspirillum seropedicae PII proteins, GlnB and GlnK. Both proteins were uridylylated by GlnD in the presence of ATP or ADP, although the uridylylation levels were higher in the presence of ATP and under high 2-OG levels. Under excess of 2-OG, the GlnB uridylylation level was higher in the presence of ATP than with ADP, while GlnK uridylylation was similar with ATP or ADP. Moreover, in the presence of ADP/ATP molar ratios varying from 10/1 to 1/10, GlnB uridylylation level decreased as ADP concentration increased, whereas GlnK uridylylation remained constant. The results suggest that uridylylation of both GlnB and GlnK responds to 2-OG levels, but only GlnB responds effectively to variation on ADP/ATP ratio.

Possible involvement of ATP-sensitive potassium channels in the antidepressant-like effects of gabapentin in mouse forced swimming test.

PubMed

Ostadhadi, Sattar; Akbarian, Reyhaneh; Norouzi-Javidan, Abbas; Nikoui, Vahid; Zolfaghari, Samira; Chamanara, Mohsen; Dehpour, Ahmad-Reza

2017-07-01

Gabapentin as an anticonvulsant drug also has beneficial effects in treatment of depression. Previously, we showed that acute administration of gabapentin produced an antidepressant-like effect in the mouse forced swimming test (FST) by a mechanism that involves the inhibition of nitric oxide (NO). Considering the involvement of NO in adenosine triphosphate (ATP)-sensitive potassium channels (K ATP ), in the present study we investigated the involvement of K ATP channels in antidepressant-like effect of gabapentin. Gabapentin at different doses (5-10 mg/kg) and fluoxetine (20 mg/kg) were administrated by intraperitoneal route, 60 and 30 min, respectively, before the test. To clarify the probable involvement of K ATP channels, mice were pretreated with K ATP channel inhibitor or opener. Gabapentin at dose 10 mg/kg significantly decreased the immobility behavior of mice similar to fluoxetine (20 mg/kg). Co-administration of subeffective dose (1 mg/kg) of glibenclamide (inhibitor of K ATP channels) with gabapentin (3 mg/kg) showed a synergistic antidepressant-like effect. Also, subeffective dose of cromakalim (opener of K ATP channels, 0.1 mg/kg) inhibited the antidepressant-like effect of gabapentin (10 mg/kg). None of the treatments had any impact on the locomotor movement. Our study, for the first time, revealed that antidepressant-like effect of gabapentin in mice is mediated by blocking the K ATP channels.

Erosion potential of the Yangtze Delta under sediment starvation and climate change.

PubMed

Yang, H F; Yang, S L; Xu, K H; Wu, H; Shi, B W; Zhu, Q; Zhang, W X; Yang, Z

2017-09-05

Deltas are widely threatened by sediment starvation and climate change. Erosion potential is an important indicator of delta vulnerability. Here, we investigate the erosion potential of the Yangtze Delta. We found that over the past half century the Yangtze's sediment discharge has decreased by 80% due to the construction of >50,000 dams and soil conservation, whereas the wind speed and wave height in the delta region have increased by 5-7%, and the sea level has risen at a rate of 3 mm/yr. According to hydrodynamic measurements and analyses of seabed sediments, the period when bed shear stress due to combined current-wave action under normal weather conditions exceeds the critical bed shear stress for erosion (τ cr ) accounts for 63% of the total observed period on average and can reach 100% during peak storms. This explains why net erosion has occurred in some areas of the subaqueous delta. We also found that the increase with depth of τ cr is very gradual in the uppermost several metres of the depositional sequence. We therefore expect that the Yangtze subaqueous delta will experience continuous erosion under sediment starvation and climate change in the next decades of this century or even a few centuries.

Tomato strigolactones are derived from carotenoids and their biosynthesis is promoted by phosphate starvation.

PubMed

López-Ráez, Juan Antonio; Charnikhova, Tatsiana; Gómez-Roldán, Victoria; Matusova, Radoslava; Kohlen, Wouter; De Vos, Ric; Verstappen, Francel; Puech-Pages, Virginie; Bécard, Guillaume; Mulder, Patrick; Bouwmeester, Harro

2008-01-01

* Strigolactones are rhizosphere signalling compounds that mediate host location in arbuscular mycorrhizal (AM) fungi and parasitic plants. Here, the regulation of the biosynthesis of strigolactones is studied in tomato (Solanum lycopersicum). * Strigolactone production under phosphate starvation, in the presence of the carotenoid biosynthesis inhibitor fluridone and in the abscisic acid (ABA) mutant notabilis were assessed using a germination bioassay with seeds of Orobanche ramosa; a hyphal branching assay with Gigaspora spp; and by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) analysis. * The root exudates of tomato cv. MoneyMaker induced O. ramosa seed germination and hyphal branching in AM fungi. Phosphate starvation markedly increased, and fluridone strongly decreased, this activity. Exudates of notabilis induced approx. 40% less germination than the wild-type. The LC-MS/MS analysis confirmed that the biological activity and changes therein were due to the presence of several strigolactones; orobanchol, solanacol and two or three didehydro-orobanchol isomers. * These results show that the AM branching factors and parasitic plant germination stimulants in tomato root exudate are strigolactones and that they are biosynthetically derived from carotenoids. The dual activity of these signalling compounds in attracting beneficial AM fungi and detrimental parasitic plants is further strengthened by environmental conditions such as phosphate availability.

A conserved two-component signal transduction system controls the response to phosphate starvation in Bifidobacterium breve UCC2003.

PubMed

Alvarez-Martin, Pablo; Fernández, Matilde; O'Connell-Motherway, Mary; O'Connell, Kerry Joan; Sauvageot, Nicolas; Fitzgerald, Gerald F; MacSharry, John; Zomer, Aldert; van Sinderen, Douwe

2012-08-01

This work reports on the identification and molecular characterization of the two-component regulatory system (2CRS) PhoRP, which controls the response to inorganic phosphate (P(i)) starvation in Bifidobacterium breve UCC2003. The response regulator PhoP was shown to bind to the promoter region of pstSCAB, specifying a predicted P(i) transporter system, as well as that of phoU, which encodes a putative P(i)-responsive regulatory protein. This interaction is assumed to cause transcriptional modulation under conditions of P(i) limitation. Our data suggest that the phoRP genes are subject to positive autoregulation and, together with pstSCAB and presumably phoU, represent the complete regulon controlled by the phoRP-encoded 2CRS in B. breve UCC2003. Determination of the minimal PhoP binding region combined with bioinformatic analysis revealed the probable recognition sequence of PhoP, designated here as the PHO box, which together with phoRP is conserved among many high-GC-content Gram-positive bacteria. The importance of the phoRP 2CRS in the response of B. breve to P(i) starvation conditions was confirmed by analysis of a B. breve phoP insertion mutant which exhibited decreased growth under phosphate-limiting conditions compared to its parent strain UCC2003.

A Conserved Two-Component Signal Transduction System Controls the Response to Phosphate Starvation in Bifidobacterium breve UCC2003

PubMed Central

Alvarez-Martin, Pablo; Fernández, Matilde; O'Connell-Motherway, Mary; O'Connell, Kerry Joan; Sauvageot, Nicolas; Fitzgerald, Gerald F.; MacSharry, John; Zomer, Aldert

2012-01-01

This work reports on the identification and molecular characterization of the two-component regulatory system (2CRS) PhoRP, which controls the response to inorganic phosphate (Pi) starvation in Bifidobacterium breve UCC2003. The response regulator PhoP was shown to bind to the promoter region of pstSCAB, specifying a predicted Pi transporter system, as well as that of phoU, which encodes a putative Pi-responsive regulatory protein. This interaction is assumed to cause transcriptional modulation under conditions of Pi limitation. Our data suggest that the phoRP genes are subject to positive autoregulation and, together with pstSCAB and presumably phoU, represent the complete regulon controlled by the phoRP-encoded 2CRS in B. breve UCC2003. Determination of the minimal PhoP binding region combined with bioinformatic analysis revealed the probable recognition sequence of PhoP, designated here as the PHO box, which together with phoRP is conserved among many high-GC-content Gram-positive bacteria. The importance of the phoRP 2CRS in the response of B. breve to Pi starvation conditions was confirmed by analysis of a B. breve phoP insertion mutant which exhibited decreased growth under phosphate-limiting conditions compared to its parent strain UCC2003. PMID:22635988

Taste-independent detection of the caloric content of sugar in Drosophila

PubMed Central

Dus, Monica; Min, SooHong; Keene, Alex C.; Lee, Ga Young; Suh, Greg S. B.

2011-01-01

Feeding behavior is influenced primarily by two factors: nutritional needs and food palatability. However, the role of food deprivation and metabolic needs in the selection of appropriate food is poorly understood. Here, we show that the fruit fly, Drosophila melanogaster, selects calorie-rich foods following prolonged food deprivation in the absence of taste-receptor signaling. Flies mutant for the sugar receptors Gr5a and Gr64a cannot detect the taste of sugar, but still consumed sugar over plain agar after 15 h of starvation. Similarly, pox-neuro mutants that are insensitive to the taste of sugar preferentially consumed sugar over plain agar upon starvation. Moreover, when given a choice between metabolizable sugar (sucrose or d-glucose) and nonmetabolizable (zero-calorie) sugar (sucralose or l-glucose), starved Gr5a; Gr64a double mutants preferred metabolizable sugars. These findings suggest the existence of a taste-independent metabolic sensor that functions in food selection. The preference for calorie-rich food correlates with a decrease in the two main hemolymph sugars, trehalose and glucose, and in glycogen stores, indicating that this sensor is triggered when the internal energy sources are depleted. Thus, the need to replenish depleted energy stores during periods of starvation may be met through the activity of a taste-independent metabolic sensing pathway. PMID:21709242

Taste-independent detection of the caloric content of sugar in Drosophila.

PubMed

Dus, Monica; Min, SooHong; Keene, Alex C; Lee, Ga Young; Suh, Greg S B

2011-07-12

Feeding behavior is influenced primarily by two factors: nutritional needs and food palatability. However, the role of food deprivation and metabolic needs in the selection of appropriate food is poorly understood. Here, we show that the fruit fly, Drosophila melanogaster, selects calorie-rich foods following prolonged food deprivation in the absence of taste-receptor signaling. Flies mutant for the sugar receptors Gr5a and Gr64a cannot detect the taste of sugar, but still consumed sugar over plain agar after 15 h of starvation. Similarly, pox-neuro mutants that are insensitive to the taste of sugar preferentially consumed sugar over plain agar upon starvation. Moreover, when given a choice between metabolizable sugar (sucrose or D-glucose) and nonmetabolizable (zero-calorie) sugar (sucralose or L-glucose), starved Gr5a; Gr64a double mutants preferred metabolizable sugars. These findings suggest the existence of a taste-independent metabolic sensor that functions in food selection. The preference for calorie-rich food correlates with a decrease in the two main hemolymph sugars, trehalose and glucose, and in glycogen stores, indicating that this sensor is triggered when the internal energy sources are depleted. Thus, the need to replenish depleted energy stores during periods of starvation may be met through the activity of a taste-independent metabolic sensing pathway.

Changes in the Expression and the Enzymic Properties of the 20S Proteasome in Sugar-Starved Maize Roots. Evidence for an in Vivo Oxidation of the Proteasome1

PubMed Central

Basset , Gilles; Raymond, Philippe; Malek, Lada; Brouquisse, Renaud

2002-01-01

The 20S proteasome (multicatalytic proteinase) was purified from maize (Zea mays L. cv DEA 1992) roots through a five-step procedure. After biochemical characterization, it was shown to be similar to most eukaryotic proteasomes. We investigated the involvement of the 20S proteasome in the response to carbon starvation in excised maize root tips. Using polyclonal antibodies, we showed that the amount of proteasome increased in 24-h-carbon-starved root tips compared with freshly excised tips, whereas the mRNA levels of α3 and β6 subunits of 20S proteasome decreased. Moreover, in carbon-starved tissues, chymotrypsin-like and caseinolytic activities of the 20S proteasome were found to increase, whereas trypsin-like activities decreased. The measurement of specific activities and kinetic parameters of 20S proteasome purified from 24-h-starved root tips suggested that it was subjected to posttranslational modifications. Using dinitrophenylhydrazine, a carbonyl-specific reagent, we observed an increase in carbonyl residues in 20S proteasome purified from starved root tips. This means that 20S proteasome was oxidized during starvation treatment. Moreover, an in vitro mild oxidative treatment of 20S proteasome from non-starved material resulted in the activation of chymotrypsin-like, peptidyl-glutamyl-peptide hydrolase and caseinolytic-specific activities and in the inhibition of trypsin-like specific activities, similar to that observed for proteasome from starved root tips. Our results provide the first evidence, to our knowledge, for an in vivo carbonylation of the 20S proteasome. They suggest that sugar deprivation induces an oxidative stress, and that oxidized 20S proteasome could be associated to the degradation of oxidatively damaged proteins in carbon starvation situations. PMID:11891269

Moderate mammalian target of rapamycin inhibition induces autophagy in HTR8/SVneo cells via O-linked β-N-acetylglucosamine signaling.

PubMed

Zhang, Qiuxia; Na, Quan; Song, Weiwei

2017-10-01

Autophagy, a highly regulated process with a dual role (pro-survival or pro-death), has been implicated in adverse pregnancy outcomes. The aim of this study was to explore the mechanism whereby mammalian target of rapamycin (mTOR) signaling regulates autophagy by modulating protein O-GlcNAcylation in human trophoblasts. HTR8/SVneo cells were incubated in serum-free medium for different time intervals or treated with varying doses of Torin1. Protein expression and cell apoptosis were detected by immunoblotting and flow cytometry, respectively. Short-term serum starvation or slight suppression of mTOR signaling promoted autophagy and decreased apoptosis in HTR8/SVneo cells. Conversely, prolonged serum starvation or excessive inhibition of mTOR reduced autophagy and enhanced cell apoptosis. Both serum starvation and mTOR signaling suppression reduced protein O-GlcNAcylation. Upregulation and downregulation of O-linked β-N-acetylglucosamine (O-GlcNAc) levels attenuated and augmented autophagy, respectively. Moderate mTOR inhibition-induced autophagy was blocked by upregulation of protein O-GlcNAcylation. Furthermore, immunoprecipitation studies revealed that Beclin1 and synaptosome associated protein 29 (SNAP29) could be O-GlcNAcylated, and that slight mTOR inhibition resulted in decreased O-GlcNAc modification of Beclin1 and SNAP29. Notably, we observed an inverse correlation between phosphorylation (Ser15) and O-GlcNAcylation of Beclin1. mTOR signaling inhibition played dual roles in regulating autophagy and apoptosis in HTR8/SVneo cells. Moderate mTOR suppression might induce autophagy via modulating O-GlcNAcylation of Beclin1 and SNAP29. Moreover, the negative interplay between Beclin1 O-GlcNAcylation and phosphorylation (Ser15) may be involved in autophagy regulation by mTOR signaling. © 2017 Japan Society of Obstetrics and Gynecology.

Evidence of the Trade-Off between Starvation and Predation Risks in Ducks

PubMed Central

Zimmer, Cédric; Boos, Mathieu; Poulin, Nicolas; Gosler, Andrew; Petit, Odile; Robin, Jean-Patrice

2011-01-01

The theory of trade-off between starvation and predation risks predicts a decrease in body mass in order to improve flight performance when facing high predation risk. To date, this trade-off has mainly been validated in passerines, birds that store limited body reserves for short-term use. In the largest avian species in which the trade-off has been investigated (the mallard, Anas platyrhynchos), the slope of the relationship between mass and flight performance was steeper in proportion to lean body mass than in passerines. In order to verify whether the same case can be applied to other birds with large body reserves, we analyzed the response to this trade-off in two other duck species, the common teal (Anas crecca) and the tufted duck (Aythya fuligula). Predation risk was simulated by disturbing birds. Ducks within disturbed groups were compared to non-disturbed control birds. In disturbed groups, both species showed a much greater decrease in food intake and body mass during the period of simulated high risk than those observed in the control group. This loss of body mass allows reaching a more favourable wing loading and increases power for flight, hence enhancing flight performances and reducing predation risk. Moreover, body mass loss and power margin gain in both species were higher than in passerines, as observed in mallards. Our results suggest that the starvation-predation risk trade-off is one of the major life history traits underlying body mass adjustments, and these findings can be generalized to all birds facing predation. Additionally, the response magnitude seems to be influenced by the strategy of body reserve management. PMID:21789252

Clusterin (Apolipoprotein J), a Molecular Chaperone That Facilitates Degradation of the Copper-ATPases ATP7A and ATP7B*

PubMed Central

Materia, Stephanie; Cater, Michael A.; Klomp, Leo W. J.; Mercer, Julian F. B.; La Fontaine, Sharon

2011-01-01

The copper-transporting P1B-type ATPases (Cu-ATPases) ATP7A and ATP7B are key regulators of physiological copper levels. They function to maintain intracellular copper homeostasis by delivering copper to secretory compartments and by trafficking toward the cell periphery to export excess copper. Mutations in the genes encoding ATP7A and ATP7B lead to copper deficiency and toxicity disorders, Menkes and Wilson diseases, respectively. This report describes the interaction between the Cu-ATPases and clusterin and demonstrates a chaperone-like role for clusterin in facilitating their degradation. Clusterin interacted with both ATP7A and ATP7B in mammalian cells. This interaction increased under conditions of oxidative stress and with mutations in ATP7B that led to its misfolding and mislocalization. A Wilson disease patient mutation (G85V) led to enhanced ATP7B turnover, which was further exacerbated when cells overexpressed clusterin. We demonstrated that clusterin-facilitated degradation of mutant ATP7B is likely to involve the lysosomal pathway. The knockdown and overexpression of clusterin increased and decreased, respectively, the Cu-ATPase-mediated copper export capacity of cells. These results highlight a new role for intracellular clusterin in mediating Cu-ATPase quality control and hence in the normal maintenance of copper homeostasis, and in promoting cell survival in the context of disease. Based on our findings, it is possible that variations in clusterin expression and function could contribute to the variable clinical expression of Menkes and Wilson diseases. PMID:21242307

Ionotropic P2X ATP Receptor Channels Mediate Purinergic Signaling in Mouse Odontoblasts

PubMed Central

Shiozaki, Yuta; Sato, Masaki; Kimura, Maki; Sato, Toru; Tazaki, Masakazu; Shibukawa, Yoshiyuki

2017-01-01

ATP modulates various functions in the dental pulp cells, such as intercellular communication and neurotransmission between odontoblasts and neurons, proliferation of dental pulp cells, and odontoblast differentiation. However, functional expression patterns and their biophysical properties of ionotropic ATP (P2X) receptors (P2X1–P2X7) in odontoblasts were still unclear. We examined these properties of P2X receptors in mouse odontoblasts by patch-clamp recordings. K+-ATP, nonselective P2X receptor agonist, induced inward currents in odontoblasts in a concentration-dependent manner. K+-ATP-induced currents were inhibited by P2X4 and P2X7 selective inhibitors (5-BDBD and KN62, respectively), while P2X1 and P2X3 inhibitors had no effects. P2X7 selective agonist (BzATP) induced inward currents dose-dependently. We could not observe P2X1, 2/3, 3 selective agonist (αβ-MeATP) induced currents. Amplitudes of K+-ATP-induced current were increased in solution without extracellular Ca2+, but decreased in Na+-free extracellular solution. In the absence of both of extracellular Na+ and Ca2+, K+-ATP-induced currents were completely abolished. K+-ATP-induced Na+ currents were inhibited by P2X7 inhibitor, while the Ca2+ currents were sensitive to P2X4 inhibitor. These results indicated that odontoblasts functionally expressed P2X4 and P2X7 receptors, which might play an important role in detecting extracellular ATP following local dental pulp injury. PMID:28163685

Adenosine triphosphate regulates the activity of guinea pig Cav1.2 channel by direct binding to the channel in a dose-dependent manner.

PubMed

Feng, Rui; Xu, Jianjun; Minobe, Etsuko; Kameyama, Asako; Yang, Lei; Yu, Lifeng; Hao, Liying; Kameyama, Masaki

2014-05-01

The present study is to investigate the mechanism by which ATP regulates Cav1.2 channel activity. Ventricular tissue was obtained from adult guinea pig hearts using collagenase. Ca(2+) channel activity was monitored using the patch-clamp technique. Proteins were purified using wheat germ agglutinin-Sepharose, and the concentration was determined using the Coomassie brilliant blue technique. ATP binding to the Cav1.2 channel was examined using the photoaffinity method. EDA-ATP-biotin maintains Ca(2+) channel activity in inside-out membrane patches. ATP directly bound to the Cav1.2 channel in a dose-dependent manner, and at least two molecules of ATP bound to one molecule of the Cav1.2 channel. Low levels of calmodulin (CaM) increased ATP binding to the Cav1.2 channel, but higher levels of CaM decreased ATP binding to the Cav1.2 channel. In addition, Ca(2+) was another regulator for ATP binding to the Cav1.2 channel. Furthermore, ATP bound to GST-fusion peptides of NH2-terminal region (amino acids 6-140) and proximal COOH-terminal region (amino acids 1,509-1,789) of the main subunit (α1C) of the Cav1.2 channel. Our data suggest that ATP might regulate Cav1.2 channel activity by directly binding to the Cav1.2 channel in a dose-dependent manner. In addition, the ATP-binding effect to the Cav1.2 channel was both CaM- and Ca(2+) dependent.

Ionotropic P2X ATP Receptor Channels Mediate Purinergic Signaling in Mouse Odontoblasts.

PubMed

Shiozaki, Yuta; Sato, Masaki; Kimura, Maki; Sato, Toru; Tazaki, Masakazu; Shibukawa, Yoshiyuki

2017-01-01

ATP modulates various functions in the dental pulp cells, such as intercellular communication and neurotransmission between odontoblasts and neurons, proliferation of dental pulp cells, and odontoblast differentiation. However, functional expression patterns and their biophysical properties of ionotropic ATP (P2X) receptors (P2X 1 -P2X 7 ) in odontoblasts were still unclear. We examined these properties of P2X receptors in mouse odontoblasts by patch-clamp recordings. K + -ATP, nonselective P2X receptor agonist, induced inward currents in odontoblasts in a concentration-dependent manner. K + -ATP-induced currents were inhibited by P2X 4 and P2X 7 selective inhibitors (5-BDBD and KN62, respectively), while P2X 1 and P2X 3 inhibitors had no effects. P2X 7 selective agonist (BzATP) induced inward currents dose-dependently. We could not observe P2X 1, 2/3, 3 selective agonist (αβ-MeATP) induced currents. Amplitudes of K + -ATP-induced current were increased in solution without extracellular Ca 2+ , but decreased in Na + -free extracellular solution. In the absence of both of extracellular Na + and Ca 2+ , K + -ATP-induced currents were completely abolished. K + -ATP-induced Na + currents were inhibited by P2X 7 inhibitor, while the Ca 2+ currents were sensitive to P2X 4 inhibitor. These results indicated that odontoblasts functionally expressed P2X 4 and P2X 7 receptors, which might play an important role in detecting extracellular ATP following local dental pulp injury.

A novel mutation m.8561C>G in MT-ATP6/8 causing a mitochondrial syndrome with ataxia, peripheral neuropathy, diabetes mellitus, and hypergonadotropic hypogonadism.

PubMed

Kytövuori, Laura; Lipponen, Joonas; Rusanen, Harri; Komulainen, Tuomas; Martikainen, Mika H; Majamaa, Kari

2016-11-01

Defects in the respiratory chain or mitochondrial ATP synthase (complex V) result in mitochondrial dysfunction that is an important cause of inherited neurological disease. Two of the subunits of complex V are encoded by MT-ATP6 and MT-ATP8 in the mitochondrial genome. Pathogenic mutations in MT-ATP6 are associated with the Leigh syndrome, the syndrome of neuropathy, ataxia, and retinitis pigmentosa (NARP), as well as with non-classical phenotypes, while MT-ATP8 is less frequently mutated in patients with mitochondrial disease. We investigated two adult siblings presenting with features of cerebellar ataxia, peripheral neuropathy, diabetes mellitus, sensorineural hearing impairment, and hypergonadotropic hypogonadism. As the phenotype was suggestive of mitochondrial disease, mitochondrial DNA was sequenced and a novel heteroplasmic mutation m.8561C>G in the overlapping region of the MT-ATP6 and MT-ATP8 was found. The mutation changed amino acids in both subunits. Mutation heteroplasmy correlated with the disease phenotype in five family members. An additional assembly intermediate of complex V and increased amount of subcomplex F 1 were observed in myoblasts of the two patients, but the total amount of complex V was unaffected. Furthermore, intracellular ATP concentration was lower in patient myoblasts indicating defective energy production. We suggest that the m.8561C>G mutation in MT-ATP6/8 is pathogenic, leads biochemically to impaired assembly and decreased ATP production of complex V, and results clinically in a phenotype with the core features of cerebellar ataxia, peripheral neuropathy, diabetes mellitus, and hypergonadotropic hypogonadism.

Requirement of the isocitrate lyase gene ICL1 for VPS41-mediated starvation response in Cryptococcus neoformans.

PubMed

Xu, Zhe; Zhi, Yafei; Dong, Jianzhang; Lin, Benfeng; Ye, Di; Liu, Xiaoguang

2016-07-01

Cryptococcus neoformans is a major cause of fungal meningitis in individuals with impaired immunity. Our previous studies have shown that the VPS41 gene plays a critical role in the survival of Cryptococcus neoformans under nitrogen starvation; however, the molecular mechanisms underlying VPS41-mediated starvation response remain to be elucidated. In the present study, we show that, under nitrogen starvation, VPS41 strongly enhanced ICL1 expression in C. neoformans and that overexpression of ICL1 in the vps41 mutant dramatically suppressed its defects in starvation response due to the loss of VPS41 function. Moreover, targeted deletion of ICL1 resulted in a dramatic decline in viability of C. neoformans cells under nitrogen deprivation. Taken together, our data suggest a model in which VPS41 up-regulates ICL1 expression, directly or indirectly, to promote survival of C. neoformans under nitrogen starvation.

Cytoplasmic dynein undergoes intracellular redistribution concomitant with phosphorylation of the heavy chain in response to serum starvation and okadaic acid.

PubMed

Lin, S X; Ferro, K L; Collins, C A

1994-11-01

Cytoplasmic dynein is a microtubule-binding protein which is considered to serve as a motor for retrograde organelle movement. In cultured fibroblasts, cytoplasmic dynein localizes primarily to lysosomes, membranous organelles whose movement and distribution in the cytoplasm have been shown to be dependent on the integrity of the microtubule cytoskeleton. We have recently identified conditions which lead to an apparent dissociation of dynein from lysosomes in vivo, indicating that alterations in membrane binding may be involved in the regulation of retrograde organelle movement (Lin, S. X. H., and C. A. Collins. 1993. J. Cell Sci. 105:579-588). Both brief serum withdrawal and low extracellular calcium levels induced this alteration, and the effect was reversed upon addition of serum or additional calcium. Here we demonstrate that the phosphorylation state of the dynein molecule is correlated with changes in its intracellular distribution in normal rat kidney fibroblasts. Dynein heavy chain phosphorylation level increased during serum starvation, and decreased back to control levels upon subsequent addition of serum. We found that okadaic acid, a phosphoprotein phosphatase inhibitor, mimicked the effects of serum starvation on both phosphorylation and the intracellular redistribution of dynein from a membrane-associated pool to one that was more soluble, with similar dose dependence for both phenomena. Cell fractionation by differential detergent extraction revealed that a higher proportion of dynein was present in a soluble pool after serum starvation than was found in comparable fractions from control cells. Our data indicate that cytoplasmic dynein is phosphorylated in vivo, and changes in phosphorylation state may be involved in a regulatory mechanism affecting the distribution of this protein among intracellular compartments.

Cooling rate and starvation affect supercooling point and cold tolerance of the Khapra beetle, Trogoderma granarium Everts fourth instar larvae (Coleoptera: Dermestidae).

PubMed

Mohammadzadeh, M; Izadi, H

2018-01-01

Trogoderma granarium Everts (Coleoptera: Dermestidae) is an important insect pest of stored products. In this study, the survival strategies of T. granarium fourth instar larvae were investigated at different sub-zero temperatures following different cooling rates, acclimation to different relative humidity (RH) and different starvation times. Our results show that larvae of T. granarium are freeze-intolerant. There was a strong link between cooling rates and supercooling point, which means the slower the decrease in temperature, the lower the supercooling point. Trehalose content was greater in insects cooled at a rate of 0.5°C/min. According to results, the RH did not affect supercooling point. However, acclimation to an RH of 25% increased mortality following exposure to - 10°C/24h. The time necessary to reach 95% mortality was 1737h and 428h at - 5°C and - 10°C. The lowest lipid and trehalose content was detected in insects acclimated to 25% RH, although, the different RH treatments did not significantly affect glycogen content of T. granarium larvae. The supercooling point of larvae was gradually increased following starvation. By contrast, fed larvae had the greatest lipid, glycogen, and trehalose content, and insects starved for eight days had the lowest energy contents. There was a sharp decline in the survival of larvae between - 11 and - 18°C after 1h exposure. Our results indicate the effects of cooling rate and starvation on energy reserves and survival of T. granarium. We conclude that T. granarium may not survive under similar stress conditions of the stored products. Copyright © 2017 Elsevier Ltd. All rights reserved.

Identification of genes associated with resilience/vulnerability to sleep deprivation and starvation in Drosophila.

PubMed

Thimgan, Matthew S; Seugnet, Laurent; Turk, John; Shaw, Paul J

2015-05-01

Flies mutant for the canonical clock protein cycle (cyc(01)) exhibit a sleep rebound that is ∼10 times larger than wild-type flies and die after only 10 h of sleep deprivation. Surprisingly, when starved, cyc(01) mutants can remain awake for 28 h without demonstrating negative outcomes. Thus, we hypothesized that identifying transcripts that are differentially regulated between waking induced by sleep deprivation and waking induced by starvation would identify genes that underlie the deleterious effects of sleep deprivation and/or protect flies from the negative consequences of waking. We used partial complementary DNA microarrays to identify transcripts that are differentially expressed between cyc(01) mutants that had been sleep deprived or starved for 7 h. We then used genetics to determine whether disrupting genes involved in lipid metabolism would exhibit alterations in their response to sleep deprivation. Laboratory. Drosophila melanogaster. Sleep deprivation and starvation. We identified 84 genes with transcript levels that were differentially modulated by 7 h of sleep deprivation and starvation in cyc(01) mutants and were confirmed in independent samples using quantitative polymerase chain reaction. Several of these genes were predicted to be lipid metabolism genes, including bubblegum, cueball, and CG4500, which based on our data we have renamed heimdall (hll). Using lipidomics we confirmed that knockdown of hll using RNA interference significantly decreased lipid stores. Importantly, genetically modifying bubblegum, cueball, or hll resulted in sleep rebound alterations following sleep deprivation compared to genetic background controls. We have identified a set of genes that may confer resilience/vulnerability to sleep deprivation and demonstrate that genes involved in lipid metabolism modulate sleep homeostasis. © 2015 Associated Professional Sleep Societies, LLC.

Analysis of function-related interactions of ATP, sodium and potassium ions with Na+- and K+-transporting ATPase studied with a thiol reagent as tool.

PubMed

Grosse, R; Eckert, K; Malur, J; Repke, K R

1978-01-01

The paper describes the interaction of ATP, Na+ and K+ with (NaK)-ATPase exploiting the inactivation by reaction with NBD-chloride as an analytical tool for the evaluation of enzyme ligandation with the various effectors. 1. The inactivation of (NaK)-ATPase by reaction with NBD-chloride showing under all conditions studied a pseudo first-order rate rests on the alkylation of thiol groups in or near catalytic centre. ATP bound to catalytic centre prevents from enzyme inactivation by NDD-chloride through protection of these thiol groups from alkylation. Na+ and K+ affect the reactivity of the thiol groups towards NBD-chloride either indirectly via influencing ATP binding or more directly via changing the conformation of catalytic centre. Proceeding from these interrelations, the interaction of the various effectors with the enzyme was analyzed. 2. The K'D-values of various nucleotides determined by our approach correspond to the values obtained by independent methods. As shown for the first time, two catalytic centres per enzyme molecule exist. They exhibit high or low affinity to both ATP and ADP apparently caused by anticooperative interaction of the half-units of the enzyme through intersubunit communication ("half-of-the-sites reactivity"). 3. In the absence of ATP, Na+ or K+ ligandation of (NaK)-ATPase produce opposite effects on the reactivity of the thiol groups of catalytic centres reflecting different changes of their conformation. This corresponds to the well-known antagonistic effect of Na+ and K+ on some partial reactions of (NaK)-ATPase. The Na+ and K+ concentrations required to change thiol reactivity are rather high, i.e. the ionophoric centres for both Na+ and K+ are not readily accessible for cation complexation in the absence of enzyme complexation with ATP. 4. Na+ being without effect on ATP binding to the enzyme also does not influence the inactivating reaction with NBD-chloride while K+ by decreasing ATP binding dramatically decreases the protective effect of ATP. The K+ affinity of the enzyme-ATP complex is by more than two orders of magnitude higher than that of free enzyme. Na+ ligandation of the K+-liganded enzyme-ATP complex reverses the effect of K+ ligandation and produces a protective effect which distinctly surpasses that of the complexation of free enzyme with ATP. Hence, the enzyme molecule carries simultaneously ionophoric centres for both Na+ and K+. 5. The findings that per enzyme molecule ionophoric centres for Na+ and K+, and two catalytic centres with anticooperative interaction coexist corroborate the corresponding basic predictions of the flip-flop concept of (NaK)-ATPase pump mechanism, and explain some peculiar kinetic features of transport and enzyme activities of (NaK)-ATPase.

Effects of catecholamines on rat myocardial metabolism. II. Influence of catecholamines on 32p-incorporation into rat myocardial adenylic nucleotides and their turn-over.

PubMed

Merouze, P; Gaudemer, Y; Gautheron, D

1975-01-01

1. The influence of catecholamines (adrenaline and noradrenaline) on 32Pi incorporation into intracellular phosphate and adenylic nucleotides has been studied on rat myocardium slices; consequently, the turn-over of nucleotides could be determined and compared under the influence of these two hormones. 2. In order to specify the site of action of these catecholamines, several inhibitors and activators of energetic metabolism were included in the incubation medium: 3'5'-AMP, caffein, ouabain, oligomycin, rotenone + antimycin. 3. Both catecholamines favour Pi exchanges between intra and extracellular spaces; ATP turn-over is greatly increased, while ADP turn-over is slightly decreased, and 32P-incorporation into ADP is increased. 4. 3'5'-AMP and caffein are without effect on Pi penetration; however, caffein increases catecholamine effects on this penetration. ATP turn-over is slightly increased by 3'5'-AMP or caffein. 5. Ouabain decreases ATP turn-over but does not prevent the adrenaline induced acceleration. Inhibitors of oxidative phosphorylation and electron transport decrease ATP-turn-over severely; this inhibition is not released by catecholamines. 6. It is concluded that the catecholamine effects observed are dependent on the oxidative phosphorylations process. The increase of Pi exchange by catecholamines may be related to the increase of extracellular space and cation translocations we observed with the hormones.

Phosphite, an analog of phosphate, suppresses the coordinated expression of genes under phosphate starvation.

PubMed

Varadarajan, Deepa K; Karthikeyan, Athikkattuvalasu S; Matilda, Paino Durzo; Raghothama, Kashchandra G

2002-07-01

Phosphate (Pi) and its analog phosphite (Phi) are acquired by plants via Pi transporters. Although the uptake and mobility of Phi and Pi are similar, there is no evidence suggesting that plants can utilize Phi as a sole source of phosphorus. Phi is also known to interfere with many of the Pi starvation responses in plants and yeast (Saccharomyces cerevisiae). In this study, effects of Phi on plant growth and coordinated expression of genes induced by Pi starvation were analyzed. Phi suppressed many of the Pi starvation responses that are commonly observed in plants. Enhanced root growth and root to shoot ratio, a hallmark of Pi stress response, was strongly inhibited by Phi. The negative effects of Phi were not obvious in plants supplemented with Pi. The expression of Pi starvation-induced genes such as LePT1, LePT2, AtPT1, and AtPT2 (high-affinity Pi transporters); LePS2 (a novel acid phosphatase); LePS3 and TPSI1 (novel genes); and PAP1 (purple acid phosphatase) was suppressed by Phi in plants and cell cultures. Expression of luciferase reporter gene driven by the Pi starvation-induced AtPT2 promoter was also suppressed by Phi. These analyses showed that suppression of Pi starvation-induced genes is an early response to addition of Phi. These data also provide evidence that Phi interferes with gene expression at the level of transcription. Synchronized suppression of multiple Pi starvation-induced genes by Phi points to its action on the early molecular events, probably signal transduction, in Pi starvation response.

Vascular ATP-sensitive potassium channels are over-expressed and partially regulated by nitric oxide in experimental septic shock.

PubMed

Collin, Solène; Sennoun, Nacira; Dron, Anne-Gaëlle; de la Bourdonnaye, Mathilde; Montemont, Chantal; Asfar, Pierre; Lacolley, Patrick; Meziani, Ferhat; Levy, Bruno

2011-05-01

To study the activation and expression of vascular (aorta and small mesenteric arteries) potassium channels during septic shock with or without modulation of the NO pathway. Septic shock was induced in rats by peritonitis. Selective inhibitors of vascular K(ATP) (PNU-37883A) or BK(Ca) [iberiotoxin (IbTX)] channels were used to demonstrate their involvement in vascular hyporeactivity. Vascular response to phenylephrine was measured on aorta and small mesenteric arteries mounted on a wire myograph. Vascular expression of potassium channels was studied by PCR and Western blot, in the presence or absence of 1400W, an inducible NO synthase (iNOS) inhibitor. Aortic activation of the transcriptional factor nuclear factor-kappaB (NF-κB) was assessed by electrophoretic mobility shift assay. Arterial pressure as well as in vivo and ex vivo vascular reactivity were reduced by sepsis and improved by PNU-37883A but not by IbTX. Sepsis was associated with an up-regulation of mRNA and protein expression of vascular K(ATP) channels, while expression of vascular BK(Ca) channels remained unchanged. Selective iNOS inhibition blunted the sepsis-induced increase in aortic NO, decreased NF-κB activation, and down-regulated vascular K(ATP) channel expression. Vascular K(ATP) but not BK(Ca) channels are activated, over-expressed, and partially regulated by NO via NF-κB activation during septic shock. Their selective inhibition restores arterial pressure and vascular reactivity and decreases lactate concentration. The present data suggest that selective vascular K(ATP) channel inhibitors offer potential therapeutic perspectives for septic shock.

Ca++-sensitizing mutations in troponin, Pi, and 2-deoxyATP alter the depressive effect of acidosis on regulated thin-filament velocity

PubMed Central

Longyear, Thomas J.; Turner, Matthew A.; Davis, Jonathan P.; Lopez, Joseph; Biesiadecki, Brandon

2014-01-01

Repeated, intense contractile activity compromises the ability of skeletal muscle to generate force and velocity, resulting in fatigue. The decrease in velocity is thought to be due, in part, to the intracellular build-up of acidosis inhibiting the function of the contractile proteins myosin and troponin; however, the underlying molecular basis of this process remains poorly understood. We sought to gain novel insight into the decrease in velocity by determining whether the depressive effect of acidosis could be altered by 1) introducing Ca++-sensitizing mutations into troponin (Tn) or 2) by agents that directly affect myosin function, including inorganic phosphate (Pi) and 2-deoxy-ATP (dATP) in an in vitro motility assay. Acidosis reduced regulated thin-filament velocity (VRTF) at both maximal and submaximal Ca++ levels in a pH-dependent manner. A truncated construct of the inhibitory subunit of Tn (TnI) and a Ca++-sensitizing mutation in the Ca++-binding subunit of Tn (TnC) increased VRTF at submaximal Ca++ under acidic conditions but had no effect on VRTF at maximal Ca++ levels. In contrast, both Pi and replacement of ATP with dATP reversed much of the acidosis-induced depression of VRTF at saturating Ca++. Interestingly, despite producing similar magnitude increases in VRTF, the combined effects of Pi and dATP were additive, suggesting different underlying mechanisms of action. These findings suggest that acidosis depresses velocity by slowing the detachment rate from actin but also by possibly slowing the attachment rate. PMID:24651988

The Effects of Simulated Micro-gravity on Cultured Chicken Embryonic Chondrocytes

NASA Astrophysics Data System (ADS)

Li, X.; Zhang, X.; Yang, S.; Li, S.; Peidong, J.; Lin, Z.

T he effects of simulated microgravity on the microtubular system, extracellular matrix, alkaline phosphatase activity, intracellular free calcium concentration, mitochondrial ATP synthase activity and oligomycin inhibition rate of cultured chicken embryonic chondrocytes were studied with a clinostat. The microtubular content decreased. The extracellualr matrix decreased significantly after rotating, and the fibers in the extracellular matrix were more tiny and disorderly. There was a time course decrease in alkaline phosphatase activity of chondrocytes, a marker of matrix mineralization. Meanwhile a significant drop in the intracellular calcium concentration happened at the beginning of rotation. No significant changes happened in the mitochondrial ATP synthase activity and oligomycin inhibition rate. The possible mechanisms about them were discussed.

The function of mitochondrial F(O)F(1) ATP-synthase from the whiteleg shrimp Litopenaeus vannamei muscle during hypoxia.

PubMed

Martinez-Cruz, O; Calderon de la Barca, A M; Uribe-Carvajal, S; Muhlia-Almazan, A

2012-08-01

The effect of hypoxia and re-oxygenation on the mitochondrial complex F(O)F(1)-ATP synthase was investigated in the whiteleg shrimp Litopenaeus vannamei. A 660 kDa protein complex isolated from mitochondria of the shrimp muscle was identified as the ATP synthase complex. After 10h at hypoxia (1.5-2.0 mg oxygen/L), the concentration of L-lactate in plasma increased significantly, but the ATP amount and the concentration of ATPβ protein remained unaffected. Nevertheless, an increase of 70% in the ATPase activity was detected, suggesting that the enzyme may be regulated at a post-translational level. Thus, during hypoxia shrimp are able to maintain ATP amounts probably by using some other energy sources as phosphoarginine when an acute lack of energy occurs. During re-oxygenation, the ATPase activity decreased significantly and the ATP production continued via the electron transport chain and oxidative phosphorylation. The results obtained showed that shrimp faces hypoxia partially by hydrolyzing the ATP through the reaction catalyzed by the mitochondrial ATPase which increases its activity. Copyright © 2012 Elsevier Inc. All rights reserved.

Piezo1 regulates mechanotransductive release of ATP from human RBCs.

PubMed

Cinar, Eyup; Zhou, Sitong; DeCourcey, James; Wang, Yixuan; Waugh, Richard E; Wan, Jiandi

2015-09-22

Piezo proteins (Piezo1 and Piezo2) are recently identified mechanically activated cation channels in eukaryotic cells and associated with physiological responses to touch, pressure, and stretch. In particular, human RBCs express Piezo1 on their membranes, and mutations of Piezo1 have been linked to hereditary xerocytosis. To date, however, physiological functions of Piezo1 on normal RBCs remain poorly understood. Here, we show that Piezo1 regulates mechanotransductive release of ATP from human RBCs by controlling the shear-induced calcium (Ca(2+)) influx. We find that, in human RBCs treated with Piezo1 inhibitors or having mutant Piezo1 channels, the amounts of shear-induced ATP release and Ca(2+) influx decrease significantly. Remarkably, a critical extracellular Ca(2+) concentration is required to trigger significant ATP release, but membrane-associated ATP pools in RBCs also contribute to the release of ATP. Our results show how Piezo1 channels are likely to function in normal RBCs and suggest a previously unidentified mechanotransductive pathway in ATP release. Thus, we anticipate that the study will impact broadly on the research of red cells, cellular mechanosensing, and clinical studies related to red cell disorders and vascular disease.

Role of the Na(+)/K(+)-ATPase beta-subunit in peptide-mediated transdermal drug delivery.

PubMed

Wang, Changli; Ruan, Renquan; Zhang, Li; Zhang, Yunjiao; Zhou, Wei; Lin, Jun; Ding, Weiping; Wen, Longping

2015-04-06

In this work, we discovered that the Na(+)/K(+)-ATPase beta-subunit (ATP1B1) on epidermal cells plays a key role in the peptide-mediated transdermal delivery of macromolecular drugs. First, using a yeast two-hybrid assay, we screened candidate proteins that have specific affinity for the short peptide TD1 (ACSSSPSKHCG) identified in our previous work. Then, we verified the specific binding of TD1 to ATP1B1 in yeast and mammalian cells by a pull-down ELISA and an immunoprecipitation assay. Finally, we confirmed that TD1 mainly interacted with the C-terminus of ATP1B1. Our results showed that the interaction between TD1 and ATP1B1 affected not only the expression and localization of ATP1B1, but also the epidermal structure. In addition, this interaction could be antagonized by the exogenous competitor ATP1B1 or be inhibited by ouabain, which results in the decreased delivery of macromolecular drugs across the skin. The discovery of a critical role of ATP1B1 in the peptide-mediated transdermal drug delivery is of great significance for the future development of new transdermal peptide enhancers.

Free fatty acids as inducers and regulators of uncoupling of oxidative phosphorylation in liver mitochondria with participation of ADP/ATP- and aspartate/glutamate-antiporter.

PubMed

Samartsev, V N; Marchik, E I; Shamagulova, L V

2011-02-01

In liver mitochondria fatty acids act as protonophoric uncouplers mainly with participation of internal membrane protein carriers - ADP/ATP and aspartate/glutamate antiporters. In this study the values of recoupling effects of carboxyatractylate and glutamate (or aspartate) were used to assess the degree of participation of ADP/ATP and aspartate/glutamate antiporters in uncoupling activity of fatty acids. These values were determined from the ability of these recoupling agents to suppress the respiration stimulated by fatty acids and to raise the membrane potential reduced by fatty acids. Increase in palmitic and lauric acid concentration was shown to increase the degree of participation of ADP/ATP antiporter and to decrease the degree of participation of aspartate/glutamate antiporter in uncoupling to the same extent. These data suggest that fatty acids are not only inducers of uncoupling of oxidative phosphorylation, but that they also act the regulators of this process. The linear dependence of carboxyatractylate and glutamate recoupling effects ratio on palmitic and lauric acids concentration was established. Comparison of the effects of fatty acids (palmitic, myristic, lauric, capric, and caprylic having 16, 14, 12, 10, and 8 carbon atoms, respectively) has shown that, as the hydrophobicity of fatty acids decreases, the effectiveness decreases to a greater degree than the respective values of their specific uncoupling activity. The action of fatty acids as regulators of uncoupling is supposed to consist of activation of transport of their anions from the internal to the external monolayer of the internal membrane with participation of ADP/ATP antiporter and, at the same time, in inhibition of this process with the participation of aspartate/glutamate antiporter.

Effect of LLLT on the level of ATP and ROS from organ of corti cells

NASA Astrophysics Data System (ADS)

Rhee, ChungKu; Chang, So-Young; Ahn, Jin-Chul; Suh, Myung-Whan; Jung, Jae Yun

2014-03-01

It is well established that ototoxic antibiotics and acoustic trauma can damage cochlear hair cells and cause hearing loss. Previous studies using transcanal LLLT (Low level laser therapy) showed that LLLT can promote recovery of hearing thresholds and cochlear hair cells. However, its mechanism has not been studied. Aim: The aim of this study is to investigate the mechanism of hearing recovery from gentamicin induced ototoxic hearing loss by LLLT. Methods: HEI- OC1 (House ear institute organ of Corti) cells were cultured for 18 hours and ototoxicity was induced by gentamicin (GM) treatment to the cells. Cultured cells were divided into 6 groups, No treatment control, LLLT only, GM 6.6 mM and GM 13.1 mM, GM 6.6 mM+LLLT and GM 13.1 mM+LLLT cells. LD laser 808 nm, 15 mW, was irradiated to the cultured cells for 15 min, at 4 hours after GM treatment to the cells. ATP was assayed using the ATP assay Kit. ROS was measured using confocal microscope after application of H2DCFDA dye. Results: ATP was decreased in GM 13.1 mM cells and increased in LLLT only cells and GM 13.1 mM+LLLT cells compared to control and 13.1 mM cells. ROS was increased in GM 6.6 mM and GM 13.1 mM cells, and decreased in GM 6.6 mM+LLLT and GM 13.1 mM+LLLT cells compared to GM 6.6 and 13.1 mM cells immediately after laser irradiation. Conclusion: This study demonstrated that LLLT on GM treated HEI-OC1 cells increased ATP and decreased ROS that may contribute to the recovery of hearing.

Liposomes as fatty acids carriers in isolated rat liver: effect on energy metabolism and on isolated mitochondria activity.

PubMed

Delmas-Beauvieux, M C; Leducq, N; Thiaudière, E; Diolez, P; Gin, H; Canioni, P; Gallis, J L

2000-02-01

The effects of fatty acids (FA)-carrier, egg-lecithin liposomes (LIPO) as alternative to BSA, on ATP, glycogen and glucose contents in isolated perfused liver of fed rats were non-invasively studied using 31P/13C nuclear magnetic resonance (NMR). Oxidative phosphorylation was studied in isolated mitochondria from the same liver consecutively to the NMR experiments. ATP content decreased slowly and ATP turnover was similar during the perfusion with saline solution (KHB) or LIPO. However, LIPO induced an enhancement of respiratory control ratio in isolated mitochondria. Tissue glycogen and glucose content decreased when FA (linoleate or linolenate) were perfused with defatted BSA (3%) or LIPO (600 mg/l) whereas glucose excretion level was unchanged and lactate excretion tended to increase, reflecting changes in the cytosolic redox state and/or an enhancement of glycolysis. Addition of FA (0.5 or 1.5 mM) to LIPO caused a dramatic fall in liver ATP, a mitochondrial uncoupling and an impairment of the phosphorylation activity. Perfusion with FA (1.5 mM) carried by BSA significantly increased the ATP degradation without change of mitochondrial function. Owing to the higher affinity of BSA than LIPO for FA, these latter could be more easily released from complex LIPO-FA, increasing their uncoupling effect. Hence, the FA concentrations have to be largely decreased from the above currently used concentrations to avoid this effect. It will then be possible to minimize the effector action of FA and to study their more specific metabolic function as fuel. It was concluded that LIPO were appropriate carriers to study the different metabolic effects of FA.

Activation of cholinergic receptors blocks non-adrenergic non-cholinergic contractions in the rat urinary bladder

PubMed Central

Henry Lai, H.; Smith, Christopher P.; Munoz, Alvaro; Boone, Timothy B.; Szigeti, Gyula P.; Somogyi, George T.

2008-01-01

In the present study, the plasticity of the non-adrenergic non-cholinergic (NANC) response was investigated. Isolated rat bladder strips were electrically stimulated and the evoked contractions were isometrically recorded. The NANC part of the contractions were unmasked by applying 500 nM 4-DAMP, a potent muscarinic antagonist. Treatment of the bladder strips with 10 μM carbachol (a cholinergic agonist) increased the muscle tone but did not alter the neurally evoked contractions. However, carbachol decreased: (1) the NANC response from 74.6% to 33.3% of control and (2) the purinergic contractile response to α,β methylene ATP (α,β mATP) (10 μM) from 97.0% to 43.4% (p<0.05). Treatment with the cholinesterase inhibitor eserine (10 μM) also significantly decreased the NANC response to 21.1% (p<0.0001). The purinergic receptor antagonist suramin (100μM) did not affect the neurally evoked contractions, however; subsequent addition of 4-DAMP decreased the contractions to 31%. Activation of the smooth muscle cholinergic receptors (with carbachol or eserine) and purinergic receptors (with α,β mATP) decreased the NANC contractions and the direct contractile response to α,β mATP. When the electrically evoked contractions were facilitated by the L-type Ca2+ channel activator, Bay-K 8644 the subsequent application of 4-DAMP did not unmask inhibited NANC contractions. We conclude that activation of muscarinic receptors by cholinergic agonist, carbachol or by endogenous acetylcholine (ACh) induce a cascade of events that leads to diminished purinergic response and consequently an inhibition of the bladder NANC response. PMID:18755252

The toxic mechanism of high lethality of herbicide butachlor in marine flatfish flounder, Paralichthys olivaceus

NASA Astrophysics Data System (ADS)

Guo, Huarong; Yin, Licheng; Zhang, Shicui; Feng, Wenrong

2010-09-01

The toxic mechanism of herbicide butachlor to induce extremely high lethality in marine flatfish flounder, Paralichthys Olivaceus, was analyzed by histopathological examination, antioxidant enzymes activities and ATP content assay. Histopathological examination of gill, liver and kidney of exposed fishes showed that gill was a target organ of butachlor. The butachlor seriously impaired the respiration of gills by a series of lesions such as edema, lifting and detachment of lamellar epithelium, breakdown of pillar cells, and blood congestion. The dysfunction of gill respiration caused suffocation to the exposed flounder with extremely high acute lethality. Antioxidant enzyme activity assay of the in vitro cultured flounder gill (FG) cells exposed to butachlor indicated that butachlor markedly inhibited the antioxidant enzyme activities of Superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX). Furthermore, along with the decline of antioxidant enzyme activities, ATP content in the exposed FG cells decreased, too. This infers that the oxidative stress induced by butachlor can inhibit the production of cellular ATP. Similar decrease of ATP content was also observed in the exposed flounder gill tissues. Taken together, as in FG cells, butachlor possibly induced a short supply of ATP in pillar cells by inhibiting the antioxidant enzyme activities and then affecting the contractibility of the pillar cells, which in turn resulted in the blood congestion and suffocation of exposed flounder.

Chloroplast ATP Synthase Modulation of the Thylakoid Proton Motive Force: Implications for Photosystem I and Photosystem II Photoprotection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kanazawa, Atsuko; Ostendorf, Elisabeth; Kohzuma, Kaori

In wild type plants, decreasing CO 2 lowers the activity of the chloroplast ATP synthase, slowing proton efflux from the thylakoid lumen resulting in buildup of thylakoid proton motive force (pmf). The resulting acidification of the lumen regulates both light harvesting, via the qE mechanism, and photosynthetic electron transfer through the cytochrome b 6f complex. Here in this paper, we show that the cfq mutant of Arabidopsis, harboring single point mutation in its γ-subunit of the chloroplast ATP synthase, increases the specific activity of the ATP synthase and disables its down-regulation under low CO 2. The increased thylakoid proton conductivitymore » (g H +) in cfq results in decreased pmf and lumen acidification, preventing full activation of qE and more rapid electron transfer through the b6f complex, particularly under low CO 2 and fluctuating light. These conditions favor the accumulation of electrons on the acceptor side of PSI, and result in severe loss of PSI activity. Comparing the current results with previous work on the pgr5 mutant suggests a general mechanism where increased PSI photodamage in both mutants is caused by loss of pmf, rather than inhibition of CEF per se. Overall, our results support a critical role for ATP synthase regulation in maintaining photosynthetic control of electron transfer to prevent photodamage.« less

Chloroplast ATP Synthase Modulation of the Thylakoid Proton Motive Force: Implications for Photosystem I and Photosystem II Photoprotection

DOE PAGES

Kanazawa, Atsuko; Ostendorf, Elisabeth; Kohzuma, Kaori; ...

2017-05-03

In wild type plants, decreasing CO 2 lowers the activity of the chloroplast ATP synthase, slowing proton efflux from the thylakoid lumen resulting in buildup of thylakoid proton motive force (pmf). The resulting acidification of the lumen regulates both light harvesting, via the qE mechanism, and photosynthetic electron transfer through the cytochrome b 6f complex. Here in this paper, we show that the cfq mutant of Arabidopsis, harboring single point mutation in its γ-subunit of the chloroplast ATP synthase, increases the specific activity of the ATP synthase and disables its down-regulation under low CO 2. The increased thylakoid proton conductivitymore » (g H +) in cfq results in decreased pmf and lumen acidification, preventing full activation of qE and more rapid electron transfer through the b6f complex, particularly under low CO 2 and fluctuating light. These conditions favor the accumulation of electrons on the acceptor side of PSI, and result in severe loss of PSI activity. Comparing the current results with previous work on the pgr5 mutant suggests a general mechanism where increased PSI photodamage in both mutants is caused by loss of pmf, rather than inhibition of CEF per se. Overall, our results support a critical role for ATP synthase regulation in maintaining photosynthetic control of electron transfer to prevent photodamage.« less

3-Monochloro-1,2-propanediol (3-MCPD) induces apoptosis via mitochondrial oxidative phosphorylation system impairment and the caspase cascade pathway.

PubMed

Peng, Xiaoli; Gan, Jing; Wang, Qian; Shi, Zhenqiang; Xia, Xiaodong

2016-11-30

3-Monochloro-1,2-propanediol (3-MCPD) is the most toxic chloropropanols compounds in foodstuff which mainly generated during thermal processing. Kidney is one of the primary target organs for 3-MCPD. Using human embryonic kidney cell (HEK293FT) as an in vitro model, we found that 3-MCPD caused concentration-dependent increase in cytoxicity as assessed by dye uptake, lactatedehydrogenase (LDH) leakage and MTT assays. HEK293FT cell treated with 3-MCPD suffered the decrease of mitochondrial membrane potential and the impairment of mitochondrial oxidative phosphorylation system, especially the reduced amount of mRNA expression and protein synthesis of electron transport chain complex II, complex IV, and complex III. More importantly, energy release (ATP synthesis) was significantly inhibited by 3-MCPD resulting from the down regulation expressions of ATP synthase (ATP6 and ATP8), as well as the loss of transmembrane potential required for synthesis of ATP. The decreased ratio of mitochondrial apoptogenic factors Bax/Bcl-2 and the cytochrome-c release from mitochondria to cytosol followed by the activation of apoptotic initiators caspase 9 and apoptotic executioners (caspase 3, caspase 6 and caspase 7) leading to apoptosis. The activation of caspase 8 and caspase 2 implied that there were probably other factors to induce the caspase-dependent apoptosis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Effect of amino acid starvation on UV sensitivity of Lactobacillus acidophilus cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Soška, J.; Nečasová, J.

1973-11-01

In Lactobacillus acidophilus cultures uv irradiated in the exponential phase of growth, the dose-survival curve was of the simple exponential type, without any shoulder. If the bacteria were subjected to amino acid starvation prior to irradiation, a shoulder corresponding to a quasi-threshold dose (D) of about 780 ergs/mm/sup 2/ appeared in the curve. The administration of protein- or RNA-synthesis inhibitors prior to irradiation had the same effect. The effect of pre-irradiation amino acid starvation was abolished by simuitaneous thymidine starvation. It was likewise abolished if amino acid starvation was followed by incubation in the presence of amino acids (without thymidine)more » and then by irradiation of the cells. Post-irradiation amino acid starvation did not lead to the formation of a shoulder but if combined with thymidine starvation it did. It can be concluded from the results that post-irradiation repair processes are facilitated or promoted if, during the post-irradiation interval DNA synthesis is delayed. This delay represents a compensation of the pre-irradiation increase of cellular DNA-content, taking place during inhibition of proteosynthesis. The post-irradiation administration of caffeine did not abolish the formation of the shoulder induced by pre-irradiation amino acid starvation; on the contrary, it induced its formation even in exponentially growing, irradiated control bacteria. (auth)« less

Increase of chromium tolerance in Scenedesmus acutus after sulfur starvation: Chromium uptake and compartmentalization in two strains with different sensitivities to Cr(VI).

PubMed

Marieschi, M; Gorbi, G; Zanni, C; Sardella, A; Torelli, A

2015-10-01

In photosynthetic organisms sulfate constitutes the main sulfur source for the biosynthesis of GSH and its precursor Cys. Hence, sulfur availability can modulate the capacity to cope with environmental stresses, a phenomenon known as SIR/SED (Sulfur Induced Resistance or Sulfur Enhanced Defence). Since chromate may compete for sulfate transport into the cells, in this study chromium accumulation and tolerance were investigated in relation to sulfur availability in two strains of the unicellular green alga Scenedesmus acutus with different Cr-sensitivities. Paradoxically, sulfur deprivation has been demonstrated to induce a transient increase of Cr-tolerance in both strains. Sulfur deprivation is known to enhance the sulfate uptake/assimilation pathway leading to important consequences on Cr-tolerance: (i) reduced chromate uptake due to the induction of high affinity sulfate transporters (ii) higher production of cysteine and GSH which can play a role both through the formation of unsoluble complexes and their sequestration in inert compartments. To investigate the role of the above mentioned mechanisms, Cr accumulation in total cells and in different cell compartments (cell wall, membranes, soluble and miscellaneous fractions) was analyzed in both sulfur-starved and unstarved cells. Both strains mainly accumulated chromium in the soluble fraction, but the uptake was higher in the wild-type. In this type a short period of sulfur starvation before Cr(VI) treatment lowered chromium accumulation to the level observed in the unstarved Cr-tolerant strain, in which Cr uptake seems instead less influenced by S-starvation, since no significant decrease was observed. The increase in Cr-tolerance following S-starvation seems thus to rely on different mechanisms in the two strains, suggesting the induction of a mechanism constitutively active in the Cr-tolerant strain, maybe a high affinity sulfate transporter also in the wild-type. Changes observed in the cell wall and membrane fractions suggest a strong involvement of these compartments in Cr-tolerance increase following S-starvation. Copyright © 2015 Elsevier B.V. All rights reserved.

Heat Stress Affects Pi-related Genes Expression and Inorganic Phosphate Deposition/Accumulation in Barley

PubMed Central

Pacak, Andrzej; Barciszewska-Pacak, Maria; Swida-Barteczka, Aleksandra; Kruszka, Katarzyna; Sega, Pawel; Milanowska, Kaja; Jakobsen, Iver; Jarmolowski, Artur; Szweykowska-Kulinska, Zofia

2016-01-01

Phosphorus (P) in plants is taken from soil as an inorganic phosphate (Pi) and is one of the most important macroelements in growth and development. Plants actively react to Pi starvation by the induced expression of Pi transporters, MIR399, MIR827, and miR399 molecular sponge – IPS1 genes and by the decreased expression of the ubiquitin-conjugating enzyme E2 (PHOSPHATE2 – PHO2) and Pi sensing and transport SPX-MFS genes. The PHO2 protein is involved in the degradation of Pi transporters PHT1;1 (from soil to roots) and PHO1 (from roots to shoots). The decreased expression of PHO2 leads to Pi accumulation in shoots. In contrast, the pho1 mutant shows a decreased level of Pi concentration in shoots. Finally, Pi starvation leads to decreased Pi concentration in all plant tissues. Little is known about plant Pi homeostasis in other abiotic stress conditions. We found that, during the first hour of heat stress, Pi accumulated in barley shoots but not in the roots, and transcriptomic data analysis as well as RT-qPCR led us to propose an explanation for this phenomenon. Pi transport inhibition from soil to roots is balanced by lower Pi efflux from roots to shoots directed by the PHO1 transporter. In shoots, the PHO2 mRNA level is decreased, leading to an increased Pi level. We concluded that Pi homeostasis in barley during heat stress is maintained by dynamic changes in Pi-related genes expression. PMID:27446155

Mechanism of nucleotide sensing in group II chaperonins.

PubMed

Pereira, Jose H; Ralston, Corie Y; Douglas, Nicholai R; Kumar, Ramya; Lopez, Tom; McAndrew, Ryan P; Knee, Kelly M; King, Jonathan A; Frydman, Judith; Adams, Paul D

2012-02-01

Group II chaperonins mediate protein folding in an ATP-dependent manner in eukaryotes and archaea. The binding of ATP and subsequent hydrolysis promotes the closure of the multi-subunit rings where protein folding occurs. The mechanism by which local changes in the nucleotide-binding site are communicated between individual subunits is unknown. The crystal structure of the archaeal chaperonin from Methanococcus maripaludis in several nucleotides bound states reveals the local conformational changes associated with ATP hydrolysis. Residue Lys-161, which is extremely conserved among group II chaperonins, forms interactions with the γ-phosphate of ATP but shows a different orientation in the presence of ADP. The loss of the ATP γ-phosphate interaction with Lys-161 in the ADP state promotes a significant rearrangement of a loop consisting of residues 160-169. We propose that Lys-161 functions as an ATP sensor and that 160-169 constitutes a nucleotide-sensing loop (NSL) that monitors the presence of the γ-phosphate. Functional analysis using NSL mutants shows a significant decrease in ATPase activity, suggesting that the NSL is involved in timing of the protein folding cycle.

A single mutation at the catalytic site of TF1-alpha3beta3gamma complex switches the kinetics of ATP hydrolysis from negative to positive cooperativity.

PubMed

Muneyuki, E; Odaka, M; Yoshida, M

1997-08-11

Previously, we reported the substitution of Tyr341 of the F1-ATPase beta subunit from a thermophilic Bacillus strain PS3 with leucine, cysteine, or alanine (M. Odaka et al. J. Biochem., 115 (1994) 789-796). These mutations resulted in a great decrease in the affinity of the isolated beta subunit for ATP-Mg and an increase in the apparent Km of the alpha3beta3gamma complex in ATP hydrolysis when examined above 0.1 mM ATP. Here, we examined the ATPase activity of the mutant complexes in a wide range of ATP concentration and found that the mutants exhibited apparent positive cooperativity in ATP hydrolysis. This is the first clear demonstration that a single mutation in the catalytic sites converts the kinetics from apparent negative cooperativity in the wild-type alpha3beta3gamma complex to apparent positive cooperativity. The conversion of apparent cooperativity could be explained in terms of a simple kinetic scheme based on the binding change model proposed by Boyer.

Endolysosomal two‐pore channels regulate autophagy in cardiomyocytes

PubMed Central

García‐Rúa, Vanessa; Feijóo‐Bandín, Sandra; Rodríguez‐Penas, Diego; Mosquera‐Leal, Ana; Abu‐Assi, Emad; Beiras, Andrés; María Seoane, Luisa; Lear, Pamela; Parrington, John; Portolés, Manuel; Roselló‐Lletí, Esther; Rivera, Miguel; Gualillo, Oreste; Parra, Valentina; Hill, Joseph A.; Rothermel, Beverly; González‐Juanatey, José Ramón

2016-01-01

Key points Two‐pore channels (TPCs) were identified as a novel family of endolysosome‐targeted calcium release channels gated by nicotinic acid adenine dinucleotide phosphate, as also as intracellular Na+ channels able to control endolysosomal fusion, a key process in autophagic flux.Autophagy, an evolutionarily ancient response to cellular stress, has been implicated in the pathogenesis of a wide range of cardiovascular pathologies, including heart failure.We report direct evidence indicating that TPCs are involved in regulating autophagy in cardiomyocytes, and that TPC knockout mice show alterations in the cardiac lysosomal system. TPC downregulation implies a decrease in the viability of cardiomyocytes under starvation conditions. In cardiac tissues from both humans and rats, TPC transcripts and protein levels were higher in females than in males, and correlated negatively with markers of autophagy.We conclude that the endolysosomal channels TPC1 and TPC2 are essential for appropriate basal and induced autophagic flux in cardiomyocytes, and also that they are differentially expressed in male and female hearts. Abstract Autophagy participates in physiological and pathological remodelling of the heart. The endolysosomal two‐pore channels (TPCs), TPC1 and TPC2, have been implicated in the regulation of autophagy. The present study aimed to investigate the role of TPC1 and TPC2 in basal and induced cardiac autophagic activity. In cultured cardiomyocytes, starvation induced a significant increase in TPC1 and TPC2 transcripts and protein levels that paralleled the increase in autophagy identified by increased LC3‐II and decreased p62 levels. Small interfering RNA depletion of TPC2 alone or together with TPC1 increased both LC3II and p62 levels under basal conditions and in response to serum starvation, suggesting that, under conditions of severe energy depletion (serum plus glucose starvation), changes in the autophagic flux (as assessed by use of bafilomycin A1) occurred either when TPC1 or TPC2 were downregulated. The knockdown of TPCs diminished cardiomyocyte viability under starvation and simulated ischaemia. Electron micrographs of hearts from TPC1/2 double knockout mice showed that cardiomyocytes contained large numbers of immature lysosomes with diameters significantly smaller than those of wild‐type mice. In cardiac tissues from humans and rats, TPC1 and TPC2 transcripts and protein levels were higher in females than in males. Furthermore, transcript levels of TPCs correlated negatively with p62 levels in heart tissues. TPC1 and TPC2 are essential for appropriate basal and induced autophagic flux in cardiomyocytes (i.e. there is a negative effect on cell viability under stress conditions in their absence) and they are differentially expressed in male and female human and murine hearts, where they correlate with markers of autophagy. PMID:26757341

Phosphate or phosphite addition promotes the proteolytic turnover of phosphate-starvation inducible tomato purple acid phosphatase isozymes.

PubMed

Bozzo, Gale G; Singh, Vinay K; Plaxton, William C

2004-08-27

Within 48 h of the addition of 2.5 mM phosphate (HPO42-, Pi) or phosphite (H2PO3-, Phi) to 8-day-old Pi-starved (-Pi) tomato suspension cells: (i) secreted and intracellular purple acid phosphatase (PAP) activities decreased by about 12- and 6-fold, respectively and (ii) immunoreactive PAP polypeptides either disappeared (secreted PAPs) or were substantially reduced (intracellular PAP). The degradation of both secreted PAP isozymes was correlated with the de novo synthesis of two extracellular serine proteases having M(r)s of 137 and 121 kDa. In vitro proteolysis of purified secreted tomato PAP isozymes occurred following their 24 h incubation with culture filtrate from Pi-resupplied cells. The results indicate that Pi or Phi addition to -Pi tomato cells induces serine proteases that degrade Pi-starvation inducible extracellular proteins.

A comparison of the transcriptome of Drosophila melanogaster in response to entomopathogenic fungus, ionizing radiation, starvation and cold shock.

PubMed

Moskalev, Alexey; Zhikrivetskaya, Svetlana; Krasnov, George; Shaposhnikov, Mikhail; Proshkina, Ekaterina; Borisoglebsky, Dmitry; Danilov, Anton; Peregudova, Darya; Sharapova, Irina; Dobrovolskaya, Eugenia; Solovev, Ilya; Zemskaya, Nadezhda; Shilova, Lyubov; Snezhkina, Anastasia; Kudryavtseva, Anna

2015-01-01

The molecular mechanisms that determine the organism's response to a variety of doses and modalities of stress factors are not well understood. We studied effects of ionizing radiation (144, 360 and 864 Gy), entomopathogenic fungus (10 and 100 CFU), starvation (16 h), and cold shock (+4, 0 and -4°C) on an organism's viability indicators (survival and locomotor activity) and transcriptome changes in the Drosophila melanogaster model. All stress factors but cold shock resulted in a decrease of lifespan proportional to the dose of treatment. However, stress-factors affected locomotor activity without correlation with lifespan. Our data revealed both significant similarities and differences in differential gene expression and the activity of biological processes under the influence of stress factors. Studied doses of stress treatments deleteriously affect the organism's viability and lead to different changes of both general and specific cellular stress response mechanisms.

Hepatic uptake of amino acids in late-pregnant rats. Effect of food deprivation.

PubMed Central

Casado, J; Remesar, X; Pastor-Anglada, M

1987-01-01

Hepatic availability, uptake and fractional extraction of amino acids were estimated in anaesthetized 21-day-pregnant and age-matched virgin rats, either fed or after 24 h starvation. Amino acid availability was unaltered in fed pregnant rats as compared with fed virgin controls. However, the hepatic uptake of these compounds was higher in the former than in the latter. These adaptations were mediated by an increase in the hepatic capability to take up amino acids in late-pregnant rats, as reflected by the changes found for the fractional extraction rates. The decrease in amino acid availability found after starvation was more pronounced in pregnant than in virgin rats. Nevertheless, the hepatic uptake was similar in both groups. These results indicate that amino acids are not limiting for ureagenesis during late pregnancy, strongly suggesting that the mechanism(s) which modulate urea synthesis may be intracellular in origin. PMID:3435433

The energy cost of triglyceride-fatty acid recycling in nonobese subjects after an overnight fast and four days of starvation.

PubMed

Elia, M; Zed, C; Neale, G; Livesey, G

1987-03-01

The basal blood glycerol concentration was determined and the rate of glycerol turnover was assessed by a nonradioactive infusion technique in six healthy nonobese adults after an overnight fast and again after four days of total starvation. Simultaneously, estimates of total energy expenditure and net fat oxidation were made from measurements of oxygen consumption, carbon dioxide production, and urinary nitrogen excretion. The data were combined to provide quantitative estimates of the activity of the triglyceride/fatty acid cycle. The basal concentration of glycerol in venous blood rose from a mean value of 54 +/- 8 mumol/L (SEM) before starvation to 154 +/- 5 mumol/L on day 4 of starvation. Glycerol turnover rates correlated well with the basal blood glycerol concentration (r = .95) and increased from a mean value of 115 +/- 17 mumol/min before starvation (equivalent to mobilization of about 3.95 kJ triglyceride/min) to 304 +/- 20 mumol/min (equivalent to mobilization of about 18.41 kJ/min). The estimated rate of net fat oxidation was 3.00 +/- 0.47 kJ/min before starvation and 4.00 +/- 0.14 kJ/min on day +4 of starvation. The rate of triglyceride energy recycling or rate of deposition of triglyceride energy into fat stores was calculated from the difference in the rate of fat energy mobilization and the rate of energy released during net fat oxidation. The values were found to be 0.94 +/- 0.26 kJ/min before starvation and 6.29 +/- 0.54 kJ/min on day +4 of starvation.(ABSTRACT TRUNCATED AT 250 WORDS)

Carbon-Starvation Induces Cross-Resistance to Thermal, Acid, and Oxidative Stress in Serratia marcescens

PubMed Central

Pittman, Joseph R.; Kline, La’Kesha C.; Kenyon, William J.

2015-01-01

The broad host-range pathogen Serratia marcescens survives in diverse host and non-host environments, often enduring conditions in which the concentration of essential nutrients is growth-limiting. In such environments, carbon and energy source starvation (carbon-starvation) is one of the most common forms of stress encountered by S. marcescens. Related members of the family Enterobacteriaceae are known to undergo substantial changes in gene expression and physiology in response to the specific stress of carbon-starvation, enabling non-spore-forming cells to survive periods of prolonged starvation and exposure to other forms of stress (i.e., starvation-induced cross-resistance). To determine if carbon-starvation also results in elevated levels of cross-resistance in S. marcescens, both log-phase and carbon-starved cultures, depleted of glucose before the onset of high cell-density stationary-phase, were grown in minimal media at either 30 °C or 37 °C and were then challenged for resistance to high temperature (50 °C), low pH (pH 2.8), and oxidative stress (15 mM H2O2). In general, carbon-starved cells exhibited a higher level of resistance to thermal stress, acid stress, and oxidative stress compared to log-phase cells. The extent of carbon-starvation-induced cross-resistance was dependent on incubation temperature and on the particular strain of S. marcescens. In addition, strain- and temperature-dependent variations in long-term starvation survival were also observed. The enhanced stress-resistance of starved S. marcescens cells could be an important factor in their survival and persistence in many non-host environments and within certain host microenvironments where the availability of carbon sources is suboptimal for growth. PMID:27682115

The adenosine triphosphate method as a quality control tool to assess 'cleanliness' of frequently touched hospital surfaces.

PubMed

Knape, L; Hambraeus, A; Lytsy, B

2015-10-01

The adenosine triphosphate (ATP) method is widely accepted as a quality control method to complement visual assessment, in the specifications of requirements, when purchasing cleaning contractors in Swedish hospitals. To examine whether the amount of biological load, as measured by ATP on frequently touched near-patient surfaces, had been reduced after an intervention; to evaluate the correlation between visual assessment and ATP levels on the same surfaces; to identify aspects of the performance of the ATP method as a tool in evaluating hospital cleanliness. A prospective intervention study in three phases was carried out in a medical ward and an intensive care unit (ICU) at a regional hospital in mid-Sweden between 2012 and 2013. Existing cleaning procedures were defined and baseline tests were sampled by visual inspection and ATP measurements of ten frequently touched surfaces in patients' rooms before and after intervention. The intervention consisted of educating nursing staff about the importance of hospital cleaning and direct feedback of ATP levels before and after cleaning. The mixed model showed a significant decrease in ATP levels after the intervention (P < 0.001). Relative light unit values were lower in the ICU. Cleanliness as judged by visual assessments improved. In the logistic regression analysis, there was a significant association between visual assessments and ATP levels. Direct feedback of ATP levels, together with education and introduction of written cleaning protocols, were effective tools to improve cleanliness. Visual assessment correlated with the level of ATP but the correlation was not absolute. The ATP method could serve as an educational tool for staff, but is not enough to assess hospital cleanliness in general as only a limited part of a large area is covered. Copyright © 2015 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

Energy demand and supply in human skeletal muscle.

PubMed

Barclay, C J

2017-04-01

The energy required for muscle contraction is provided by the breakdown of ATP but the amount of ATP in muscles cells is sufficient to power only a short duration of contraction. Buffering of ATP by phosphocreatine, a reaction catalysed by creatine kinase, extends the duration of activity possible but sustained activity depends on continual regeneration of PCr. This is achieved using ATP generated by oxidative processes and, during intense activity, by anaerobic glycolysis. The rate of ATP breakdown ranges from 70 to 140 mM min -1 during isometric contractions of various intensity to as much as 400 mM min -1 during intense, dynamic activity. The maximum rate of oxidative energy supply in untrained people is ~50 mM min -1 which, if the contraction duty cycle is 0.5 as is often the case in cyclic activity, is sufficient to match an ATP breakdown rate during contraction of 100 mM min -1 . During brief, intense activity the rate of ATP turnover can exceed the rates of PCr regeneration by combined oxidative and glycolytic energy supply, resulting in a net decrease in PCr concentration. Glycolysis has the capacity to produce between 30 and 50 mM of ATP so that, for example, anaerobic glycolysis could provide ATP at an average of 100 mM min -1 over 30 s of exhausting activity. The creatine kinase reaction plays an important role not only in buffering ATP but also in communicating energy demand from sites of ATP breakdown to the mitochondria. In that role, creatine kinases acts to slow and attenuate the response of mitochondria to changes in energy demand.

The metal chaperone Atox1 regulates the activity of the human copper transporter ATP7B by modulating domain dynamics.

PubMed

Yu, Corey H; Yang, Nan; Bothe, Jameson; Tonelli, Marco; Nokhrin, Sergiy; Dolgova, Natalia V; Braiterman, Lelita; Lutsenko, Svetlana; Dmitriev, Oleg Y

2017-11-03

The human transporter ATP7B delivers copper to the biosynthetic pathways and maintains copper homeostasis in the liver. Mutations in ATP7B cause the potentially fatal hepatoneurological disorder Wilson disease. The activity and intracellular localization of ATP7B are regulated by copper, but the molecular mechanism of this regulation is largely unknown. We show that the copper chaperone Atox1, which delivers copper to ATP7B, and the group of the first three metal-binding domains (MBD1-3) are central to the activity regulation of ATP7B. Atox1-Cu binding to ATP7B changes domain dynamics and interactions within the MBD1-3 group and activates ATP hydrolysis. To understand the mechanism linking Atox1-MBD interactions and enzyme activity, we have determined the MBD1-3 conformational space using small angle X-ray scattering and identified changes in MBD dynamics caused by apo -Atox1 and Atox1-Cu by solution NMR. The results show that copper transfer from Atox1 decreases domain interactions within the MBD1-3 group and increases the mobility of the individual domains. The N-terminal segment of MBD1-3 was found to interact with the nucleotide-binding domain of ATP7B, thus physically coupling the domains involved in copper binding and those involved in ATP hydrolysis. Taken together, the data suggest a regulatory mechanism in which Atox1-mediated copper transfer activates ATP7B by releasing inhibitory constraints through increased freedom of MBD1-3 motions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Fe deficiency differentially affects the vacuolar proton pumps in cucumber and soybean roots

PubMed Central

Dell’Orto, Marta; Nisi, Patrizia De; Vigani, Gianpiero; Zocchi, Graziano

2013-01-01

Iron uptake in dicots depends on their ability to induce a set of responses in root cells including rhizosphere acidification through H+ extrusion and apoplastic Fe(III) reduction by Fe(III)-chelate reductase. These responses must be sustained by metabolic rearrangements aimed at providing the required NAD(P)H, ATP and H+. Previous results in Fe-deficient cucumber roots showed that high H+ extrusion is accompanied by increased phosphoenolpyruvate carboxylase (PEPC) activity, involved in the cytosol pH-stat; moreover 31P-NMR analysis revealed increased vacuolar pH and decreased vacuolar [inorganic phosphate (Pi)]. The opposite was found in soybean: low rhizosphere acidification, decreased PEPC activity, vacuole acidification, and increased vacuolar [Pi]. These findings, highlighting a different impact of the Fe deficiency responses on cytosolic pH in the two species, lead to hypothesize different roles for H+ and Pi movements across the tonoplast in pH homeostasis. The role of vacuole in cytosolic pH-stat involves the vacuolar H+-ATPase (V-ATPase) and vacuolar H+-pyrophosphatase (V-PPase) activities, which generating the ΔpH and ΔΨ, mediate the transport of solutes, among which Pi, across the tonoplast. Fluxes of Pi itself in its two ionic forms, H2PO4- predominating in the vacuole and HPO42- in the cytosol, may be involved in pH homeostasis owing to its pH-dependent protonation/deprotonation reactions. Tonoplast enriched fractions were obtained from cucumber and soybean roots grown with or without Fe. Both V-ATPase and V-PPase activities were analyzed and the enrichment and localization of the corresponding proteins in root tissues were determined by Western blot and immunolocalization. V-ATPase did not change its activity and expression level in response to Fe starvation in both species. V-PPase showed a different behavior: in cucumber roots its activity and abundance were decreased, while in Fe-deficient soybean roots they were increased. The distinct role of the two H+ pumps in Pi fluxes between cytoplasm and vacuole in Fe-deficient cucumber and soybean root cells is discussed. PMID:23986768

Rapid and precise determination of ATP using a modified photometer

USGS Publications Warehouse

Shultz, David J.; Stephens, Doyle W.

1980-01-01

An inexpensive delay timer was designed to modify a commercially available ATP photometer which allows a disposable tip pipette to be used for injecting either enzyme or sample into the reaction cuvette. The disposable tip pipette is as precise and accurate as a fixed-needle syringe but eliminates the problem of sample contamination and decreases analytical time. (USGS)

Serum-deprivation stimulates cap-binding by PARN at the expense of eIF4E, consistent with the observed decrease in mRNA stability

PubMed Central

Seal, Ruth; Temperley, Richard; Wilusz, Jeffrey; Lightowlers, Robert N.; Chrzanowska-Lightowlers, Zofia M. A.

2005-01-01

PARN, a poly(A)-specific ribonuclease, binds the 5′ cap-structure of mRNA and initiates deadenylation-dependent decay. Eukaryotic initiation factor 4E (eIF4E) also binds to the cap structure, an interaction that is critical for initiating cap-dependent translation. The stability of various mRNA transcripts in human cell lines is reduced under conditions of serum starvation as determined by both functional and chemical half-lives. Serum starvation also leads to enhanced cap association by PARN. In contrast, the 5′ cap occupancy by eIF4E decreases under serum-deprivation, as does the translation of reporter transcripts. Further, we show that PARN is a phosphoprotein and that this modification can be modulated by serum status. Taken together, these data are consistent with a natural competition existing at the 5′ cap structure between PARN and eIF4E that may be regulated by changes in post-translational modifications. These phosphorylation-induced changes in the interplay of PARN and eIF4E may determine whether the mRNA is translated or decayed. PMID:15653638

Consequences of lower food intake on the digestive enzymes activities, the energy reserves and the reproductive outcome in Gammarus fossarum.

PubMed

Charron, Laetitia; Geffard, Olivier; Chaumot, Arnaud; Coulaud, Romain; Jaffal, Ali; Gaillet, Véronique; Dedourge-Geffard, Odile; Geffard, Alain

2015-01-01

Digestive enzyme activity is often used as a sensitive response to environmental pollution. However, only little is known about the negative effects of stress on digestive capacities and their consequences on energy reserves and reproduction, although these parameters are important for the maintenance of populations. To highlight if changes in biochemical responses (digestive enzymes and reserves) led to impairments at an individual level (fertility), Gammarus fossarum were submitted to a lower food intake throughout a complete female reproductive cycle (i.e. from ovogenesis to offspring production). For both males and females, amylase activity was inhibited by the diet stress, whereas trypsin activity was not influenced. These results underline similar sensitivity of males and females concerning their digestive capacity. Energy reserves decreased with food starvation in females, and remained stable in males. The number of embryos per female decreased with food starvation. Lower digestive activity in males and females therefore appears as an early response. These results underline the ecological relevance of digestive markers, as they make it possible to anticipate upcoming consequences on reproduction in females, a key biological variable for population dynamics.

Hypoxia decreases creatine uptake in cardiomyocytes, while creatine supplementation enhances HIF activation.

PubMed

Santacruz, Lucia; Arciniegas, Antonio Jose Luis; Darrabie, Marcus; Mantilla, Jose G; Baron, Rebecca M; Bowles, Dawn E; Mishra, Rajashree; Jacobs, Danny O

2017-08-01

Creatine (Cr), phosphocreatine (PCr), and creatine kinases (CK) comprise an energy shuttle linking ATP production in mitochondria with cellular consumption sites. Myocytes cannot synthesize Cr: these cells depend on uptake across the cell membrane by a specialized creatine transporter (CrT) to maintain intracellular Cr levels. Hypoxia interferes with energy metabolism, including the activity of the creatine energy shuttle, and therefore affects intracellular ATP and PCr levels. Here, we report that exposing cultured cardiomyocytes to low oxygen levels rapidly diminishes Cr transport by decreasing V max and K m Pharmacological activation of AMP-activated kinase (AMPK) abrogated the reduction in Cr transport caused by hypoxia. Cr supplementation increases ATP and PCr content in cardiomyocytes subjected to hypoxia, while also significantly augmenting the cellular adaptive response to hypoxia mediated by HIF-1 activation. Our results indicate that: (1) hypoxia reduces Cr transport in cardiomyocytes in culture, (2) the cytoprotective effects of Cr supplementation are related to enhanced adaptive physiological responses to hypoxia mediated by HIF-1, and (3) Cr supplementation increases the cellular ATP and PCr content in RNCMs exposed to hypoxia. © 2017 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.

Leucine-Enriched Essential Amino Acids Augment Muscle Glycogen Content in Rats Seven Days after Eccentric Contraction

PubMed Central

Kato, Hiroyuki; Miura, Kyoko; Suzuki, Katsuya; Bannai, Makoto

2017-01-01

Eccentric contractions induce muscle damage, which impairs recovery of glycogen and adenosine tri-phosphate (ATP) content over several days. Leucine-enriched essential amino acids (LEAAs) enhance the recovery in muscles that are damaged after eccentric contractions. However, the role of LEAAs in this process remains unclear. We evaluated the content in glycogen and high energy phosphates molecules (phosphocreatine (PCr), adenosine di-phosphate (ADP) and ATP) in rats that were following electrically stimulated eccentric contractions. Muscle glycogen content decreased immediately after the contraction and remained low for the first three days after the stimulation, but increased seven days after the eccentric contraction. LEAAs administration did not change muscle glycogen content during the first three days after the contraction. Interestingly, however, it induced a further increase in muscle glycogen seven days after the stimulation. Contrarily, ATP content decreased immediately after the eccentric contraction, and remained lower for up to seven days after. Additionally, LEAAs administration did not affect the ATP content over the experimental period. Finally, ADP and PCr levels did not significantly change after the contractions or LEAA administration. LEAAs modulate the recovery of glycogen content in muscle after damage-inducing exercise. PMID:29065533

Leucine-Enriched Essential Amino Acids Augment Muscle Glycogen Content in Rats Seven Days after Eccentric Contraction.

PubMed

Kato, Hiroyuki; Miura, Kyoko; Suzuki, Katsuya; Bannai, Makoto

2017-10-23

Eccentric contractions induce muscle damage, which impairs recovery of glycogen and adenosine tri-phosphate (ATP) content over several days. Leucine-enriched essential amino acids (LEAAs) enhance the recovery in muscles that are damaged after eccentric contractions. However, the role of LEAAs in this process remains unclear. We evaluated the content in glycogen and high energy phosphates molecules (phosphocreatine (PCr), adenosine di-phosphate (ADP) and ATP) in rats that were following electrically stimulated eccentric contractions. Muscle glycogen content decreased immediately after the contraction and remained low for the first three days after the stimulation, but increased seven days after the eccentric contraction. LEAAs administration did not change muscle glycogen content during the first three days after the contraction. Interestingly, however, it induced a further increase in muscle glycogen seven days after the stimulation. Contrarily, ATP content decreased immediately after the eccentric contraction, and remained lower for up to seven days after. Additionally, LEAAs administration did not affect the ATP content over the experimental period. Finally, ADP and PCr levels did not significantly change after the contractions or LEAA administration. LEAAs modulate the recovery of glycogen content in muscle after damage-inducing exercise.

[Mechanism of changes in the rate of glycolysis and levels of ATP and 2,3-diphosphoglycerate in human erythrocytes during aging].

PubMed

Bogatskaia, L N; Pisaruk, A V

1987-01-01

Reasons which have induced changes in the glycolysis rate, ATP and 2,3-diphosphoglycerate content in human erythrocytes with ageing are studied. A fall of the hexokinase activity is shown to be one of the reasons of a significant decrease in the glycolysis rate. The total ATPase activity in erythrocytes does not change with the age. At the same time the decay rate of 2,3-diphosphoglycerate increases, that, evidently, is one of the reasons of the 2,3-diphosphoglycerate content decrease in erythrocytes with ageing.

Human macrophage ATP7A is localized in the trans-Golgi apparatus, controls intracellular copper levels, and mediates macrophage responses to dermal wounds.

PubMed

Kim, Ha Won; Chan, Qilin; Afton, Scott E; Caruso, Joseph A; Lai, Barry; Weintraub, Neal L; Qin, Zhenyu

2012-02-01

The copper transporter ATP7A has attracted significant attention since the discovery of its gene mutation leading to human Menkes disease. We previously reported that ATP7A is highly expressed in the human vasculature and identified a novel vascular function of ATP7A in modulation of the expression and activity of extracellular superoxide dismutase. We recently identified that ATP7A expression in THP-1 cells (a monocyte/macrophage model cell line) plays a role in the oxidation of low density lipoproteins, indicating that it is necessary to further investigate its expression and function in monocytes/macrophages. In the current study, we demonstrated the protein and mRNA expression of ATP7A in human peripheral blood mononuclear cell (PBMC)-derived macrophages and alveolar macrophages. ATP7A was strongly co-localized with the trans-Golgi apparatus in PBMC-derived macrophages. Intracellular copper, detected by synchrotron X-ray fluorescence microscopy, was found to be distributed to the nucleus and cytoplasm in human THP-1 cells. To confirm the role of endogenous ATP7A in macrophage copper homeostasis, we performed inductively coupled plasma mass spectrometry in murine peritoneal macrophages, which showed markedly increased intracellular copper levels in macrophages isolated from ATP7A-deficient mice versus control mice. Moreover, the role of ATP7A in regulating macrophage responses to dermal wounds was studied by introduction of control and ATP7A-downregulated THP-1 cells into dermal wounds of nude mice. Infiltration of THP-1 cells into the wounded area (detected by expression of human macrophage markers MAC2 and CD68) was reduced in response to downregulation of ATP7A, hinting decreased macrophage accumulation subsequent to dermal wounds. In summary, alongside our previous studies, these findings indicate that human macrophage ATP7A is localized in the trans-Golgi apparatus, regulates intracellular copper levels, and mediates macrophage responses to a dermal wound.

Renal epithelial cells can release ATP by vesicular fusion

PubMed Central

Bjaelde, Randi G.; Arnadottir, Sigrid S.; Overgaard, Morten T.; Leipziger, Jens; Praetorius, Helle A.

2013-01-01

Renal epithelial cells have the ability to release nucleotides as paracrine factors. In the intercalated cells of the collecting duct, ATP is released by connexin30 (cx30), which is selectively expressed in this cell type. However, ATP is released by virtually all renal epithelia and the aim of the present study was to identify possible alternative nucleotide release pathways in a renal epithelial cell model. We used MDCK (type1) cells to screen for various potential ATP release pathways. In these cells, inhibition of the vesicular H+-ATPases (bafilomycin) reduced both the spontaneous and hypotonically (80%)-induced nucleotide release. Interference with vesicular fusion using N-ethylamide markedly reduced the spontaneous nucleotide release, as did interference with trafficking from the endoplasmic reticulum to the Golgi apparatus (brefeldin A1) and vesicular transport (nocodazole). These findings were substantiated using a siRNA directed against SNAP-23, which significantly reduced spontaneous ATP release. Inhibition of pannexin and connexins did not affect the spontaneous ATP release in this cell type, which consists of ~90% principal cells. TIRF-microscopy of either fluorescently-labeled ATP (MANT-ATP) or quinacrine-loaded vesicles, revealed that spontaneous release of single vesicles could be promoted by either hypoosmolality (50%) or ionomycin. This vesicular release decreased the overall cellular fluorescence by 5.8 and 7.6% respectively. In summary, this study supports the notion that spontaneous and induced ATP release can occur via exocytosis in renal epithelial cells. PMID:24065923

Deficiency of ATP6V1H Causes Bone Loss by Inhibiting Bone Resorption and Bone Formation through the TGF-β1 Pathway

PubMed Central

Duan, Xiaohong; Liu, Jin; Zheng, Xueni; Wang, Zhe; Zhang, Yanli; Hao, Ying; Yang, Tielin; Deng, Hongwen

2016-01-01

Vacuolar-type H +-ATPase (V-ATPase) is a highly conserved, ancient enzyme that couples the energy of ATP hydrolysis to proton transport across vesicular and plasma membranes of eukaryotic cells. Previously reported mutations of various V-ATPase subunits are associated with increased bone density. We now show that haploinsufficiency for the H subunit of the V1 domain (ATP6V1H) is associated with osteoporosis in humans and mice. A genome-wide SNP array analysis of 1625 Han Chinese found that 4 of 15 tag SNPs (26.7%) within ATP6V1H were significantly associated with low spine bone mineral density. Atp6v1h+/- knockout mice generated by the CRISPR/Cas9 technique had decreased bone remodeling and a net bone matrix loss. Atp6v1h+/- osteoclasts showed impaired bone formation and increased bone resorption. The increased intracellular pH of Atp6v1h+/- osteoclasts downregulated TGF-β1 activation, thereby reducing induction of osteoblast formation but the bone mineralization was not altered. However, bone formation was reduced more than bone resorption. Our data provide evidence that partial loss of ATP6V1H function results in osteoporosis/osteopenia. We propose that defective osteoclast formation triggers impaired bone formation by altering bone remodeling. In the future, ATP6V1H might, therefore, serve as a target for the therapy of osteoporosis. PMID:27924156

Global Proteomics Analysis of the Response to Starvation in C. elegans*

PubMed Central

Larance, Mark; Pourkarimi, Ehsan; Wang, Bin; Brenes Murillo, Alejandro; Kent, Robert; Lamond, Angus I.; Gartner, Anton

2015-01-01

Periodic starvation of animals induces large shifts in metabolism but may also influence many other cellular systems and can lead to adaption to prolonged starvation conditions. To date, there is limited understanding of how starvation affects gene expression, particularly at the protein level. Here, we have used mass-spectrometry-based quantitative proteomics to identify global changes in the Caenorhabditis elegans proteome due to acute starvation of young adult animals. Measuring changes in the abundance of over 5,000 proteins, we show that acute starvation rapidly alters the levels of hundreds of proteins, many involved in central metabolic pathways, highlighting key regulatory responses. Surprisingly, we also detect changes in the abundance of chromatin-associated proteins, including specific linker histones, histone variants, and histone posttranslational modifications associated with the epigenetic control of gene expression. To maximize community access to these data, they are presented in an online searchable database, the Encyclopedia of Proteome Dynamics (http://www.peptracker.com/epd/). PMID:25963834

Butyltin exposure causes a rapid decrease in cyclic AMP levels in human lymphocytes.

PubMed

Whalen, M M; Loganathan, B G

2001-03-15

Natural killer (NK) cells are a subset of lymphocytes that are capable of killing tumor cells, virally infected cells, and antibody-coated cells. Butyltins (BTs) are used in a variety of consumer products and industrial applications. Tributyltin (TBT) is found in dairy products, meat, and fish. Dibutyltin (DBT) is found in plastic products, beverages stored in PVC pipes during manufacturing, and poultry products. BTs appear to increase the risk of cancer and viral infections in exposed individuals. This increased risk may be due in part to the inhibitory effect of these compounds on the cytotoxic function of NK cells. A 24-h exposure of NK cells to 200 nM TBT or 1.5 microM DBT decreased the cytotoxic function of NK cells by greater than 90%. Higher concentrations of TBT and DBT decreased the cytotoxic function of NK cells (by greater than 90%) after only a 1-h exposure. A 24-h exposure to either TBT or DBT decreased intracellular ATP levels by about 30%. However, as much as a 1-h exposure to either 300 nM TBT or 10 microM DBT caused no significant decrease in ATP levels. Thus, a decrease in ATP levels is a longer-term consequence of BT exposure. Intracellular levels of cAMP are decreased by as much as 80% within 5 min of exposure to either TBT or DBT. This rapid decline in cAMP levels in NK cells may be a consequence of BT exposure that is related to the rapid decrease in the cytotoxic function of NK cells. Copyright 2001 Academic Press.

Dynamics of shear-induced ATP release from red blood cells.

PubMed

Wan, Jiandi; Ristenpart, William D; Stone, Howard A

2008-10-28

Adenosine triphosphate (ATP) is a regulatory molecule for many cell functions, both for intracellular and, perhaps less well known, extracellular functions. An important example of the latter involves red blood cells (RBCs), which help regulate blood pressure by releasing ATP as a vasodilatory signaling molecule in response to the increased shear stress inside arterial constrictions. Although shear-induced ATP release has been observed widely and is believed to be triggered by deformation of the cell membrane, the underlying mechanosensing mechanism inside RBCs is still controversial. Here, we use an in vitro microfluidic approach to investigate the dynamics of shear-induced ATP release from human RBCs with millisecond resolution. We demonstrate that there is a sizable delay time between the onset of increased shear stress and the release of ATP. This response time decreases with shear stress, but surprisingly does not depend significantly on membrane rigidity. Furthermore, we show that even though the RBCs deform significantly in short constrictions (duration of increased stress <3 ms), no measurable ATP is released. This critical timescale is commensurate with a characteristic membrane relaxation time determined from observations of the cell deformation by using high-speed video. Taken together our results suggest a model wherein the retraction of the spectrin-actin cytoskeleton network triggers the mechanosensitive ATP release and a shear-dependent membrane viscosity controls the rate of release.

Model study of ATP and ADP buffering, transport of Ca(2+) and Mg(2+), and regulation of ion pumps in ventricular myocyte

NASA Technical Reports Server (NTRS)

Michailova, A.; McCulloch, A.

2001-01-01

We extended the model of the ventricular myocyte by Winslow et al. (Circ. Res 1999, 84:571-586) by incorporating equations for Ca(2+) and Mg(2+) buffering and transport by ATP and ADP and equations for MgATP regulation of ion transporters (Na(+)-K(+) pump, sarcolemmal and sarcoplasmic Ca(2+) pumps). The results indicate that, under normal conditions, Ca(2+) binding by low-affinity ATP and diffusion of CaATP may affect the amplitude and time course of intracellular Ca(2+) signals. The model also suggests that a fall in ATP/ADP ratio significantly reduces sarcoplasmic Ca(2+) content, increases diastolic Ca(2+), lowers systolic Ca(2+), increases Ca(2+) influx through L-type channels, and decreases the efficiency of the Na(+)/Ca(2+) exchanger in extruding Ca(2+) during periodic voltage-clamp stimulation. The analysis suggests that the most important reason for these changes during metabolic inhibition is the down-regulation of the sarcoplasmic Ca(2+)-ATPase pump by reduced diastolic MgATP levels. High Ca(2+) concentrations developed near the membrane might have a greater influence on Mg(2+), ATP, and ADP concentrations than that of the lower Ca(2+) concentrations in the bulk myoplasm. The model predictions are in general agreement with experimental observations measured under normal and pathological conditions.

Ectonucleotidase NTPDase3 is abundant in pancreatic β-cells and regulates glucose-induced insulin secretion.

PubMed

Syed, Samreen K; Kauffman, Audra L; Beavers, Lisa S; Alston, James T; Farb, Thomas B; Ficorilli, James; Marcelo, Marialuisa C; Brenner, Martin B; Bokvist, Krister; Barrett, David G; Efanov, Alexander M

2013-11-15

Extracellular ATP released from pancreatic β-cells acts as a potent insulinotropic agent through activation of P2 purinergic receptors. Ectonucleotidases, a family of membrane-bound nucleotide-metabolizing enzymes, regulate extracellular ATP levels by degrading ATP and related nucleotides. Ectonucleotidase activity affects the relative proportion of ATP and its metabolites, which in turn will impact the level of purinergic receptor stimulation exerted by extracellular ATP. Therefore, we investigated the expression and role of ectonucleotidases in pancreatic β-cells. Of the ectonucleotidases studied, only ENTPD3 (gene encoding the NTPDase3 enzyme) mRNA was detected at fairly abundant levels in human and mouse pancreatic islets as well as in insulin-secreting MIN6 cells. ARL67156, a selective ectonucleotidase inhibitor, blocked degradation of extracellular ATP that was added to MIN6 cells. The compound also decreased degradation of endogenous ATP released from cells. Measurements of insulin secretion in MIN6 cells as well as in mouse and human pancreatic islets demonstrated that ARL67156 potentiated glucose-dependent insulin secretion. Downregulation of NTPDase3 expression in MIN6 cells with the specific siRNA replicated the effects of ARL67156 on extracellular ATP hydrolysis and insulin secretion. Our results demonstrate that NTPDase3 is the major ectonucleotidase in pancreatic β-cells in multiple species and that it modulates insulin secretion by controlling activation of purinergic receptors.

ATP Synthase Repression in Tobacco Restricts Photosynthetic Electron Transport, CO2 Assimilation, and Plant Growth by Overacidification of the Thylakoid Lumen[OA

PubMed Central

Rott, Markus; Martins, Nádia F.; Thiele, Wolfram; Lein, Wolfgang; Bock, Ralph; Kramer, David M.; Schöttler, Mark A.

2011-01-01

Tobacco (Nicotiana tabacum) plants strictly adjust the contents of both ATP synthase and cytochrome b6f complex to the metabolic demand for ATP and NADPH. While the cytochrome b6f complex catalyzes the rate-limiting step of photosynthetic electron flux and thereby controls assimilation, the functional significance of the ATP synthase adjustment is unknown. Here, we reduced ATP synthase accumulation by an antisense approach directed against the essential nuclear-encoded γ-subunit (AtpC) and by the introduction of point mutations into the translation initiation codon of the plastid-encoded atpB gene (encoding the essential β-subunit) via chloroplast transformation. Both strategies yielded transformants with ATP synthase contents ranging from 100 to <10% of wild-type levels. While the accumulation of the components of the linear electron transport chain was largely unaltered, linear electron flux was strongly inhibited due to decreased rates of plastoquinol reoxidation at the cytochrome b6f complex (photosynthetic control). Also, nonphotochemical quenching was triggered at very low light intensities, strongly reducing the quantum efficiency of CO2 fixation. We show evidence that this is due to an increased steady state proton motive force, resulting in strong lumen overacidification, which in turn represses photosynthesis due to photosynthetic control and dissipation of excitation energy in the antenna bed. PMID:21278125

Five months of fasting in king penguin chicks: body mass loss and fuel metabolism.

PubMed

Cherel, Y; Le Maho, Y

1985-10-01

When king penguin chicks are 3-4 mo old, they enter a season of interrupted growth due to long periods of fasting, because they are irregularly fed in winter. Nine captive chicks [mean initial body mass (m) = 12.5 kg] had fasted an average of 5 mo at the end of the experiment; m was then 4.0 kg, a 68% decrease. They probably could have fasted longer, since chicks of parents delayed in the return to the colony die from starvation at an m of 3.0 kg. The long fast could be divided into three periods based on the changes in rate of decrease of m. The remarkable resistance of king penguin chicks to starvation may be partly explained by their ability to maintain protein sparing for as much as 4 mo, the duration of period II; plasma concentrations of uric acid, urea, and alanine were then minimum, 0.1, 0.4, and 0.4 mmol X l-1 respectively. Particular changes during this period, i.e., progressive increase of beta-hydroxybutyrate and decrease of glucose concentrations, might contribute to the efficiency of protein sparing. Period III was marked by a rise in protein utilization, plasma concentrations of uric acid, urea, and alanine increasing to 0.7, 1.5, and 0.8 mmol X l-1, respectively.

Temporal pattern of feeding response of Chaoborus larvae to starvation

Treesearch

Rakesh Minocha; James F. Haney

1986-01-01

The effect of starvation on the feeding rate of larval Chaoborus (Diptera. Chaoboridae) was investigated using Daphnia rosea as prey. The starvation period varied from 12 h to 22 days. The starved Chaoborus were individually incubated with 10 Daphnia under controlled light and temperature...

Fragrance chemicals lyral and lilial decrease viability of HaCat cells' by increasing free radical production and lowering intracellular ATP level: protection by antioxidants.

PubMed

Usta, Julnar; Hachem, Yassmine; El-Rifai, Omar; Bou-Moughlabey, Yolla; Echtay, Karim; Griffiths, David; Nakkash-Chmaisse, Hania; Makki, Rajaa Fakhoury

2013-02-01

We investigate in this study the biochemical effects on cells in culture of two commonly used fragrance chemicals: lyral and lilial. Whereas both chemicals exerted a significant effect on primary keratinocyte(s), HaCat cells, no effect was obtained with any of HepG2, Hek293, Caco2, NIH3T3, and MCF7 cells. Lyral and lilial: (a) decreased the viability of HaCat cells with a 50% cell death at 100 and 60 nM respectively; (b) decreased significantly in a dose dependant manner the intracellular ATP level following 12-h of treatment; (c) inhibited complexes I and II of electron transport chain in liver sub-mitochondrial particles; and (d) increased reactive oxygen species generation that was reversed by N-acetyl cysteine and trolox and the natural antioxidant lipoic acid, without influencing the level of free and/or oxidized glutathione. Lipoic acid protected HaCat cells against the decrease in viability induced by either compound. Dehydrogenation of lyral and lilial produce α,β-unsaturated aldehydes, that reacts with lipoic acid requiring proteins resulting in their inhibition. We propose lyral and lilial as toxic to mitochondria that have a direct effect on electron transport chain, increase ROS production, derange mitochondrial membrane potential, and decrease cellular ATP level, leading thus to cell death. Copyright © 2012 Elsevier Ltd. All rights reserved.

Combined inhibition of glycolysis and AMPK induces synergistic breast cancer cell killing

PubMed Central

Wu, Yong; Sarkissyan, Marianna; Mcghee, Eva; Lee, Sangkyu

2015-01-01

Targeting glycolysis for cancer treatment has been investigated as a therapeutic method but has not offered a feasible chemotherapeutic strategy. Our aim was to examine whether AMP-activated protein kinase (AMPK), a conditional oncogene, rescues the energetic stress and cytotoxicity induced by 2-deoxyglucose (2-DG), a glycolytic inhibitor, and the related mechanisms. Luciferin/luciferase adenosine triphosphate (ATP) determination, Western analysis, qRT-PCR analyses, MTT growth assay, clonogenic assay, and statistical analysis were performed in this study. 2-DG decreased ATP levels and subsequently activated AMPK, which contribute to intracellular ATP recovery in MCF-7 cells thus exhibiting no apparent cytotoxicity. Compound C, an AMPK inhibitor, further potentiates 2-DG-induced decrease in ATP levels and inhibits their recovery. 2-DG, via AMPK activation, stimulated cAMP response element-binding protein (CREB) phosphorylation and activity and promoted nuclear peroxisome proliferator-activated receptor gamma coactivator-1-beta (PGC-1β) and estrogen-related receptor α (ERRα) protein expression, leading to augmented mitochondrial biogenesis and expression of fatty acid oxidation (FAO) genes including PPARα, MCAD, CPT1C, and ACO. This metabolic adaptation elicited by AMPK counteracts the ATP-depleting and cancer cell-killing effect of 2-DG. However, 2-DG in combination with AMPK antagonists or small interfering RNA caused a dramatic increase in cytotoxicity in MCF-7 but not in MCF-10A cells. Similarly, when combined with inhibition of CREB/PGC-1β/ERRα pathway, 2-DG saliently suppressed mitochondrial biogenesis and the expression of FAO genes, depleted ATP production, and enhanced cytotoxicity in cancer cells. Collectively, the combination of 2-DG and AMPK inhibition synergistically enhanced the cytotoxic potential in breast cancer cells with a relative nontoxicity to normal cells and may offer a promising, safe, and effective breast cancer therapeutic strategy. PMID:25975952

Gualou Xiebai Decoction, a Traditional Chinese Medicine, Prevents Cardiac Reperfusion Injury of Hyperlipidemia Rat via Energy Modulation.

PubMed

Yan, Lu-Lu; Zhang, Wei-Yang; Wei, Xiao-Hong; Yan, Li; Pan, Chun-Shui; Yu, Yang; Fan, Jing-Yu; Liu, Yu-Ying; Zhou, Hua; Han, Jing-Yan; Yao, Xin-Sheng

2018-01-01

Background: Gualou Xiebai Decoction (GLXB) is a classic prescription of Chinese medicine used for the treatment of cardiac problems. The present study was designed to explore the effect and mechanism of GLXB on ischemia/reperfusion (I/R) induced disorders in myocardial structure and function, focusing on the regulation of energy metabolism and the RhoA/ROCK pathway. Methods: After hyperlipidemic rat model was established by oral administration of high fat diet, the rats were treated with GLXB for 6 weeks and subjected to 30 min occlusion of the left anterior descending coronary artery (LADCA) followed by 90 min reperfusion to elicit I/R challenge. Myocardial infarct size was assessed by Evans blue-TTC staining. Myocardial blood flow (MBF) and cardiac function were evaluated. Enzyme-linked immunosorbent assay was performed to examine the content of ATP, ADP, AMP, CK, CK-MB, LDH, cTnT, cTnI, and IL-6. Double staining of F-actin and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling was conducted to assess myocardial apoptosis. Expressions of ATP synthase subunit δ (ATP 5D), and RhoA and ROCK were determined by Western blotting. Results: Administration with GLXB at high dose for 6 weeks protected heart against I/R-induced MBF decrease, myocardial infarction and apoptosis, ameliorated I/R-caused impairment of cardiac function and myocardial structure, restored the decrease in the ratio of ADP/ATP and AMP/ATP, and the expression of ATP 5D with inhibiting the expression of RhoA and ROCK. Conclusions: Treatment with GLXB effectively protects myocardial structure and function from I/R challenge, possibly via regulating energy metabolism involving inactivation of RhoA/ROCK signaling pathway.

Thiamethoxam induced hepatic energy changes in silver catfish via impairment of the phosphoryl transfer network pathway: Toxicological effects on energetics homeostasis.

PubMed

Baldissera, Matheus D; Souza, Carine F; Golombieski, Jaqueline I; Seben, Débora; Sippert, Letícia R; Salbego, Joseânia; Zanella, Renato; Baldisserotto, Bernardo

2018-06-01

Precise coupling of spatially separated intracellular adenosine triphosphate (ATP)-producing and ATP-consuming processes exerts a pivotal role in bioenergetic homeostasis of living organisms, and the phosphotransfer network pathway, catalyzed by adenylate kinase (AK) and pyruvate kinase (PK), is fundamental in cellular and tissue energetic homeostasis. Measurement of the phosphotransfer network can provide new information for understanding the alterations in hepatic energetic metabolism during exposition to insecticides, such as thiamethoxam. Therefore, the aim of this study was to evaluate whether exposition to thiamethoxam negatively affects the hepatic enzymes of the phosphotransfer network in silver catfish (Rhamdia quelen). Hepatic AK and PK activities were inhibited at 3.75 μg L -1 after 24 h of exposure and at 1.125 and 3.75 μg L -1 after 96 h of exposure compared with the control group. The hepatic ATP levels were decreased following 3.75 μg L -1 thiamethoxam treatment after 24 h of exposure and at 1.125 and 3.75 μg L -1 after 96 h of exposure compared with the control group. The enzymatic activity of the phosphotransfer network and ATP levels did not recover after 48 h of recovery in clean water. Thus, the inhibition of hepatic AK and PK activities by thiamethoxam caused impairment of energy homeostasis in liver tissue, decreasing hepatic ATP availability. Moreover, the absence of a mutual compensatory mechanism between these enzymes directly contributes to ATP depletion and to a severe energetic dysregulation, which may contribute to toxic effects caused by thiamethoxam. Copyright © 2018 Elsevier B.V. All rights reserved.

Detailed conformation dynamics and activation process of wild type c-Abl and T315I mutant

NASA Astrophysics Data System (ADS)

Yang, Li-Jun; Zhao, Wen-Hua; Liu, Qian

2014-10-01

Bcr-Abl is an important target for therapy against chronic myelogenous leukemia (CML) and acute lymphocytic leukemia (ALL). The synergistic effect between myristyl pocket and the ATP pocket has been found. But its detailed information based on molecular level still has not been achieved. In this study, conventional molecular dynamics (CMD) and target molecular dynamics (TMD) simulations were performed to explore the effect of T315I mutation on dynamics and activation process of Abl containing the N-terminal cap (Ncap). The CMD simulation results reveal the increasing flexibility of ATP pocket in kinase domain (KD) after T315I mutation which confirms the disability of ATP-pocket inhibitors to the Abl-T315I mutant. On the contrary, the T315I mutation decreased the flexibility of remote helix αI which suggests the synergistic effect between them. The mobility of farther regions containing Ncap, SH3, SH2 and SH2-KD linker were not affected by T315I mutation. The TMD simulation results show that the activation process of wild type Abl and Abl-T315I mutant experienced global conformation change. Their differences were elucidated by the activation motion of subsegments including A-loop, P-loop and Ncap. Besides, the T315I mutation caused decreasing energy barrier and increasing intermediate number in activation process, which results easier activation process. The TMD and CMD results indicate that a drug targeting only the ATP pocket is not enough to inhibit the Abl-T315I mutant. An effective way to inhibit the abnormal activity of Abl-T315I mutant is to combine the ATP-pocket inhibitors with inhibitors binding at non-ATP pockets mainly related to Ncap, SH2-KD linker and myristyl pocket.

Transgenic expression of Bcl-2 modulates energy metabolism, prevents cytosolic acidification during ischemia, and reduces ischemia/reperfusion injury.

PubMed

Imahashi, Kenichi; Schneider, Michael D; Steenbergen, Charles; Murphy, Elizabeth

2004-10-01

The antiapoptotic protein Bcl-2 is targeted to the mitochondria, but it is uncertain whether Bcl-2 affects only myocyte survival after ischemia, or whether it also affects metabolic functions of mitochondria during ischemia. Hearts from mice overexpressing human Bcl-2 and from their wild-type littermates (WT) were subjected to 24 minutes of global ischemia followed by reperfusion. During ischemia, the decrease in pH(i) and the initial rate of decline in ATP were significantly reduced in Bcl-2 hearts compared with WT hearts (P<0.05). The reduced acidification during ischemia was dependent on the activity of mitochondrial F1F0-ATPase. In the presence of oligomycin (Oligo), an F1F0-ATPase inhibitor, the decrease in pH(i) was attenuated in WT hearts, but in Bcl-2 hearts, Oligo had no additional effect on pH(i) during ischemia. Likewise, addition of Oligo to WT hearts slowed the rate of decline in ATP during ischemia to a level similar to that observed in Bcl-2 hearts, but addition of Oligo had no significant effect on the rate of decline in ATP in Bcl-2 hearts during ischemia. These data are consistent with Bcl-2-mediated inhibition of consumption of glycolytic ATP. Furthermore, mitochondria from Bcl-2 hearts have a reduced rate of consumption of ATP on uncoupler addition. This could be accomplished by limiting ATP entry into the mitochondria through the voltage-dependent anion channel, and/or the adenine nucleotide transporter, or by direct inhibition of the F1F0-ATPase. Immunoprecipitation showed greater interaction between Bcl-2 and voltage-dependent anion channel during ischemia. These data indicate that Bcl-2 modulation of metabolism contributes to cardioprotection.

Polymorphisms within the COL5A1 gene and regulators of the extracellular matrix modify the risk of Achilles tendon pathology in a British case-control study.

PubMed

Brown, Karryn L; Seale, Kirsten B; El Khoury, Louis Y; Posthumus, Michael; Ribbans, William J; Raleigh, Stuart M; Collins, Malcolm; September, Alison V

2017-08-01

Several genetic loci have been associated with risk of Achilles tendon pathology (ATP) within South African and Australian populations. The aim of this study was, therefore, to evaluate eight previously implicated genetic variants in an independent British population. A total of 130 asymptomatic controls (CON) and 112 participants clinically diagnosed with ATP comprising 87 individuals with chronic Achilles tendinopathy (TEN) and 25 with Achilles tendon ruptures (RUP) were included. All participants were genotyped for variants within the COL5A1, MIR608, IL-1β, IL-6 and CASP8 genes. Primary findings implicated COL5A1 and CASP8. Three inferred allele combinations constructed from COL5A1 rs12722, rs3196378 and rs71746744 were identified as risk modifiers. The T-C-D combination was associated with increased risk of ATP (P = 0.023) and RUP (P < 0.001), the C-A-I combination was associated with increased risk of ATP (P = 0.011), TEN (P = 0.011) and RUP (P = 0.011) and the C-C-D combination was associated with decreased risk of ATP (P = 0.011) and RUP (P = 0.004). The CASP8 rs3834129 DD genotype was associated with decreased risk of TEN (P = 0.020, odds ratio: 0.45, 95% confidence interval: 0.22-0.90) and the CASP8 I-G (rs3834129-rs1045485) inferred allele combination was associated with increased risk of TEN (P = 0.031). This study further highlights the importance of polymorphisms within COL5A1 and CASP8 in the aetiology of ATP.

Phosphorylation of Eukaryotic Initiation Factor-2α during Stress and Encystation in Entamoeba Species

PubMed Central

Hendrick, Holland M.; Welter, Brenda H.; Sykes, Steven E.; Sullivan, William J.; Temesvari, Lesly A.

2016-01-01

Entamoeba histolytica is an enteric pathogen responsible for amoebic dysentery and liver abscess. It alternates between the host-restricted trophozoite form and the infective environmentally-stable cyst stage. Throughout its lifecycle E. histolytica experiences stress, in part, from host immune pressure. Conversion to cysts is presumed to be a stress-response. In other systems, stress induces phosphorylation of a serine residue on eukaryotic translation initiation factor-2α (eIF2α). This inhibits eIF2α activity resulting in a general decline in protein synthesis. Genomic data reveal that E. histolytica possesses eIF2α (EheIF2α) with a conserved phosphorylatable serine at position 59 (Ser59). Thus, this pathogen may have the machinery for stress-induced translational control. To test this, we exposed cells to different stress conditions and measured the level of total and phospho-EheIF2α. Long-term serum starvation, long-term heat shock, and oxidative stress induced an increase in the level of phospho-EheIF2α, while short-term serum starvation, short-term heat shock, or glucose deprivation did not. Long-term serum starvation also caused a decrease in polyribosome abundance, which is in accordance with the observation that this condition induces phosphorylation of EheIF2α. We generated transgenic cells that overexpress wildtype EheIF2α, a non-phosphorylatable variant of eIF2α in which Ser59 was mutated to alanine (EheIF2α-S59A), or a phosphomimetic variant of eIF2α in which Ser59 was mutated to aspartic acid (EheIF2α-S59D). Consistent with the known functions of eIF2α, cells expressing wildtype or EheIF2α-S59D exhibited increased or decreased translation, respectively. Surprisingly, cells expressing EheIF2α-S59A also exhibited reduced translation. Cells expressing EheIF2α-S59D were more resistant to long-term serum starvation underscoring the significance of EheIF2α phosphorylation in managing stress. Finally, phospho-eIF2α accumulated during encystation in E. invadens, a model encystation system. Together, these data demonstrate that the eIF2α-dependent stress response system is operational in Entamoeba species. PMID:27930733

Interaction between ATP, metal ions, glycine, and several minerals

NASA Technical Reports Server (NTRS)

Rishpon, J.; Ohara, P. J.; Lawless, J. G.; Lahav, N.

1982-01-01

Interactions between ATP, glycine and montmorillonite and kaolinite clay minerals in the presence of various metal cations are investigated. The adsorption of adenine nucleotides on clays and Al(OH)3 was measured as a function of pH, and glycine condensation was followed in the presence of ATP, ZnCl2, MgCl2 and either kaolinite or montmorillonite. The amounts of ATP and ADP adsorbed are found to decrease with increasing Ph, and to be considerably enhanced in experiments with Mg(2+)- and Zn(2+)-montmorillonite with respect to Na(+)-montmorillonite. The effects of divalent cations are less marked in kaolinite. Results for Al(OH)3 show the importance of adsorption at clay platelet edges at high pH. The decomposition of ATP during drying at high temperature is observed to be inhibited by small amounts of clay, vacuum, or Mg(2+) or Zn(2+) ions, and to be accompanied by peptide formation in the presence of glycine. Results suggest the importance of Zn(2+) and Mg(2+) in chemical evolution.

POLI-mix functional food enhances steady-state bioenergetic status independently of age: an experimental study.

PubMed

Rastmanesh, Reza; Marotta, Francesco; Kantah, Makoto Keiichi; Nagpal, Ravinder; Lorenzetti, Aldo; Takadanohara, Hiroshi; Mashizume, Hiroshi; Kobayashi, Riyichi; Chui, De Hua

2012-04-01

BALB/c mice were divided into young, middle-aged, and aged groups, and each group was given 3 weeks of oral treatments: (1) 1 mL of VBC1-99 (a mixture of 42 fruits and vegetables extracts) or (2) 1 mL of same amount of antioxidant vitamins as control. Steady-state hepatic adenosine triphosphate (ATP) was assessed by phosphorus-31 nuclear magnetic resonance ((31)P-NMR) spectroscopy as: β-ATP/reference peak, inorganic phosphorus (Pi)/peak and β-ATP/Pi. As compared to untreated control, VBC1-99 significantly enhanced β-ATP/peak and β-ATP/Pi ratios (p<0.01) in all age groups and throughout the observation period (p<0.05) together with a significant decrease of Pi/ref peak ratio (p<0.05). However, this value in middle-aged and aged mice was comparable to antioxidant control mice. These NMR data demonstrate that VBC1-99 has a beneficial effect on hepatic energy metabolism, irrespective of age.

ATP-induced noncooperative thermal unfolding of hen lysozyme

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Honglin; Yin, Peidong; He, Shengnan

To understand the role of ATP underlying the enhanced amyloidosis of hen egg white lysozyme (HEWL), the synchrotron radiation circular dichroism, combined with tryptophan fluorescence, dynamic light-scattering, and differential scanning calorimetry, is used to examine the alterations of the conformation and thermal unfolding pathway of the HEWL in the presence of ATP, Mg{sup 2+}-ATP, ADP, AMP, etc. It is revealed that the binding of ATP to HEWL through strong electrostatic interaction changes the secondary structures of HEWL and makes the exposed residue W62 move into hydrophobic environments. This alteration of W62 decreases the {beta}-domain stability of HEWL, induces a noncooperativemore » unfolding of the secondary structures, and produces a partially unfolded intermediate. This intermediate containing relatively rich {alpha}-helix and less {beta}-sheet structures has a great tendency to aggregate. The results imply that the ease of aggregating of HEWL is related to the extent of denaturation of the amyloidogenic region, rather than the electrostatic neutralizing effect or monomeric {beta}-sheet enriched intermediate.« less

Capture and quality control mechanisms for adenosine-5'-triphosphate binding.

PubMed

Li, Li; Martinis, Susan A; Luthey-Schulten, Zaida

2013-04-24

The catalytic events in members of the nucleotidylyl transferase superfamily are initiated by a millisecond binding of ATP in the active site. Through metadynamics simulations on a class I aminoacyl-tRNA synthetase (aaRSs), the largest group in the superfamily, we calculate the free energy landscape of ATP selection and binding. Mutagenesis studies and fluorescence spectroscopy validated the identification of the most populated intermediate states. The rapid first binding step involves formation of encounter complexes captured through a fly casting mechanism that acts upon the triphosphate moiety of ATP. In the slower nucleoside binding step, a conserved histidine in the HxxH motif orients the incoming ATP through base-stacking interactions resulting in a deep minimum in the free energy surface. Mutation of this histidine significantly decreases the binding affinity measured experimentally and computationally. The metadynamics simulations further reveal an intermediate quality control state that the synthetases and most likely other members of the superfamily use to select ATP over other nucleoside triphosphates.

Resistance of soil microorganisms to starvation.

NASA Technical Reports Server (NTRS)

Chen, M.; Alexander, M.

1972-01-01

Most groups of soil microorganisms died when exposed to prolonged starvation in a carbon-free solution, but the relative abundance of Bacillus and actinomycetes increased with time. Certain nonspore-forming bacteria also persisted. The ability of individual soil isolates to endure starvation in solution was not correlated with their glycogen content or rate of endogenous respiration. However, cells of the resistant populations were rich in poly-beta-hydroxybutyrate, whereas the starvation-susceptible bacteria generally contained little of this substance. Poly-beta-hydroxybutyrate was used rapidly in cells deprived of exogenous sources of carbon.

P2Y2 receptor activation inhibits the expression of the sodium-chloride cotransporter NCC in distal convoluted tubule cells.

PubMed

Gailly, P; Szutkowska, M; Olinger, E; Debaix, H; Seghers, F; Janas, S; Vallon, V; Devuyst, O

2014-11-01

Luminal nucleotide stimulation is known to reduce Na(+) transport in the distal nephron. Previous studies suggest that this mechanism may involve the thiazide-sensitive Na(+)-Cl(-) cotransporter (NCC), which plays an essential role in NaCl reabsorption in the cells lining the distal convoluted tubule (DCT). Here we show that stimulation of mouse DCT (mDCT) cells with ATP or UTP promoted Ca(2+) transients and decreased the expression of NCC at both mRNA and protein levels. Specific siRNA-mediated silencing of P2Y2 receptors almost completely abolished ATP/UTP-induced Ca(2+) transients and significantly reduced ATP/UTP-induced decrease of NCC expression. To test whether local variations in the intracellular Ca(2+) concentration ([Ca(2+)]i) may control NCC transcription, we overexpressed the Ca(2+)-binding protein parvalbumin selectively in the cytosol or in the nucleus of mDCT cells. The decrease in NCC mRNA upon nucleotide stimulation was abolished in cells overexpressing cytosolic PV but not in cells overexpressing either a nuclear-targeted PV or a mutated PV unable to bind Ca(2+). Using a firefly luciferase reporter gene strategy, we observed that the activity of NCC promoter region from -1 to -2,200 bp was not regulated by changes in [Ca(2+)]i. In contrast, high cytosolic calcium level induced instability of NCC mRNA. We conclude that in mDCT cells: (1) P2Y2 receptor is essential for the intracellular Ca(2+) signaling induced by ATP/UTP stimulation; (2) P2Y2-mediated increase of cytoplasmic Ca(2+) concentration down-regulates the expression of NCC; (3) the decrease of NCC expression occurs, at least in part, via destabilization of its mRNA.

Mechanisms for monovalent cation-dependent depletion of intracellular Mg2+:Na(+)-independent Mg2+ pathways in guinea-pig smooth muscle.

PubMed

Nakayama, Shinsuke; Nomura, Hideki; Smith, Lorraine M; Clark, Joseph F; Uetani, Tadayuki; Matsubara, Tatsuaki

2003-09-15

It has been suggested that magnesium deficiency is correlated with many diseases. 31P NMR experiments were carried out in order to investigate the effects of Na+ substitution on Mg2+ depletion in smooth muscle under divalent cation-free conditions. In the taenia of guinea-pig caeci, the intracellular free Mg2+ concentration ([Mg2+]i) was estimated from the chemical shifts of (1) the beta-ATP peak alone and (2) beta- and gamma-ATP peaks. Both estimations indicated that [Mg2+]i decreased only very slowly in Mg(2+)-free, Ca(2+)-free solutions in which Na+ was substituted with large cations such as NMDG (N-methyl-D-glucamine) and choline. Furthermore, the measurements of tension development supported the suggestion of preservation of intracellular Mg2+ with NMDG substitution. Substituting extracellular Na+ with the small cation, Li+, also shifted the beta-ATP peak towards a lower frequency, but the frequency shift was significantly less than that seen upon Na+ substitution with K+. The estimated [Mg2+]i depletion was, however, comparable with that seen after Na+ substitution with K+ using the titration curves of metal-free and Mg(2+)-bound ATP obtained in Li(+)-based model solutions. It was concluded that Mg2+ rapidly decreases only when small cations were the major electrolyte of the extracellular medium. Na+ substitutions with NMDG, choline or Li+ had little effect on intracellular ATP concentration after 100 min treatment.

Extracellular ATP Acts on Jasmonate Signaling to Reinforce Plant Defense.

PubMed

Tripathi, Diwaker; Zhang, Tong; Koo, Abraham J; Stacey, Gary; Tanaka, Kiwamu

2018-01-01

Damaged cells send various signals to stimulate defense responses. Recent identification and genetic studies of the plant purinoceptor, P2K1 (also known as DORN1), have demonstrated that extracellular ATP is a signal involved in plant stress responses, including wounding, perhaps to evoke plant defense. However, it remains largely unknown how extracellular ATP induces plant defense responses. Here, we demonstrate that extracellular ATP induces plant defense mediated through activation of the intracellular signaling of jasmonate (JA), a well-characterized defense hormone. In Arabidopsis ( Arabidopsis thaliana ) leaves, ATP pretreatment induced resistance against the necrotrophic fungus, Botrytis cinerea The induced resistance was enhanced in the P2K1 receptor overexpression line, but reduced in the receptor mutant, dorn1 - 3 Mining the transcriptome data revealed that ATP induces a set of JA-induced genes. In addition, the P2K1-associated coexpression network contains defense-related genes, including those encoding jasmonate ZIM-domain (JAZ) proteins, which play key roles as repressors of JA signaling. We examined whether extracellular ATP impacts the stability of JAZ1 in Arabidopsis. The results showed that the JAZ1 stability decreased in response to ATP addition in a proteasome-dependent manner. This reduction required intracellular signaling via second messengers-cytosolic calcium, reactive oxygen species, and nitric oxide. Interestingly, the ATP-induced JAZ1 degradation was attenuated in the JA receptor mutant, coi1 , but not in the JA biosynthesis mutant, aos , or upon addition of JA biosynthesis inhibitors. Immunoprecipitation analysis demonstrated that ATP increases the interaction between COI1 and JAZ1, suggesting direct cross talk between extracellular ATP and JA in intracellular signaling events. Taken together, these results suggest that extracellular ATP signaling directly impacts the JA signaling pathway to maximize plant defense responses. © 2018 American Society of Plant Biologists. All Rights Reserved.

Purinergic modulation of preBötzinger complex inspiratory rhythm in rodents: the interaction between ATP and adenosine

PubMed Central

Zwicker, J D; Rajani, V; Hahn, L B; Funk, G D

2011-01-01

Abstract ATP signalling in the CNS is mediated by a three-part system comprising the actions of ATP (and ADP) at P2 receptors (P2Rs), adenosine (ADO) at P1 receptors (P1Rs), and ectonucleotidases that degrade ATP into ADO. ATP excites preBötzinger complex (preBötC) inspiratory rhythm-generating networks where its release attenuates the hypoxic depression of breathing. Its metabolite, ADO, inhibits breathing through unknown mechanisms that may involve the preBötC. Our objective is to understand the dynamics of this signalling system and its influence on preBötC networks. We show that the preBötC of mouse and rat is sensitive to P2Y1 purinoceptor (P2Y1R) activation, responding with a >2-fold increase in frequency. Remarkably, the mouse preBötC is insensitive to ATP. Only after block of A1 ADORs is the ATP-evoked, P2Y1R-mediated frequency increase observed. This demonstrates that ATP is rapidly degraded to ADO, which activates inhibitory A1Rs, counteracting the P2Y1R-mediated excitation. ADO sensitivity of mouse preBötC was confirmed by a frequency decrease that was absent in rat. Differential ectonucleotidase activities are likely to contribute to the negligible ATP sensitivity of mouse preBötC. Real-time PCR analysis of ectonucleotidase isoforms in preBötC punches revealed TNAP (degrades ATP to ADO) or ENTPDase2 (favours production of excitatory ADP) as the primary constituent in mouse and rat, respectively. These data further establish the sensitivity of this vital network to P2Y1R-mediated excitation, emphasizing that individual components of the three-part signalling system dramatically alter network responses to ATP. Data also suggest therapeutic potential may derive from methods that alter the ATP–ADO balance to favour the excitatory actions of ATP. PMID:21788352

Extracellular adenosine triphosphate increases cation permeability of chronic lymphocytic leukemic lymphocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wiley, J.S.; Dubyak, G.R.

Extracellular adenosine triphosphate (ATP) is known to reversibly increase the cation permeability of a variety of freshly isolated and cultured cell types. In this study the effects of extracellular ATP were studied using peripheral blood lymphocytes (PBL) isolated from both normal subjects and from patients with chronic lymphocytic leukemia (CLL). Changes in the permeability to Na+, Rb+, and Li+ ions were measured using conventional isotope and flame photometry techniques. In addition, changes in cytosolic (Ca2+) were fluorimetrically monitored to assess possible changes in net Ca2+ influx. ATP produced a 12-fold increase in 22Na+ influx into CLL cells but only amore » 3.5-fold increase in this flux in PBL cells. A maximal response was produced by 0.1 mmol/L ATP in the absence of Mg2+, while a twofold molar excess of Mg2+ over ATP abolished the response. ATP had no effect on the passive (ouabain-insensitive) 86Rb+ influx into PBL cells but stimulated this flux by fivefold in the CLL cells. Li+ influx into CLL cells was also stimulated threefold by ATP. Under these same conditions ATP also produced a net increase in total cell Na and a decrease in total cell K in the CLL cells. Exclusion of two normally impermeable dyes, trypan blue and ethidium bromide, was not altered in the ATP-treated CLL cells. Finally, extracellular ATP (3 mmol/L) produced no significant change in the cytosolic (Ca2+) of normal, monocyte-depleted populations of PBL. Conversely, this same concentration of ATP produced a very rapid and a significant (an average threefold peak change) increase in the cytosolic (Ca2+) of cell preparations derived from five out of nine CLL patients. In these latter CLL cells, the ATP-induced elevation in cytosolic (Ca2+) appeared to be due to a net increase in Ca2+ influx, since no elevations were observed when the extracellular (Ca2+) was reduced to less than 0.1 mmol/L.« less

The Influence of Age, Sex, Pregnancy, Starvation, and Other Factors on the Cytoplasmic Ribonucleoproteins of Rat Liver

PubMed Central

Petermann, Mary L.; Hamilton, Mary G.

1958-01-01

Rat liver was homogenized in 0.88 M sucrose. The DNA and total RNA were determined, and the homogenate was fractionated by differential centrifugation. The pellets obtained between 30 minutes at 20,000 g and 180 minutes at 105,000 g were analyzed for RNA and nitrogen. The ribonucleoproteins were determined in the analytical ultracentrifuge. The non-pellet RNA was calculated by difference. The results are reported as amounts per 6.7 x 10-9 mg. of DNA. In young, growing male rats the amounts of microsomal protein and ribonucleoprotein B (83S) increased with age. Non-pregnant adult females showed less non-pellet RNA and much more ribonucleoprotein C (63S) than did adult males. During pregnancy both of these cell constituents reverted to levels characteristic for male animals. Starvation for 5 days resulted in a reduction in the mass of liver tissue, the non-pellet RNA, the microsomal protein, and ribonucleoproteins B and C. During recovery from starvation the return of the liver to normal paralleled the rate at which body weight was restored. Treatment with cortisone, 25 mg. per rat per day for 5 days, caused an increase in microsomal protein and a decrease in ribonucleoprotein B. Treatment with 6-mercapto-purine, 50 mg. per kilo per day for 5 days, caused little change in liver composition in either males or females. PMID:13610943

Unravelling the cross-talk between iron starvation and oxidative stress responses highlights the key role of PerR (alr0957) in peroxide signalling in the cyanobacterium Nostoc PCC 7120.

PubMed

Yingping, Fan; Lemeille, Sylvain; Talla, Emmanuel; Janicki, Annick; Denis, Yann; Zhang, Cheng-Cai; Latifi, Amel

2014-10-01

The cyanobacterial phylum includes oxygenic photosynthetic prokaryotes of a wide variety of morphologies, metabolisms and ecologies. Their adaptation to their various ecological niches is mainly achieved by sophisticated regulatory mechanisms and depends on a fine cross-talk between them. We assessed the global transcriptomic response of the filamentous cyanobacterium Nostoc PCC 7120 to iron starvation and oxidative stress. More than 20% of the differentially expressed genes in response to iron stress were also responsive to oxidative stress. These transcripts include antioxidant proteins-encoding genes that confirms that iron depletion leads to reactive oxygen accumulation. The activity of the Fe-superoxide dismutase was not significantly decreased under iron starvation, indicating that the oxidative stress generated under iron deficiency is not a consequence of (SOD) deficiency. The transcriptional data indicate that the adaptation of Nostoc to iron-depleted conditions displays important differences with what has been shown in unicellular cyanobacteria. While the FurA protein that regulates the response to iron deprivation has been well characterized in Nostoc, the regulators in charge of the oxidative stress response are unknown. Our study indicates that the alr0957 (perR) gene encodes the master regulator of the peroxide stress. PerR is a peroxide-sensor repressor that senses peroxide by metal-catalysed oxidation.

Emodin Inhibits ATP-Induced Proliferation and Migration by Suppressing P2Y Receptors in Human Lung Adenocarcinoma Cells.

PubMed

Wang, Xia; Li, Long; Guan, Ruijuan; Zhu, Danian; Song, Nana; Shen, Linlin

2017-01-01

Extracellular ATP performs multiple important functions via activation of P2 receptors on the cell surface. P2Y receptors play critical roles in ATP evoked response in human lung adenocarcinoma cells (A549 cells). Emodin is an anthraquinone derivative originally isolated from Chinese rhubarb, possesses anticancer properties. In this study we examined the inhibiting effects of emodin on proliferation, migration and epithelial-mesenchymal transition (EMT) by suppressing P2Y receptors-dependent Ca2+ increase and nuclear factor-κB (NF-KB) signaling in A549 cells. A549 cells were pretreated with emodin before stimulation with ATP for the indicated time. Then, intracellular Ca2+ concentration ([Ca2+]i) was measured by Fluo-8/AM staining. Cell proliferation and cell cycle progression were tested by CCK8 assay and flow cytometry In addition, wound healing and western blot were performed to determine cell migration and related protein levels (Bcl-2, Bax, claudin-1, NF-κB). Emodin blunted ATP/UTP-induced increase of [Ca2+]i and cell proliferation concentration-dependently Meanwhile, it decreased ATP-induced cells accumulation in the S phase. Furthermore, emodin altered protein abundance of Bcl-2, Bax and claudin-1 and attenuated EMT caused by ATP. Such ATP-induced cellular reactions were also inhibited by a nonselective P2Y receptors antagonist, suramin, in a similar way to emodin. Besides, emodin could inhibit activation of NF-κB, thus suppressed ATP-induced proliferation, migration and EMT. Our results demonstrated that emodin inhibits ATP-induced proliferation, migration, EMT by suppressing P2Y receptors-mediated [Ca2+]i increase and NF-κB signaling in A549 cells. © 2017 The Author(s). Published by S. Karger AG, Basel.

ATP and Mg2+ Promote the Reversible Oligomerization and Aggregation of Chloroplast 2-Cys Peroxiredoxin*

PubMed Central

Aran, Martín; Ferrero, Diego; Wolosiuk, Alejandro; Mora-García, Santiago; Wolosiuk, Ricardo A.

2011-01-01

2-Cys peroxiredoxins (2-Cys Prxs) are ubiquitous peroxidases with important roles in cellular antioxidant defense and hydrogen peroxide-mediated signaling. Post-translational modifications of conserved cysteines cause the transition from low to high molecular weight oligomers, triggering the functional change from peroxidase to molecular chaperone. However, it remains unclear how non-covalent interactions of 2-Cys Prx with metabolites modulate the quaternary structure. Here, we disclose that ATP and Mg2+ (ATP/Mg) promote the self-polymerization of chloroplast 2-Cys Prx (polypeptide 23.5 kDa) into soluble higher order assemblies (>2 MDa) that proceed to insoluble aggregates beyond 5 mm ATP. Remarkably, the withdrawal of ATP or Mg2+ brings soluble oligomers and insoluble aggregates back to the native conformation without compromising the associated functions. As confirmed by transmission electron microscopy, ATP/Mg drive the toroid-like decamers (diameter 13 nm) to the formation of large sphere-like particles (diameter ∼30 nm). Circular dichroism studies on ATP-labeled 2-Cys Prx reveal that ATP/Mg enhance the proportion of β-sheets with the concurrent decrease in the content of α-helices. In line with this observation, the formation of insoluble aggregates is strongly prevented by 2,2,2-trifluoroethanol, a cosolvent employed to induce α-helical conformations. We further find that the response of self-polymerization to ATP/Mg departs abruptly from that of the associated peroxidase and chaperone activities when two highly conserved residues, Arg129 and Arg152, are mutated. Collectively, our data uncover that non-covalent interactions of ATP/Mg with 2-Cys Prx modulate dynamically the quaternary structure, thereby coupling the non-redox chemistry of cell energy with redox transformations at cysteine residues. PMID:21525006

Energy status of ripening and postharvest senescent fruit of litchi (Litchi chinensis Sonn.)

PubMed Central

2013-01-01

Background Recent studies have demonstrated that cellular energy is a key factor switching on ripening and senescence of fruit. However, the factors that influence fruit energy status remain largely unknown. Results HPLC profiling showed that ATP abundance increased significantly in developing preharvest litchi fruit and was strongly correlated with fruit fresh weight. In contrast, ATP levels declined significantly during postharvest fruit senescence and were correlated with the decrease in the proportion of edible fruit. The five gene transcripts isolated from the litchi fruit pericarp were highly expressed in vegetative tissues and peaked at 70 days after flowering (DAF) consistent with fruit ADP concentrations, except for uncoupling mitochondrial protein 1 (UCP1), which was predominantly expressed in the root, and ATP synthase beta subunit (AtpB), which was up-regulated significantly before harvest and peaked 2 days after storage. These results indicated that the color-breaker stage at 70 DAF and 2 days after storage may be key turning points in fruit energy metabolism. Transcript abundance of alternative oxidase 1 (AOX1) increased after 2 days of storage to significantly higher levels than those of LcAtpB, and was down-regulated significantly by exogenous ATP. ATP supplementation had no significant effect on transcript abundance of ADP/ATP carrier 1 (AAC1) and slowed the changes in sucrose non-fermenting-1-related kinase 2 (SnRK2) expression, but maintained ATP and energy charge levels, which were correlated with delayed senescence. Conclusions Our results suggest that senescence of litchi fruit is closely related with energy. A surge of LcAtpB expression marked the beginning of fruit senescence. The findings may provide a new strategy to extend fruit shelf life by regulating its energy level. PMID:23547657

ATP-induced current in isolated outer hair cells of guinea pig cochlea.

PubMed

Nakagawa, T; Akaike, N; Kimitsuki, T; Komune, S; Arima, T

1990-05-01

1. Electrical and pharmacologic properties of ATP-induced current in outer hair cells isolated from guinea pig cochlea were investigated in the whole-cell recording mode by the use of a conventional patch-clamp technique. 2. Under current-clamp conditions, rapid application of ATP depolarized the outer hair cells resulting in an increase in conductance. The ATP-induced response did not show any desensitization during a continuous application. 3. At a holding potential of -70 mV, the ATP-induced inward current increased in a sigmoidal fashion over the concentration range between 3 microM and 1 mM. The half-maximum concentration (EC50) was 12 microM and the Hill coefficient was 0.93. 4. The ATP-induced current had a reversal potential near 6 mV, which was close to the theoretical value (1 mV) calculated from the Goldman-Hodgkin-Katz equation for permeable intra- and extracellular cations. 5. In the current-voltage (I-V) relationship for the ATP response, a slight inward-going rectification was observed at more positive potentials than the reversal potential. 6. The substitution of extracellular Na+ by equimolar choline+ shifted the reversal potential of the ATP-induced current to more negative values. The substitution of Cs+ in the internal solution by N-methyl-D-glucamine+ (NMG+) shifted it in the positive direction. The reversal potential of ATP-induced current was also shifted to positive values with increasing extracellular Ca2+ concentration. A decrease of intracellular Cl- by gluconate- did not affect the reversal potential, thereby indicating that the ATP-induced current is carried through a large cation channel.(ABSTRACT TRUNCATED AT 250 WORDS)

Hypophosphatemia promotes lower rates of muscle ATP synthesis.

PubMed

Pesta, Dominik H; Tsirigotis, Dimitrios N; Befroy, Douglas E; Caballero, Daniel; Jurczak, Michael J; Rahimi, Yasmeen; Cline, Gary W; Dufour, Sylvie; Birkenfeld, Andreas L; Rothman, Douglas L; Carpenter, Thomas O; Insogna, Karl; Petersen, Kitt Falk; Bergwitz, Clemens; Shulman, Gerald I

2016-10-01

Hypophosphatemia can lead to muscle weakness and respiratory and heart failure, but the mechanism is unknown. To address this question, we noninvasively assessed rates of muscle ATP synthesis in hypophosphatemic mice by using in vivo saturation transfer [ 31 P]-magnetic resonance spectroscopy. By using this approach, we found that basal and insulin-stimulated rates of muscle ATP synthetic flux (V ATP ) and plasma inorganic phosphate (P i ) were reduced by 50% in mice with diet-induced hypophosphatemia as well as in sodium-dependent P i transporter solute carrier family 34, member 1 (NaPi2a)-knockout (NaPi2a -/- ) mice compared with their wild-type littermate controls. Rates of V ATP normalized in both hypophosphatemic groups after restoring plasma P i concentrations. Furthermore, V ATP was directly related to cellular and mitochondrial P i uptake in L6 and RC13 rodent myocytes and isolated muscle mitochondria. Similar findings were observed in a patient with chronic hypophosphatemia as a result of a mutation in SLC34A3 who had a 50% reduction in both serum P i content and muscle V ATP After oral P i repletion and normalization of serum P i levels, muscle V ATP completely normalized in the patient. Taken together, these data support the hypothesis that decreased muscle ATP synthesis, in part, may be caused by low blood P i concentrations, which may explain some aspects of muscle weakness observed in patients with hypophosphatemia.-Pesta, D. H., Tsirigotis, D. N., Befroy, D. E., Caballero, D., Jurczak, M. J., Rahimi, Y., Cline, G. W., Dufour, S., Birkenfeld, A. L., Rothman, D. L., Carpenter, T. O., Insogna, K., Petersen, K. F., Bergwitz, C., Shulman, G. I. Hypophosphatemia promotes lower rates of muscle ATP synthesis. © The Author(s).

Unexpected role of the copper transporter ATP7A in PDGF-induced vascular smooth muscle cell migration.

PubMed

Ashino, Takashi; Sudhahar, Varadarajan; Urao, Norifumi; Oshikawa, Jin; Chen, Gin-Fu; Wang, Huan; Huo, Yuqing; Finney, Lydia; Vogt, Stefan; McKinney, Ronald D; Maryon, Edward B; Kaplan, Jack H; Ushio-Fukai, Masuko; Fukai, Tohru

2010-09-17

Copper, an essential nutrient, has been implicated in vascular remodeling and atherosclerosis with unknown mechanism. Bioavailability of intracellular copper is regulated not only by the copper importer CTR1 (copper transporter 1) but also by the copper exporter ATP7A (Menkes ATPase), whose function is achieved through copper-dependent translocation from trans-Golgi network (TGN). Platelet-derived growth factor (PDGF) promotes vascular smooth muscle cell (VSMC) migration, a key component of neointimal formation. To determine the role of copper transporter ATP7A in PDGF-induced VSMC migration. Depletion of ATP7A inhibited VSMC migration in response to PDGF or wound scratch in a CTR1/copper-dependent manner. PDGF stimulation promoted ATP7A translocation from the TGN to lipid rafts, which localized at the leading edge, where it colocalized with PDGF receptor and Rac1, in migrating VSMCs. Mechanistically, ATP7A small interfering RNA or CTR small interfering RNA prevented PDGF-induced Rac1 translocation to the leading edge, thereby inhibiting lamellipodia formation. In addition, ATP7A depletion prevented a PDGF-induced decrease in copper level and secretory copper enzyme precursor prolysyl oxidase (Pro-LOX) in lipid raft fraction, as well as PDGF-induced increase in LOX activity. In vivo, ATP7A expression was markedly increased and copper accumulation was observed by synchrotron-based x-ray fluorescence microscopy at neointimal VSMCs in wire injury model. These findings suggest that ATP7A plays an important role in copper-dependent PDGF-stimulated VSMC migration via recruiting Rac1 to lipid rafts at the leading edge, as well as regulating LOX activity. This may contribute to neointimal formation after vascular injury. Our findings provide insight into ATP7A as a novel therapeutic target for vascular remodeling and atherosclerosis.

Intestinal alkaline phosphatase regulates protective surface microclimate pH in rat duodenum.

PubMed

Mizumori, Misa; Ham, Maggie; Guth, Paul H; Engel, Eli; Kaunitz, Jonathan D; Akiba, Yasutada

2009-07-15

Regulation of localized extracellular pH (pH(o)) maintains normal organ function. An alkaline microclimate overlying the duodenal enterocyte brush border protects the mucosa from luminal acid. We hypothesized that intestinal alkaline phosphatase (IAP) regulates pH(o) due to pH-sensitive ATP hydrolysis as part of an ecto-purinergic pH regulatory system, comprised of cell-surface P2Y receptors and ATP-stimulated duodenal bicarbonate secretion (DBS). To test this hypothesis, we measured DBS in a perfused rat duodenal loop, examining the effect of the competitive alkaline phosphatase inhibitor glycerol phosphate (GP), the ecto-nucleoside triphosphate diphosphohydrolase inhibitor ARL67156, and exogenous nucleotides or P2 receptor agonists on DBS. Furthermore, we measured perfusate ATP concentration with a luciferin-luciferase bioassay. IAP inhibition increased DBS and luminal ATP output. Increased luminal ATP output was partially CFTR dependent, but was not due to cellular injury. Immunofluorescence localized the P2Y(1) receptor to the brush border membrane of duodenal villi. The P2Y(1) agonist 2-methylthio-ADP increased DBS, whereas the P2Y(1) antagonist MRS2179 reduced ATP- or GP-induced DBS. Acid perfusion augmented DBS and ATP release, further enhanced by the IAP inhibitor l-cysteine, and reduced by the exogenous ATPase apyrase. Furthermore, MRS2179 or the highly selective P2Y(1) antagonist MRS2500 co-perfused with acid induced epithelial injury, suggesting that IAP/ATP/P2Y signalling protects the mucosa from acid injury. Increased DBS augments IAP activity presumably by raising pH(o), increasing the rate of ATP degradation, decreasing ATP-mediated DBS, forming a negative feedback loop. The duodenal epithelial brush border IAP-P2Y-HCO(3-) surface microclimate pH regulatory system effectively protects the mucosa from acid injury.

Reconsideration of the sequence of rigor mortis through postmortem changes in adenosine nucleotides and lactic acid in different rat muscles.

PubMed

Kobayashi, M; Takatori, T; Iwadate, K; Nakajima, M

1996-10-25

We examined the changes in adenosine triphosphate (ATP), lactic acid, adenosine diphosphate (ADP) and adenosine monophosphate (AMP) in five different rat muscles after death. Rigor mortis has been thought to occur simultaneously in dead muscles and hence to start in small muscles sooner than in large muscles. In this study we found that the rate of decrease in ATP was significantly different in each muscle. The greatest drop in ATP was observed in the masseter muscle. These findings contradict the conventional theory of rigor mortis. Similarly, the rates of change in ADP and lactic acid, which are thought to be related to the consumption or production of ATP, were different in each muscle. However, the rate of change of AMP was the same in each muscle.

Regulation of glucose and ketone-body metabolism in brain of anaesthetized rats

PubMed Central

Ruderman, Neil B.; Ross, Peter S.; Berger, Michael; Goodman, Michael N.

1974-01-01

1. The effects of starvation and diabetes on brain fuel metabolism were examined by measuring arteriovenous differences for glucose, lactate, acetoacetate and 3-hydroxybutyrate across the brains of anaesthetized fed, starved and diabetic rats. 2. In fed animals glucose represented the sole oxidative fuel of the brain. 3. After 48h of starvation, ketone-body concentrations were about 2mm and ketone-body uptake accounted for 25% of the calculated O2 consumption: the arteriovenous difference for glucose was not diminished, but lactate release was increased, suggesting inhibition of pyruvate oxidation. 4. In severe diabetic ketosis, induced by either streptozotocin or phlorrhizin (total blood ketone bodies >7mm), the uptake of ketone bodies was further increased and accounted for 45% of the brain's oxidative metabolism, and the arteriovenous difference for glucose was decreased by one-third. The arteriovenous difference for lactate was increased significantly in the phlorrhizin-treated rats. 5. Infusion of 3-hydroxybutyrate into starved rats caused marked increases in the arteriovenous differences for lactate and both ketone bodies. 6. To study the mechanisms of these changes, steady-state concentrations of intermediates and co-factors of the glycolytic pathway were determined in freeze-blown brain. 7. Starved rats had increased concentrations of acetyl-CoA. 8. Rats with diabetic ketosis had increased concentrations of fructose 6-phosphate and decreased concentrations of fructose 1,6-diphosphate, indicating an inhibition of phosphofructokinase. 9. The concentrations of acetyl-CoA, glycogen and citrate, a potent inhibitor of phosphofructokinase, were increased in the streptozotocin-treated rats. 10. The data suggest that cerebral glucose uptake is decreased in diabetic ketoacidosis owing to inhibition of phosphofructokinase as a result of the increase in brain citrate. 11. The inhibition of brain pyruvate oxidation in starvation and diabetes can be related to the accelerated rate of ketone-body metabolism; however, we found no correlation between the decrease in glucose uptake in the diabetic state and the arteriovenous difference for ketone bodies. 12. The data also suggest that the rates of acetoacetate and 3-hydroxybutyrate utilization by brain are governed by their concentrations in plasma. 13. The finding of very low concentrations of acetoacetate and 3-hydroxybutyrate in brain compared with plasma suggests that diffusion across the blood–brain barrier may be the rate-limiting step in their metabolism. PMID:4275704

Resuscitation of starved anaerobic ammonium oxidation sludge system: Impacts of repeated short-term starvation.

PubMed

Ye, Lihong; Li, Dong; Zhang, Jie; Zeng, Huiping

2018-05-04

Starvation of biomass is common during underloading of bioreactors or sludge storage in biological wastewater treatment industries. The aim of this work was to study the impact of starvation modes on the nitrogen removal capacity of anaerobic ammonium oxidation (anammox) process in sequencing batch reactor (SBR). The repeated short-term starvation and reactivation experiments were performed to evaluate the response of anammox sludge system in the condition of 27 ± 1.5 °C and 320 min HRT. Moreover, the nitrogen removal ability of the anammox process was reactivated rapidly in the low substrate condition, then the total nitrogen (TN) removal efficiency reached 82.5%, with the effluent TN of around 14.6 mgNL -1 . The repeated short-term starvation (1 day-4 days) and recovery mode could improve the tolerance and apparent activity of anammox sludge system. The dominant species of general anaerobic ammonium oxidation bacteria (AnAOB) was Candidatus Brocadia, which had better self-adaption to repeated starvation. Copyright © 2018 Elsevier Ltd. All rights reserved.

An analysis of the effects of Mn{sup 2+} on oxidative phosphorylation in liver, brain, and heart mitochondria using state 3 oxidation rate assays

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gunter, Thomas E., E-mail: thomas_gunter@urmc.rochester.ed; Gerstner, Brent, E-mail: brent_gerstner@urmc.rochester.ed; Lester, Tobias, E-mail: Tlester200@gmail.co

2010-11-15

Manganese (Mn) toxicity is partially mediated by reduced ATP production. We have used oxidation rate assays-a measure of ATP production-under rapid phosphorylation conditions to explore sites of Mn{sup 2+} inhibition of ATP production in isolated liver, brain, and heart mitochondria. This approach has several advantages. First, the target tissue for Mn toxicity in the basal ganglia is energetically active and should be studied under rapid phosphorylation conditions. Second, Mn may inhibit metabolic steps which do not affect ATP production rate. This approach allows identification of inhibitions that decrease this rate. Third, mitochondria from different tissues contain different amounts of themore » components of the metabolic pathways potentially resulting in different patterns of ATP inhibition. Our results indicate that Mn{sup 2+} inhibits ATP production with very different patterns in liver, brain, and heart mitochondria. The primary Mn{sup 2+} inhibition site in liver and heart mitochondria, but not in brain mitochondria, is the F{sub 1}F{sub 0} ATP synthase. In mitochondria fueled by either succinate or glutamate + malate, ATP production is much more strongly inhibited in brain than in liver or heart mitochondria; moreover, Mn{sup 2+} inhibits two independent sites in brain mitochondria. The primary site of Mn-induced inhibition of ATP production in brain mitochondria when succinate is substrate is either fumarase or complex II, while the likely site of the primary inhibition when glutamate plus malate are the substrates is either the glutamate/aspartate exchanger or aspartate aminotransferase.« less

Why does starvation make bones fat?

PubMed Central

Devlin, Maureen J.

2011-01-01

Body fat, or adipose tissue, is a crucial energetic buffer against starvation in humans and other mammals, and reserves of white adipose tissue (WAT) rise and fall in parallel with food intake. Much less is known about the function of bone marrow adipose tissue (BMAT), which are fat cells found in bone marrow. BMAT mass actually increases during starvation, even as other fat depots are being mobilized for energy. Here I review the possible reasons for this poorly understood phenomenon. Is BMAT a passive filler that occupies spaces left by dying bone cells, a pathological consequence of suppressed bone formation, or potentially an adaptation for surviving starvation? To evaluate these possibilities, here I review what is known about the effects of starvation on the body, particularly the skeleton, and the mechanisms involved in storing and metabolizing BMAT during negative energy balance. PMID:21793093

Why does starvation make bones fat?

PubMed

Devlin, Maureen J

2011-01-01

Body fat, or adipose tissue, is a crucial energetic buffer against starvation in humans and other mammals, and reserves of white adipose tissue (WAT) rise and fall in parallel with food intake. Much less is known about the function of bone marrow adipose tissue (BMAT), which are fat cells found in bone marrow. BMAT mass actually increases during starvation, even as other fat depots are being mobilized for energy. This review considers several possible reasons for this poorly understood phenomenon. Is BMAT a passive filler that occupies spaces left by dying bone cells, a pathological consequence of suppressed bone formation, or potentially an adaptation for surviving starvation? These possibilities are evaluated in terms of the effects of starvation on the body, particularly the skeleton, and the mechanisms involved in storing and metabolizing BMAT during negative energy balance. Copyright © 2011 Wiley-Liss, Inc.

The adenosine salvage pathway as an alternative to mitochondrial production of ATP in maturing mammalian oocytes.

PubMed

Scantland, Sara; Tessaro, Irene; Macabelli, Carolina H; Macaulay, Angus D; Cagnone, Gaël; Fournier, Éric; Luciano, Alberto M; Robert, Claude

2014-09-01

Although the oocyte is the largest cell in the body and an unavoidable phase in life, its physiology is still poorly understood, and other cell types provide little insight into its unique nature. Even basic cellular functions in the oocyte such as energy metabolism are not yet fully understood. It is known that the mitochondria of the female gamete exhibit an immature form characterized by limited energy production from glucose and oxidative phosphorylation. We show that the bovine oocyte uses alternative means to maintain ATP production during maturation, namely, the adenosine salvage pathway. Meiosis resumption is triggered by destruction of cyclic AMP by phosphodiesterases producing adenosine monophosphate that is converted into ATP by adenylate kinases and creatine kinases. Inhibition of these enzymes decreased ATP production, and addition of their substrates restored ATP production in denuded oocytes. Addition of phosphocreatine to the oocyte maturation medium influenced the phenotype of the resulting blastocysts. We propose a model in which adenylate kinases and creatine kinases act as drivers of ATP production from added AMP during oocyte maturation. © 2014 by the Society for the Study of Reproduction, Inc.

The pH dependence of the allosteric response of human liver pyruvate kinase to fructose-1,6-bisphosphate, ATP, and alanine

PubMed Central

Fenton, Aron W.; Hutchinson, Myra

2009-01-01

The allosteric regulation of human liver pyruvate kinase (hL-PYK) by fructose-1,6-bisphosphate (Fru-1,6-BP; activator), ATP (inhibitor) and alanine (Ala; inhibitor) was monitored over a pH range from 6.5 to 8.0 at 37°C. As a function of increasing pH, hL-PYK's affinity for the substrate phosphoenolpyruvate (PEP), and for Fru-1,6-BP decreases, while affinities for ATP and Ala slightly increases. At pH 6.5, Fru-1,6-BP and ATP elicit only small allosteric impacts on PEP affinity. As pH increases, Fru-1,6-BP and ATP elicit greater allosteric responses, but the response to Ala is relatively constant. Since the magnitudes of the allosteric coupling for ATP and for Ala inhibition are different and the pH dependences of these magnitudes are not similar, these inhibitors likely elicit their responses using different molecular mechanisms. In addition, our results fail to support a general correlation between pH dependent changes in effector affinity and pH dependent changes in the corresponding allosteric response. PMID:19467627

Aptamer-Based Carboxyl-Terminated Nanocrystalline Diamond Sensing Arrays for Adenosine Triphosphate Detection

PubMed Central

Suaebah, Evi; Naramura, Takuro; Myodo, Miho; Hasegawa, Masataka; Shoji, Shuichi; Buendia, Jorge J.; Kawarada, Hiroshi

2017-01-01

Here, we propose simple diamond functionalization by carboxyl termination for adenosine triphosphate (ATP) detection by an aptamer. The high-sensitivity label-free aptamer sensor for ATP detection was fabricated on nanocrystalline diamond (NCD). Carboxyl termination of the NCD surface by vacuum ultraviolet excimer laser and fluorine termination of the background region as a passivated layer were investigated by X-ray photoelectron spectroscopy. Single strand DNA (amide modification) was used as the supporting biomolecule to immobilize into the diamond surface via carboxyl termination and become a double strand with aptamer. ATP detection by aptamer was observed as a 66% fluorescence signal intensity decrease of the hybridization intensity signal. The sensor operation was also investigated by the field-effect characteristics. The shift of the drain current–drain voltage characteristics was used as the indicator for detection of ATP. From the field-effect characteristics, the shift of the drain current–drain voltage was observed in the negative direction. The negative charge direction shows that the aptamer is capable of detecting ATP. The ability of the sensor to detect ATP was investigated by fabricating a field-effect transistor on the modified NCD surface. PMID:28753998

Aptamer-Based Carboxyl-Terminated Nanocrystalline Diamond Sensing Arrays for Adenosine Triphosphate Detection.

PubMed

Suaebah, Evi; Naramura, Takuro; Myodo, Miho; Hasegawa, Masataka; Shoji, Shuichi; Buendia, Jorge J; Kawarada, Hiroshi

2017-07-21

Here, we propose simple diamond functionalization by carboxyl termination for adenosine triphosphate (ATP) detection by an aptamer. The high-sensitivity label-free aptamer sensor for ATP detection was fabricated on nanocrystalline diamond (NCD). Carboxyl termination of the NCD surface by vacuum ultraviolet excimer laser and fluorine termination of the background region as a passivated layer were investigated by X-ray photoelectron spectroscopy. Single strand DNA (amide modification) was used as the supporting biomolecule to immobilize into the diamond surface via carboxyl termination and become a double strand with aptamer. ATP detection by aptamer was observed as a 66% fluorescence signal intensity decrease of the hybridization intensity signal. The sensor operation was also investigated by the field-effect characteristics. The shift of the drain current-drain voltage characteristics was used as the indicator for detection of ATP. From the field-effect characteristics, the shift of the drain current-drain voltage was observed in the negative direction. The negative charge direction shows that the aptamer is capable of detecting ATP. The ability of the sensor to detect ATP was investigated by fabricating a field-effect transistor on the modified NCD surface.

Energy metabolism of intervertebral disc under mechanical loading.

PubMed

Wang, Chong; Gonzales, Silvia; Levene, Howard; Gu, Weiyong; Huang, Chun-Yuh Charles

2013-11-01

Intervertebral disc (IVD) degeneration is closely associated with low back pain (LBP), which is a major health concern in the U.S. Cellular biosynthesis of extracellular matrix (ECM), which is important for maintaining tissue integrity and preventing tissue degeneration, is an energy demanding process. Due to impaired nutrient support in avascular IVD, adenosine triphosphate (ATP) supply could be a limiting factor for maintaining normal ECM synthesis. Therefore, the objective of this study was to investigate the energy metabolism in the annulus fibrosus (AF) and nucleus pulposus (NP) of porcine IVD under static and dynamic compressions. Under compression, pH decreased and the contents of lactate and ATP increased significantly in both AF and NP regions, suggesting that compression can promote ATP production via glycolysis and reduce pH by increasing lactate accumulation. A high level of extracellular ATP content was detected in the NP region and regulated by compressive loading. Since ATP can serve not only as an intra-cellular energy currency, but also as a regulator of a variety of cellular activities extracellularly through the purinergic signaling pathway, our findings suggest that compression-mediated ATP metabolism could be a novel mechanobiological pathway for regulating IVD metabolism. © 2013 Orthopaedic Research Society.

Mitochondrial membrane potential and reactive oxygen species in liquid stored and cryopreserved turkey (Meleagris gallopavo) spermatozoa.

PubMed

Slowinska, M; Liszewska, E; Judycka, S; Konopka, M; Ciereszko, A

2018-06-01

The extensive use of artificial insemination in turkeys has led to the development of in vitro semen storage. However, fertility rates from liquid stored and frozen/thawed turkey semen are still unsatisfactory. The aim of the study was to assess spermatozoa viability, mitochondrial membrane potential (MMP), and reactive oxygen species production (ROS) in liquid stored and cryopreserved turkey semen with the use of flow cytometry. Moreover, motility and adenosine triphosphate (ATP) content in sperm were monitored at the same time to link flow cytometry data with sperm movement and energetics. Liquid storage led to a decrease in sperm motility (80.6 vs. 55.6%, for fresh and stored for 48 h), live sperm with an intact MMP (59.9 vs. 30.5% for fresh and stored for 48 h), and a 20-fold decrease in ATP content after 24 h of storage. A 3-fold increase in ROS+ sperm was observed after 48 h of storage (9.3 vs. 26.8% for fresh and stored for 48 h). Semen equilibration before cryopreservation affected only ATP content. However, freezing/thawing led to a dramatic decrease in all of the studied semen quality parameters. A 5-fold decrease in live sperm with intact MMP (59.8 vs. 11.9%) and a 7-fold increase in sperm ROS+ (10.8 vs. 74.4%) were recorded between fresh and frozen/thawed semen. The results strongly suggested that a significant loss of MMP and a disturbance in sperm ATP production during semen storage can be the main reason for the decline in sperm motility. The disturbance of mitochondria activity during storage seems to be associated with the increase in oxidative stress in turkey semen. Turkey sperm mitochondria also appear to be very sensitive to cryodamage. Diminished energy production in turkey spermatozoa, visible as the low percentage of sperm with an intact MMP and low level of ATP after freezing/thawing, which is associated with high ROS generation, could be responsible for the low fertilizing ability of cryopreserved turkey semen.

The Molecular Mechanism of Ethylene-Mediated Root Hair Development Induced by Phosphate Starvation

PubMed Central

Song, Li; Yu, Haopeng; Dong, Jinsong; Liu, Dong

2016-01-01

Enhanced root hair production, which increases the root surface area for nutrient uptake, is a typical adaptive response of plants to phosphate (Pi) starvation. Although previous studies have shown that ethylene plays an important role in root hair development induced by Pi starvation, the underlying molecular mechanism is not understood. In this work, we characterized an Arabidopsis mutant, hps5, that displays constitutive ethylene responses and increased sensitivity to Pi starvation due to a mutation in the ethylene receptor ERS1. hps5 accumulates high levels of EIN3 protein, a key transcription factor involved in the ethylene signaling pathway, under both Pi sufficiency and deficiency. Pi starvation also increases the accumulation of EIN3 protein. Combined molecular, genetic, and genomic analyses identified a group of genes that affect root hair development by regulating cell wall modifications. The expression of these genes is induced by Pi starvation and is enhanced in the EIN3-overexpressing line. In contrast, the induction of these genes by Pi starvation is suppressed in ein3 and ein3eil1 mutants. EIN3 protein can directly bind to the promoter of these genes, some of which are also the immediate targets of RSL4, a key transcription factor that regulates root hair development. Based on these results, we propose that under normal growth conditions, the level of ethylene is low in root cells; a group of key transcription factors, including RSL4 and its homologs, trigger the transcription of their target genes to promote root hair development; Pi starvation increases the levels of the protein EIN3, which directly binds to the promoters of the genes targeted by RSL4 and its homologs and further increase their transcription, resulting in the enhanced production of root hairs. This model not only explains how ethylene mediates root hair responses to Pi starvation, but may provide a general mechanism for how ethylene regulates root hair development under both stress and non-stress conditions. PMID:27427911

High fat, high sucrose diet causes cardiac mitochondrial dysfunction due in part to oxidative post-translational modification of mitochondrial complex II.

PubMed

Sverdlov, Aaron L; Elezaby, Aly; Behring, Jessica B; Bachschmid, Markus M; Luptak, Ivan; Tu, Vivian H; Siwik, Deborah A; Miller, Edward J; Liesa, Marc; Shirihai, Orian S; Pimentel, David R; Cohen, Richard A; Colucci, Wilson S

2015-01-01

Diet-induced obesity leads to metabolic heart disease (MHD) characterized by increased oxidative stress that may cause oxidative post-translational modifications (OPTM) of cardiac mitochondrial proteins. The functional consequences of OPTM of cardiac mitochondrial proteins in MHD are unknown. Our objective was to determine whether cardiac mitochondrial dysfunction in MHD due to diet-induced obesity is associated with cysteine OPTM. Male C57BL/6J mice were fed either a high-fat, high-sucrose (HFHS) or control diet for 8months. Cardiac mitochondria from HFHS-fed mice (vs. control diet) had an increased rate of H2O2 production, a decreased GSH/GSSG ratio, a decreased rate of complex II substrate-driven ATP synthesis and decreased complex II activity. Complex II substrate-driven ATP synthesis and complex II activity were partially restored ex-vivo by reducing conditions. A biotin switch assay showed that HFHS feeding increased cysteine OPTM in complex II subunits A (SDHA) and B (SDHB). Using iodo-TMT multiplex tags we found that HFHS feeding is associated with reversible oxidation of cysteines 89 and 231 in SDHA, and 100, 103 and 115 in SDHB. MHD due to consumption of a HFHS "Western" diet causes increased H2O2 production and oxidative stress in cardiac mitochondria associated with decreased ATP synthesis and decreased complex II activity. Impaired complex II activity and ATP production are associated with reversible cysteine OPTM of complex II. Possible sites of reversible cysteine OPTM in SDHA and SDHB were identified by iodo-TMT tag labeling. Mitochondrial ROS may contribute to the pathophysiology of MHD by impairing the function of complex II. This article is part of a Special Issue entitled "Mitochondria: From Basic Mitochondrial Biology to Cardiovascular Disease". Copyright © 2014 Elsevier Ltd. All rights reserved.

Potentiation of lead-induced cell death in PC12 cells by glutamate: Protection by N-acetylcysteine amide (NACA), a novel thiol antioxidant

DOE Office of Scientific and Technical Information (OSTI.GOV)

Penugonda, Suman; Mare, Suneetha; Lutz, P.

2006-10-15

Oxidative stress has been implicated as an important factor in many neurological diseases. Oxidative toxicity in a number of these conditions is induced by excessive glutamate release and subsequent glutamatergic neuronal stimulation. This, in turn, causes increased generation of reactive oxygen species (ROS), oxidative stress, excitotoxicity, and neuronal damage. Recent studies indicate that the glutamatergic neurotransmitter system is involved in lead-induced neurotoxicity. Therefore, this study aimed to (1) investigate the potential effects of glutamate on lead-induced PC12 cell death and (2) elucidate whether the novel thiol antioxidant N-acetylcysteine amide (NACA) had any protective abilities against such cytotoxicity. Our results suggestmore » that glutamate (1 mM) potentiates lead-induced cytotoxicity by increased generation of ROS, decreased proliferation (MTS), decreased glutathione (GSH) levels, and depletion of cellular adenosine-triphosphate (ATP). Consistent with its ability to decrease ATP levels and induce cell death, lead also increased caspase-3 activity, an effect potentiated by glutamate. Exposure to glutamate and lead elevated the cellular malondialdehyde (MDA) levels and phospholipase-A{sub 2} (PLA{sub 2}) activity and diminished the glutamine synthetase (GS) activity. NACA protected PC12 cells from the cytotoxic effects of glutamate plus lead, as evaluated by MTS assay. NACA reduced the decrease in the cellular ATP levels and restored the intracellular GSH levels. The increased levels of ROS and MDA in glutamate-lead treated cells were significantly decreased by NACA. In conclusion, our data showed that glutamate potentiated the effects of lead-induced PC12 cell death by a mechanism involving mitochondrial dysfunction (ATP depletion) and oxidative stress. NACA had a protective role against the combined toxic effects of glutamate and lead by inhibiting lipid peroxidation and scavenging ROS, thus preserving intracellular GSH.« less

Does autophagy have a license to kill mammalian cells?

PubMed

Scarlatti, F; Granata, R; Meijer, A J; Codogno, P

2009-01-01

Macroautophagy is an evolutionarily conserved vacuolar, self-digesting mechanism for cellular components, which end up in the lysosomal compartment. In mammalian cells, macroautophagy is cytoprotective, and protects the cells against the accumulation of damaged organelles or protein aggregates, the loss of interaction with the extracellular matrix, and the toxicity of cancer therapies. During periods of nutrient starvation, stimulating macroautophagy provides the fuel required to maintain an active metabolism and the production of ATP. Macroautophagy can inhibit the induction of several forms of cell death, such as apoptosis and necrosis. However, it can also be part of the cascades of events that lead to cell death, either by collaborating with other cell death mechanisms or by causing cell death on its own. Loss of the regulation of bulk macroautophagy can prime self-destruction by cells, and some forms of selective autophagy and non-canonical forms of macroautophagy have been shown to be associated with cell demise. There is now mounting evidence that autophagy and apoptosis share several common regulatory elements that are crucial in any attempt to understand the dual role of autophagy in cell survival and cell death.

ATP Dependence of Na+/H+ Exchange

PubMed Central

Demaurex, Nicolas; Romanek, Robert R.; Orlowski, John; Grinstein, Sergio

1997-01-01

We studied the ATP dependence of NHE-1, the ubiquitous isoform of the Na+/H+ antiporter, using the whole-cell configuration of the patch-clamp technique to apply nucleotides intracellularly while measuring cytosolic pH (pHi) by microfluorimetry. Na+/H+ exchange activity was measured as the Na+-driven pHi recovery from an acid load, which was imposed via the patch pipette. In Chinese hamster ovary (CHO) fibroblasts stably transfected with NHE-1, omission of ATP from the pipette solution inhibited Na+/H+ exchange. Conversely, ATP perfusion restored exchange activity in cells that had been metabolically depleted by 2-deoxy-d-glucose and oligomycin. In cells dialyzed in the presence of ATP, no “run-down” was observed even after extended periods, suggesting that the nucleotide is the only diffusible factor required for optimal NHE-1 activity. Half-maximal activation of the antiporter was obtained at ∼5 mM Mg-ATP. Submillimolar concentrations failed to sustain Na+/H+ exchange even when an ATP regenerating system was included in the pipette solution. High ATP concentrations are also known to be required for the optimal function of other cation exchangers. In the case of the Na/Ca2+ exchanger, this requirement has been attributed to an aminophospholipid translocase, or “flippase.” The involvement of this enzyme in Na+/H+ exchange was examined using fluorescent phosphatidylserine, which is actively translocated by the flippase. ATP depletion decreased the transmembrane uptake of NBD-labeled phosphatidylserine (NBD-PS), indicating that the flippase was inhibited. Diamide, an agent reported to block the flippase, was as potent as ATP depletion in reducing NBD-PS uptake. However, diamide had no effect on Na+/H+ exchange, implying that the effect of ATP is not mediated by changes in lipid distribution across the plasma membrane. K-ATP and ATPγS were as efficient as Mg-ATP in sustaining NHE-1 activity, while AMP-PNP and AMP-PCP only partially substituted for ATP. In contrast, GTPγS was ineffective. We conclude that ATP is the only soluble factor necessary for optimal activity of the NHE-1 isoform of the antiporter. Mg2+ does not appear to be essential for the stimulatory effect of ATP. We propose that two mechanisms mediate the activation of the antiporter by ATP: one requires hydrolysis and is likely an energy-dependent event. The second process does not involve hydrolysis of the γ-phosphate, excluding mediation by protein or lipid kinases. We suggest that this effect is due to binding of ATP to an as yet unidentified, nondiffusible effector that activates the antiporter. PMID:9041442

Cellular, physiological, and molecular adaptive responses of Erwinia amylovora to starvation.

PubMed

Santander, Ricardo D; Oliver, James D; Biosca, Elena G

2014-05-01

Erwinia amylovora causes fire blight, a destructive disease of rosaceous plants distributed worldwide. This bacterium is a nonobligate pathogen able to survive outside the host under starvation conditions, allowing its spread by various means such as rainwater. We studied E. amylovora responses to starvation using water microcosms to mimic natural oligotrophy. Initially, survivability under optimal (28 °C) and suboptimal (20 °C) growth temperatures was compared. Starvation induced a loss of culturability much more pronounced at 28 °C than at 20 °C. Natural water microcosms at 20 °C were then used to characterize cellular, physiological, and molecular starvation responses of E. amylovora. Challenged cells developed starvation-survival and viable but nonculturable responses, reduced their size, acquired rounded shapes and developed surface vesicles. Starved cells lost motility in a few days, but a fraction retained flagella. The expression of genes related to starvation, oxidative stress, motility, pathogenicity, and virulence was detected during the entire experimental period with different regulation patterns observed during the first 24 h. Further, starved cells remained as virulent as nonstressed cells. Overall, these results provide new knowledge on the biology of E. amylovora under conditions prevailing in nature, which could contribute to a better understanding of the life cycle of this pathogen. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Starvation and Imidacloprid Exposure Influence Immune Response by Anoplophora glabripennis (Coleoptera: Cerambycidae) to a Fungal Pathogen.

PubMed

Fisher, Joanna J; Castrillo, Louela A; Donzelli, Bruno G G; Hajek, Ann E

2017-08-01

In several insect systems, fungal entomopathogens synergize with neonicotinoid insecticides which results in accelerated host death. Using the Asian longhorned beetle, Anoplophora glabripennis (Motschulsky), an invasive woodborer inadvertently introduced into North America and Europe, we investigated potential mechanisms in the synergy between the entomopathogenic fungus Metarhizium brunneum Petch and the insecticide imidacloprid. A potential mechanism underlying this synergy could be imidacloprid's ability to prevent feeding shortly after administration. We investigated whether starvation would have an impact similar to imidacloprid exposure on the mortality of fungal-inoculated beetles. Using real-time PCR to quantify fungal load in inoculated beetles, we determined how starvation and pesticide exposure impacted beetles' ability to tolerate or resist a fungal infection. The effect of starvation and pesticide exposure on the encapsulation and melanization immune responses of the beetles was also quantified. Starvation had a similar impact on the survival of M. brunneum-inoculated beetles compared to imidacloprid exposure. The synergy, however, was not completely due to starvation, as imidacloprid reduced the beetles' melanotic encapsulation response and capsule area, while starvation did not significantly reduce these immune responses. Our results suggest that there are multiple interacting mechanisms involved in the synergy between M. brunneum and imidacloprid. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Identification of Genes in Saccharomyces cerevisiae that Are Haploinsufficient for Overcoming Amino Acid Starvation

PubMed Central

Bae, Nancy S.; Seberg, Andrew P.; Carroll, Leslie P.; Swanson, Mark J.

2017-01-01

The yeast Saccharomyces cerevisiae responds to amino acid deprivation by activating a pathway conserved in eukaryotes to overcome the starvation stress. We have screened the entire yeast heterozygous deletion collection to identify strains haploinsufficient for growth in the presence of sulfometuron methyl, which causes starvation for isoleucine and valine. We have discovered that cells devoid of MET15 are sensitive to sulfometuron methyl, and loss of heterozygosity at the MET15 locus can complicate screening the heterozygous deletion collection. We identified 138 cases of loss of heterozygosity in this screen. After eliminating the issues of the MET15 loss of heterozygosity, strains isolated from the collection were retested on sulfometuron methyl. To determine the general effect of the mutations for a starvation response, SMM-sensitive strains were tested for the ability to grow in the presence of canavanine, which induces arginine starvation, and strains that were MET15 were also tested for growth in the presence of ethionine, which causes methionine starvation. Many of the genes identified in our study were not previously identified as starvation-responsive genes, including a number of essential genes that are not easily screened in a systematic way. The genes identified span a broad range of biological functions, including many involved in some level of gene expression. Several unnamed proteins have also been identified, giving a clue as to possible functions of the encoded proteins. PMID:28209762

Comparison of the effects of continuous and pulsatile left ventricular-assist devices on ventricular unloading using a cardiac electromechanics model

PubMed Central

Lim, Ki Moo; Constantino, Jason; Gurev, Viatcheslav; Zhu, Renjun; Trayanova, Natalia A.

2012-01-01

Left ventricular-assist devices (LVADs) are used to supply blood to the body of patients with heart failure. Pressure unloading is greater for counter-pulsating LVADs than for continuous LVADs. However, several clinical trials have demonstrated that myocardial recovery is similar for both types of LVAD. This study examined the contractile energy consumption of the myocardium with continuous and counter-pulsating LVAD support to ascertain the effect of the different LVADs on myocardial recovery. We used a three-dimensional electromechanical model of canine ventricles, with models of the circulatory system and an LVAD. We compared the left ventricular peak pressure (LVPP) and contractile ATP consumption between pulsatile and continuous LVADs. With the continuous and counter-pulsating LVAD, the LVPP decreased to 46 and 10%, respectively, and contractile ATP consumption decreased to 60 and 50%. The small difference between the contractile ATP consumption of these two types of LVAD may explain the comparable effects of the two types on myocardial recovery. PMID:22076841

Role of Orthophosphate and Other Factors in the Regulation of Starch Formation in Leaves and Isolated Chloroplasts

PubMed Central

Heldt, Hans W.; Chon, Chong Ja; Maronde, Dorothea; Herold, Alice; Stankovic, Zivko S.; Walker, David A.; Kraminer, Anna; Kirk, Martha R.; Heber, Ulrich

1977-01-01

Starch synthesis in leaves was increased by phosphate starvation or by treatments which decreased cytoplasmic orthophosphate levels (such as mannose feeding). Usually less than 30% of the total carbon fixed during CO2 assimilation was incorporated into starch in spinach (Spinacia oleracea L.), spinach beet (Beta vulgaris), and tobacco (Nicotiana tabacum) leaves. In isolated spinach chloroplasts, formation of starch from CO2 was usually less than in leaves. In the absence of significant levels of 3-phosphoglycerate, concentrations of phosphate as low as 1 mm (in the medium) or 10 mm (in the stroma) almost completely inhibited starch synthesis. The inhibitory action of phosphate could be overcome by 3-phosphoglycerate. The controlling factor of starch synthesis appeared to be the ratio of phosphoglycerate to orthophosphate rather than the stromal hexose monophosphate concentration, and it is suggested that this control is exerted via the phosphate translocator and the known allosteric regulation of ADP-glucose pyrophosphorylase. Starch synthesis was also favored by the presence of dihydroxyacetone phosphate and by high light and high temperature. Oxygen was inhibitory, probably owing to carbon drain into glycolate. Starch formation by intact chloroplasts could not be promoted by added glucose or glucose 6-phosphate. Starch mobilization in the dark was promoted by orthophosphate and phosphate-dependent mobilization was inhibited by phosphoglycerate. The principal products of starch breakdown in the presence of phosphate were the transport metabolites dihydroxyacetone phosphate and 3-phosphoglycerate. Formation of these compounds from starch was stimulated by ATP or oxaloacetate. In a phosphate-independent reaction, starch was also converted to neutral products such as maltose and glucose. The rates of phosphate-dependent starch degradation phosphorolysis were very much higher than those of starch hydrolysis for which there was no phosphate requirement. PMID:16660011

[Molecular and functional diversity of ATP-sensitive K+ channels: the pathophysiological roles and potential drug targets].

PubMed

Nakaya, Haruaki; Miki, Takashi; Seino, Susumu; Yamada, Katsuya; Inagaki, Nobuya; Suzuki, Masashi; Sato, Toshiaki; Yamada, Mitsuhiko; Matsushita, Kenji; Kurachi, Yoshihisa; Arita, Makoto

2003-09-01

ATP-sensitive K(+) (K(ATP)) channels comprise the pore-forming subunit (Kir6.1 or Kir6.2) and the regulatory subunit sulfonylurea receptors (SUR1 or SUR2). K(ATP) channels with different combinations of these subunits are present in various tissues and regulate cellular functions. From the analysis of mouse models with targeted deletion of the gene encoding the pore-forming subunit Kir6.1 or Kir6.2, functional roles of K(ATP) channels in various organs have been clarified. Kir6.1(-/-) mice showed sudden death associated with ST elevation and atrioventricular block in ECG, a phenotype resembling Prinzmetal angina in humans. Kir6.2(-/-) mice were more susceptible to generalized seizure during hypoxia than wild-type (WT) mice, suggesting that neuronal K(ATP) channels, probably composed of Kir6.2 and SUR1, play a crucial role for the protection of the brain against lethal damage due to seizure. In Kir6.2(-/-) mice lacking the sarcolemmal K(ATP) channel activity in cardiac cells, ischemic preconditioning failed to reduce the infarct size, suggesting that sarcolemmal K(ATP) channels play an important role in cardioprotection against ischemia/reperfusion injuries in the heart. Mitochondrial K(ATP) channels have been also proposed to play a crucial role in cardioprotection, although the molecular identity of the channel has not been established. Nicorandil and minoxidil, K(+) channel openers activating mitochondrial K(ATP) channels, decreased the mitochondrial membrane potential, thereby preventing the Ca(2+) overload in the mitochondria of guinea-pig ventricular cells. SURs are the receptors for K(+) channel openers and the activating effects on sarcolemmal K(ATP) channels in cardiovascular tissues could be modulated by the interaction of nucleotides. Due to the molecular diversity of the accessory and pore subunits of K(ATP) channels, there would be considerable differences in the tissue selectivity of K(ATP) channel-acting drugs. Studies of Kir6.1 and Kir6.2 knockout mice indicate that K(ATP) channels are involved in the mechanisms of the protection against metabolic stress. Further clarification of physiological as well as pathophysiological roles of K(ATP) channels may lead to a new therapeutic strategy to improve the quality of life.

Ensuring US National Aeronautics Test Capabilities

NASA Technical Reports Server (NTRS)

Marshall, Timothy J.

2010-01-01

U.S. leadership in aeronautics depends on ready access to technologically advanced, efficient, and affordable aeronautics test capabilities. These systems include major wind tunnels and propulsion test facilities and flight test capabilities. The federal government owns the majority of the major aeronautics test capabilities in the United States, primarily through the National Aeronautics and Space Administration (NASA) and the Department of Defense (DoD). However, changes in the Aerospace landscape, primarily the decrease in demand for testing over the last 20 years required an overarching strategy for management of these national assets. Therefore, NASA established the Aeronautics Test Program (ATP) as a two-pronged strategic initiative to: (1) retain and invest in NASA aeronautics test capabilities considered strategically important to the agency and the nation, and (2) establish a strong, high level partnership with the DoD. Test facility utilization is a critical factor for ATP because it relies on user occupancy fees to recover a substantial part of the operations costs for its facilities. Decreasing utilization is an indicator of excess capacity and in some cases low-risk redundancy (i.e., several facilities with basically the same capability and overall low utilization). However, low utilization does not necessarily translate to lack of strategic importance. Some facilities with relatively low utilization are nonetheless vitally important because of the unique nature of the capability and the foreseeable aeronautics testing needs. Unfortunately, since its inception, the customer base for ATP has continued to shrink. Utilization of ATP wind tunnels has declined by more than 50% from the FY 2006 levels. This significant decrease in customer usage is attributable to several factors, including the overall decline in new programs and projects in the aerospace sector; the impact of computational fluid dynamics (CFD) on the design, development, and research process; and the reductions in wind tunnel testing requirements within the largest consumer of ATP wind tunnel test time, the Aeronautics Research Mission Directorate (ARMD). Retirement of the Space Shuttle Program and recent perturbations of NASA's Constellation Program will exacerbate this downward trend. Therefore it is crucial that ATP periodically revisit and determine which of its test capabilities are strategically important, which qualify as low-risk redundancies that could be put in an inactive status or closed, and address the challenges associated with both sustainment and improvements to the test capabilities that must remain active. This presentation will provide an overview of the ATP vision, mission, and goals as well as the challenges and opportunities the program is facing both today and in the future. We will discuss the strategy ATP is taking over the next five years to address the National aeronautics test capability challenges and what the program will do to capitalize on its opportunities to ensure a ready, robust and relevant portfolio of National aeronautics test capabilities.

Effects of starvation and molting on the metabolic rate of the bed bug (Cimex lectularius L.).

PubMed

DeVries, Zachary C; Kells, Stephen A; Appel, Arthur G

2015-01-01

The bed bug (Cimex lectularius L.) is a common hematophagous pest in the urban environment and is capable of surviving extended periods of starvation. However, the relationship between starvation and metabolism in bed bugs is not well understood. To better understand this relationship, we measured the metabolism of all life stages for >900 h after feeding (starvation) using closed-system respirometry. Measurements were made around molting for the immature life stages, which occurs only after a blood meal. In addition, both mated and unmated adults were measured. Starvation and molting had significant effects on the metabolism of the bed bug. Mass-specific metabolic rate (V(O2); mL g(-1) h(-1)) declined in a curvilinear fashion with the period of starvation for adults and with the postmolting period for immature bed bugs (used to standardize all immature life stages). A standard curve was developed to depict the generalized pattern of metabolic decline observed in all life stages that molted. Individual metabolic comparisons among life stages that molted revealed some differences in metabolic rate between unmated males and females. In addition, the mass scaling coefficient was found to decline with starvation time (postmolting time) for all life stages that molted. In most life stages, the ratio of V(CO2) to V(O2) (respiratory exchange ratio) declined over time, indicating a change in metabolic substrate with starvation. Finally, daily percent loss in body mass declined in a pattern similar to that of V(O2). The observed patterns in metabolic decline are evaluated in relation to the life history of bed bugs. In addition, the evolutionary development of these patterns is discussed. The metabolic pattern after feeding was also found to share several similarities with that of other ectothermic species.

Cerebral utilization of glucose, ketone bodies and oxygen in starving infant rats and the effect of intrauterine growth retardation.

PubMed

Dahlquist, G

1976-10-01

Cerebral arteriovenous differences of acetoacetate, D-beta-hydroxybutyrate, glucose, lactate and oxygen and brain DNA content was measured at 20 days of age in intrauterine growth retarded (IUGR) rats and normal littermates after 48 and 72 h of starvation. Cerebral blood flow (CBF) was measured with labeled microspheres in other comparable groups of IUGR and control rats. CBF was similar in IUGR and normal littermates (0.57+/-0.09 and 0.58+/-0.10 ml/min respectively). After 48 h of starvation, arterial glucose was significantly lower in IUGR than control animals but the arterial concentrations of ketone bodies were similar. After 48 h of starvation, cerebral arteriovenous difference of beta-hydroxybutyrate was significantly higher in control than IUGR rats also when expressed per mg brain DNA as was the fractional uptake of D-beta-hydroxybutyrate. After 72 h of starvation, arterial concentrations of ketone bodies were significantly lower in IUGR rats than controls but the fractional uptake of D-beta-hydroxybutyrate was increased compared to IUGR rats starved for 48 h. The average percentage of calculated total substrate uptake (mumol/min) accounted for by ketone bodies increased in control animals from 31.1% after 48 h of starvation to 41.0% after 72 h of starvation. In IUGR rats these percentage values were 26.5 and 25.7 respectively. After 72 h of starvation the fraction of total cerebral uptake of substrates accounted for by ketone bodies was significantly higher in control that IUGR rats. As total cerebral uptake of substrates was similar between IUGR and control animals it is concluded that IUGR rats are more dependent on glucose as a substrate for the brain during starvation.

Local potential evolutions during proton exchange membrane fuel cell operation with dead-ended anode - Part I: Impact of water diffusion and nitrogen crossover

NASA Astrophysics Data System (ADS)

Abbou, S.; Dillet, J.; Maranzana, G.; Didierjean, S.; Lottin, O.

2017-02-01

Operating a PEMFC with a dead-ended anode may lead to local fuel-starvation because of water and possibly nitrogen accumulation in the anode compartment. In previous works, we used a segmented linear cell with reference electrodes to monitor simultaneously the local potentials and current densities during dead-ended anode operation. The results indicated that water transport as well as nitrogen crossover through the membrane were most probably the two key factors governing fuel starvation. In this first from a set of two papers, we evaluated with more details the contributions of nitrogen crossover and water transport to hydrogen starvation. To assess nitrogen contribution, the fuel cell cathode compartment was first supplied with pure oxygen instead of air. The results showed that in the absence of nitrogen (in the cathode side) the fuel starvation was much slower than with air, suggesting that nitrogen contribution cannot be neglected. On the other hand, the contribution of water flooding to hydrogen starvation was investigated by using different cooling temperature on the cathode and anode sides in order to drive water toward the colder plate. The results showed that with a colder anode side, fuel starvation was faster. In the opposite case of a hotter anode plate, water accumulation in the anode compartment was limited, nitrogen crossover through the membrane was the main reason for hydrogen starvation in this case. To fully assess the impact of the thermal configurations on membrane-electrode assembly (MEA) degradation, aging protocols with a dead-ended anode and a fixed closing time were also performed. The results showed that operation with a hotter anode could help to limit significantly cathode ElectroChemical Surface Area (ECSA) losses along the cell area and performance degradation induced by hydrogen starvation.

Differential contribution of key metabolic substrates and cellular oxygen in HIF signalling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhdanov, Alexander V., E-mail: a.zhdanov@ucc.ie; Waters, Alicia H.C.; Golubeva, Anna V.

2015-01-01

Changes in availability and utilisation of O{sub 2} and metabolic substrates are common in ischemia and cancer. We examined effects of substrate deprivation on HIF signalling in PC12 cells exposed to different atmospheric O{sub 2}. Upon 2–4 h moderate hypoxia, HIF-α protein levels were dictated by the availability of glutamine and glucose, essential for deep cell deoxygenation and glycolytic ATP flux. Nuclear accumulation of HIF-1α dramatically decreased upon inhibition of glutaminolysis or glutamine deprivation. Elevation of HIF-2α levels was transcription-independent and associated with the activation of Akt and Erk1/2. Upon 2 h anoxia, HIF-2α levels strongly correlated with cellular ATP,more » produced exclusively via glycolysis. Without glucose, HIF signalling was suppressed, giving way to other regulators of cell adaptation to energy crisis, e.g. AMPK. Consequently, viability of cells deprived of O{sub 2} and glucose decreased upon inhibition of AMPK with dorsomorphin. The capacity of cells to accumulate HIF-2α decreased after 24 h glucose deprivation. This effect, associated with increased AMPKα phosphorylation, was sensitive to dorsomorphin. In chronically hypoxic cells, glutamine played no major role in HIF-2α accumulation, which became mainly glucose-dependent. Overall, the availability of O{sub 2} and metabolic substrates intricately regulates HIF signalling by affecting cell oxygenation, ATP levels and pathways involved in production of HIF-α. - Highlights: • Gln and Glc regulate HIF levels in hypoxic cells by maintaining low O{sub 2} and high ATP. • HIF-α levels under anoxia correlate with cellular ATP and critically depend on Glc. • Gln and Glc modulate activity of Akt, Erk and AMPK, regulating HIF production. • HIF signalling is differentially inhibited by prolonged Glc and Gln deprivation. • Unlike Glc, Gln plays no major role in HIF signalling in chronically hypoxic cells.« less

Oxidative stress alters mitochondrial bioenergetics and modifies pancreatic cell death independently of cyclophilin D, resulting in an apoptosis-to-necrosis shift

PubMed Central

Armstrong, Jane A.; Cash, Nicole J.; Ouyang, Yulin; Morton, Jack C.; Chvanov, Michael; Latawiec, Diane; Awais, Muhammad; Tepikin, Alexei V.; Sutton, Robert; Criddle, David N.

2018-01-01

Mitochondrial dysfunction lies at the core of acute pancreatitis (AP). Diverse AP stimuli induce Ca2+-dependent formation of the mitochondrial permeability transition pore (MPTP), a solute channel modulated by cyclophilin D (CypD), the formation of which causes ATP depletion and necrosis. Oxidative stress reportedly triggers MPTP formation and is elevated in clinical AP, but how reactive oxygen species influence cell death is unclear. Here, we assessed potential MPTP involvement in oxidant-induced effects on pancreatic acinar cell bioenergetics and fate. H2O2 application promoted acinar cell apoptosis at low concentrations (1–10 μm), whereas higher levels (0.5–1 mm) elicited rapid necrosis. H2O2 also decreased the mitochondrial NADH/FAD+ redox ratio and ΔΨm in a concentration-dependent manner (10 μm to 1 mm H2O2), with maximal effects at 500 μm H2O2. H2O2 decreased the basal O2 consumption rate of acinar cells, with no alteration of ATP turnover at <50 μm H2O2. However, higher H2O2 levels (≥50 μm) diminished spare respiratory capacity and ATP turnover, and bioenergetic collapse, ATP depletion, and cell death ensued. Menadione exerted detrimental bioenergetic effects similar to those of H2O2, which were inhibited by the antioxidant N-acetylcysteine. Oxidant-induced bioenergetic changes, loss of ΔΨm, and cell death were not ameliorated by genetic deletion of CypD or by its acute inhibition with cyclosporine A. These results indicate that oxidative stress alters mitochondrial bioenergetics and modifies pancreatic acinar cell death. A shift from apoptosis to necrosis appears to be associated with decreased mitochondrial spare respiratory capacity and ATP production, effects that are independent of CypD-sensitive MPTP formation. PMID:29626097

Exposure to a Northern Contaminant Mixture (NCM) Alters Hepatic Energy and Lipid Metabolism Exacerbating Hepatic Steatosis in Obese JCR Rats

PubMed Central

Mailloux, Ryan J.; Florian, Maria; Chen, Qixuan; Yan, Jin; Petrov, Ivan; Coughlan, Melanie C.; Laziyan, Mahemuti; Caldwell, Don; Lalande, Michelle; Patry, Dominique; Gagnon, Claude; Sarafin, Kurtis; Truong, Jocelyn; Chan, Hing Man; Ratnayake, Nimal; Li, Nanqin; Willmore, William G.; Jin, Xiaolei

2014-01-01

Non-alcoholic fatty liver disease (NAFLD), defined by the American Liver Society as the buildup of extra fat in liver cells that is not caused by alcohol, is the most common liver disease in North America. Obesity and type 2 diabetes are viewed as the major causes of NAFLD. Environmental contaminants have also been implicated in the development of NAFLD. Northern populations are exposed to a myriad of persistent organic pollutants including polychlorinated biphenyls, organochlorine pesticides, flame retardants, and toxic metals, while also affected by higher rates of obesity and alcohol abuse compared to the rest of Canada. In this study, we examined the impact of a mixture of 22 contaminants detected in Inuit blood on the development and progression of NAFLD in obese JCR rats with or without co-exposure to10% ethanol. Hepatosteatosis was found in obese rat liver, which was worsened by exposure to 10% ethanol. NCM treatment increased the number of macrovesicular lipid droplets, total lipid contents, portion of mono- and polyunsaturated fatty acids in the liver. This was complemented by an increase in hepatic total cholesterol and cholesterol ester levels which was associated with changes in the expression of genes and proteins involved in lipid metabolism and transport. In addition, NCM treatment increased cytochrome P450 2E1 protein expression and decreased ubiquinone pool, and mitochondrial ATP synthase subunit ATP5A and Complex IV activity. Despite the changes in mitochondrial physiology, hepatic ATP levels were maintained high in NCM-treated versus control rats. This was due to a decrease in ATP utilization and an increase in creatine kinase activity. Collectively, our results suggest that NCM treatment decreases hepatic cholesterol export, possibly also increases cholesterol uptake from circulation, and promotes lipid accumulation and alters ATP homeostasis which exacerbates the existing hepatic steatosis in genetically obese JCR rats with or without co-exposure to ethanol. PMID:25222487

HPS4/SABRE regulates plant responses to phosphate starvation through antagonistic interaction with ethylene signalling

PubMed Central

Yu, Hailan; Luo, Nan; Sun, Lichao; Liu, Dong

2012-01-01

The phytohormone ethylene plays important roles in regulating plant responses to phosphate (Pi) starvation. To date, however, no molecular components have been identified that interact with ethylene signalling in regulating such responses. In this work, an Arabidopsis mutant, hps4, was characterized that exhibits enhanced responses to Pi starvation, including increased inhibition of primary root growth, enhanced expression of Pi starvation-induced genes, and overproduction of root-associated acid phosphatases. Molecular cloning indicated that hps4 is a new allele of SABRE, which was previously identified as an important regulator of cell expansion in Arabidopsis. HPS4/SABRE antagonistically interacts with ethylene signalling to regulate plant responses to Pi starvation. Furthermore, it is shown that Pi-starved hps4 mutants accumulate more auxin in their root tips than the wild type, which may explain the increased inhibition of their primary root growth when grown under Pi deficiency. PMID:22615140

SU-E-T-269: Differential Hazard Analysis For Conventional And New Linac Acceptance Testing Procedures

DOE Office of Scientific and Technical Information (OSTI.GOV)

Harry, T; Yaddanapudi, S; Mutic, S

Purpose: New techniques and materials have recently been developed to expedite the conventional Linac Acceptance Testing Procedure (ATP). The new ATP method uses the Electronic Portal Imaging Device (EPID) for data collection and is presented separately. This new procedure is meant to be more efficient then conventional methods. While not clinically implemented yet, a prospective risk assessment is warranted for any new techniques. The purpose of this work is to investigate the risks and establish the pros and cons between the conventional approach and the new ATP method. Methods: ATP tests that were modified and performed with the EPID weremore » analyzed. Five domain experts (Medical Physicists) comprised the core analysis team. Ranking scales were adopted from previous publications related to TG 100. The number of failure pathways for each ATP test procedure were compared as well as the number of risk priority numbers (RPN’s) greater than 100 were compared. Results: There were fewer failure pathways with the new ATP compared to the conventional, 262 and 556, respectively. There were fewer RPN’s > 100 in the new ATP compared to the conventional, 41 and 115. Failure pathways and RPN’s > 100 for individual ATP tests on average were 2 and 3.5 times higher in the conventional ATP compared to the new, respectively. The pixel sensitivity map of the EPID was identified as a key hazard to the new ATP procedure with an RPN of 288 for verifying beam parameters. Conclusion: The significant decrease in failure pathways and RPN’s >100 for the new ATP mitigates the possibilities of a catastrophic error occurring. The Pixel Sensitivity Map determining the response and inherent characteristics of the EPID is crucial as all data and hence results are dependent on that process. Grant from Varian Medical Systems Inc.« less

A new strategy for the detection of adenosine triphosphate by aptamer/quantum dot biosensor based on chemiluminescence resonance energy transfer.

PubMed

Zhou, Zi-Ming; Yu, Yong; Zhao, Yuan-Di

2012-09-21

We designed an aptasensor for the detection of adenosine triphosphate (ATP) based on chemiluminescence resonance energy transfer (CRET). An adenosine aptamer was cut into two pieces of ssDNA, which were attached to quantum dots (QDs) and horse radish peroxidase (HRP), respectively. They could reassemble into specific structures in the presence of ATP and then decrease the distance of HRP and QDs. ATP detection can be easily realized according to the fluorescent intensity of QDs, which is excited by CRET between luminol and QDs. Results show that the concentration of ATP is linear relation with the fluorescent intensity of the peak of QDs emission and the linear range for the linear equation is from 50 μM to 231 μM and the detection limit was 185 nM. When the concentration of ATP was 2 mM, the efficiency of CRET is 13.6%. Good specificity for ATP had been demonstrated compared to thymidine triphosphate (TTP), cytidine triphosphate (CTP) and guanosine triphosphate (GTP), when 1 mM of each was added, respectively. This method needs no external light source and can avoid autofluorescence and photobleaching, and ATP can be detected selectively, specifically, and sensitively in a low micromolar range, which means that the strategy reported here can be applicable to the detection of several other target molecules.

Quantum dots electrochemical aptasensor based on three-dimensionally ordered macroporous gold film for the detection of ATP.

PubMed

Zhou, Jinjun; Huang, Haiping; Xuan, Jie; Zhang, Jianrong; Zhu, Jun-Jie

2010-10-15

A sensitive electrochemical aptasensor was successfully fabricated for the detection of adenosine triphosphate (ATP) by combining three-dimensionally ordered macroporous (3DOM) gold film and quantum dots (QDs). The 3DOM gold film was electrochemically fabricated with an inverted opal template, making the active surface area of the electrode up to 9.52 times larger than that of a classical bare flat one. 5′-thiolated ATP-binding aptamer (ABA) was first assembled onto the 3DOM gold film via sulfur–gold affinity. Then, 5′-biotinated complementary strand (BCS) was immobilized via hybridization reaction to form the DNA/DNA duplex. Since the tertiary structure of the aptamer was stabilized in the presence of target ATP, the duplex can be denatured to liberate BCS. The reaction was monitored by electrochemical stripping analysis of dissolved QDs which were bound to the residual BCS through biotin-streptavidin system. The decrease of peak current was proportional to the amount of ATP. The unique interconnected structure in 3DOM gold film along with the "built-in" preconcentration remarkably improved the sensitivity. ATP detection with high selectivity, wide linear dynamic range of 4 orders of magnitude and high sensitivity down to 0.01 nm were achieved. The results demonstrated that the novel strategy was feasible for sensitive ATP assay and provided a promising model for the detection of small molecules.

Gravity loading induces adenosine triphosphate release and phosphorylation of extracellular signal-regulated kinases in human periodontal ligament cells.

PubMed

Ito, Mai; Arakawa, Toshiya; Okayama, Miki; Shitara, Akiko; Mizoguchi, Itaru; Takuma, Taishin

2014-11-01

The periodontal ligament (PDL) receives mechanical stress (MS) from dental occlusion or orthodontic tooth movement. Mechanical stress is thought to be a trigger for remodeling of the PDL and alveolar bone, although its signaling mechanism is still unclear. So we investigated the effect of MS on adenosine triphosphate (ATP) release and extracellular signal-regulated kinases (ERK) phosphorylation in PDL cells. Mechanical stress was applied to human PDL cells as centrifugation-mediated gravity loading. Apyrase, Ca(2+)-free medium and purinergic receptor agonists and antagonists were utilized to analyze the contribution of purinergic receptors to ERK phosphorylation. Gravity loading and ATP increased ERK phosphorylation by 5 and 2.5 times, respectively. Gravity loading induced ATP release from PDL cells by tenfold. Apyrase and suramin diminished ERK phosphorylation induced by both gravity loading and ATP. Under Ca(2+)-free conditions the phosphorylation by gravity loading was partially decreased, whereas ATP-induced phosphorylation was unaffected. Receptors P2Y4 and P2Y6 were prominently expressed in the PDL cells. Gravity loading induced ATP release and ERK phosphorylation in PDL fibroblasts, and ATP signaling via P2Y receptors was partially involved in this phosphorylation, which in turn would enhance gene expression for the remodeling of PDL tissue during orthodontic tooth movement. © 2013 Wiley Publishing Asia Pty Ltd.

Determination of adenosine phosphates in rat gastrocnemius at various postmortem intervals using high performance liquid chromatography.

PubMed

Huang, Hong; Yan, Youyi; Zuo, Zhong; Yang, Lin; Li, Bin; Song, Yu; Liao, Linchuan

2010-09-01

Although the change in adenosine phosphate levels in muscles may contribute to the development of rigor mortis, the relationship between their levels and the onset and development of rigor mortis has not been well elucidated. In the current study, levels of the adenosine phosphates including adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) in gastrocnemius at various postmortem intervals of 180 rats from different death modes were detected by high performance liquid chromatography. The results showed that the levels of ATP and ADP significantly decreased along with the postmortem period of rats from different death mode whereas the AMP level remained the same. In addition, it was found that changes in the ATP levels in muscles after death correlated well with the development of rigor mortis. Therefore, the ATP level could serve as a reference parameter for the deduction of rigor mortis in forensic science.

Energy metabolism in neuroblastoma and Wilms tumor

PubMed Central

Aminzadeh, Sepideh; Vidali, Silvia; Sperl, Wolfgang; Feichtinger, René G.

2015-01-01

To support high proliferation, the majority of cancer cells undergo fundamental metabolic changes such as increasing their glucose uptake and shifting to glycolysis for ATP production at the expense of far more efficient mitochondrial energy production by oxidative phosphorylation (OXPHOS), which at first glance is a paradox. This phenomenon is known as the Warburg effect. However, enhanced glycolysis is necessary to provide building blocks for anabolic growth. Apart from the generation of ATP, intermediates of glycolysis serve as precursors for a variety of biosynthetic pathways essential for cell proliferation. In the last 10-15 years the field of tumor metabolism has experienced an enormous boom in interest. It is now well established that tumor suppressor genes and oncogenes often play a central role in the regulation of cellular metabolism. Therefore, they significantly contribute to the manifestation of the Warburg effect. While much attention has focused on adult solid tumors, so far there has been comparatively little effort directed at elucidation of the mechanism responsible for the Warburg effect in childhood cancers. In this review we focus on metabolic pathways in neuroblastoma (NB) and Wilms tumor (WT), the two most frequent solid tumors in children. Both tumor types show alterations of the OXPHOS system and glycolytic features. Chromosomal alterations and activation of oncogenes like MYC or inactivation of tumor suppressor genes like TP53 can in part explain the changes of energy metabolism in these cancers. The strict dependence of cancer cells on glucose metabolism is a fairly common feature among otherwise biologically diverse types of cancer. Therefore, inhibition of glycolysis or starvation of cancer cells through glucose deprivation via a high-fat low-carbohydrate diet may be a promising avenue for future adjuvant therapeutic strategies. PMID:26835356

How low can you go? Assessing minimum concentrations of NSC in carbon limited tree saplings

NASA Astrophysics Data System (ADS)

Hoch, Guenter; Hartmann, Henrik; Schwendener, Andrea

2016-04-01

Tissue concentrations of non-structural carbohydrates (NSC) are frequently used to determine the carbon balance of plants. Over the last years, an increasing number of studies have inferred carbon starvation in trees under environmental stress like drought from low tissue NSC concentrations. However, such inferences are limited by the fact that minimum concentrations of NSC required for survival are not known. So far, it was hypothesized that even under lethal carbon starvation, starch and low molecular sugar concentrations cannot be completely depleted and that minimum NSC concentrations at death vary across tissues and species. Here we present results of an experiment that aimed to determine minimum NSC concentrations in different tissues of saplings of two broad-leaved tree species (Acer pseudoplatanus and Quercus petratea) exposed to lethal carbon starvation via continuous darkening. In addition, we investigated recovery rates of NSC concentrations in saplings that had been darkened for different periods of time and were then re-exposed to light. Both species survived continuous darkening for about 12 weeks (confirmed by testing the ability to re-sprout after darkness). In all investigated tissues, starch concentrations declined close to zero within three to six weeks of darkness. Low molecular sugars also decreased strongly within the first weeks of darkness, but seemed to stabilize at low concentrations of 0.5 to 2 % dry matter (depending on tissue and species) almost until death. NSC concentrations recovered surprisingly fast in saplings that were re-exposed to light. After 3 weeks of continuous darkness, tissue NSC concentrations recovered within 6 weeks to levels of unshaded control saplings in all tissues and in both species. To our knowledge, this study represents the first experimental attempt to quantify minimum tissue NSC concentrations at lethal carbon starvation. Most importantly, our results suggest that carbon-starved tree saplings are able to survive several weeks without starch reserves and with extremely low sugar concentrations in all organs. Although it remains to be tested whether our findings are also valid for mature trees, these results show that NSC pools in trees are very sensitive to carbon limitation and that lethal carbon starvation is preceded by a significant (almost complete) depletion of starch and sugars in all tree organs.

The effect of aerobic exercise and starvation on growth performance and postprandial metabolic response in juvenile southern catfish (Silurus meridionalis).

PubMed

Li, Xiu-Ming; Liu, Li; Yuan, Jian-Ming; Xiao, Yuan-Yuan; Fu, Shi-Jian; Zhang, Yao-Guang

2016-03-01

To investigate the effects of aerobic exercise and starvation on growth performance, postprandial metabolic response and their interaction in a sedentary fish species, either satiation-fed or starved juvenile southern catfish (Silurus meridionalis) were exercised at 25 °C under three water velocities, i.e., nearly still water (control), 1 body length (bl) s(-1) and 2 bl s(-1), for eight weeks. Then, the feed intake (FI), food conversion efficiency (FCE), specific growth rate (SGR), morphological parameters, resting ṀO2 (ṀO2rest) and postprandial ṀO2 responses of the experimental fish were measured. Exercise at a low velocity (1 bl s(-1)) showed no effect on any growth performance parameter, whereas exercise at a high velocity (2 bl s(-1)) exhibited higher FI but similar SGR due to the extra energy expenditure from swimming and consequent decreased FCE. Starvation led to a significant body mass loss, whereas the effect intensified in both exercise groups. Exercise resulted in improved cardio-respiratory capacity, as indicated by increased gill and heart indexes, whereas it exhibited no effect on resting and postprandial metabolism in S. meridionalis. The starved fish displayed significantly larger heart, gill and digestive tract indexes compared with the feeding fish, suggesting selective maintenance of cardio-respiratory and digestive function in this fish species during starvation. However, starved fish still exhibited impaired digestive performance, as evidenced by the prolonged duration and low postprandial metabolic increase, and this effect was further exacerbated in both the 1 and 2 bl s(-1) exercise groups. These data suggest the following: (1) aerobic exercise produced no improvement in growth performance but may have led to the impairment of growth under insufficient food conditions; (2) the mass of different organs and tissues responded differently to aerobic exercise and starvation due to the different physiological roles they play; and (3) aerobic exercise had no effect on the postprandial metabolic response under a "normal feeding" situation, whereas it may have resulted in the impairment of the digestive capacity when food availability was low due to the competition of energy and oxygen under unfavorable conditions in juvenile S. meridionalis. Copyright © 2016 Elsevier Inc. All rights reserved.

Starvation effects on the hydrodynamic lubrication of rigid nonconformal contacts in combined rolling and normal motion

NASA Technical Reports Server (NTRS)

Ghosh, M. K.; Hamrock, B. J.; Brewe, D. E.

1986-01-01

The effect of inlet starvation on the hydrodynamic lubrication of lightly loaded rigid nonconformal contacts in combined rolling and normal motion is determined through a numerical solution of the Reynolds' equation for an isoviscous, incompressible lubricant. Starvation is effected by systematically reducing the fluid inlet level. The pressures are taken to be ambient at the inlet meniscus boundary and Reynolds' boundary condition is applied for film rupture in the exit region. Results are presented for the dynamic performance of the starved contacts in combined rolling and normal motion for both normal approach and separation. During normal approach the dynamic load ratio (i.e. ratio of dynamic to steady state load capacity) increases considerably with increase in the inlet starvation. The effect of starvation on the dynamic peak pressure ratio is relatively small. Further, it has been observed that with increasing starvation, film thickness effects become significant in the dynamic behavior of the nonconformal contacts. For significantly starved contacts the dynamic load ratio increases with increase in film thickness during normal approach and a similar reduction is observed during separation. A similar effect is noted for the dynamic peak pressure ratio.

Evaluating a strategy for maintaining nitrifier activity during long-term starvation in a moving bed biofilm reactor (MBBR) treating reverse osmosis concentrate.

PubMed

Ye, Liu; Hu, Shihu; Poussade, Yvan; Keller, Jurg; Yuan, Zhiguo

2012-01-01

A two-stage moving bed biofilm reactor (MBBR) was applied at the Bundamba advanced water treatment plant (AWTP) (Queensland, Australia) to treat the reverse osmosis concentrate (ROC) for inorganic nutrient removal. One of the operational challenges for the system was to cope with the large fluctuations of the ROC flow. This study investigated the decay rates of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) and biofilm detachment in MBBR during starvation for up to one month. An intermittent aeration strategy of 15 min aeration every 6 h was applied. This study also evaluated the activity recovery of both AOB and NOB after normal operation was resumed. The results showed that the activity loss of AOB and NOB was relatively minor (<20%) within 10 days of starvation, which ensured relatively quick recovery of ammonium removal when normal operation resumed. In contrast, the AOB and NOB activity loss reached 60-80% when the starvation time was longer than 20 days, resulting in slower recovery of ammonium removal after starvation. Starvation for less than 20 days didn't result in an apparent biomass detachment from carriers.

CYP2E1 overexpression inhibits microsomal Ca2+-ATPase activity in HepG2 cells.

PubMed

Caro, Andres A; Evans, Kerry L; Cederbaum, Arthur I

2009-01-31

Cytochrome P450 2E1 (CYP2E1) is a microsomal enzyme that generates reactive oxygen species during its catalytic cycle. We previously found an important role for calcium in CYP2E1-potentiated injury in HepG2 cells. The possibility that CYP2E1 may oxidatively damage and inactivate the microsomal Ca2+-ATPase in intact liver cells was evaluated, in order to explain why calcium is elevated during CYP2E1 toxicity. Microsomes were isolated by differential centrifugation from two liver cell line: E47 cells (HepG2 cells transfected with the pCI neo expression vector containing the human CYP2E1 cDNA, which overexpress active microsomal CYP2E1), and control C34 cells (HepG2 cells transfected with the pCI neo expression vector alone, which do not express significantly any cytochrome P450). The Ca2+-dependent ATPase activity was determined by measuring the accumulation of inorganic phosphate from ATP hydrolysis. CYP2E1 overexpression produced a 45% decrease in Ca2+-dependent ATPase activity (8.6 nmol Pi/min/mg protein in C34 microsomes versus 4.7 nmol Pi/min/mg protein in microsomes). Saturation curves with Ca2+ or ATP showed that CYP2E1 overexpression produced a decrease in Vmax but did not affect the Km for either Ca2+ or ATP. The decrease in activity was not associated with a decrease in SERCA protein levels. The ATP-dependent microsomal calcium uptake was evaluated by fluorimetry using fluo-3 as the fluorogenic probe. Calcium uptake rate in E47 microsomes was 28% lower than in C34 microsomes. Treatment of E47 cells with 2mM N-acetylcysteine prevented the decrease in microsomal Ca2+-ATPase found in E47 cells. These results suggest that CYP2E1 overexpression produces a decrease in microsomal Ca2+-ATPase activity in HepG2 cells mediated by reactive oxygen species. This may contribute to elevated cytosolic calcium and to CYP2E1-potentiated injury.

Enhanced Production of Adenosine Triphosphate by Pharmacological Activation of Adenosine Monophosphate-Activated Protein Kinase Ameliorates Acetaminophen-Induced Liver Injury

PubMed Central

Hwang, Jung Hwan; Kim, Yong-Hoon; Noh, Jung-Ran; Choi, Dong-Hee; Kim, Kyoung-Shim; Lee, Chul-Ho

2015-01-01

The hepatic cell death induced by acetaminophen (APAP) is closely related to cellular adenosine triphosphate (ATP) depletion, which is mainly caused by mitochondrial dysfunction. Adenosine monophosphate (AMP)-activated protein kinase (AMPK) is a key sensor of low energy status. AMPK regulates metabolic homeostasis by stimulating catabolic metabolism and suppressing anabolic pathways to increase cellular energy levels. We found that the decrease in active phosphorylation of AMPK in response to APAP correlates with decreased ATP levels, in vivo. Therefore, we hypothesized that the enhanced production of ATP via AMPK stimulation can lead to amelioration of APAP-induced liver failure. A769662, an allosteric activator of AMPK, produced a strong synergistic effect on AMPK Thr172 phosphorylation with APAP in primary hepatocytes and liver tissue. Interestingly, activation of AMPK by A769662 ameliorated the APAP-induced hepatotoxicity in C57BL/6N mice treated with APAP at a dose of 400 mg/kg intraperitoneally. However, mice treated with APAP alone developed massive centrilobular necrosis, and APAP increased their serum alanine aminotransferase and aspartate aminotransferase levels. Furthermore, A769662 administration prevented the loss of intracellular ATP without interfering with the APAP-mediated reduction of mitochondrial dysfunction. In contrast, inhibition of glycolysis by 2-deoxy-glucose eliminated the beneficial effects of A769662 on APAP-mediated liver injury. In conclusion, A769662 can effectively protect mice against APAP-induced liver injury through ATP synthesis by anaerobic glycolysis. Furthermore, stimulation of AMPK may have potential therapeutic application for APAP overdose. PMID:26434492

Hemoglobin isoform differentiation and allosteric regulation of oxygen binding in the turtle, Trachemys scripta

PubMed Central

Damsgaard, Christian; Storz, Jay F.; Hoffmann, Federico G.

2013-01-01

When freshwater turtles acclimatize to winter hibernation, there is a gradual transition from aerobic to anaerobic metabolism, which may require adjustments of blood O2 transport before turtles become anoxic. Here, we report the effects of protons, anionic cofactors, and temperature on the O2-binding properties of isolated hemoglobin (Hb) isoforms, HbA and HbD, in the turtle Trachemys scripta. We determined the primary structures of the constituent subunits of the two Hb isoforms, and we related the measured functional properties to differences in O2 affinity between untreated hemolysates from turtles that were acclimated to normoxia and anoxia. Our data show that HbD has a consistently higher O2 affinity compared with HbA, whereas Bohr and temperature effects, as well as thiol reactivity, are similar. Although sequence data show amino acid substitutions at two known β-chain ATP-binding site positions, we find high ATP affinities for both Hb isoforms, suggesting an alternative and stronger binding site for ATP. The high ATP affinities indicate that, although ATP levels decrease in red blood cells of turtles acclimating to anoxia, the O2 affinity would remain largely unchanged, as confirmed by O2-binding measurements of untreated hemolysates from normoxic and anoxic turtles. Thus, the increase in blood-O2 affinity that accompanies winter acclimation is mainly attributable to a decrease in temperature rather than in concentrations of organic phosphates. This is the first extensive study on freshwater turtle Hb isoforms, providing molecular evidence for adaptive changes in O2 transport associated with acclimation to severe hypoxia. PMID:23986362

Enhanced Production of Adenosine Triphosphate by Pharmacological Activation of Adenosine Monophosphate-Activated Protein Kinase Ameliorates Acetaminophen-Induced Liver Injury.

PubMed

Hwang, Jung Hwan; Kim, Yong-Hoon; Noh, Jung-Ran; Choi, Dong-Hee; Kim, Kyoung-Shim; Lee, Chul-Ho

2015-10-01

The hepatic cell death induced by acetaminophen (APAP) is closely related to cellular adenosine triphosphate (ATP) depletion, which is mainly caused by mitochondrial dysfunction. Adenosine monophosphate (AMP)-activated protein kinase (AMPK) is a key sensor of low energy status. AMPK regulates metabolic homeostasis by stimulating catabolic metabolism and suppressing anabolic pathways to increase cellular energy levels. We found that the decrease in active phosphorylation of AMPK in response to APAP correlates with decreased ATP levels, in vivo. Therefore, we hypothesized that the enhanced production of ATP via AMPK stimulation can lead to amelioration of APAP-induced liver failure. A769662, an allosteric activator of AMPK, produced a strong synergistic effect on AMPK Thr172 phosphorylation with APAP in primary hepatocytes and liver tissue. Interestingly, activation of AMPK by A769662 ameliorated the APAP-induced hepatotoxicity in C57BL/6N mice treated with APAP at a dose of 400 mg/kg intraperitoneally. However, mice treated with APAP alone developed massive centrilobular necrosis, and APAP increased their serum alanine aminotransferase and aspartate aminotransferase levels. Furthermore, A769662 administration prevented the loss of intracellular ATP without interfering with the APAP-mediated reduction of mitochondrial dysfunction. In contrast, inhibition of glycolysis by 2-deoxy-glucose eliminated the beneficial effects of A769662 on APAP-mediated liver injury. In conclusion, A769662 can effectively protect mice against APAP-induced liver injury through ATP synthesis by anaerobic glycolysis. Furthermore, stimulation of AMPK may have potential therapeutic application for APAP overdose.

Oxygen Transport and Root Respiration of Maize Seedlings

PubMed Central

Saglio, Pierre H.; Raymond, Philippe; Pradet, Alain

1983-01-01

Oxygen uptake and ATP/ADP ratio were simultaneously monitored during incubation of excised maize (Zea mays L. INRA 508) root tips under varying O2 partial pressure. Both variables were independent of O2 tension until a critical O2 pressure was reached. Below this pressure, ATP/ADP ratio and respiratory rate declined. However, in tissues having a high glycolytic capacity, the correlation between the ATP/ADP ratio and the respiratory rate breaks down as O2 tension decreases, due to the increasing contribution of fermentative processes. In presence of 2 millimolar NaF, the ATP/ADP ratio varied solely as a function of the O2 tension, without interference by fermentative activity, and a close correlation links the ATP/ADP ratio and the respiratory rate of excised maize root tips over the whole range of O2 tensions tested. Using this correlation, a method is proposed for the quantitative determination of the relative cellular respiratory rate permitted by O2 transport from the aerial part of young maize seedlings along the seminal root placed in an anoxic environment. Data are presented which demonstrate the preeminent part played by the cortical air spaces in O2 transport. Their contribution to respiration was high in the first few centimeters nearest the seed and decreased rapidly as the distance from the aerated source increased. It is concluded that O2 transport might contribute to the survival or to adaptive responses of root tissues in flooded soils but that the ventilation of the apical growing zone was inadequate to sustain the growth. PMID:16663116

Tomato Fruit Chromoplasts Behave as Respiratory Bioenergetic Organelles during Ripening1[W][OPEN

PubMed Central

Renato, Marta; Pateraki, Irini; Boronat, Albert; Azcón-Bieto, Joaquín

2014-01-01

During tomato (Solanum lycopersicum) fruit ripening, chloroplasts differentiate into photosynthetically inactive chromoplasts. It was recently reported that tomato chromoplasts can synthesize ATP through a respiratory process called chromorespiration. Here we show that chromoplast oxygen consumption is stimulated by the electron donors NADH and NADPH and is sensitive to octyl gallate (Ogal), a plastidial terminal oxidase inhibitor. The ATP synthesis rate of isolated chromoplasts was dependent on the supply of NAD(P)H and was fully inhibited by Ogal. It was also inhibited by the proton uncoupler carbonylcyanide m-chlorophenylhydrazone, suggesting the involvement of a chemiosmotic gradient. In addition, ATP synthesis was sensitive to 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone, a cytochrome b6f complex inhibitor. The possible participation of this complex in chromorespiration was supported by the detection of one of its components (cytochrome f) in chromoplasts using immunoblot and immunocytochemical techniques. The observed increased expression of cytochrome c6 during ripening suggests that it could act as electron acceptor of the cytochrome b6f complex in chromorespiration. The effects of Ogal on respiration and ATP levels were also studied in tissue samples. Oxygen uptake of mature green fruit and leaf tissues was not affected by Ogal, but was inhibited increasingly in fruit pericarp throughout ripening (up to 26% in red fruit). Similarly, Ogal caused a significant decrease in ATP content of red fruit pericarp. The number of energized mitochondria, as determined by confocal microscopy, strongly decreased in fruit tissue during ripening. Therefore, the contribution of chromoplasts to total fruit respiration appears to increase in late ripening stages. PMID:25125503

Dissecting nutrient-related co-expression networks in phosphate starved poplars.

PubMed

Kavka, Mareike; Polle, Andrea

2017-01-01

Phosphorus (P) is an essential plant nutrient, but its availability is often limited in soil. Here, we studied changes in the transcriptome and in nutrient element concentrations in leaves and roots of poplars (Populus × canescens) in response to P deficiency. P starvation resulted in decreased concentrations of S and major cations (K, Mg, Ca), in increased concentrations of N, Zn and Al, while C, Fe and Mn were only little affected. In roots and leaves >4,000 and >9,000 genes were differently expressed upon P starvation. These genes clustered in eleven co-expression modules of which seven were correlated with distinct elements in the plant tissues. One module (4.7% of all differentially expressed genes) was strongly correlated with changes in the P concentration in the plant. In this module the GO term "response to P starvation" was enriched with phosphoenolpyruvate carboxylase kinases, phosphatases and pyrophosphatases as well as regulatory domains such as SPX, but no phosphate transporters. The P-related module was also enriched in genes of the functional category "galactolipid synthesis". Galactolipids substitute phospholipids in membranes under P limitation. Two modules, one correlated with C and N and the other with biomass, S and Mg, were connected with the P-related module by co-expression. In these modules GO terms indicating "DNA modification" and "cell division" as well as "defense" and "RNA modification" and "signaling" were enriched; they contained phosphate transporters. Bark storage proteins were among the most strongly upregulated genes in the growth-related module suggesting that N, which could not be used for growth, accumulated in typical storage compounds. In conclusion, weighted gene coexpression network analysis revealed a hierarchical structure of gene clusters, which separated phosphate starvation responses correlated with P tissue concentrations from other gene modules, which most likely represented transcriptional adjustments related to down-stream nutritional changes and stress.

Metabolite Profiling of Low-P Tolerant and Low-P Sensitive Maize Genotypes under Phosphorus Starvation and Restoration Conditions

PubMed Central

Ganie, Arshid Hussain; Ahmad, Altaf; Pandey, Renu; Aref, Ibrahim M.; Yousuf, Peerzada Yasir; Ahmad, Sayeed; Iqbal, Muhammad

2015-01-01

Background Maize (Zea mays L.) is one of the most widely cultivated crop plants. Unavoidable economic and environmental problems associated with the excessive use of phosphatic fertilizers demands its better management. The solution lies in improving the phosphorus (P) use efficiency to sustain productivity even at low P levels. Untargeted metabolomic profiling of contrasting genotypes provides a snap shot of whole metabolome which differs under specific conditions. This information provides an understanding of the mechanisms underlying tolerance to P stress and the approach for increasing P-use-efficiency. Methodology/Principal Findings A comparative metabolite-profiling approach based on gas chromatography-mass spectrometry (GC/MS) was applied to investigate the effect of P starvation and its restoration in low-P sensitive (HM-4) and low-P tolerant (PEHM-2) maize genotypes. A comparison of the metabolite profiles of contrasting genotypes in response to P-deficiency revealed distinct differences among low-P sensitive and tolerant genotypes. Another set of these genotypes were grown under P-restoration condition and sampled at different time intervals (3, 5 and 10 days) to investigate if the changes in metabolite profile under P-deficiency was restored. Significant variations in the metabolite pools of these genotypes were observed under P-deficiency which were genotype specific. Out of 180 distinct analytes, 91 were identified. Phosphorus-starvation resulted in accumulation of di- and trisaccharides and metabolites of ammonium metabolism, specifically in leaves, but decreased the levels of phosphate-containing metabolites and organic acids. A sharp increase in the concentrations of glutamine, asparagine, serine and glycine was observed in both shoots and roots under low-P condition. Conclusion The new insights generated on the maize metabolome in resposne to P-starvation and restoration would be useful towards improvement of the P-use efficiency in maize. PMID:26090681

Disruption of copper-dependent signaling pathway in the nitrofen-induced congenital diaphragmatic hernia.

PubMed

Takahashi, Toshiaki; Friedmacher, Florian; Takahashi, Hiromizu; Hofmann, Alejandro Daniel; Puri, Prem

2015-01-01

Normal development of the fetal diaphragm requires muscularization of the diaphragm as well as the structural integrity of its underlying connective tissue components. Developmental mutations that inhibit the formation of extracellular matrix (ECM) have been shown to result in congenital diaphragmatic hernia (CDH). Copper (Cu) is an important element during diaphragm morphogenesis by participating in cross-linking of collagen and elastin fibers. Cu transport is strictly regulated by two membrane proteins: Cu-uptake transporter 1 (CTR1) and the Cu-efflux pump ATP7A. Animals lacking Cu-dependent enzymes exhibit abnormal connective tissue with diaphragmatic defects. However, the molecular basis of disruptions in Cu-mediated ECM formation in CDH remains unclear. We designed this study to investigate the hypothesis that diaphragmatic expression of CTR1 and ATP7A is decreased in the nitrofen-induced CDH model. Timed-pregnant rats were exposed to either nitrofen or vehicle on gestational day 9 (D9), and fetuses were harvested on selected time-points D15 and D18. Microdissected fetal diaphragms (n = 48) were divided into control and nitrofen-induced CDH samples (n = 12 per experimental group and time-point). Diaphragmatic gene expression levels of CTR1 and ATP7A were analyzed by quantitative real-time polymerase chain reaction. Immunohistochemistry was performed to evaluate CTR1 and ATP7A protein expression in fetal diaphragms, which was combined with specific rhodanine staining to determine diaphragmatic Cu content. Relative mRNA levels of CTR1 and ATP7A were significantly reduced in diaphragms of nitrofen-exposed fetuses on D15 (0.06 ± 0.02 vs. 0.18 ± 0.08; p < 0.05 and 0.04 ± 0.02 vs. 0.08 ± 0.02; p < 0.05) and D18 (0.10 ± 0.03 vs. 0.17 ± 0.02; p < 0.05 and 0.09 ± 0.03 vs. 0.16 ± 0.04; p < 0.05) compared to controls. Immunoreactivity of CTR1 and ATP7A was markedly decreased in the malformed diaphragmatic ECM of nitrofen-exposed fetuses on D15 and D18, which was associated with a significantly decreased diaphragmatic Cu content on D15 (7.22 ± 2.91 vs. 17.50 ± 3.09; p < 0.05) and D18 (17.60 ± 3.54 vs. 28.20 ± 4.63; p < 0.05) compared to controls. Reduced diaphragmatic expression of CTR1 and ATP7A during morphogenesis may impair the activity of Cu-dependent enzymes and thus contribute to defective ECM during diaphragmatic development.

Effects of H2 and Formate on Growth Yield and Regulation of Methanogenesis in Methanococcus maripaludis

PubMed Central

Costa, Kyle C.; Yoon, Sung Ho; Pan, Min; Burn, June A.; Baliga, Nitin S.

2013-01-01

Hydrogenotrophic methanogenic Archaea are defined by an H2 requirement for growth. Despite this requirement, many hydrogenotrophs are also capable of growth with formate as an electron donor for methanogenesis. While certain responses of these organisms to hydrogen availability have been characterized, responses to formate starvation have not been reported. Here we report that during continuous culture of Methanococcus maripaludis under defined nutrient conditions, growth yields relative to methane production decreased markedly with either H2 excess or formate excess. Analysis of the growth yields of several mutants suggests that this phenomenon occurs independently of the storage of intracellular carbon or a transcriptional response to methanogenesis. Using microarray analysis, we found that the expression of genes encoding coenzyme F420-dependent steps of methanogenesis, including one of two formate dehydrogenases, increased with H2 starvation but with formate occurred at high levels regardless of limitation or excess. One gene, encoding H2-dependent methylene-tetrahydromethanopterin dehydrogenase, decreased in expression with either H2 limitation or formate limitation. Expression of genes for the second formate dehydrogenase, molybdenum-dependent formylmethanofuran dehydrogenase, and molybdenum transport increased specifically with formate limitation. Of the two formate dehydrogenases, only the first could support growth on formate in batch culture where formate was in excess. PMID:23335420

Influence of temperature acclimation and gut content on the supercooling ability of the land snail Cornu aspersum.

PubMed

Ansart, Armelle; Aulne, Pierre-Aymeric; Madec, Luc; Vernon, Philippe

2008-05-01

The invasive land snail Cornu aspersum possesses a low ability to supercool (c. -5 degrees C in winter) and survives only minimal ice formation in its body fluids, what may limit its expansion to colder environments. In the present study, we investigated the influence of acclimation and starvation on its supercooling ability. During eight weeks, individuals were maintained at 20 degrees C, fed or starved, or placed at 5 degrees C, directly or with progressive acclimation to cold and shorter photoperiod. Temperature of crystallisation of whole individual (Tc(I)) and hemolymph (Tc(H)), mass data and gut content were recorded every two weeks. Hemolymphatic glucose and glycerol were measured at the end of experiment and occurrence of intestinal ice-nucleating agents (INA) was researched. Acclimation had no effect on Tc(I) but stimulated purging of the gut. Starvation induced a slight decrease of Tc(I) whereas a high quantity of alimentary particles in the digestive tract limited the supercooling ability. Glucose and glycerol were not synthesized in cold conditions. Mean Tc(H) was low (c. -17 degrees C), some INA being present in hemolymph of fed animals. Intestinal content of starved individuals exhibited a mean Tc of c. -6 degrees C, decreasing to c. -12 degrees after heating, suggesting the presence of organic INA.

Central roles of iron in the regulation of oxidative stress in the yeast Saccharomyces cerevisiae.

PubMed

Matsuo, Ryo; Mizobuchi, Shogo; Nakashima, Maya; Miki, Kensuke; Ayusawa, Dai; Fujii, Michihiko

2017-10-01

Oxygen is essential for aerobic organisms but causes cytotoxicity probably through the generation of reactive oxygen species (ROS). In this study, we screened for the genes that regulate oxidative stress in the yeast Saccharomyces cerevisiae, and found that expression of CTH2/TIS11 caused an increased resistance to ROS. CTH2 is up-regulated upon iron starvation and functions to remodel metabolism to adapt to iron starvation. We showed here that increased resistance to ROS by CTH2 would likely be caused by the decreased ROS production due to the decreased activity of mitochondrial respiration, which observation is consistent with the fact that CTH2 down-regulates the mitochondrial respiratory proteins. We also found that expression of CTH1, a paralog of CTH2, also caused an increased resistance to ROS. This finding supported the above view, because mitochondrial respiratory proteins are the common targets of CTH1 and CTH2. We further showed that supplementation of iron in medium augmented the growth of S. cerevisiae under oxidative stress, and expression of CTH2 and supplementation of iron collectively enhanced its growth under oxidative stress. Since CTH2 is regulated by iron, these findings suggested that iron played crucial roles in the regulation of oxidative stress in S. cerevisiae.

The Lumenal Loop Met672–Pro707 of Copper-transporting ATPase ATP7A Binds Metals and Facilitates Copper Release from the Intramembrane Sites*

PubMed Central

Barry, Amanda N.; Otoikhian, Adenike; Bhatt, Sujata; Shinde, Ujwal; Tsivkovskii, Ruslan; Blackburn, Ninian J.; Lutsenko, Svetlana

2011-01-01

The copper-transporting ATPase ATP7A has an essential role in human physiology. ATP7A transfers the copper cofactor to metalloenzymes within the secretory pathway; inactivation of ATP7A results in an untreatable neurodegenerative disorder, Menkes disease. Presently, the mechanism of ATP7A-mediated copper release into the secretory pathway is not understood. We demonstrate that the characteristic His/Met-rich segment Met672–Pro707 (HM-loop) that connects the first two transmembrane segments of ATP7A is important for copper release. Mutations within this loop do not prevent the ability of ATP7A to form a phosphorylated intermediate during ATP hydrolysis but inhibit subsequent dephosphorylation, a step associated with copper release. The HM-loop inserted into a scaffold protein forms two structurally distinct binding sites and coordinates copper in a mixed His-Met environment with an ∼2:1 stoichiometry. Binding of either copper or silver, a Cu(I) analog, induces structural changes in the loop. Mutations of 4 Met residues to Ile or two His-His pairs to Ala-Gly decrease affinity for copper. Altogether, the data suggest a two-step process, where copper released from the transport sites binds to the first His(Met)2 site, triggering a structural change and binding to a second 2-coordinate His-His or His-Met site. We also show that copper binding within the HM-loop stabilizes Cu(I) and protects it from oxidation, which may further aid the transfer of copper from ATP7A to acceptor proteins. The mechanism of copper entry into the secretory pathway is discussed. PMID:21646353

The ADP/ATP Carrier and Its Relationship to Oxidative Phosphorylation in Ancestral Protist Trypanosoma brucei

PubMed Central

Gnipová, Anna; Šubrtová, Karolína; Panicucci, Brian; Horváth, Anton; Lukeš, Julius

2015-01-01

The highly conserved ADP/ATP carrier (AAC) is a key energetic link between the mitochondrial (mt) and cytosolic compartments of all aerobic eukaryotic cells, as it exchanges the ATP generated inside the organelle for the cytosolic ADP. Trypanosoma brucei, a parasitic protist of medical and veterinary importance, possesses a single functional AAC protein (TbAAC) that is related to the human and yeast ADP/ATP carriers. However, unlike previous studies performed with these model organisms, this study showed that TbAAC is most likely not a stable component of either the respiratory supercomplex III+IV or the ATP synthasome but rather functions as a physically separate entity in this highly diverged eukaryote. Therefore, TbAAC RNA interference (RNAi) ablation in the insect stage of T. brucei does not impair the activity or arrangement of the respiratory chain complexes. Nevertheless, RNAi silencing of TbAAC caused a severe growth defect that coincides with a significant reduction of mt ATP synthesis by both substrate and oxidative phosphorylation. Furthermore, TbAAC downregulation resulted in a decreased level of cytosolic ATP, a higher mt membrane potential, an elevated amount of reactive oxygen species, and a reduced consumption of oxygen in the mitochondria. Interestingly, while TbAAC has previously been demonstrated to serve as the sole ADP/ATP carrier for ADP influx into the mitochondria, our data suggest that a second carrier for ATP influx may be present and active in the T. brucei mitochondrion. Overall, this study provides more insight into the delicate balance of the functional relationship between TbAAC and the oxidative phosphorylation (OXPHOS) pathway in an early diverged eukaryote. PMID:25616281

Mutations in valosin-containing protein (VCP) decrease ADP/ATP translocation across the mitochondrial membrane and impair energy metabolism in human neurons

PubMed Central

Arber, Charles; Bartolome, Fernando; de Vicente, Macarena; Houlden, Henry

2017-01-01

Mutations in the gene encoding valosin-containing protein (VCP) lead to multisystem proteinopathies including frontotemporal dementia. We have previously shown that patient-derived VCP mutant fibroblasts exhibit lower mitochondrial membrane potential, uncoupled respiration, and reduced ATP levels. This study addresses the underlying basis for mitochondrial uncoupling using VCP knockdown neuroblastoma cell lines, induced pluripotent stem cells (iPSCs), and iPSC-derived cortical neurons from patients with pathogenic mutations in VCP. Using fluorescent live cell imaging and respiration analysis we demonstrate a VCP mutation/knockdown-induced dysregulation in the adenine nucleotide translocase, which results in a slower rate of ADP or ATP translocation across the mitochondrial membranes. This deregulation can explain the mitochondrial uncoupling and lower ATP levels in VCP mutation-bearing neurons via reduced ADP availability for ATP synthesis. This study provides evidence for a role of adenine nucleotide translocase in the mechanism underlying altered mitochondrial function in VCP-related degeneration, and this new insight may inform efforts to better understand and manage neurodegenerative disease and other proteinopathies. PMID:28360103

A multifunctional label-free electrochemical impedance biosensor for Hg(2+), adenosine triphosphate and thrombin.

PubMed

Chen, Lifen; Chen, Zhong-Ning

2015-01-01

A multifunctional label-free biosensor for the detection of Hg(2+), adenosine triphosphate and thrombin has been developed based on the changing of the electrochemical impedance spectroscopy (EIS) from the modified electrodes when nucleic acid subunits interacting with different targets. The modified electrode consists of three interaction sections, including DNA with T-T mismatch recognizing Hg(2+) to form T-Hg(2+)-T complex, split DNA chip against ATP, and DNA domin against thrombin to form G-quadruplex. Upon DNA interaction with thrombin or ATP, an increased charge transfer resistance (Rct) had been detected. However, a decreased Rct against Hg(2+) was obtained. The Rct difference (ΔRct) has relationship with the concentration of the different targets, Hg(2+), ATP and thrombin can be selectively detected with the detection limit of 0.03, 0.25, and 0.20 nmol L(-1), respectively. To separately detect the three analytes existing in the same sample, ATP aptamer, G-rich DNA strands and EDTA were applied to mask ATP, Hg(2+) or thrombin separately. Copyright © 2014 Elsevier B.V. All rights reserved.

Inhibition of aldolase A blocks biogenesis of ATP and attenuates Japanese encephalitis virus production.

PubMed

Tien, Chih-Feng; Cheng, Shih-Ching; Ho, Yen-Peng; Chen, Yi-Shiuan; Hsu, Jung-Hsin; Chang, Ruey-Yi

2014-01-10

Viral replication depends on host proteins to supply energy and replication accessories for the sufficient production of viral progeny. In this study, we identified fructose-bisphosphate aldolase A as a binding partner of Japanese encephalitis virus (JEV) untranslated regions (UTRs) on the antigenome via RNA affinity capture and mass spectrometry. Direct interaction of aldolase A with JEV RNAs was confirmed by gel mobility shift assay and colocalization with active replication of double-stranded RNA in JEV-infected cells. Infection of JEV caused an increase in aldolase A expression of up to 33%. Knocking down aldolase A reduced viral translation, genome replication, and viral production significantly. Furthermore, JEV infection consumed 50% of cellular ATP, and the ATP level decreased by 70% in the aldolase A-knockdown cells. Overexpression of aldolase A in aldolase A-knockdown cells increased ATP levels significantly. Taken together, these results indicate that JEV replication requires aldolase A and consumes ATP. This is the first report of direct involvement of a host metabolic enzyme, aldolase A protein, in JEV replication. Copyright © 2013 Elsevier Inc. All rights reserved.