Prenylated flavonol glycosides Epimedium grandiflorum: Cytotoxicity and evaluation against inflammation and metabolic disorders

USDA-ARS?s Scientific Manuscript database

Two new prenylated flavonol glycosides, epimedigrandiosides A and B (1 and 2), and 28 previously known compounds including prenylated flavonol derivatives, flavonol glycoside, megastigmanes, phenyl alkanoids, sesquiterpenoid glycoside, lignan, and hexene glucoside were isolated from the methanol ext...

Expression of Genes Involved in Anthocyanin Biosynthesis in Relation to Anthocyanin, Proanthocyanidin, and Flavonol Levels during Bilberry Fruit Development1

PubMed Central

Jaakola, Laura; Määttä, Kaisu; Pirttilä, Anna Maria; Törrönen, Riitta; Kärenlampi, Sirpa; Hohtola, Anja

2002-01-01

The production of anthocyanins in fruit tissues is highly controlled at the developmental level. We have studied the expression of flavonoid biosynthesis genes during the development of bilberry (Vaccinium myrtillus) fruit in relation to the accumulation of anthocyanins, proanthocyanidins, and flavonols in wild berries and in color mutants of bilberry. The cDNA fragments of five genes from the flavonoid pathway, phenylalanine ammonia-lyase, chalcone synthase, flavanone 3-hydroxylase, dihydroflavonol 4-reductase, and anthocyanidin synthase, were isolated from bilberry using the polymerase chain reaction technique, sequenced, and labeled with a digoxigenin-dUTP label. These homologous probes were used for determining the expression of the flavonoid pathway genes in bilberries. The contents of anthocyanins, proanthocyanidins, and flavonols in ripening bilberries were analyzed with high-performance liquid chromatography-diode array detector and were identified using a mass spectrometry interface. Our results demonstrate a correlation between anthocyanin accumulation and expression of the flavonoid pathway genes during the ripening of berries. At the early stages of berry development, procyanidins and quercetin were the major flavonoids, but the levels decreased dramatically during the progress of ripening. During the later stages of ripening, the content of anthocyanins increased strongly and they were the major flavonoids in the ripe berry. The expression of flavonoid pathway genes in the color mutants of bilberry was reduced. A connection between flavonol and anthocyanin synthesis in bilberry was detected in this study and also in previous data collected from flavonol and anthocyanin analyses from other fruits. In accordance with this, models for the connection between flavonol and anthocyanin syntheses in fruit tissues are presented. PMID:12376640

Flavonols Accumulate Asymmetrically and Affect Auxin Transport in Arabidopsis1[C][W][OA

PubMed Central

Kuhn, Benjamin M.; Geisler, Markus; Bigler, Laurent; Ringli, Christoph

2011-01-01

Flavonoids represent a class of secondary metabolites with diverse functions in plants including ultraviolet protection, pathogen defense, and interspecies communication. They are also known as modulators of signaling processes in plant and animal systems and therefore are considered to have beneficial effects as nutraceuticals. The rol1-2 (for repressor of lrx1) mutation of Arabidopsis (Arabidopsis thaliana) induces aberrant accumulation of flavonols and a cell-growth phenotype in the shoot. The hyponastic cotyledons, aberrant shape of pavement cells, and deformed trichomes in rol1-2 mutants are suppressed by blocking flavonoid biosynthesis, suggesting that the altered flavonol accumulation in these plants induces the shoot phenotype. Indeed, the identification of several transparent testa, myb, and fls1 (for flavonol synthase1) alleles in a rol1-2 suppressor screen provides genetic evidence that flavonols interfere with shoot development in rol1-2 seedlings. The increased accumulation of auxin in rol1-2 seedlings appears to be caused by a flavonol-induced modification of auxin transport. Quantification of auxin export from mesophyll protoplasts revealed that naphthalene-1-acetic acid but not indole-3-acetic acid transport is affected by the rol1-2 mutation. Inhibition of flavonol biosynthesis in rol1-2 fls1-3 restores naphthalene-1-acetic acid transport to wild-type levels, indicating a very specific mode of action of flavonols on the auxin transport machinery. PMID:21502189

Three-dimensional quantitative structure-activity relationship studies on UGT1A9-mediated 3-O-glucuronidation of natural flavonols using a pharmacophore-based comparative molecular field analysis model.

PubMed

Wu, Baojian; Morrow, John Kenneth; Singh, Rashim; Zhang, Shuxing; Hu, Ming

2011-02-01

Glucuronidation is often recognized as one of the rate-determining factors that limit the bioavailability of flavonols. Hence, design and synthesis of more bioavailable flavonols would benefit from the establishment of predictive models of glucuronidation using kinetic parameters [e.g., K(m), V(max), intrinsic clearance (CL(int)) = V(max)/K(m)] derived for flavonols. This article aims to construct position (3-OH)-specific comparative molecular field analysis (CoMFA) models to describe UDP-glucuronosyltransferase (UGT) 1A9-mediated glucuronidation of flavonols, which can be used to design poor UGT1A9 substrates. The kinetics of recombinant UGT1A9-mediated 3-O-glucuronidation of 30 flavonols was characterized, and kinetic parameters (K(m), V(max), CL(int)) were obtained. The observed K(m), V(max), and CL(int) values of 3-O-glucuronidation ranged from 0.04 to 0.68 μM, 0.04 to 12.95 nmol/mg/min, and 0.06 to 109.60 ml/mg/min, respectively. To model UGT1A9-mediated glucuronidation, 30 flavonols were split into the training (23 compounds) and test (7 compounds) sets. These flavonols were then aligned by mapping the flavonols to specific common feature pharmacophores, which were used to construct CoMFA models of V(max) and CL(int), respectively. The derived CoMFA models possessed good internal and external consistency and showed statistical significance and substantive predictive abilities (V(max) model: q(2) = 0.738, r(2) = 0.976, r(pred)(2) = 0.735; CL(int) model: q(2) = 0.561, r(2) = 0.938, r(pred)(2) = 0.630). The contour maps derived from CoMFA modeling clearly indicate structural characteristics associated with rapid or slow 3-O-glucuronidation. In conclusion, the approach of coupling CoMFA analysis with a pharmacophore-based structural alignment is viable for constructing a predictive model for regiospecific glucuronidation rates of flavonols by UGT1A9.

Flavonol Glycosides from Gaura Biennis

USDA-ARS?s Scientific Manuscript database

Phytochemical investigation of the native American plant Gaura biennis led to the isolation of three new flavonol glycosides (1-3), along with eight known ones. Their structures were established primarily by spectroscopic data as quercetin 3-O-(2"-O-a-L-rhamnopyranosyl-6"-O-E-p-coumaroyl)-ß-D- gluco...

Fluorescent analysis of interaction of flavonols with hemoglobin and bovine serum albumin

NASA Astrophysics Data System (ADS)

Sentchouk, V. V.; Bondaryuk, E. V.

2007-09-01

We have studied the fluorescent properties of flavonols (quercetin, fisetin, morin, rutin) with the aim of studying possible interaction with hemoglobin and bovine serum albumin (BSA). We observed an increase in the intensity of intrinsic fluorescence for all the flavonols except rutin in the presence of BSA. From the changes in the fluorescence spectra, we concluded that tautomeric forms are formed on interaction with hemoglobin. We determined the interconnection between the structure of related flavonols and their fluorescent properties on interaction with proteins, and we determined the binding constants for binding with BSA and hemoglobin.

Optimization of β-cyclodextrin-based flavonol extraction from apple pomace using response surface methodology.

PubMed

Parmar, Indu; Sharma, Sowmya; Rupasinghe, H P Vasantha

2015-04-01

The present study investigated five cyclodextrins (CDs) for the extraction of flavonols from apple pomace powder and optimized β-CD based extraction of total flavonols using response surface methodology. A 2(3) central composite design with β-CD concentration (0-5 g 100 mL(-1)), extraction temperature (20-72 °C), extraction time (6-48 h) and second-order quadratic model for the total flavonol yield (mg 100 g(-1) DM) was selected to generate the response surface curves. The optimal conditions obtained were: β-CD concentration, 2.8 g 100 mL(-1); extraction temperature, 45 °C and extraction time, 25.6 h that predicted the extraction of 166.6 mg total flavonols 100 g(-1) DM. The predicted amount was comparable to the experimental amount of 151.5 mg total flavonols 100 g(-1) DM obtained from optimal β-CD based parameters, thereby giving a low absolute error and adequacy of fitted model. In addition, the results from optimized extraction conditions showed values similar to those obtained through previously established solvent based sonication assisted flavonol extraction procedure. To the best of our knowledge, this is the first study to optimize aqueous β-CD based flavonol extraction which presents an environmentally safe method for value-addition to under-utilized bio resources.

Fisetin, a bioactive flavonol, attenuates allergic airway inflammation through negative regulation of NF-κB.

PubMed

Goh, Fera Y; Upton, Nadine; Guan, Shouping; Cheng, Chang; Shanmugam, Muthu K; Sethi, Gautam; Leung, Bernard P; Wong, W S Fred

2012-03-15

Persistent activation of nuclear factor-κB (NF-κB) has been associated with the development of asthma. Fisetin (3,7,3',4'-tetrahydroxyflavone), a naturally occurring bioactive flavonol, has been shown to inhibit NF-κB activity. We hypothesized that fisetin may attenuate allergic asthma via negative regulation of the NF-κB activity. Female BALB/c mice sensitized and challenged with ovalbumin developed airway inflammation. Bronchoalveolar lavage fluid was assessed for total and differential cell counts, and cytokine and chemokine levels. Lung tissues were examined for cell infiltration and mucus hypersecretion, and the expression of inflammatory biomarkers. Airway hyperresponsiveness was monitored by direct airway resistance analysis. Fisetin dose-dependently inhibited ovalbumin-induced increases in total cell count, eosinophil count, and IL-4, IL-5 and IL-13 levels recovered in bronchoalveolar lavage fluid. It attenuated ovalbumin-induced lung tissue eosinophilia and airway mucus production, mRNA expression of adhesion molecules, chitinase, IL-17, IL-33, Muc5ac and inducible nitric oxide synthase in lung tissues, and airway hyperresponsiveness to methacholine. Fisetin blocked NF-κB subunit p65 nuclear translocation and DNA-binding activity in the nuclear extracts from lung tissues of ovalbumin-challenged mice. In normal human bronchial epithelial cells, fisetin repressed TNF-α-induced NF-κB-dependent reporter gene expression. Our findings implicate a potential therapeutic value of fisetin in the treatment of asthma through negative regulation of NF-κB pathway. Copyright © 2012 Elsevier B.V. All rights reserved.

Effect of flavonol and its dimethoxy derivatives on paclitaxel-induced peripheral neuropathy in mice.

PubMed

Sayeli, Vijaykumar; Nadipelly, Jagan; Kadhirvelu, Parimala; Cheriyan, Binoy Varghese; Shanmugasundaram, Jaikumar; Subramanian, Viswanathan

2018-04-13

Peripheral neuropathy is the dose limiting side effect of many anticancer drugs. Flavonoids exhibit good antinociceptive effect in animal models. Their efficacy against different types of nociception has been documented. The present study investigated the effect of flavonol (3-hydroxy flavone), 3',4'-dimethoxy flavonol, 6,3'-dimethoxy flavonol, 7,2'-dimethoxy flavonol and 7,3'-dimethoxy flavonol against paclitaxel-induced peripheral neuropathy in mice. A single dose of paclitaxel (10 mg/kg, i.p.) was administered to induce peripheral neuropathy in mice and the manifestations of peripheral neuropathy such as tactile allodynia, cold allodynia and thermal hyperalgesia were assessed 24 h later by employing Von Frey hair aesthesiometer test, acetone bubble test and hot water tail immersion test, respectively. The test compounds were prepared as a suspension in 0.5% carboxymethyl cellulose and were administered s.c. in various doses (25, 50, 100 and 200 mg/kg). The above behavioral responses were assessed prior to and 30 min after drug treatment. In addition, the effect of test compounds on proinflammatory cytokines like tumor necrosis factor-alpha (TNF-α), interleukin-1-beta (IL-1β) and free radicals was investigated by using suitable in vitro assays. A dose-dependent attenuation of tactile allodynia, cold allodynia and thermal hyperalgesia was evidenced in mice treated with flavonol derivatives. The test compounds inhibited TNF-α, IL-1β and free radicals in a concentration-dependent manner. These results revealed that flavonol and its dimethoxy derivatives ameliorated the manifestations of paclitaxel-induced peripheral neuropathy in mice. The inhibition of proinflammatory cytokines and free radicals could contribute to this beneficial effect.

Quantification of flavonol glycosides in Camellia sinensis by MRM mode of UPLC-QQQ-MS/MS.

PubMed

Wu, Yahui; Jiang, Xiaolan; Zhang, Shuxiang; Dai, Xinlong; Liu, Yajun; Tan, Huarong; Gao, Liping; Xia, Tao

2016-04-01

Phenolic compounds are major components of tea flavour, in which catechins and flavonol glycosides play important roles in the astringent taste of tea infusion. However, the flavonol glycosides are difficult to quantify because of the large variety, as well as the inefficient seperation on chromatography. In this paper, a total of 15 flavonol glycosides in the tea plant (Camellia sinensis) were identified by the high performance liquid chromatography (HPLC) coupled to a time-of-flight mass spectrometer (TOF-MS), and a quantitative method was established based on multiple reaction monitoring (MRM) mode of ultra-high performance liquid chromatography (UPLC) coupled to a triple quadrupole mass spectrometer (QQQ-MS/MS). It provided the limit of detection and quantification to the order of picogram, which was more sensitive than the HPLC detection of the order of nanogram. The relative standard deviations of the intra- and inter-day variations in retention time and signal intensity (peak area) of six analytes were less than 0.26% and 4%, respectively. The flavonol glycosides of four tea cultivars were relatively quantified using the signal intensity (peak area) of product ion, in which six flavonol glycosides were quantified by the authentic standards. The results showed that the flavonol mono-, di- and tri-glycoside mostly accumulated in young leaves of the four tea cultivars. Notably, the myricetin 3-O-galactoside was the major component among the six flavonol glycosides detected. Copyright © 2016 Elsevier B.V. All rights reserved.

SEPARATION OF THE MINOR FLAVONOLS FROM FLOS GOSSYPII BY HIGH-SPEED COUNTERCURRENT CHROMATOGRAPHY

PubMed Central

Yang, Yi; Zhao, Yongxin; Gu, Dongyu; Ayupbek, Amatjan; Huang, Yun; Dou, Jun; Ito, Yoichiro; Zhang, Tianyou; Aisa, Haji Akber

2010-01-01

An effective high-speed countercurrent chromatography (HSCCC) method was established for further separation and purification of four minor flavonols in addition to five major flavonols which were reported by our previous study from extracts of Flos Gossypii. HSCCC was performed with three two-phase solvent systems composed of n-hexane-ethyl acetate-methanol-water (7.5:15:6:7, v/v), (2.5:15:2:7, v/v) and (0:1:0:1, v/v). The separation was repeated 3 times, and 3.8 mg of 8-methoxyl-kaempferol-7-O-β-D-rhamnoside (HPLC purity 98.27%), 6.7 mg of astragalin (HPLC purity 94.18%), 3.3 mg of 4′-methoxyl-quercetin-7-O-β-D-glucoside (HPLC purity 94.30%) and 8.2 mg of hyperoside (HPLC purity 93.48%) were separated from 150 mg of the crude sample. The chemical structures of the flavonols were confirmed by MS, 1H NMR and 13C NMR. Meanwhile, the results indicated that the target compound with smaller K value (<0.5) can be separated by increasing column length of HSCCC. And four separation rules of flavonols according to the present study and references were summarized, which can be used as a useful guide for separation of flavonols by HSCCC. PMID:21494318

Cytotoxicity of flavones and flavonols to a human esophageal squamous cell carcinoma cell line (KYSE-510) by induction of G2/M arrest and apoptosis.

PubMed

Zhang, Qiang; Zhao, Xin-Huai; Wang, Zhu-Jun

2009-08-01

In this study, cytotoxic effects of structurally related flavones and flavonols on a human esophageal squamous cell carcinoma cell line (KYSE-510) were determined, and the molecular mechanisms responsible for their cytotoxic effects were studied. The results of MTT assay showed that flavones (luteolin, apigenin, chrysin) and flavonols (quercetin, kaempferol, myricetin) were able to induce cytotoxicity in KYSE-510 cells in a dose- and time-dependent manner, and the cytotoxic potency of these compounds was in the order of: luteolin>quercetin>chrysin>kaempferol>apigenin>myricetin. Flow cytometry and DNA fragmentation analysis indicated that the cytotoxicity induced by flavones and flavonols was mediated by G(2)/M cell cycle arrest and apoptosis. Furthermore, the expression of genes related to cell cycle arrest and apoptosis was assessed by oligonucleotide microarray, real-time RT-PCR and Western blot. It was shown that the treatment of KYSE-510 cells with these compounds caused G(2)/M arrest through up-regulation of p21(waf1) and down-regulation of cyclin B1 at the mRNA and protein levels, and induced p53-independent mitochondrial-mediated apoptosis through up-regulation of PIG3 and cleavage of caspase-9 and caspase-3. The results of western blot analysis further showed that increases of p63 and p73 protein translation or stability might be contributed to the regulation of p21(waf1), cyclin B1 and PIG3.

Regulation of Staphylococcal Penicillinase Synthesis

PubMed Central

Imsande, John; Zyskind, Judith W.; Mile, Imre

1972-01-01

5-Methyl tryptophan was found to be an efficient inducer of penicillinase synthesis in Staphylococcus aureus. Addition of actinomycin D or tryptophan to the culture medium shuts off the 5-methyl tryptophan-induced synthesis of penicillinase with an apparent half-life of approximately 1 to 2 min, respectively. Hence, in the induction of penicillinase synthesis, 5-methyl tryptophan seems to function as a structural analogue of penicillin rather than by becoming incorporated in proteins and thereby creating faulty penicillinase repressor or antirepressor. This conclusion is supported by similarities in the structures of the two compounds as revealed by solid atomic models. The fact that S. aureus exposed to 14C-penicillin in the absence of protein synthesis failed to synthesize penicillinase at an increased level when cell growth was resumed strongly suggests that a protein involved in the regulation of penicillinase synthesis must be synthesized in the presence of the penicillinase inducer. In turn, this observation suggests that the penicillinase inducer promotes penicillinase synthesis by directing the penicillinase regulatory protein (i.e., the penicillinase antirepressor) to acquire a different conformation when it is synthesized in the presence of the penicillinase inducer. A working model for the regulation of penicillinase synthesis based on these and other data has been constructed and is presented. Images PMID:4333374

Flavonol-rich RVHxR from Rhus verniciflua Stokes and its major compound fisetin inhibits inflammation-related cytokines and angiogenic factor in rheumatoid arthritic fibroblast-like synovial cells and in vivo models.

PubMed

Lee, Jae-Dong; Huh, Jeong-Eun; Jeon, GeumSeon; Yang, Ha-Ru; Woo, Hyun-Su; Choi, Do-Young; Park, Dong-Suk

2009-03-01

Rheumatoid arthritis (RA) is an aggressive inflammatory disease in which cytokines/chemokines are thought to recruit leukocytes and induce angiogenesis. The aim of this study is to investigate the effect of flavonol-rich residual layer of hexane fraction from Rhus verniciflua Stokes (RVHxR) and its major compound fisetin on inflammatory cytokine/chemokine production and angiogenic factor in IL-1beta-stimulated RA fibroblast-like synovial cells (FLS) and inflammatory in vivo models. Flavonol-rich RVHxR and its major compound fisetin significantly inhibited IL-1beta-induced FLS proliferation in a dose-dependent manner. Flavonol-rich RVHxR and fisetin significantly decreased IL-1beta-induced inflammatory cytokines (TNF-alpha, interleukin (IL)-6)/chemokines (IL-8, monocyte chemoattractant protein (MCP)-1), and vascular endothelial growth factor (VEGF) of RA FLS. Flavonol-rich RVHxR dose dependently diminished the phophorylation of extracellular signal regulated kinase (ERK) and phospho-Jun NH((2))-terminal kinase (JNK), and its down regulation induced by RVHxR at nontoxic concentrations, while activated the phosphorylation of p38 MAPK in IL-1beta-stimulated RA FLS. The p38 specific inhibitor SB203580 cotreatment with RVHxR effectively increased the expression of VEGF and blocked the phosphorylation of p38 MAPK in IL-1beta-stimulated RA FLS, confirming a critical role of p38 MAPK pathway in angiogenesis inhibition. In experimental inflammation-related models, flavonol-rich RVHxR and fisetin have shown significant anti-inflammatory activities on vascular permeability, leukocyte migration and cellular immunity. Also, flavonol-rich RVHxR and fisetin treatments significantly reduced the incidence and severity of collagen-induced arthritis model. These results suggest that RVHxR and its major compound fisetin have shown potent suppressive effects on some inflammatory cytokines/chemokines and angiogenic factor in IL-1beta-stimulated RA FLS and inflammatory in vivo models. We

Further acylated flavonol bisdesmosides from Sinocrassula indica.

PubMed

Xie, Hai-Hui; Yoshikawa, Masayuki

2013-01-01

Further investigation on the whole herbs of Sinocrassula indica (Crassulaceae) led to the isolation of four new acylated flavonol bisdesmosides, sinocrassosides A₁₃, B₆, B₇, and D₄, together with kaempferol 3-O-β-D-(6-O-acetyl)glucopyranosyl-7-O-α-L-rhamnopyranoside. Their structures were established by spectral and chemical methods.

Anti-androgenic effects of flavonols in prostate cancer

PubMed Central

Boam, Tristan

2015-01-01

Dietary-derived agents, such as the flavonoids, are of particular interest for prostate cancer (PCa) chemoprevention as they may offer a favourable safety and side-effect profile. An agent that demonstrates action on the androgen receptor (AR) axis may have value for preventing or treating castrate-resistant PCa. Four main flavonols – quercetin, myricetin, kaempferol, and fisetin – have been demonstrated in laboratory studies to have chemopreventive action in both castrate-resistant and castrate-sensitive PCa models. Mechanisms of flavonol action on the AR axis in PCa have been proposed to be inhibition of the 5α-reductase enzymes, direct androgen competition, suppression of the AR complex and transactivation by coregulators such as c-Jun, Sp1, and the PI3K/Akt pathway. It is, however, still unclear with current levels of evidence whether AR axis-mediated effects can fully account for the flavonols’ chemopreventive action. PMID:26557883

Involvement of three putative glucosyltransferases from the UGT72 family in flavonol glucoside/rhamnoside biosynthesis in Lotus japonicus seeds

PubMed Central

Yin, Qinggang; Shen, Guoan; Chang, Zhenzhan; Tang, Yuhong; Gao, Hongwen

2017-01-01

Abstract Flavonols are one of the largest groups of flavonoids that confer benefits for the health of plants and animals. Flavonol glycosides are the predominant flavonoids present in the model legume Lotus japonicus. The molecular mechanisms underlying the biosynthesis of flavonol glycosides as yet remain unknown in L. japonicus. In the present study, we identified a total of 188 UDP-glycosyltransferases (UGTs) in L. japonicus by genome-wide searching. Notably, 12 UGTs from the UGT72 family were distributed widely among L. japonicus chromosomes, expressed in all tissues, and showed different docking scores in an in silico bioinformatics docking analysis. Further enzymatic assays showed that five recombinant UGTs (UGT72AD1, UGT72AF1, UGT72AH1, UGT72V3, and UGT72Z2) exhibit activity toward flavonol, flavone, and isoflavone aglycones. In particular, UGT72AD1, UGT72AH1, and UGT72Z2 are flavonol-specific UGTs with different kinetic properties. In addition, the overexpression of UGT72AD1 and UGT72Z2 led to increased accumulation of flavonol rhamnosides in L. japonicus and Arabidopsis thaliana. Moreover, the increase of kaempferol 3-O-rhamnoside-7-O-rhamnoside in transgenic A. thaliana inhibited root growth as compared with the wild-type control. These results highlight the significance of the UGT72 family in flavonol glycosylation and the role of flavonol rhamnosides in plant growth. PMID:28204516

Flavonol glycosides of sea buckthorn (Hippophaë rhamnoides ssp. sinensis) and lingonberry (Vaccinium vitis-idaea) are bioavailable in humans and monoglucuronidated for excretion.

PubMed

Lehtonen, Henna-Maria; Lehtinen, Outi; Suomela, Jukka-Pekka; Viitanen, Matti; Kallio, Heikki

2010-01-13

Glucuronidation and excretion of sea buckthorn and lingonberry flavonols were investigated in a postprandial trial by analyzing the intact forms of flavonol glycosides as well as glucuronides in plasma, urine, and feces. Four study subjects consumed sea buckthorn (study day 1) and lingonberry (study day 2) breakfasts, and blood, urine, and fecal samples were collected for 8, 24, and 48 h, respectively. Both glycosides and glucuronides of the flavonol quercetin as well as kaempferol glucuronides were detected in urine and plasma samples after the consumption of lingonberries; 14% of flavonols in urine were glycosides, and 86% were glucuronidated forms (wt %). After the consumption of sea buckthorn, 5% of flavonols excreted in urine were detected intact, and 95% as the glucuronides (wt %). Solely glucuronides of flavonols isorhamnetin and quercetin were found in plasma after the consumption of sea buckthorn berries. Only glycosides were detected in the feces after each berry trial. Flavonol glycosides and glucuronides remained in blood and urine quite long, and the peak concentrations appeared slightly later than previously described. The berries seemed to serve as a good flavonol supply, providing steady flavonol input for the body for a relatively long time.

Impact of different stages of juice processing on the anthocyanin, flavonol, and procyanidin contents of cranberries.

PubMed

White, Brittany L; Howard, Luke R; Prior, Ronald L

2011-05-11

Juice is the most common form in which cranberries are consumed; however there is limited information on the changes of polyphenolic content of the berries during juice processing. This study investigated the effects of three different pretreatments (grinding plus blanching; only grinding; only blanching) for cranberry juice processing on the concentrations of anthocyanins, flavonols, and procyanidins throughout processing. Flavonols and procyanidins were retained in the juice to a greater extent than anthocyanins, and pressing resulted in the most significant losses in polyphenolics due to removal of the seeds and skins. Flavonol aglycones were formed during processing as a result of heat treatment. Drying of cranberry pomace resulted in increased extraction of flavonols and procyanidin oligomers but lower extraction of polymeric procyanidins. The results indicate that cranberry polyphenolics are relatively stable during processing compared to other berries; however, more work is needed to determine their fate during storage of juices.

Complementary action of jasmonic acid on salicylic acid in mediating fungal elicitor-induced flavonol glycoside accumulation of Ginkgo biloba cells.

PubMed

Xu, Maojun; Dong, Jufang; Wang, Huizhong; Huang, Luqi

2009-08-01

The antagonistic action between jasmonic acid (JA) and salicylic acid (SA) in plant defence responses has been well documented. However, their relationship in secondary metabolite production is largely unknown. Here, we report that PB90, a protein elicitor from Phytophthora boehmeriae, triggers JA generation, SA accumulation and flavonol glycoside production of Ginkgo biloba cells. JA inhibitors suppress not only PB90-triggered JA generation, but also the elicitor-induced flavonol glycoside production. However, the elicitor can still enhance flavonol glycoside production even though the JA generation is totally inhibited. Over-expression of SA hydrolase gene NahG not only abolishes SA accumulation, but also suppresses the elicitor-induced flavonol glycoside production when JA signalling is inhibited. Interestingly, expression of NahG does not inhibit the elicitor-induced flavonol glycoside accumulation in the absence of JA inhibitors. Moreover, JA levels are significantly enhanced when SA accumulation is impaired in the transgenic cells. Together, the data suggest that both JA and SA are involved in PB90-induced flavonol glycoside production. Furthermore, we demonstrate that JA signalling might be enhanced to substitute for SA to mediate the elicitor-induced flavonol glycoside accumulation when SA signalling is impaired, which reveals an unusual complementary relationship between JA and SA in mediating plant secondary metabolite production.

Induction of antioxidant flavonol biosynthesis in fresh-cut potatoes. Effect of domestic cooking.

PubMed

Tudela, Juan A; Cantos, Emma; Espín, Juan C; Tomás-Barberán, Francisco A; Gil, María I

2002-10-09

The effect of fresh-cutting and subsequent cold storage on phenolic compounds from five long-term-stored potato cultivars (Agria, Cara, Liseta, Monalisa, and Spunta) was studied. Fresh-cutting induced the biosynthesis of three flavonols, which were identified by HPLC-DAD-ESIMS as quercetin 3-rutinoside, quercetin 3-diglucoside, and quercetin 3-glucosylrutinoside. The flavonols were detected after a lag period of 3 days of cold storage. The content ranged from 6 to 14 mg/100 g of fresh weight depending on the cultivar after 6 days of storage. Chlorogenic acid as the main caffeic acid derivative and the amino acids tyrosine and tryptophan were also quantified. The effect of cold storage under light or in dark was studied with new-season-harvested Monalisa potatoes. The flavonol induction was higher in fresh-cut potatoes stored under light than in the dark. However, caffeic acid derivatives were not affected. Domestic cooking such as boiling, microwaving, and frying provoked a partial loss of the flavonols, which were retained in the range of 4-16 mg per serving (213 g). Steam-cooking resulted in the highest retention of caffeic acid derivatives and aromatic amino acids compared with the other cooking methods studied. This means that due to the large amount of potatoes consumed in the Western diet, fresh-cut potatoes can be a significant source of health-promoting phenolics.

Nitrogen form and mycorrhizal inoculation amount and timing affect flavonol biosynthesis in onion (Allium cepa L.).

PubMed

Mollavali, Mohanna; Perner, Henrike; Rohn, Sascha; Riehle, Peer; Hanschen, Franziska S; Schwarz, Dietmar

2018-01-01

Mycorrhizal symbiosis is known to be the most prevalent form of fungal symbiosis with plants. Although some studies focus on the importance of mycorrhizal symbiosis for enhanced flavonoids in the host plants, a comprehensive understanding of the relationship still is lacking. Therefore, we studied the effects of mycorrhizal inoculation of onions (Allium cepa L.) regarding flavonol concentration and the genes involved in flavonol biosynthesis when different forms of nitrogen were supplied. We hypothesized that mycorrhizal inoculation can act as a biotic stress and might lead to an increase in flavonols and expression of related genes. The three main quercetin compounds [quercetin-3,4'-di-O-β-D-glucoside (QDG), quercetin-4'-O-β-D-glucoside (QMG), and isorhamnetin-4'-O-β-D-glucoside (IMG)] of onion bulbs were identified and analyzed after inoculating with increasing amounts of mycorrhizal inocula at two time points and supplying either predominantly NO 3 - or NH 4 + nitrogen. We also quantified plant dry mass, nutrient element uptake, chalcone synthase (CHS), flavonol synthase (FLS), and phenyl alanine lyase (PAL) gene expression as key enzymes for flavonol biosynthesis. Inoculation with arbuscular mycorrhizal fungi (highest amount) and colonization at late development stages (bulb growth) increased QDG and QMG concentrations if plants were additionally supplied with predominantly NH 4 + . No differences were observed in the IMG content. RNA accumulation of CHS, FLS, and PAL was affected by the stage of the mycorrhizal symbiosis and the nitrogen form. Accumulation of flavonols was not correlated, however, with either the percentage of myorrhization or the abundance of transcripts of flavonoid biosynthesis genes. We found that in plants at late developmental stages, RNA accumulation as a reflection of a current physiological situation does not necessarily correspond with the content of metabolites that accumulate over a long period. Our findings suggest that

Triterpenes and flavonol glucuronides from Oenothera cheiranthifolia.

PubMed

Nakanishi, Tsutomu; Inatomi, Yuka; Murata, Hiroko; Ishida, Syun-Suke; Fujino, Yuri; Miura, Kanako; Yasuno, Yoshito; Inada, Akira; Lang, Frank A; Murata, Jin

2007-02-01

A new ursane-type triterpene, named as cheiranthic acid (1), was isolated from the MeOH extract of whole plants of Oenothera cheiranthifolia (Onagraceae) along with an isomeric pair of known oleanane- and ursane-type triterpenes (arjunolic acid and asiatic acid) and three flavonol glucuronide analogues (quercetin 3-O-glucuronide, its n-butyl ester, and myricetin 3-O-glucuronide). Their structures were elucidated based on spectroscopic evidence.

Bioavailability and health effects of dietary flavonols in man.

PubMed

Hollman, P C; Katan, M B

1998-01-01

Flavonoids are polyphenolic compounds that occur ubiquitously in foods of plant origin. Over 4000 different flavonoids have been described, and they are categorized into flavonols, flavones, catechins, flavanones, anthocyanidins, and isoflavonoids. Flavonoids have a variety of biological effects in numerous mammalian cell systems, as well as in vivo. Recently much attention has been paid to their antioxidant properties and to their inhibitory role in various stages of tumour development in animal studies. Quercetin, the major representative of the flavonol subclass, is a strong antioxidant, and prevents oxidation of low density lipoproteins in vitro. Oxidized low density lipoproteins are atherogenic, and are considered to be a crucial intermediate in the formation of atherosclerotic plaques. This agrees with observations in epidemiological studies that the intake of flavonols and flavones was inversely associated with subsequent coronary heart disease. However, no effects of flavonols on cancer were found in these studies. The extent of absorption of flavonoids is an important unsolved problem in judging their many alleged health effects. Flavonoids present in foods were considered non-absorbable because they are bound to sugars as beta-glycosides. Only free flavonoids without a sugar molecule, the so-called aglycones were thought to be able to pass through the gut wall. Hydrolysis only occurs in the colon by microorganisms, which at the same time degrade flavonoids. We performed a study to quantify absorption of various dietary forms of quercetin. To our surprise, the quercetin glycosides from onions were absorbed far better than the pure aglycone. Subsequent pharmacokinetic studies with dietary quercetin glycosides showed marked differences in absorption rate and bioavailability. Absorbed quercetin was eliminated only slowly from the blood. The metabolism of flavonoids has been studied frequently in various animals, but very few data in humans are available. Two

Flavonols and Carotenoids in Yellow Petals of Rose Cultivar ( Rosa 'Sun City'): A Possible Rich Source of Bioactive Compounds.

PubMed

Wan, Huihua; Yu, Chao; Han, Yu; Guo, Xuelian; Ahmad, Sagheer; Tang, Aoying; Wang, Jia; Cheng, Tangren; Pan, Huitang; Zhang, Qixiang

2018-04-25

Rose flowers have received increasing interest as rich sources of bioactive compounds. The composition of flavonols and carotenoids in yellow petals of Rosa 'Sun City' was determined by high-performance liquid chromatography coupled with photodiode array and mass spectrometric detectors (HPLC-PDA-MS). In total, 19 flavonols and 16 carotenoids were identified, some of which were first discovered in rose petals. Significant changes were observed in their profiles during seven blooming stages. Total flavonol contents showed the highest levels at stage 2 (S2; 1152.29 μg/g, FW). Kaempferol 7- O-glucoside and kaempferol 3- O-rhamnoside were the predominant individual flavonols. Total carotenoid concentration was highest at S4 (142.71 μg/g, FW). Violaxanthins with different geometrical configurations appeared as the major carotenoids across all blooming stages. These results indicated that 'Sun City' petals are rich sources of flavonols and carotenoids. Moreover, it is important to choose the appropriate harvest time on the basis of the targeted compounds.

Kaempferol, a mutagenic flavonol from Helichrysum simillimum.

PubMed

Elgorashi, Ee; van Heerden, Fr; van Staden, J

2008-11-01

Helichrysum simillimum is native to South Africa. It is used for the treatment of coughs, colds, fever, infections, headache, and menstrual pain. Extracts of this species showed mutagenic effects in the Salmonella/microsome assay. The aim of this study was to isolate and determine the mutagenic constituents of H. simillimum. Bioassay-guided fractionation of 90% aqueous methanol extracts, using Salmonella typhimurium TA98, led to the isolation of the flavonol kaempferol.

Flavonol glycosides in berries of two major subspecies of sea buckthorn (Hippophaë rhamnoides L.) and influence of growth sites.

PubMed

Ma, Xueying; Laaksonen, Oskar; Zheng, Jie; Yang, Wei; Trépanier, Martin; Kallio, Heikki; Yang, Baoru

2016-06-01

Flavonol glycosides of wild sea buckthorn (Hippophaë rhamnoides ssp. sinensis) berries from China and cultivated berries (H. rhamnoides ssp. mongolica) from Finland and Canada were identified and quantified. Twenty-six flavonol glycosides were found with isorhamnetin and quercetin as the major aglycones. The contents of flavonol glycosides ranged 23-250 mg/100 g fresh berries and were significantly higher in the berries of ssp. sinensis than in those of ssp. mongolica. Among the cultivars of ssp. mongolica, the berries of 'Oranzhevaya' had the lowest (23 mg/100 g) content, and those of 'Prevoshodnaya' the highest content of flavonol glycosides (80 mg/100 g). Within the ssp. mongolica, the samples from Kittilä (Northern Finland) had higher levels of most flavonol glycosides than those from Turku (Southern Finland) and Québec. Among the ssp. sinensis berries of different growth sites, increasing trends were detected in the contents of most of the compounds as the altitude increased and as the latitude decreased. The wild berries (ssp. sinensis) from Sichuan had remarkably high contents and unique profiles of flavonol glycosides. Copyright © 2016 Elsevier Ltd. All rights reserved.

Flavonols, alkaloids, and antioxidant capacity of edible wild berberis species from patagonia.

PubMed

Ruiz, Antonieta; Zapata, Moises; Sabando, Constanza; Bustamante, Luis; von Baer, Dietrich; Vergara, Carola; Mardones, Claudia

2014-12-24

There are 20 species of the Berberidaceae family described in Chile, whose fruits are edible and show high anthocyanin and hydroxycinnamic acid levels. Berberis microphylla G. Forst, commonly known as calafate, is the most extensively distributed. Flavonols and alkaloids in seed, pulp, skin, and whole calafate berry extracts and other Berberis were studied using HPLC-DAD-ESI-MS/MS and HPLC with fluorescence detector. Berry samples from different locations in Chilean Patagonia, including different phenological stages, were systematically addressed. Results were compared with other organs of the plant and with other Berberis species. Total flavonol concentration in calafate (n = 65) was 1.33 ± 0.54 μmol/g. Glycosyl metabolites of quercetin and isorhamnetin were the most abundant. Similar profiles were observed in calafate from distinct locations, but important differences were observed for the other edible Berberis species. Calafate pulp and skin have higher flavonol concentrations than seeds, and the maturation process reduced its levels. TEACCUPRAC and TEACABTS of whole calafate extracts and fractions are also explored. Finally, only berberine was detected in the fruit (0.001%), mainly in seeds. Results contribute to the promotion of this berry as a superfruit from Patagonia.

Negative Regulation of Anthocynanin Biosynthesis in Arabidopsis by a miR156-Targeted SPL Transcription Factor

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

Gou, J.Y.; Liu, C.; Felippes, F. F.

2011-04-01

Flavonoids are synthesized through an important metabolic pathway that leads to the production of diverse secondary metabolites, including anthocyanins, flavonols, flavones, and proanthocyanidins. Anthocyanins and flavonols are derived from Phe and share common precursors, dihydroflavonols, which are substrates for both flavonol synthase and dihydroflavonol 4-reductase. In the stems of Arabidopsis thaliana, anthocyanins accumulate in an acropetal manner, with the highest level at the junction between rosette and stem. We show here that this accumulation pattern is under the regulation of miR156-targeted SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) genes, which are deeply conserved and known to have important roles in regulating phasemore » change and flowering. Increased miR156 activity promotes accumulation of anthocyanins, whereas reduced miR156 activity results in high levels of flavonols. We further provide evidence that at least one of the miR156 targets, SPL9, negatively regulates anthocyanin accumulation by directly preventing expression of anthocyanin biosynthetic genes through destabilization of a MYB-bHLH-WD40 transcriptional activation complex. Our results reveal a direct link between the transition to flowering and secondary metabolism and provide a potential target for manipulation of anthocyanin and flavonol content in plants.« less

Solar UV light regulates flavonoid metabolism in apple (Malus x domestica).

PubMed

Henry-Kirk, Rebecca A; Plunkett, Blue; Hall, Miriam; McGhie, Tony; Allan, Andrew C; Wargent, Jason J; Espley, Richard V

2018-03-01

Ultraviolet-B light (UV-B) is one environmental signal perceived by plants that affects the flavonoid pathway and influences the levels of anthocyanins, flavonols, and proanthocyanidins. To understand the mechanisms underlying UV exposure, apple trees were grown under spectral filters that altered transmission of solar UV light. Fruit analysis showed that UV induced changes in physiology, metabolism, and gene expression levels during development over a season. These changes were sustained after storage. Under low UV, ripening was delayed, fruit size decreased, and anthocyanin and flavonols were reduced. Expression analysis showed changes in response to UV light levels for genes in the regulation and biosynthesis of anthocyanin and flavonols. Transcription of flavonol synthase (FLS), ELONGATED HYPOCOTYL 5 (HY5), MYB10, and MYB22 were down-regulated throughout fruit development under reduced UV. Functional testing showed that the FLS promoter was activated by HY5, and this response was enhanced by the presence of MYB22. The MYB22 promoter can also be activated by the anthocyanin regulator, MYB10. As ambient levels of UV light vary around the globe, this study has implications for future crop production, the quality of which can be determined by the response to UV. © 2018 John Wiley & Sons Ltd.

Effect of a low dose of sea buckthorn berries on circulating concentrations of cholesterol, triacylglycerols, and flavonols in healthy adults.

PubMed

Larmo, Petra S; Yang, Baoru; Hurme, Saija A M; Alin, Jouni A; Kallio, Heikki P; Salminen, Eeva K; Tahvonen, Raija L

2009-08-01

Epidemiological studies indicate beneficial effects of flavonoids on cardiovascular disease (CVD) risk. To study the effect of flavonoid-rich sea buckthorn berry (SBB) on circulating lipid markers associated with CVD risk and plasma flavonol concentration. Also investigated was whether changes in the circulating flavonol concentrations correlate with the SBB induced changes in C-reactive protein (CRP) concentration observed previously. In all 229 healthy participants completed the randomized double-blind study and consumed daily 28 g of SBB or placebo for 3 months. Fasting blood samples for the analysis of lipid markers and flavonols were obtained at the beginning and end of the study. Compared to the placebo, the consumption of SBB increased the plasma concentration of the flavonols quercetin and isorhamnetin significantly [treatment differences 3.0 ng/ml (P = 0.03) and 3.9 ng/ml (P < 0.01), respectively]. The increase of kaempferol concentration was not significant [treatment difference 0.7 ng/ml (P = 0.08)]. SBB did not affect the serum total, HDL, LDL cholesterol, or the serum triacylglycerol concentrations. There was no correlation between the changes in flavonol and CRP concentrations of participants. The consumption of SBB significantly increased the fasting plasma concentration of quercetin and isorhamnetin indicating that it is a good dietary source of flavonols. However, this did not convert to affecting the circulating concentrations of lipid markers in healthy, normolipidemic adults having healthy diets.

Enhanced profiling of flavonol glycosides in the fruits of sea buckthorn (Hippophae rhamnoides).

PubMed

Fang, Rui; Veitch, Nigel C; Kite, Geoffrey C; Porter, Elaine A; Simmonds, Monique S J

2013-04-24

Use of enhanced LC-MS/MS methods to identify common glycosyl groups of flavonoid glycosides enabled better characterization of the flavonoids in fruits of sea buckthorn (Hippophae rhamnoides). The saccharide moieties of 48 flavonol O-glycosides detected in a methanol extract were identified by these methods. Several of the flavonol glycosides were acylated, two of which were isolated and found to be new compounds. Their structures were determined using spectroscopic and chemical methods as isorhamnetin 3-O-(6-O-E-sinapoyl-β-D-glucopyranosyl)-(1→2)-β-D-glucopyranoside-7-O-α-L-rhamnopyranoside (24) and isorhamnetin 3-O-(6-O-E-feruloyl-β-D-glucopyranosyl)-(1→2)-β-D-glucopyranoside-7-O-α-L-rhamnopyranoside (30). Analysis of the acylated glycosyl groups of 24 and 30 by serial mass spectrometry provided evidence to suggest the acylation position of 11 other minor flavonol glycosides acylated with hydroxycinnamic or hydroxybenzoic acids. The nitric oxide scavenging activities of 24 and 30 were compared with those of other flavonoids and with ascorbic acid and the potassium salt of 2-(4-carboxyphenyl)-4,5-dihydro-4,4,5,5-tetramethyl-1H-imidazolyl-1-oxy-3-oxide (carboxy-PTIO).

A new flavonol glycoside from the medicinal halophyte Suaeda fruticosa.

PubMed

Oueslati, Samia; Ksouri, Riadh; Pichette, André; Lavoie, Serge; Girard-Lalancette, Karl; Mshvildadze, Vakhtang; Abdelly, Chedly; Legault, Jean

2014-01-01

A new flavonol glycoside, namely 3-(α-rhamnopyranosyl-(1 → 2)-[β-xylopyranosyl-(1 → 6)]-β-glucopyranosyloxy) isorhamnetin was reported from methanol extracts of aerial parts of Suaeda fruticosa for the first time. In this work, liquid chromatography coupled to atmospheric pressure chemical ionisation mass spectrometry, high-resolution mass spectrometry and nuclear magnetic resonance spectroscopy were used to identify this new compound. Structure was elucidated on the basis of extensive spectroscopic analysis, including HSQC, HMBC and (1)H-(1)H COSY. Antioxidant potentialities of a pure compound were evaluated. The estimation of antioxidant capacities using oxygen radical absorbance capacity (ORAC method) and a cell based-assay (WS1) indicated that this new flavonol exhibited the highest antioxidant activities with an ORAC value of 5.0 ± 0.3 μmol Trolox/μmol and inhibited the tBH-induced oxidation of 2',7'-dichlorofluorescin with an IC50 value of 4.9 ± 0.6 μM.

Dietary flavonols contribute to false-positive elevation of homovanillic acid, a marker of catecholamine-secreting tumors.

PubMed

Combet, Emilie; Lean, Michael E J; Boyle, James G; Crozier, Alan; Davidson, D Fraser

2011-01-14

Urinary homovanillic acid (HVA) measurement is used routinely as a marker of the first test for the screening of catecholamine-secreting tumors and dopamine metabolism, but generates a large number of false-positive results. With no guidelines for dietary restrictions prior to the test, we hypothesize that consumption of flavonol-rich foods (such as onions, tomatoes, tea) prior to urinary catecholamine screening could be responsible for false-positive urinary HVA in healthy subjects. A randomized, crossover dietary intervention was carried out in healthy subjects (n=17). Volunteers followed either a low or high-flavonol diet, for a duration of 3 days, prior to providing a 24-h urine sample for HVA measurement using a routine, validated liquid chromatography method as well as a gas chromatography-mass spectrometry method. Dietary flavonol intake significantly increased urinary HVA excretion (p < 0.001), with 3 out of 17 volunteers (20%) exceeding the 40 μmol/24 h upper limit of normal for HVA excretion (false-positive result). Dietary flavonols commonly found in foodstuff such as tomatoes, onions, and tea, interfered with the routine urinary HVA screening test and should be avoided in the three-day run-up to the test. Copyright © 2010 Elsevier B.V. All rights reserved.

Absorption of flavonols derived from sea buckthorn (Hippophaë rhamnoides L.) and their effect on emerging risk factors for cardiovascular disease in humans.

PubMed

Suomela, Jukka-Pekka; Ahotupa, Markku; Yang, Baoru; Vasankari, Tommi; Kallio, Heikki

2006-09-20

Sea buckthorn (Hippophaë rhamnoides L.) is a rich source of flavonols, especially isorhamnetin. Most prospective cohort studies have indicated some degree of inverse association between flavonoid intake and coronary heart disease. Animal and human studies suggest that sea buckthorn flavonoids may scavenge free radicals, lower blood viscosity, and enhance cardiac function. The effects of flavonol aglycones derived from sea buckthorn on the risk factors of cardiovascular disease as well as their absorption were studied in humans. The flavonols, ingested with oatmeal porridge, did not have a significant effect on the levels of oxidized low-density lipoprotein, C-reactive protein, and homocysteine, on the plasma antioxidant potential, or on the paraoxonase activity. Flavonols at two dosages in oatmeal porridge were rapidly absorbed, and a relatively small amount of sea buckthorn oil added to the porridge seemed to have increased the bioavailability of sea buckthorn flavonols consumed at the higher dose.

Do Interleukin Polymorphisms Play a Role in the Prevention of Colorectal Adenoma Recurrence by Dietary Flavonols?

PubMed Central

Bobe, Gerd; Murphy, Gwen; Albert, Paul S.; Sansbury, Leah B.; Young, Matthew R.; Lanza, Elaine; Schatzkin, Arthur; Colburn, Nancy H.; Cross, Amanda J.

2010-01-01

Chemopreventive dietary compounds, such as flavonols, may inhibit colorectal carcinogenesis partly by altering cytokine expression and attenuating inflammation. Single nucleotide polymorphisms (SNPs) in the promoter regions of genes encoding cytokines may influence flavonol-induced changes in cytokine expression and consequently cancer risk. Using logistic regression, we estimated odds ratios (OR) and 95% confidence intervals (CI) for the association between SNPs of interleukin (IL)-1β, 6, 8, and 10, alone or combined with flavonol intake or serum IL concentration changes, and adenoma recurrence in 808 participants from the intervention arm of the Polyp Prevention Trial, a 4-year intervention study evaluating the effectiveness of a low-fat, high-fiber, high-fruit and vegetable diet on adenoma recurrence.. Overall, SNPs in genes encoding IL-1β, 6, 8, and 10 were not associated with their corresponding serum concentrations or adenoma recurrence. However, individuals homozygous for IL-10 -592 C (OR = 2.23, 95% CI: 1.07–4.66, P interaction = 0.03) or IL-10 -819 C (OR = 2.18, 95% CI: 1.05–4.51, P interaction = 0.05) had an elevated risk of high risk adenoma recurrence when their serum IL-10 concentrations increased during the trial. In addition, IL-6 -174 GG in combination with above median flavonol intake (OR = 0.14, 95% CI: 0.03–0.66) or with decreased IL 6 concentrations (OR = 0.14, 95% CI: 0.03–0.65) reduced the risk of advanced adenoma recurrence, although the interaction term was not statistically significant. In conclusion, our results suggest that IL SNPs, in combination with a flavonol-rich diet or decreased serum IL, may lower the risk of adenoma recurrence. PMID:21160427

Functional Characterization of CsBGlu12, a β-Glucosidase from Crocus sativus, Provides Insights into Its Role in Abiotic Stress through Accumulation of Antioxidant Flavonols.

PubMed

Baba, Shoib Ahmad; Vishwakarma, Ram A; Ashraf, Nasheeman

2017-03-17

Glycosylation and deglycosylation are impressive mechanisms that allow plants to regulate the biological activity of an array of secondary metabolites. Although glycosylation improves solubility and renders the metabolites suitable for transport and sequestration, deglycosylation activates them to carry out biological functions. Herein, we report the functional characterization of Cs BGlu12, a β-glucosidase from Crocus sativus. Cs BGlu12 has a characteristic glucoside hydrolase 1 family (α/β) 8 triose-phosphate isomerase (TIM) barrel structure with a highly conserved active site. In vitro enzyme activity revealed that Cs BGlu12 catalyzes the hydrolysis of flavonol β-glucosides and cello-oligosaccharides. Site-directed mutagenesis of any of the two conserved catalytic glutamic acid residues (Glu 200 and Glu 414 ) of the active site completely abolishes the β-glucosidase activity. Transcript analysis revealed that Csbglu12 is highly induced in response to UV-B, dehydration, NaCl, methyl jasmonate, and abscisic acid treatments indicating its possible role in plant stress response. Transient overexpression of Cs BGlu12 leads to the accumulation of antioxidant flavonols in Nicotiana benthamiana and confers tolerance to abiotic stresses. Antioxidant assays indicated that accumulation of flavonols alleviated the accretion of reactive oxygen species during abiotic stress conditions. β-Glucosidases are known to play a role in abiotic stresses, particularly dehydration through abscisic acid; however, their role through accumulation of reactive oxygen species (ROS) scavenging flavonols has not been established. Furthermore, only one β-glucosidase 12 homolog has been characterized so far. Therefore, this work presents an important report on characterization of Cs BGlu12 and its role in abiotic stress through ROS scavenging. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Functional Characterization of CsBGlu12, a β-Glucosidase from Crocus sativus, Provides Insights into Its Role in Abiotic Stress through Accumulation of Antioxidant Flavonols*

PubMed Central

Baba, Shoib Ahmad; Vishwakarma, Ram A.; Ashraf, Nasheeman

2017-01-01

Glycosylation and deglycosylation are impressive mechanisms that allow plants to regulate the biological activity of an array of secondary metabolites. Although glycosylation improves solubility and renders the metabolites suitable for transport and sequestration, deglycosylation activates them to carry out biological functions. Herein, we report the functional characterization of CsBGlu12, a β-glucosidase from Crocus sativus. CsBGlu12 has a characteristic glucoside hydrolase 1 family (α/β)8 triose-phosphate isomerase (TIM) barrel structure with a highly conserved active site. In vitro enzyme activity revealed that CsBGlu12 catalyzes the hydrolysis of flavonol β-glucosides and cello-oligosaccharides. Site-directed mutagenesis of any of the two conserved catalytic glutamic acid residues (Glu200 and Glu414) of the active site completely abolishes the β-glucosidase activity. Transcript analysis revealed that Csbglu12 is highly induced in response to UV-B, dehydration, NaCl, methyl jasmonate, and abscisic acid treatments indicating its possible role in plant stress response. Transient overexpression of CsBGlu12 leads to the accumulation of antioxidant flavonols in Nicotiana benthamiana and confers tolerance to abiotic stresses. Antioxidant assays indicated that accumulation of flavonols alleviated the accretion of reactive oxygen species during abiotic stress conditions. β-Glucosidases are known to play a role in abiotic stresses, particularly dehydration through abscisic acid; however, their role through accumulation of reactive oxygen species (ROS) scavenging flavonols has not been established. Furthermore, only one β-glucosidase 12 homolog has been characterized so far. Therefore, this work presents an important report on characterization of CsBGlu12 and its role in abiotic stress through ROS scavenging. PMID:28154174

Profiling of primary metabolites and flavonols in leaves of two table grape varieties collected from semiarid and temperate regions.

PubMed

Harb, Jamil; Alseekh, Saleh; Tohge, Takayuki; Fernie, Alisdair R

2015-09-01

Cultivation of grapes in West Bank - Palestine is very old and a large number of grape varieties exist as a result of continuous domestication over thousands of years. This rich biodiversity has highly influenced the consumer behavior of local people, who consume both grape berries and leaves. However, studies that address the contents of health-promoting metabolites in leaves are scarce. Accordingly the aim of this study is to assess metabolite levels in leaves of two grape varieties that were collected from semiarid and temperate regions. Metabolic profiling was conducted using GC-MS and LC-MS. The obtained results show that abiotic stresses in the semiarid region led to clear changes in primary metabolites, in particular in amino acids, which exist at very high levels. By contrast, qualitative and genotype-dependent differences in secondary metabolites were observed, whereas abiotic stresses appear to have negligible effect on the content of these metabolites. The qualitative difference in the flavonol profiles between the two genotypes is most probably related to differential expression of specific genes, in particular flavonol 3-O-rhamnosyltransferase, flavonol-3-O-glycoside pentosyltransferases and flavonol-3-O-d-glucosidel-rhamnosyltransferase by 'Beituni' grape leaves, which led to much higher levels of flavonols with rutinoside, pentoside, and rhamnoside moieties with this genotype. Copyright © 2015 Elsevier Ltd. All rights reserved.

Antidiabetic complications and anti-Alzheimer activities of sophoflavescenol, a prenylated flavonol from Sophora flavescens, and its structure-activity relationship.

PubMed

Jung, Hyun Ah; Jin, Seong Eun; Park, Jun-Seong; Choi, Jae Sue

2011-05-01

It was previously reported that prenylated flavonols from Sophora flavescens are inhibitors of rat lens aldose reductase (RLAR), human recombinant aldose reductase (HRAR), advanced glycation endproducts (AGE), β-secretase (BACE1) and cholinesterases (ChE). Based upon structure-activity relationships, 3,4'-dihydroxy flavonols with a prenyl or lavandulyl group substitution at the C-8 position, and a hydroxy group at the C-5, are important for such inhibition. In our ongoing study to isolate active principles from S. flavescens by an activity-guided isolation procedure, further detailed phytochemical investigations of the CH(2)Cl(2) fraction were conducted via repeated chromatography over silica gel and Sephadex LH-20 columns. This ultimately resulted in the isolation of a promising active sophoflavescenol with higher inhibitory activities among the current prenylated flavonols isolated from S. flavescens against RLAR, HRAR, AGE, BACE1 and ChEs. The results further support that 3,4'-dihydroxy flavonols with a prenyl or lavandulyl substitution at the C-8 position and a methoxy group at C-5 represent a new class of RLAR, HRAR and AGE inhibitors. Nevertheless, the C-5 hydroxyl group of prenylated flavonoids is important for inhibition of BACE1 and ChEs, indicating that the hydroxyl group at C-5 might be the main contributor to the augmentation and/or modification of prenylated flavonol activity. Copyright © 2010 John Wiley & Sons, Ltd.

Interleukin-6 as a Potential Indicator for Prevention of High Risk Adenoma Recurrence by Dietary Flavonols in the Polyp Prevention Trial

PubMed Central

Bobe, Gerd; Albert, Paul S.; Sansbury, Leah B.; Lanza, Elaine; Schatzkin, Arthur; Colburn, Nancy H.; Cross, Amanda J.

2010-01-01

Serum interleukin (IL)-6, a pro-inflammatory cytokine, is considered an indicator of inflammation and may be an indicator of colorectal carcinogenesis given that inflammation can promote carcinogenesis. Flavonols, which can be found in fruits and vegetables, may inhibit colorectal carcinogenesis partly by inhibiting inflammation. We estimated odds ratios (ORs) and 95% confidence intervals (CIs) to determine whether serum IL-6 was associated with colorectal adenoma recurrence and flavonol intake and, thus may serve as a risk indicator and as a response indicator to dietary flavonols. Serum IL-6 concentrations at baseline, year 1 and 3 were measured in 872 participants from the intervention arm of the Polyp Prevention Trial, a 4-year trial that examined the effectiveness of a low-fat, high-fiber, high-fruit and vegetable diet on adenoma recurrence. Intake of flavonols, especially of isorhamnetin, kaempferol, and quercetin, was inversely associated with serum IL-6 concentrations (highest vs. lowest flavonol intake quartile, 1.80 vs. 2.20 pg/mL) and high risk (OR = 0.51, 95% CI: 0.26–0.98) and advanced adenoma recurrence (OR = 0.17, 95% CI: 0.06–0.50). A decrease in IL-6 concentration during the trial was inversely associated with high risk (OR = 0.44, 95% CI: 0.23–0.84) and advanced adenoma recurrence (OR = 0.47, 95% CI: 0.19–1.18). Individuals with above median flavonol intake and equal or below median IL-6 change after baseline had the lowest risk of recurrence of high risk and advanced adenoma. Our results suggest that serum IL-6 may serve as a risk indicator and as a response indicator to dietary flavonols for colorectal cancer prevention. PMID:20484173

Change of supercooling capability in solutions containing different kinds of ice nucleators by flavonol glycosides from deep supercooling xylem parenchyma cells in trees.

PubMed

Kuwabara, Chikako; Kasuga, Jun; Wang, Donghui; Fukushi, Yukiharu; Arakawa, Keita; Koyama, Toshie; Inada, Takaaki; Fujikawa, Seizo

2011-12-01

Deep supercooling xylem parenchyma cells (XPCs) in Katsura tree contain flavonol glycosides with high supercooling-facilitating capability in solutions containing the ice nucleation bacterium (INB) Erwinia ananas, which is thought to have an important role in deep supercooling of XPCs. The present study, in order to further clarify the roles of these flavonol glycosides in deep supercooling of XPCs, the effects of these supercooling-facilitating (anti-ice nucleating) flavonol glycosides, kaempferol 3-O-β-D-glucopyranoside (K3Glc), kaempferol 7-O-β-D-glucopyranoside (K7Glc) and quercetin 3-O-β-D-glucopyranoside (Q3Glc), in buffered Milli-Q water (BMQW) containing different kinds of ice nucleators, including INB Xanthomonas campestris, silver iodide and phloroglucinol, were examined by a droplet freezing assay. The results showed that all of the flavonol glycosides promoted supercooling in all solutions containing different kinds of ice nucleators, although the magnitudes of supercooling capability of each flavonol glycoside changed in solutions containing different kinds of ice nucleators. On the other hand, these flavonol glycosides exhibited complicated nucleating reactions in BMQW, which did not contain identified ice nucleators but contained only unidentified airborne impurities. Q3Glc exhibited both supercooling-facilitating and ice nucleating capabilities depending on the concentrations in such water. Both K3Glc and K7Glc exhibited only ice nucleation capability in such water. It was also shown by an emulsion freezing assay in BMQW that K3Glc and Q3Glc had no effect on homogeneous ice nucleation temperature, whereas K7Glc increased ice nucleation temperature. The results indicated that each flavonol glycoside affected ice nucleation by very complicated and varied reactions. More studies are necessary to determine the exact roles of these flavonol glycosides in deep supercooling of XPCs in which unidentified heterogeneous ice nucleators may exist. Copyright

Mechanism and Regulation of Protein Synthesis in Saccharomyces cerevisiae

PubMed Central

Dever, Thomas E.; Kinzy, Terri Goss; Pavitt, Graham D.

2016-01-01

In this review, we provide an overview of protein synthesis in the yeast Saccharomyces cerevisiae. The mechanism of protein synthesis is well conserved between yeast and other eukaryotes, and molecular genetic studies in budding yeast have provided critical insights into the fundamental process of translation as well as its regulation. The review focuses on the initiation and elongation phases of protein synthesis with descriptions of the roles of translation initiation and elongation factors that assist the ribosome in binding the messenger RNA (mRNA), selecting the start codon, and synthesizing the polypeptide. We also examine mechanisms of translational control highlighting the mRNA cap-binding proteins and the regulation of GCN4 and CPA1 mRNAs. PMID:27183566

Acute myotube protein synthesis regulation by IL-6-related cytokines.

PubMed

Gao, Song; Durstine, J Larry; Koh, Ho-Jin; Carver, Wayne E; Frizzell, Norma; Carson, James A

2017-11-01

IL-6 and leukemia inhibitory factor (LIF), members of the IL-6 family of cytokines, play recognized paradoxical roles in skeletal muscle mass regulation, being associated with both growth and atrophy. Overload or muscle contractions can induce a transient increase in muscle IL-6 and LIF expression, which has a regulatory role in muscle hypertrophy. However, the cellular mechanisms involved in this regulation have not been completely identified. The induction of mammalian target of rapamycin complex 1 (mTORC1)-dependent myofiber protein synthesis is an established regulator of muscle hypertrophy, but the involvement of the IL-6 family of cytokines in this process is poorly understood. Therefore, we investigated the acute effects of IL-6 and LIF administration on mTORC1 signaling and protein synthesis in C2C12 myotubes. The role of glycoprotein 130 (gp130) receptor and downstream signaling pathways, including phosphoinositide 3-kinase (PI3K)-Akt-mTORC1 and signal transducer and activator of transcription 3 (STAT3)-suppressor of cytokine signaling 3 (SOCS3), was investigated by administration of specific siRNA or pharmaceutical inhibitors. Acute administration of IL-6 and LIF induced protein synthesis, which was accompanied by STAT3 activation, Akt-mTORC1 activation, and increased SOCS3 expression. This induction of protein synthesis was blocked by both gp130 siRNA knockdown and Akt inhibition. Interestingly, STAT3 inhibition or Akt downstream mTORC1 signaling inhibition did not fully block the IL-6 or LIF induction of protein synthesis. SOCS3 siRNA knockdown increased basal protein synthesis and extended the duration of the protein synthesis induction by IL-6 and LIF. These results demonstrate that either IL-6 or LIF can activate gp130-Akt signaling axis, which induces protein synthesis via mTORC1-independent mechanisms in cultured myotubes. However, IL-6- or LIF-induced SOCS3 negatively regulates the activation of myotube protein synthesis. Copyright © 2017 the

Flavonols Protect Against UV Radiation-Induced Thymine Dimer Formation in an Artificial Skin Mimic.

PubMed

Maini, Sabia; Fahlman, Brian M; Krol, Ed S

2015-01-01

Exposure of skin to ultraviolet light has been shown to have a number of deleterious effects including photoaging, photoimmunosuppression and photoinduced DNA damage which can lead to the development of skin cancer. In this paper we present a study on the ability of three flavonols to protect EpiDerm™, an artificial skin mimic, against UV-induced damage. EpiDerm™ samples were treated with flavonol in acetone and exposed to UVA (100 kJ/m(2) at 365 nm) and UVB (9000 J/m(2) at 310 nm) radiation. Secretion of matrix metalloproteinase-1 (MMP-1) and tumor necrosis factor-α (TNF-a) were determined by ELISA, cyclobutane pyrimidine dimers were quantified using LC-APCI-MS. EpiDerm™ treated topically with quercetin significantly decreased MMP-1 secretion induced by UVA (100 µM) or UVB (200 µM) and TNF-a secretion was significantly reduced at 100 µM quercetin for both UVA and UVB radiation. In addition, topically applied quercetin was found to be photostable over the duration of the experiment. EpiDerm™ samples were treated topically with quercetin, kaempferol or galangin (52 µM) immediately prior to UVA or UVB exposure, and the cyclobutane thymine dimers (T-T (CPD)) were quantified using an HPLC-APCI MS/MS method. All three flavonols significantly decreased T-T (CPD) formation in UVB irradiated EpiDerm™, however no effect could be observed for the UVA irradiation experiments as thymine dimer formation was below the limit of quantitation. Our results suggest that flavonols can provide protection against UV radiation-induced skin damage through both antioxidant activity and direct photo-absorption. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Two new flavonol glycosides from Gymnema sylvestre and Euphorbia ebracteolata.

PubMed

Liu, Xin; Ye, Wencai; Yu, Biao; Zhao, Shouxun; Wu, Houming; Che, Chuntao

2004-03-15

Two new flavonol glycosides, namely kaempferol 3-O-beta-D-glucopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->6)-beta-D-galactopyranoside (1) and quercetin 3-O-6"-(3-hydroxyl-3-methylglutaryl)-beta-D-glucopyranoside (2), have been isolated from the aerial parts of Gymnema sylvestre and Euphorbia ebracteolata, respectively. Their structures were determined on the basis of chemical and spectroscopic methods.

Regulation of Phospholipid Synthesis in Escherichia coli by Guanosine Tetraphosphate

PubMed Central

Merlie, John P.; Pizer, Lewis I.

1973-01-01

Phospholipid synthesis has been reported to be subject to stringent control in Escherichia coli. We present evidence that demonstrates a strict correlation between guanosine tetraphosphate accumulation and inhibition of phospholipid synthesis. In vivo experiments designed to examine the pattern of phospholipid labeling with 32P-inorganic phosphate and 32P-sn-glycerol-3-phosphate suggest that regulation must occur at the glycerol-3-phosphate acyltransferase step. Assay of phospholipid synthesis by cell-free extracts and semipurified preparations revealed that guanosine tetraphosphate inhibits at least two enzymes specific for the biosynthetic pathway, sn-glycerol-3-phosphate acyltransferase as well as sn-glycerol-3-phosphate phosphatidyl transferase. These findings provide a biochemical basis for the stringent control of lipid synthesis as well as regulation of steady-state levels of phospholipid in growing cells. Images PMID:4583220

Opposite effects on regulation of urea synthesis by early and late uraemia in rats.

PubMed

Nielsen, Susanne Schouw; Grøfte, Thorbjørn; Grønbaek, Henning; Tygstrup, Niels; Vilstrup, Hendrik

2007-04-01

Acute and chronic kidney failure lead to catabolism with loss of lean body mass. Up-regulation of hepatic urea synthesis may play a role for the loss of body nitrogen and for the level of uraemia. The aims were to investigate the effects of early and late experimental renal failure on the regulation of hepatic urea synthesis and the expression of urea cycle enzyme genes in the liver. We examined the in vivo capacity of urea nitrogen synthesis, mRNA levels of urea cycle enzyme genes, and N-balances 6 days and 21 days after 5/6th partial nephrectomy in rats, and compared these data with pair- and free-fed control animals. Compared with pair-fed animals, early uraemia halved the in vivo urea synthesis capacity and decreased urea gene expressions (P<0.05). In contrast, late uraemia up-regulated in vivo urea synthesis and expression of all urea genes (P<0.05), save that of the flux-generating enzyme carbamoyl phosphate synthetase. The N-balance in rats with early uraemia was markedly negative (P<0.05) and near zero in late uraemia. Early uraemia down-regulated urea synthesis, so hepatic ureagenesis was not in itself involved in the negative N-balance. In contrast, late uraemia up-regulated urea synthesis, which probably contributed towards the reduced N-balance of this condition. These time-dependent, opposite effects on the uraemia-induced regulation of urea synthesis in vivo were not related to food restriction and probably mostly reflected regulation on gene level.

Molecular insight into the inclusion of the dietary plant flavonol fisetin and its chromophore within a chemically modified γ-cyclodextrin: Multi-spectroscopic, molecular docking and solubility studies.

PubMed

Pahari, Biswapathik; Chakraborty, Sandipan; Sengupta, Pradeep K

2018-09-15

We explored the encapsulation of dietary plant flavonols fisetin and its chromophore 3-hydroxyflavone, within 2-hydroxypropyl-γ-cyclodextrin (HPγ-CDx) nano-cavity in aqueous solution using multi-spectroscopic approaches and molecular docking. Upon addition of HPγ-CDx, dramatic changes occur in the intrinsic 'two color' fluorescence behavior of the fluorophores. This is manifested by significant increase in the steady state fluorescence intensities, anisotropies, average fluorescence lifetimes and rotational correlation times. Furthermore, in the CDx environment, intrinsically achiral flavonols exhibit prominent induced circular dichroism bands. These findings indicate that the flavonol molecules spontaneously enter the relatively hydrophobic, chiral environment of the HPγ-CDx nano-cavities. Molecular docking computations corroborate the spectroscopic findings, and predict selectivity in orientation of the encapsulated flavonols. HPγ-CDx inclusion increases the aqueous solubility of individual flavonols ∼100-1000 times. The present study demonstrates that the hydroxypropyl substituent in γ-CDx controls the inclusion mode of the flavonols, leading to their enhanced solubilization and altered spectral signatures. Copyright © 2018 Elsevier Ltd. All rights reserved.

Regulation of Flavivirus RNA synthesis and replication

PubMed Central

Selisko, Barbara; Wang, Chunling; Harris, Eva; Canard, Bruno

2014-01-01

RNA synthesis and replication of the members of the Flavivirus genus (including dengue, West Nile and Japanese encephalitis viruses) is regulated by a wide variety of mechanisms and actors. These include the sequestration of the RNA-dependent RNA polymerase (RdRp) for functions other than RNA synthesis, regulatory interactions with other viral and host proteins within the replication complex (RC), and regulatory elements within the RNA genome itself. In this review, we discuss our current knowledge of the multiple levels at which Flavivirus RNA synthesis is controlled. We aim to bring together two active research fields: the structural and functional biology of individual proteins of the RC and the impressive wealth of knowledge acquired regarding the viral genomic RNA. PMID:25462437

Antioxidant activities and structural characterization of flavonol O-glycosides from seeds of Japanese horse chestnut (Aesculus turbinata BLUME).

PubMed

Kimura, Hideto; Ogawa, Satoshi; Ishihara, Tomoe; Maruoka, Mahoko; Tokuyama-Nakai, Shota; Jisaka, Mitsuo; Yokota, Kazushige

2017-08-01

We attempted to evaluate the contents and distribution of antioxidants in the whole seeds, seed shells, and peeled seeds of the Japanese horse chestnut. The seed shells exhibited the highest antioxidant activities due to the presence of highly polymeric proanthocyanidins as we have reported recently. On the other hand, the peeled seeds predominantly contained flavonols such as quercetin and kaempferol at a high level of 66.7% of total polyphenols, also contributing to the predominant antioxidant activities. The instrumental analysis of the extract from the whole seeds revealed the identification of eight flavonol O-glycosides, including six compounds with quercetin and two species with kaempferol as aglycones. The isolated species exhibited different antioxidant activities depending on the types of aglycones, glycosides, and acylated moieties. The results indicate that the peeled seeds are a good source of flavonol O-glycosides serving as antioxidants to be used for food additives and dietary supplements. Copyright © 2017 Elsevier Ltd. All rights reserved.

Systemic and cerebral exposure to and pharmacokinetics of flavonols and terpene lactones after dosing standardized Ginkgo biloba leaf extracts to rats via different routes of administration

PubMed Central

Chen, Feng; Li, Li; Xu, Fang; Sun, Yan; Du, Feifei; Ma, Xutao; Zhong, Chenchun; Li, Xiuxue; Wang, Fengqing; Zhang, Nating; Li, Chuan

2013-01-01

BACKGROUND AND PURPOSE Flavonols and terpene lactones are putatively responsible for the properties of Ginkgo biloba leaf extracts that relate to prevention and treatment of cardiovascular disease and cerebral insufficiency. Here, we characterized rat systemic and cerebral exposure to these ginkgo compounds after dosing, as well as the compounds’ pharmacokinetics. EXPERIMENTAL APPROACH Rats received single or multiple doses of ShuXueNing injection (prepared from GBE50 for intravenous administration) or GBE50 (a standardized extract of G. biloba leaves for oral administration). Brain delivery of the ginkgo compounds was assessed with microdialysis. Various rat samples were analysed using liquid chromatography/mass spectrometry. KEY RESULTS Slow terminal elimination features of the flavonols counterbalanced the influence of poor oral bioavailability on their systemic exposure levels, which also resulted in significant accumulation of the compounds in plasma during the subchronic treatment with ShuXueNing injection and GBE50. Unlike the flavonols, the terpene lactones had poor enterohepatic circulation due to their rapid renal excretion and unknown metabolism. The flavonol glycosides occurred as major forms in plasma after dosing with ShuXueNing injection, while the flavonol aglycone conjugates were predominant in plasma after dosing with GBE50. Cerebral exposure was negligible for the flavonols and low for the terpene lactones. CONCLUSION AND IMPLICATIONS Unlike the significant systemic exposure levels, the levels of cerebral exposure to the flavonols and terpene lactones are low. The elimination kinetic differences between the two classes of ginkgo compounds influence their relative systemic exposure levels. The information gained is relevant to linking ginkgo administration to the medicinal effects. PMID:23808355

Attenuation of airway smooth muscle contractility via flavonol-mediated inhibition of phospholipase-Cβ

PubMed Central

Brown, Amy; Danielsson, Jennifer; Townsend, Elizabeth A.; Zhang, Yi; Perez-Zoghbi, Jose F.; Emala, Charles W.

2016-01-01

Enhanced contractility of airway smooth muscle (ASM) is a major pathophysiological characteristic of asthma. Expanding the therapeutic armamentarium beyond β-agonists that target ASM hypercontractility would substantially improve treatment options. Recent studies have identified naturally occurring phytochemicals as candidates for acute ASM relaxation. Several flavonoids were evaluated for their ability to acutely relax human and murine ASM ex vivo and murine airways in vivo and were evaluated for their ability to inhibit procontractile signaling pathways in human ASM (hASM) cells. Two members of the flavonol subfamily, galangin and fisetin, significantly relaxed acetylcholine-precontracted murine tracheal rings ex vivo (n = 4 and n = 5, respectively, P < 0.001). Galangin and fisetin also relaxed acetylcholine-precontracted hASM strips ex vivo (n = 6–8, P < 0.001). Functional respiratory in vivo murine studies demonstrated that inhaled galangin attenuated the increase in lung resistance induced by inhaled methacholine (n = 6, P < 0.01). Both flavonols, galangin and fisetin, significantly inhibited purified phosphodiesterase-4 (PDE4) (n = 7, P < 0.05; n = 7, P < 0.05, respectively), and PLCβ enzymes (n = 6, P < 0.001 and n = 6, P < 0.001, respectively) attenuated procontractile Gq agonists' increase in intracellular calcium (n = 11, P < 0.001), acetylcholine-induced increases in inositol phosphates, and CPI-17 phosphorylation (n = 9, P < 0.01) in hASM cells. The prorelaxant effect retained in these structurally similar flavonols provides a novel pharmacological method for dual inhibition of PLCβ and PDE4 and therefore may serve as a potential treatment option for acute ASM constriction. PMID:26773068

Attenuation of airway smooth muscle contractility via flavonol-mediated inhibition of phospholipase-Cβ.

PubMed

Brown, Amy; Danielsson, Jennifer; Townsend, Elizabeth A; Zhang, Yi; Perez-Zoghbi, Jose F; Emala, Charles W; Gallos, George

2016-04-15

Enhanced contractility of airway smooth muscle (ASM) is a major pathophysiological characteristic of asthma. Expanding the therapeutic armamentarium beyond β-agonists that target ASM hypercontractility would substantially improve treatment options. Recent studies have identified naturally occurring phytochemicals as candidates for acute ASM relaxation. Several flavonoids were evaluated for their ability to acutely relax human and murine ASM ex vivo and murine airways in vivo and were evaluated for their ability to inhibit procontractile signaling pathways in human ASM (hASM) cells. Two members of the flavonol subfamily, galangin and fisetin, significantly relaxed acetylcholine-precontracted murine tracheal rings ex vivo (n = 4 and n = 5, respectively, P < 0.001). Galangin and fisetin also relaxed acetylcholine-precontracted hASM strips ex vivo (n = 6-8, P < 0.001). Functional respiratory in vivo murine studies demonstrated that inhaled galangin attenuated the increase in lung resistance induced by inhaled methacholine (n = 6, P < 0.01). Both flavonols, galangin and fisetin, significantly inhibited purified phosphodiesterase-4 (PDE4) (n = 7, P < 0.05; n = 7, P < 0.05, respectively), and PLCβ enzymes (n = 6, P < 0.001 and n = 6, P < 0.001, respectively) attenuated procontractile Gq agonists' increase in intracellular calcium (n = 11, P < 0.001), acetylcholine-induced increases in inositol phosphates, and CPI-17 phosphorylation (n = 9, P < 0.01) in hASM cells. The prorelaxant effect retained in these structurally similar flavonols provides a novel pharmacological method for dual inhibition of PLCβ and PDE4 and therefore may serve as a potential treatment option for acute ASM constriction. Copyright © 2016 the American Physiological Society.

Competition between anthocyanin and flavonol biosynthesis produces spatial pattern variation of floral pigments between Mimulus species

PubMed Central

Yuan, Yao-Wu; Rebocho, Alexandra B.; Sagawa, Janelle M.; Stanley, Lauren E.; Bradshaw, Harvey D.

2016-01-01

Flower color patterns have long served as a model for developmental genetics because pigment phenotypes are visually striking, yet generally not required for plant viability, facilitating the genetic analysis of color and pattern mutants. The evolution of novel flower colors and patterns has played a key role in the adaptive radiation of flowering plants via their specialized interactions with different pollinator guilds (e.g., bees, butterflies, birds), motivating the search for allelic differences affecting flower color pattern in closely related plant species with different pollinators. We have identified LIGHT AREAS1 (LAR1), encoding an R2R3-MYB transcription factor, as the causal gene underlying the spatial pattern variation of floral anthocyanin pigmentation between two sister species of monkeyflower: the bumblebee-pollinated Mimulus lewisii and the hummingbird-pollinated Mimulus cardinalis. We demonstrated that LAR1 positively regulates FLAVONOL SYNTHASE (FLS), essentially eliminating anthocyanin biosynthesis in the white region (i.e., light areas) around the corolla throat of M. lewisii flowers by diverting dihydroflavonol into flavonol biosynthesis from the anthocyanin pigment pathway. FLS is preferentially expressed in the light areas of the M. lewisii flower, thus prepatterning the corolla. LAR1 expression in M. cardinalis flowers is much lower than in M. lewisii, explaining the unpatterned phenotype and recessive inheritance of the M. cardinalis allele. Furthermore, our gene-expression analysis and genetic mapping results suggest that cis-regulatory change at the LAR1 gene played a critical role in the evolution of different pigmentation patterns between the two species. PMID:26884205

Competition between anthocyanin and flavonol biosynthesis produces spatial pattern variation of floral pigments between Mimulus species.

PubMed

Yuan, Yao-Wu; Rebocho, Alexandra B; Sagawa, Janelle M; Stanley, Lauren E; Bradshaw, Harvey D

2016-03-01

Flower color patterns have long served as a model for developmental genetics because pigment phenotypes are visually striking, yet generally not required for plant viability, facilitating the genetic analysis of color and pattern mutants. The evolution of novel flower colors and patterns has played a key role in the adaptive radiation of flowering plants via their specialized interactions with different pollinator guilds (e.g., bees, butterflies, birds), motivating the search for allelic differences affecting flower color pattern in closely related plant species with different pollinators. We have identified LIGHT AREAS1 (LAR1), encoding an R2R3-MYB transcription factor, as the causal gene underlying the spatial pattern variation of floral anthocyanin pigmentation between two sister species of monkeyflower: the bumblebee-pollinated Mimulus lewisii and the hummingbird-pollinated Mimulus cardinalis. We demonstrated that LAR1 positively regulates FLAVONOL SYNTHASE (FLS), essentially eliminating anthocyanin biosynthesis in the white region (i.e., light areas) around the corolla throat of M. lewisii flowers by diverting dihydroflavonol into flavonol biosynthesis from the anthocyanin pigment pathway. FLS is preferentially expressed in the light areas of the M. lewisii flower, thus prepatterning the corolla. LAR1 expression in M. cardinalis flowers is much lower than in M. lewisii, explaining the unpatterned phenotype and recessive inheritance of the M. cardinalis allele. Furthermore, our gene-expression analysis and genetic mapping results suggest that cis-regulatory change at the LAR1 gene played a critical role in the evolution of different pigmentation patterns between the two species.

Impact of Flavonols on Cardiometabolic Biomarkers: A Meta-Analysis of Randomized Controlled Human Trials to Explore the Role of Inter-Individual Variability

PubMed Central

Menezes, Regina; Rodriguez-Mateos, Ana; Kaltsatou, Antonia; González-Sarrías, Antonio; Greyling, Arno; Giannaki, Christoforos; Andres-Lacueva, Cristina; Milenkovic, Dragan; Gibney, Eileen R.; Dumont, Julie; Schär, Manuel; Garcia-Aloy, Mar; Palma-Duran, Susana Alejandra; Ruskovska, Tatjana; Maksimova, Viktorija; Combet, Emilie; Pinto, Paula

2017-01-01

Several epidemiological studies have linked flavonols with decreased risk of cardiovascular disease (CVD). However, some heterogeneity in the individual physiological responses to the consumption of these compounds has been identified. This meta-analysis aimed to study the effect of flavonol supplementation on biomarkers of CVD risk such as, blood lipids, blood pressure and plasma glucose, as well as factors affecting their inter-individual variability. Data from 18 human randomized controlled trials were pooled and the effect was estimated using fixed or random effects meta-analysis model and reported as difference in means (DM). Variability in the response of blood lipids to supplementation with flavonols was assessed by stratifying various population subgroups: age, sex, country, and health status. Results showed significant reductions in total cholesterol (DM = −0.10 mmol/L; 95% CI: −0.20, −0.01), LDL cholesterol (DM = −0.14 mmol/L; 95% CI: −0.21, 0.07), and triacylglycerol (DM = −0.10 mmol/L; 95% CI: −0.18, 0.03), and a significant increase in HDL cholesterol (DM = 0.05 mmol/L; 95% CI: 0.02, 0.07). A significant reduction was also observed in fasting plasma glucose (DM = −0.18 mmol/L; 95% CI: −0.29, −0.08), and in blood pressure (SBP: DM = −4.84 mmHg; 95% CI: −5.64, −4.04; DBP: DM = −3.32 mmHg; 95% CI: −4.09, −2.55). Subgroup analysis showed a more pronounced effect of flavonol intake in participants from Asian countries and in participants with diagnosed disease or dyslipidemia, compared to healthy and normal baseline values. In conclusion, flavonol consumption improved biomarkers of CVD risk, however, country of origin and health status may influence the effect of flavonol intake on blood lipid levels. PMID:28208791

Changes in tissue protein synthesis are involved in regulating urea synthesis in rats given proteins of different quality.

PubMed

Tujioka, Kazuyo; Lyou, Sunok; Sano, Atushi; Hayase, Kazutoshi; Yokogoshi, Hidehiko

2004-10-01

The purpose of present study was to determine whether the regulation of urea synthesis is mediated through changes in supply of amino acids by protein synthesis and whether the concentration of ammonia, or activities of amino acid catabolizing enzymes, regulate urea synthesis when the dietary protein quality is manipulated. Experiments were done on three groups of rats given diets containing 10 g gluten, 10 g casein or 10 g whole egg protein/100 g for 10 d. The urinary excretion of urea, and the liver concentrations of glutamate, serine and alanine increased with a decrease in quality of dietary protein. The fractional and absolute rates of protein synthesis in tissues declined with the decrease in quality of dietary protein quality. The ammonia concentration in plasma and liver, and activities of hepatic amino acid catabolizing enzymes was not related to urea excretion under these conditions. These results suggest that the lower protein synthesis seen in tissues of rats given the lower quality of protein is likely to be one of the factors to increasing the supply of amino acids and stimulating urea synthesis.

Myostatin inhibits eEF2K-eEF2 by regulating AMPK to suppress protein synthesis.

PubMed

Deng, Zhao; Luo, Pei; Lai, Wen; Song, Tongxing; Peng, Jian; Wei, Hong-Kui

2017-12-09

Growth of skeletal muscle is dependent on the protein synthesis, and the rate of protein synthesis is mainly regulated in the stage of translation initiation and elongation. Myostatin, a member of the transforming growth factor-β (TGF-β) superfamily, is a negative regulator of protein synthesis. C2C12 myotubes was incubated with 0, 0.01, 0.1, 1, 2, 3 μg/mL myostatin recombinant protein, and then we detected the rates of protein synthesis by the method of SUnSET. We found that high concentrations of myostatin (2 and 3 μg/mL) inhibited protein synthesis by blocking mTOR and eEF2K-eEF2 pathway, while low concentration of myostatin (0.01, 0.1 and 1 μg/mL) regulated eEF2K-eEF2 pathway activity to block protein synthesis without affected mTOR pathway, and myostatin inhibited eEF2K-eEF2 pathway through regulating AMPK pathway to suppress protein synthesis. It provided a new mechanism for myostatin regulating protein synthesis and treating muscle atrophy. Copyright © 2017. Published by Elsevier Inc.

UHPLC/PDA-ESI/MS analysis of the main berry and leaf flavonol glycosides from different Carpathian Hippophaë rhamnoides L. varieties.

PubMed

Pop, Raluca Maria; Socaciu, Carmen; Pintea, Adela; Buzoianu, Anca Dana; Sanders, Mark Gerardus; Gruppen, Harry; Vincken, Jean-Paul

2013-01-01

Sea buckthorn (Hippophaë rhamnoides L.) is known to be rich in many bioactive compounds (such as vitamins, phenolics, carotenoids) important for human health and nutrition. Among the phenolics, berries and leaves contain a wide range of flavonols that are good quality and authenticity biomarkers. To compare the composition of the main flavonols of Romanian sea buckthorn berry and leaf varieties and to identify the specific biomarkers that contribute to sample differentiation among varieties. Six varieties of cultivated sea buckthorn (ssp. Carpatica) berries and leaves were analysed by UHPLC/PDA-ESI/MS. Berries and leaves contained mainly isorhamnetin (I) glycosides in different ratios. Whereas I-3-neohesperidoside, I-3-glucoside, I-3-rhamnosylglucoside, I-3-sophoroside-7-rhamnoside and free isorhamnetin were predominant for berries (out of 17 compounds identified), I-3-rhamnosylglucoside, I-3-neohesperidoside, I-3-glucoside, quercetin-3-pentoside, kaempferol-3-rutinoside, and quercetin-3-glucoside were predominant in leaves (out of 19 compounds identified). Berries contained, on average, 917 mg/100 g DW flavonol glycosides. Leaves had higher content of flavonol glycosides than berries, on average 1118 mg/100 g DW. The variation of the quantitative dataset analysed using principal component analysis accounted for 91% of the total variance in the case of berries and 73% in case of leaves, demonstrating a good discrimination among samples. Based on quantitative analysis, by principal component analysis, the flavonol derivatives can be considered as biomarkers to discriminate among varieties and to recognise specifically the berry versus leaf composition. Copyright © 2013 John Wiley & Sons, Ltd.

Effect of flavonols on wine astringency and their interaction with human saliva.

PubMed

Ferrer-Gallego, Raúl; Brás, Natércia F; García-Estévez, Ignacio; Mateus, Nuno; Rivas-Gonzalo, Julián C; de Freitas, Victor; Escribano-Bailón, M Teresa

2016-10-15

The addition of external phenolic compounds to wines in order to improve their sensory quality is an established winemaking practice. This study was aimed at evaluating the effect of the addition of quercetin 3-O-glucoside on the astringency and bitterness of wines. Sensory results showed that the addition of this flavonol to wines results in an increase in astringency and bitterness. Additionally, flavonol-human salivary protein interactions were studied using fluorescence spectroscopy, dynamic light scattering and molecular dynamic simulations (MD). The apparent Stern-Volmer (KsvApp) and the apparent bimolecular quenching constants (kqApp) were calculated from fluorescence spectra. The KsvApp was 12620±390M(-1), and the apparent biomolecular constant was 3.94×10(12)M(-1)s(-1), which suggests that a complex was formed between the human salivary proteins and quercetin 3-O-glucoside. MD simulations showed that the quercetin 3-O-glucoside molecules have the ability to bind to the IB937 model peptide. Copyright © 2016 Elsevier Ltd. All rights reserved.

Systemic and cerebral exposure to and pharmacokinetics of flavonols and terpene lactones after dosing standardized Ginkgo biloba leaf extracts to rats via different routes of administration.

PubMed

Chen, Feng; Li, Li; Xu, Fang; Sun, Yan; Du, Feifei; Ma, Xutao; Zhong, Chenchun; Li, Xiuxue; Wang, Fengqing; Zhang, Nating; Li, Chuan

2013-09-01

Flavonols and terpene lactones are putatively responsible for the properties of Ginkgo biloba leaf extracts that relate to prevention and treatment of cardiovascular disease and cerebral insufficiency. Here, we characterized rat systemic and cerebral exposure to these ginkgo compounds after dosing, as well as the compounds' pharmacokinetics. Rats received single or multiple doses of ShuXueNing injection (prepared from GBE50 for intravenous administration) or GBE50 (a standardized extract of G. biloba leaves for oral administration). Brain delivery of the ginkgo compounds was assessed with microdialysis. Various rat samples were analysed using liquid chromatography/mass spectrometry. Slow terminal elimination features of the flavonols counterbalanced the influence of poor oral bioavailability on their systemic exposure levels, which also resulted in significant accumulation of the compounds in plasma during the subchronic treatment with ShuXueNing injection and GBE50. Unlike the flavonols, the terpene lactones had poor enterohepatic circulation due to their rapid renal excretion and unknown metabolism. The flavonol glycosides occurred as major forms in plasma after dosing with ShuXueNing injection, while the flavonol aglycone conjugates were predominant in plasma after dosing with GBE50. Cerebral exposure was negligible for the flavonols and low for the terpene lactones. Unlike the significant systemic exposure levels, the levels of cerebral exposure to the flavonols and terpene lactones are low. The elimination kinetic differences between the two classes of ginkgo compounds influence their relative systemic exposure levels. The information gained is relevant to linking ginkgo administration to the medicinal effects. © 2013 The Authors. British Journal of Pharmacology published by John Wiley &. Sons Ltd on behalf of The British Pharmacological Society.

Variation of Select Flavonols and Chlorogenic Acid Content of Elderberry Collected Throughout the Eastern United States

PubMed Central

Mudge, Elizabeth; Applequist, Wendy L.; Finley, Jamie; Lister, Patience; Townesmith, Andrew K.; Walker, Karen M.; Brown, Paula N.

2016-01-01

American elderberries are commonly collected from wild plants for use as food and medicinal products. The degree of phytochemical variation amongst wild populations has not been established and might affect the overall quality of elderberry dietary supplements. The three major flavonols identified in elderberries are rutin, quercetin and isoquercetin. Variation in the flavonols and chlorogenic acid was determined for 107 collections of elderberries from throughout the eastern United States using an optimized high performance liquid chromatography with ultraviolet detection method. The mean content was 71.9 mg per 100g fresh weight with variation ranging from 7.0 to 209.7 mg per 100 g fresh weight within the collected population. Elderberries collected from southeastern regions had significantly higher contents in comparison with those in more northern regions. The variability of the individual flavonol and chlorogenic acid profiles of the berries was complex and likely influenced by multiple factors. Several outliers were identified based on unique phytochemical profiles in comparison with average populations. This is the first study to determine the inherent variability of American elderberries from wild collections and can be used to identify potential new cultivars that may produce fruits of unique or high-quality phytochemical content for the food and dietary supplement industries. PMID:26877585

Proteomic and functional analyses reveal MAPK1 regulates milk protein synthesis.

PubMed

Lu, Li-Min; Li, Qing-Zhang; Huang, Jian-Guo; Gao, Xue-Jun

2012-12-27

L-Lysine (L-Lys) is an essential amino acid that plays fundamental roles in protein synthesis. Many nuclear phosphorylated proteins such as Stat5 and mTOR regulate milk protein synthesis. However, the details of milk protein synthesis control at the transcript and translational levels are not well known. In this current study, a two-dimensional gel electrophoresis (2-DE)/MS-based proteomic technology was used to identify phosphoproteins responsible for milk protein synthesis in dairy cow mammary epithelial cells (DCMECs). The effect of L-Lys on DCMECs was analyzed by CASY technology and reversed phase high performance liquid chromatography (RP-HPLC). The results showed that cell proliferation ability and β-casein expression were enhanced in DCMECs treated with L-Lys. By phosphoproteomics analysis, six proteins, including MAPK1, were identified up-expressed in DCMECs treated with 1.2 mM L-Lys for 24 h, and were verified by quantitative real-time PCR (qRT-PCR) and western blot. Overexpression and siRNA inhibition of MAPK1 experiments showed that MAPK1 upregulated milk protein synthesis through Stat5 and mTOR pathway. These findings that MAPK1 involves in regulation of milk synthesis shed new insights for understanding the mechanisms of milk protein synthesis.

Regulation of protein synthesis by amino acids in muscle of neonates

PubMed Central

Suryawan, Agus; Davis, Teresa A.

2011-01-01

The marked increase in skeletal muscle mass during the neonatal period is largely due to a high rate of postprandial protein synthesis that is modulated by an enhanced sensitivity to insulin and amino acids. The amino acid signaling pathway leading to the stimulation of protein synthesis has not been fully elucidated. Among the amino acids, leucine is considered to be a principal anabolic agent that regulates protein synthesis. mTORC1, which controls protein synthesis, has been implicated as a target for leucine. Until recently, there have been few studies exploring the role of amino acids in enhancing muscle protein synthesis in vivo. In this review, we discuss amino acid-induced protein synthesis in muscle in the neonate, focusing on current knowledge of the role of amino acids in the activation of mTORC1 leading to mRNA translation. The role of the amino acid transporters, SNAT2, LAT1, and PAT, in the modulation of mTORC1 activation and the role of amino acids in the activation of putative regulators of mTORC1, i.e., raptor, Rheb, MAP4K3, Vps34, and Rag GTPases, are discussed. PMID:21196241

A Gibberellin-Mediated DELLA-NAC Signaling Cascade Regulates Cellulose Synthesis in Rice.

PubMed

Huang, Debao; Wang, Shaogan; Zhang, Baocai; Shang-Guan, Keke; Shi, Yanyun; Zhang, Dongmei; Liu, Xiangling; Wu, Kun; Xu, Zuopeng; Fu, Xiangdong; Zhou, Yihua

2015-06-01

Cellulose, which can be converted into numerous industrial products, has important impacts on the global economy. It has long been known that cellulose synthesis in plants is tightly regulated by various phytohormones. However, the underlying mechanism of cellulose synthesis regulation remains elusive. Here, we show that in rice (Oryza sativa), gibberellin (GA) signals promote cellulose synthesis by relieving the interaction between SLENDER RICE1 (SLR1), a DELLA repressor of GA signaling, and NACs, the top-layer transcription factors for secondary wall formation. Mutations in GA-related genes and physiological treatments altered the transcription of CELLULOSE SYNTHASE genes (CESAs) and the cellulose level. Multiple experiments demonstrated that transcription factors NAC29/31 and MYB61 are CESA regulators in rice; NAC29/31 directly regulates MYB61, which in turn activates CESA expression. This hierarchical regulation pathway is blocked by SLR1-NAC29/31 interactions. Based on the results of anatomical analysis and GA content examination in developing rice internodes, this signaling cascade was found to be modulated by varied endogenous GA levels and to be required for internode development. Genetic and gene expression analyses were further performed in Arabidopsis thaliana GA-related mutants. Altogether, our findings reveal a conserved mechanism by which GA regulates secondary wall cellulose synthesis in land plants and provide a strategy for manipulating cellulose production and plant growth. © 2015 American Society of Plant Biologists. All rights reserved.

Regulation of Polyhydroxybutyrate Synthesis in the Soil Bacterium Bradyrhizobium diazoefficiens

PubMed Central

Quelas, J. I.; Mesa, S.; Mongiardini, E. J.; Jendrossek, D.

2016-01-01

ABSTRACT Polyhydroxybutyrate (PHB) is a carbon and energy reserve polymer in various prokaryotic species. We determined that, when grown with mannitol as the sole carbon source, Bradyrhizobium diazoefficiens produces a homopolymer composed only of 3-hydroxybutyrate units (PHB). Conditions of oxygen limitation (such as microoxia, oxic stationary phase, and bacteroids inside legume nodules) were permissive for the synthesis of PHB, which was observed as cytoplasmic granules. To study the regulation of PHB synthesis, we generated mutations in the regulator gene phaR and the phasin genes phaP1 and phaP4. Under permissive conditions, mutation of phaR impaired PHB accumulation, and a phaP1 phaP4 double mutant produced more PHB than the wild type, which was accumulated in a single, large cytoplasmic granule. Moreover, PhaR negatively regulated the expression of phaP1 and phaP4 as well as the expression of phaA1 and phaA2 (encoding a 3-ketoacyl coenzyme A [CoA] thiolases), phaC1 and phaC2 (encoding PHB synthases), and fixK2 (encoding a cyclic AMP receptor protein [CRP]/fumarate and nitrate reductase regulator [FNR]-type transcription factor of genes for microoxic lifestyle). In addition to the depressed PHB cycling, phaR mutants accumulated more extracellular polysaccharides and promoted higher plant shoot dry weight and competitiveness for nodulation than the wild type, in contrast to the phaC1 mutant strain, which is defective in PHB synthesis. These results suggest that phaR not only regulates PHB granule formation by controlling the expression of phasins and biosynthetic enzymes but also acts as a global regulator of excess carbon allocation and symbiosis by controlling fixK2. IMPORTANCE In this work, we investigated the regulation of polyhydroxybutyrate synthesis in the soybean-nodulating bacterium Bradyrhizobium diazoefficiens and its influence in bacterial free-living and symbiotic lifestyles. We uncovered a new interplay between the synthesis of this carbon reserve

Fatty acid, flavonol, and mineral composition variability among seven macrotyloma uniflorum (Lam.) verdc. accessions

USDA-ARS?s Scientific Manuscript database

Horse gram [Macrotyloma uniflorum (Lam.) Verdc.] seeds containing high concentrations of fatty acids, flavonols and minerals will provide government, public and private organizations with a nutritious and healthy food for use by malnourished and food deprived people worldwide. Seeds from seven horse...

Cloning and characterization of soybean gene Fg1 encoding flavonol 3-O-glucoside/galactoside (1→6) glucosyltransferase.

PubMed

Rojas Rodas, Felipe; Di, Shaokang; Murai, Yoshinori; Iwashina, Tsukasa; Sugawara, Satoko; Mori, Tetsuya; Nakabayashi, Ryo; Yonekura-Sakakibara, Keiko; Saito, Kazuki; Takahashi, Ryoji

2016-11-01

Flavonoids are important secondary metabolites in plants. Sugar-sugar glycosyltransferases are involved in the final step of flavonoid biosynthesis and contribute to the structural diversity of flavonoids. This manuscript describes the first cloning of a sugar-sugar glucosyltransferase gene in the UGT family that attaches glucose to the 6″-position of sugar bound to a flavonol. The results provide a glimpse on the possible evolution of sugar-sugar glycosyltransferase genes and identify putative amino acids responsible for the recognition of the hydroxyl group of the sugar moiety and specification of sugar. A scheme for the genetic control of flavonol glycoside biosynthesis is proposed. Flavonol glycosides (FGs) are predominant in soybean leaves and they show substantial differences among genotypes. In previous studies, we identified two flavonoid glycoside glycosyltransferase genes that segregated in recombinant inbred lines developed from a cross between cultivars Nezumisaya and Harosoy; one was responsible for the attachment of glucose to the 2″-position of glucose or galactose that is bound to the 3-position of kaempferol and the other was involved in the attachment of glucose to the 6″-position. This study was conducted to clone and characterize the 6″-glucosyltransferase gene. Linkage mapping indicated that the gene was located in the molecular linkage group I (chromosome 20). Based on the genome sequence, we cloned a candidate cDNA, GmF3G6"Gt from Harosoy but the corresponding cDNA could not be amplified by PCR from Nezumisaya. The coding region of GmF3G6″Gt in Harosoy is 1386 bp long encoding 462 amino acids. This gene was not expressed in leaves of Nezumisaya. The GmF3G6″Gt recombinant protein converted UDP-glucose and kaempferol 3-O-glucoside or kaempferol 3-O-galactoside to kaempferol 3-O-glucosyl-(1→6)-glucoside or kaempferol 3-O-glucosyl-(1→6)-galactoside, respectively. These results indicate that GmF3G6″Gt encodes a flavonol 3-O

Stimulation of secretion by the T84 colonic epithelial cell line with dietary flavonols.

PubMed

Nguyen, T D; Canada, A T; Heintz, G G; Gettys, T W; Cohn, J A

1991-06-15

Flavonols are dietary compounds widely distributed in plants and characterized by a 2-phenyl-benzo(alpha)pyrane nucleus possessing hydroxyl and ketone groups at positions 3 and 4, respectively. Kaempferol, quercetin, and myricetin are flavonols that are further mono-, di-, or trihydroxylated on the phenyl ring, respectively. To test whether these ingested flavonols might exert a direct secretory effect on intestinal epithelial cells, monolayers of the T84 colonocyte cell line were mounted in Ussing chambers and examined for ion transport response. Twenty minutes after addition of 100 microM quercetin to either the serosal or mucosal side, the short-circuit current change was maximal at 16.6 microA/cm2. Kaempferol was less potent than quercetin, while myricetin and glycosylated quercetin (rutin) did not induce secretion. The secretion induced by quercetin did not seem to be mediated by the reactive oxygen species generated by quercetin through auto-oxidation and/or redox cycling (superoxide, hydrogen peroxide, and the hydroxyl radical) because it was neither enhanced by iron, nor inhibited by desferroxamine B or catalase (alone or in combination with superoxide dismutase). Like vasoactive intestinal peptide, quercetin induced a secretory response that was inhibited by barium chloride and bumetanide, and which exhibited synergism with carbachol. Quercetin also stimulated a modest increase in intracellular cAMP levels and the phosphorylation of endogenous protein substrates for cAMP-dependent protein kinase. Thus, quercetin is a potent stimulus of colonocyte secretion that resembles secretagogues which act via a cAMP-mediated signaling pathway.

Reassessment of the Genetic Regulation of Fatty Acid Synthesis in Escherichia coli: Global Positive Control by the Dual Functional Regulator FadR

PubMed Central

My, L.; Ghandour Achkar, N.; Viala, J. P.

2015-01-01

ABSTRACT In Escherichia coli, the FadR transcriptional regulator represses the expression of fatty acid degradation (fad) genes. However, FadR is also an activator of the expression of fabA and fabB, two genes involved in unsaturated fatty acid synthesis. Therefore, FadR plays an important role in maintaining the balance between saturated and unsaturated fatty acids in the membrane. We recently showed that FadR also activates the promoter upstream of the fabH gene (L. My, B. Rekoske, J. J. Lemke, J. P. Viala, R. L. Gourse, and E. Bouveret, J Bacteriol 195:3784–3795, 2013, doi:10.1128/JB.00384-13). Furthermore, recent transcriptomic and proteomic data suggested that FadR activates the majority of fatty acid (FA) synthesis genes. In the present study, we tested the role of FadR in the expression of all genes involved in FA synthesis. We found that FadR activates the transcription of all tested FA synthesis genes, and we identified the FadR binding site for each of these genes. This necessitated the reassessment of the transcription start sites for accA and accB genes described previously, and we provide evidence for the presence of multiple promoters driving the expression of these genes. We showed further that regulation by FadR impacts the amount of FA synthesis enzymes in the cell. Our results show that FadR is a global regulator of FA metabolism in E. coli, acting both as a repressor of catabolism and an activator of anabolism, two directly opposing pathways. IMPORTANCE In most bacteria, a transcriptional regulator tunes the level of FA synthesis enzymes. Oddly, such a global regulator still was missing for E. coli, which nonetheless is one of the prominent model bacteria used for engineering biofuel production using the FA synthesis pathway. Our work identifies the FadR functional dual regulator as a global activator of almost all FA synthesis genes in E. coli. Because FadR also is the repressor of FA degradation, FadR acts both as a repressor and an activator

A Gibberellin-Mediated DELLA-NAC Signaling Cascade Regulates Cellulose Synthesis in Rice[OPEN

PubMed Central

Huang, Debao; Wang, Shaogan; Zhang, Baocai; Shang-Guan, Keke; Shi, Yanyun; Zhang, Dongmei; Liu, Xiangling; Wu, Kun; Xu, Zuopeng; Fu, Xiangdong; Zhou, Yihua

2015-01-01

Cellulose, which can be converted into numerous industrial products, has important impacts on the global economy. It has long been known that cellulose synthesis in plants is tightly regulated by various phytohormones. However, the underlying mechanism of cellulose synthesis regulation remains elusive. Here, we show that in rice (Oryza sativa), gibberellin (GA) signals promote cellulose synthesis by relieving the interaction between SLENDER RICE1 (SLR1), a DELLA repressor of GA signaling, and NACs, the top-layer transcription factors for secondary wall formation. Mutations in GA-related genes and physiological treatments altered the transcription of CELLULOSE SYNTHASE genes (CESAs) and the cellulose level. Multiple experiments demonstrated that transcription factors NAC29/31 and MYB61 are CESA regulators in rice; NAC29/31 directly regulates MYB61, which in turn activates CESA expression. This hierarchical regulation pathway is blocked by SLR1-NAC29/31 interactions. Based on the results of anatomical analysis and GA content examination in developing rice internodes, this signaling cascade was found to be modulated by varied endogenous GA levels and to be required for internode development. Genetic and gene expression analyses were further performed in Arabidopsis thaliana GA-related mutants. Altogether, our findings reveal a conserved mechanism by which GA regulates secondary wall cellulose synthesis in land plants and provide a strategy for manipulating cellulose production and plant growth. PMID:26002868

Syndecan-2 regulates melanin synthesis via protein kinase C βII-mediated tyrosinase activation.

PubMed

Jung, Hyejung; Chung, Heesung; Chang, Sung Eun; Choi, Sora; Han, Inn-Oc; Kang, Duk-Hee; Oh, Eok-Soo

2014-05-01

Syndecan-2, a transmembrane heparan sulfate proteoglycan that is highly expressed in melanoma cells, regulates melanoma cell functions (e.g. migration). Since melanoma is a malignant tumor of melanocytes, which largely function to synthesize melanin, we investigated the possible involvement of syndecan-2 in melanogenesis. Syndecan-2 expression was increased in human skin melanoma tissues compared with normal skin. In both mouse and human melanoma cells, siRNA-mediated knockdown of syndecan-2 was associated with reduced melanin synthesis, whereas overexpression of syndecan-2 increased melanin synthesis. Similar effects were also detected in human primary epidermal melanocytes. Syndecan-2 expression did not affect the expression of tyrosinase, a key enzyme in melanin synthesis, but instead enhanced the enzymatic activity of tyrosinase by increasing the membrane and melanosome localization of its regulator, protein kinase CβII. Furthermore, UVB caused increased syndecan-2 expression, and this up-regulation of syndecan-2 was required for UVB-induced melanin synthesis. Taken together, these data suggest that syndecan-2 regulates melanin synthesis and could be a potential therapeutic target for treating melanin-associated diseases. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

A new ultra-high pressure liquid chromatography method for the determination of antioxidant flavonol aglycones in six Lysimachia species.

PubMed

Tóth, Anita; Végh, Krisztina; Alberti, Ágnes; Béni, Szabolcs; Kéry, Ágnes

2016-10-01

UPLC-DAD method was developed and validated for the quantitative determination of free flavonol aglycones (kaempferol, quercetin and myricetin) after acidic hydrolysis in six Lysimachia species. Quantitative analyses showed that the amounts of various flavonol aglycones were significantly different in Lysimachia vulgaris, Lysimachia nummularia, Lysimachia punctata, Lysimachia christinae, Lysimachia ciliata and Lysimachia clethroides. The L. clethroides sample was found to be the richest in kaempferol (25.77 ± 1.29 μg/mg extract) and quercetin (97.67 ± 4.61 μg/mg extract), while the L. nummularia sample contained the highest amount of myricetin (20.79 ± 1.00 μg/mg extract). The antioxidant capacity of hydrolysed extracts was evaluated using in vitro DPPH(•) (2,2-diphenyl-1-picrylhydrazyl) and ABTS(•+) [2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid)] decolourisation tests. The observed radical scavenging capacities of the extracts showed a relationship with the measured flavonol aglycone content and composition. The acidic treatment resulted in an increased free radical scavenging activity compared to the untreated methanol extract.

Glucosinolates, myrosinase hydrolysis products, and flavonols found in rocket (Eruca sativa and Diplotaxis tenuifolia).

PubMed

Bell, Luke; Wagstaff, Carol

2014-05-21

Rocket species have been shown to have very high concentrations of glucosinolates and flavonols, which have numerous positive health benefits with regular consumption. This review highlights how breeders and processors of rocket species can utilize genomic and phytochemical research to improve varieties and enhance the nutritive benefits to consumers. Plant breeders are increasingly looking to new technologies such as HPLC, UPLC, LC-MS, and GC-MS to screen populations for their phytochemical content to inform plant selections. This paper collates the research that has been conducted to date in rocket and summarizes all glucosinolate and flavonol compounds identified in the species. The paper emphasizes the importance of the broad screening of populations for phytochemicals and myrosinase degradation products, as well as unique traits that may be found in underutilized gene bank resources. This review also stresses that collaboration with industrial partners is becoming essential for long-term plant breeding goals through research.

Physico-chemical and Biological Evaluation of Flavonols: Fisetin, Quercetin and Kaempferol Alone and Incorporated in beta Cyclodextrins.

PubMed

Corina, Danciu; Bojin, Florina; Ambrus, Rita; Muntean, Delia; Soica, Codruta; Paunescu, Virgil; Cristea, Mirabela; Pinzaru, Iulia; Dehelean, Cristina

2017-01-01

Fisetin,quercetin and kaempferol are among the important representatives of flavonols, biological active phytocomounds, with low water solubility. To evaluate the antimicrobial effect, respectively the antiproliferative and pro apoptotic activity on the B164A5 murine melanoma cell line of pure flavonols and their beta cyclodextrins complexes. Incorporation of fisetin, quercetin and kaempferol in beta cyclodextrins was proved by scanning electron microscopy (SEM), differencial scanning calorimetry (DSC) and X-ray powder diffraction (XRPD). Pure compounds and their complexes were tested for antiproliferative (MTT) and pro-apoptotic activity (Annexin V-PI) on the B164A5 murine melanoma cell line and for the antimicrobial properties (Disk Diffusion Method) on the selected strains. The phytocompounds presented in a different manner in vitro chemopreventive activity against B164A5 murine melanoma cell line and weak antimicrobial effect. The three flavonols: fisetin, quercetin and kaempferol were successfully incorporated in beta-cyclodextrin (BCD) and hydroxylpropyl-beta-cyclodextrin (HPBCD). Incorporation in beta cyclodextrins had a mix effect on the biological activity conducing to decrease, increase or consistent effect compared to pure phytocompound, depending on the screened process and on the chosen combination. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Phosphatidate Phosphatase Plays Role in Zinc-mediated Regulation of Phospholipid Synthesis in Yeast*

PubMed Central

Soto-Cardalda, Aníbal; Fakas, Stylianos; Pascual, Florencia; Choi, Hyeon-Son; Carman, George M.

2012-01-01

In the yeast Saccharomyces cerevisiae, the synthesis of phospholipids is coordinately regulated by mechanisms that control the homeostasis of the essential mineral zinc (Carman, G.M., and Han, G. S. (2007) Regulation of phospholipid synthesis in Saccharomyces cerevisiae by zinc depletion. Biochim. Biophys. Acta 1771, 322–330; Eide, D. J. (2009) Homeostatic and adaptive responses to zinc deficiency in Saccharomyces cerevisiae. J. Biol. Chem. 284, 18565–18569). The synthesis of phosphatidylcholine is balanced by the repression of CDP-diacylglycerol pathway enzymes and the induction of Kennedy pathway enzymes. PAH1-encoded phosphatidate phosphatase catalyzes the penultimate step in triacylglycerol synthesis, and the diacylglycerol generated in the reaction may also be used for phosphatidylcholine synthesis via the Kennedy pathway. In this work, we showed that the expression of PAH1-encoded phosphatidate phosphatase was induced by zinc deficiency through a mechanism that involved interaction of the Zap1p zinc-responsive transcription factor with putative upstream activating sequence zinc-responsive elements in the PAH1 promoter. The pah1Δ mutation resulted in the derepression of the CHO1-encoded phosphatidylserine synthase (CDP-diacylglycerol pathway enzyme) and loss of the zinc-mediated regulation of the enzyme. Loss of phosphatidate phosphatase also resulted in the derepression of the CKI1-encoded choline kinase (Kennedy pathway enzyme) but decreased the synthesis of phosphatidylcholine when cells were deficient of zinc. This result confirmed the role phosphatidate phosphatase plays in phosphatidylcholine synthesis via the Kennedy pathway. PMID:22128164

Influence of Light and Temperature on Gene Expression Leading to Accumulation of Specific Flavonol Glycosides and Hydroxycinnamic Acid Derivatives in Kale (Brassica oleracea var. sabellica).

PubMed

Neugart, Susanne; Krumbein, Angelika; Zrenner, Rita

2016-01-01

Light intensity and temperature are very important signals for the regulation of plant growth and development. Plants subjected to less favorable light or temperature conditions often respond with accumulation of secondary metabolites. Some of these metabolites have been identified as bioactive compounds, considered to exert positive effects on human health when consumed regularly. In order to test a typical range of growth parameters for the winter crop Brassica oleracea var. sabellica, plants were grown either at 400 μmol m(-2) s(-1) or 100 μmol m(-2) s(-1) at 10°C, or at 400 μmol m(-2) s(-1) with 5 or 15°C. The higher light intensity overall increased flavonol content of leaves, favoring the main quercetin glycosides, a caffeic acid monoacylated kaempferol triglycoside, and disinapoyl-gentiobiose. The higher temperature mainly increased the hydroxycinnamic acid derivative disinapoyl-gentiobiose, while at lower temperature synthesis is in favor of very complex sinapic acid acylated flavonol tetraglycosides such as kaempferol-3-O-sinapoyl-sophoroside-7-O-diglucoside. A global analysis of light and temperature dependent alterations of gene expression in B. oleracea var. sabellica leaves was performed with the most comprehensive Brassica microarray. When compared to the light experiment much less genes were differentially expressed in kale leaves grown at 5 or 15°C. A structured evaluation of differentially expressed genes revealed the expected enrichment in the functional categories of e.g. protein degradation at different light intensities or phytohormone metabolism at different temperature. Genes of the secondary metabolism namely phenylpropanoids are significantly enriched with both treatments. Thus, the genome of B. oleracea was screened for predicted genes putatively involved in the biosynthesis of flavonoids and hydroxycinnamic acid derivatives. All identified B. oleracea genes were analyzed for their most specific 60-mer oligonucleotides present on the

Influence of Light and Temperature on Gene Expression Leading to Accumulation of Specific Flavonol Glycosides and Hydroxycinnamic Acid Derivatives in Kale (Brassica oleracea var. sabellica)

PubMed Central

Neugart, Susanne; Krumbein, Angelika; Zrenner, Rita

2016-01-01

Light intensity and temperature are very important signals for the regulation of plant growth and development. Plants subjected to less favorable light or temperature conditions often respond with accumulation of secondary metabolites. Some of these metabolites have been identified as bioactive compounds, considered to exert positive effects on human health when consumed regularly. In order to test a typical range of growth parameters for the winter crop Brassica oleracea var. sabellica, plants were grown either at 400 μmol m−2 s−1 or 100 μmol m−2 s−1 at 10°C, or at 400 μmol m−2 s−1 with 5 or 15°C. The higher light intensity overall increased flavonol content of leaves, favoring the main quercetin glycosides, a caffeic acid monoacylated kaempferol triglycoside, and disinapoyl-gentiobiose. The higher temperature mainly increased the hydroxycinnamic acid derivative disinapoyl-gentiobiose, while at lower temperature synthesis is in favor of very complex sinapic acid acylated flavonol tetraglycosides such as kaempferol-3-O-sinapoyl-sophoroside-7-O-diglucoside. A global analysis of light and temperature dependent alterations of gene expression in B. oleracea var. sabellica leaves was performed with the most comprehensive Brassica microarray. When compared to the light experiment much less genes were differentially expressed in kale leaves grown at 5 or 15°C. A structured evaluation of differentially expressed genes revealed the expected enrichment in the functional categories of e.g. protein degradation at different light intensities or phytohormone metabolism at different temperature. Genes of the secondary metabolism namely phenylpropanoids are significantly enriched with both treatments. Thus, the genome of B. oleracea was screened for predicted genes putatively involved in the biosynthesis of flavonoids and hydroxycinnamic acid derivatives. All identified B. oleracea genes were analyzed for their most specific 60-mer oligonucleotides present on the

Roles of Transcriptional and Translational Control Mechanisms in Regulation of Ribosomal Protein Synthesis in Escherichia coli.

PubMed

Burgos, Hector L; O'Connor, Kevin; Sanchez-Vazquez, Patricia; Gourse, Richard L

2017-11-01

Bacterial ribosome biogenesis is tightly regulated to match nutritional conditions and to prevent formation of defective ribosomal particles. In Escherichia coli , most ribosomal protein (r-protein) synthesis is coordinated with rRNA synthesis by a translational feedback mechanism: when r-proteins exceed rRNAs, specific r-proteins bind to their own mRNAs and inhibit expression of the operon. It was recently discovered that the second messenger nucleotide guanosine tetra and pentaphosphate (ppGpp), which directly regulates rRNA promoters, is also capable of regulating many r-protein promoters. To examine the relative contributions of the translational and transcriptional control mechanisms to the regulation of r-protein synthesis, we devised a reporter system that enabled us to genetically separate the cis -acting sequences responsible for the two mechanisms and to quantify their relative contributions to regulation under the same conditions. We show that the synthesis of r-proteins from the S20 and S10 operons is regulated by ppGpp following shifts in nutritional conditions, but most of the effect of ppGpp required the 5' region of the r-protein mRNA containing the target site for translational feedback regulation and not the promoter. These results suggest that most regulation of the S20 and S10 operons by ppGpp following nutritional shifts is indirect and occurs in response to changes in rRNA synthesis. In contrast, we found that the promoters for the S20 operon were regulated during outgrowth, likely in response to increasing nucleoside triphosphate (NTP) levels. Thus, r-protein synthesis is dynamic, with different mechanisms acting at different times. IMPORTANCE Bacterial cells have evolved complex and seemingly redundant strategies to regulate many high-energy-consuming processes. In E. coli , synthesis of ribosomal components is tightly regulated with respect to nutritional conditions by mechanisms that act at both the transcription and translation steps. In

The Onion (Allium cepa L.) R2R3-MYB Gene MYB1 Regulates Anthocyanin Biosynthesis

PubMed Central

Schwinn, Kathy E.; Ngo, Hanh; Kenel, Fernand; Brummell, David A.; Albert, Nick W.; McCallum, John A.; Pither-Joyce, Meeghan; Crowhurst, Ross N.; Eady, Colin; Davies, Kevin M.

2016-01-01

Bulb color is an important consumer trait for onion (Allium cepa L., Allioideae, Asparagales). The bulbs accumulate a range of flavonoid compounds, including anthocyanins (red), flavonols (pale yellow), and chalcones (bright yellow). Flavonoid regulation is poorly characterized in onion and in other plants belonging to the Asparagales, despite being a major plant order containing many important crop and ornamental species. R2R3-MYB transcription factors associated with the regulation of distinct branches of the flavonoid pathway were isolated from onion. These belonged to sub-groups (SGs) that commonly activate anthocyanin (SG6, MYB1) or flavonol (SG7, MYB29) production, or repress phenylpropanoid/flavonoid synthesis (SG4, MYB4, MYB5). MYB1 was demonstrated to be a positive regulator of anthocyanin biosynthesis by the induction of anthocyanin production in onion tissue when transiently overexpressed and by reduction of pigmentation when transiently repressed via RNAi. Furthermore, ectopic red pigmentation was observed in garlic (Allium sativum L.) plants stably transformed with a construct for co-overexpression of MYB1 and a bHLH partner. MYB1 also was able to complement the acyanic petal phenotype of a defined R2R3-MYB anthocyanin mutant in Antirrhinum majus of the asterid clade of eudicots. The availability of sequence information for flavonoid-related MYBs from onion enabled phylogenetic groupings to be determined across monocotyledonous and dicotyledonous species, including the identification of characteristic amino acid motifs. This analysis suggests that divergent evolution of the R2R3-MYB family has occurred between Poaceae/Orchidaceae and Allioideae species. The DNA sequences identified will be valuable for future analysis of classical flavonoid genetic loci in Allium crops and will assist the breeding of these important crop species. PMID:28018399

The Onion (Allium cepa L.) R2R3-MYB Gene MYB1 Regulates Anthocyanin Biosynthesis.

PubMed

Schwinn, Kathy E; Ngo, Hanh; Kenel, Fernand; Brummell, David A; Albert, Nick W; McCallum, John A; Pither-Joyce, Meeghan; Crowhurst, Ross N; Eady, Colin; Davies, Kevin M

2016-01-01

Bulb color is an important consumer trait for onion ( Allium cepa L., Allioideae, Asparagales). The bulbs accumulate a range of flavonoid compounds, including anthocyanins (red), flavonols (pale yellow), and chalcones (bright yellow). Flavonoid regulation is poorly characterized in onion and in other plants belonging to the Asparagales, despite being a major plant order containing many important crop and ornamental species. R2R3-MYB transcription factors associated with the regulation of distinct branches of the flavonoid pathway were isolated from onion. These belonged to sub-groups (SGs) that commonly activate anthocyanin (SG6, MYB1) or flavonol (SG7, MYB29) production, or repress phenylpropanoid/flavonoid synthesis (SG4, MYB4, MYB5). MYB1 was demonstrated to be a positive regulator of anthocyanin biosynthesis by the induction of anthocyanin production in onion tissue when transiently overexpressed and by reduction of pigmentation when transiently repressed via RNAi. Furthermore, ectopic red pigmentation was observed in garlic ( Allium sativum L.) plants stably transformed with a construct for co-overexpression of MYB1 and a bHLH partner. MYB1 also was able to complement the acyanic petal phenotype of a defined R2R3-MYB anthocyanin mutant in Antirrhinum maju s of the asterid clade of eudicots. The availability of sequence information for flavonoid-related MYBs from onion enabled phylogenetic groupings to be determined across monocotyledonous and dicotyledonous species, including the identification of characteristic amino acid motifs. This analysis suggests that divergent evolution of the R2R3-MYB family has occurred between Poaceae/Orchidaceae and Allioideae species. The DNA sequences identified will be valuable for future analysis of classical flavonoid genetic loci in Allium crops and will assist the breeding of these important crop species.

Flower color changes in three Japanese hibiscus species: further quantitative variation of anthocyanin and flavonols.

PubMed

Shimokawa, Satoshi; Iwashina, Tsukasa; Murakami, Noriaki

2015-03-01

One anthocyanin and four flavonols were detected from the petals of Hibiscus hamabo, H. tiliaceus and H. glaber. They were identified as cyanidin 3-0- sambubioside, gossypetin 3-O-glucuronide-8-O-glucoside, quercetin 7-O-rutinoside, gossypetin 3-O-glucoside and gossypetin 8-O-glucuronide by UV spectra, LC-MS, acid hydrolysis and HPLC. The flavonoid composition was essentially the same among the petals ofH. hamabo, H. tiliaceus and H. glaber, and there was little quantitative variation, except for cyanidin 3-O-sambubioside, the content of which in the petals ofH. tiliaceus and H. glaber was much higher than in that of H. hamabo. Flower colors of H. tiliaceus and H. glaber change from yellow to red, and that of H. hamabo changes from yellow to orange. These changes were caused by contents of anthocyanin and flavonols, which increased after flowering of H. hamabo, H. tiliaceus and H. glaber.

Eukaryotic elongation factor 2 kinase regulates the synthesis of microtubule-related proteins in neurons.

PubMed

Kenney, Justin W; Genheden, Maja; Moon, Kyung-Mee; Wang, Xuemin; Foster, Leonard J; Proud, Christopher G

2016-01-01

Modulation of the elongation phase of protein synthesis is important for numerous physiological processes in both neurons and other cell types. Elongation is primarily regulated via eukaryotic elongation factor 2 kinase (eEF2K). However, the consequence of altering eEF2K activity on the synthesis of specific proteins is largely unknown. Using both pharmacological and genetic manipulations of eEF2K combined with two protein-labeling techniques, stable isotope labeling of amino acids in cell culture and bio-orthogonal non-canonical amino acid tagging, we identified a subset of proteins whose synthesis is sensitive to inhibition of eEF2K in murine primary cortical neurons. Gene ontology (GO) analyses indicated that processes related to microtubules are particularly sensitive to eEF2K inhibition. Our findings suggest that eEF2K likely contributes to neuronal function by regulating the synthesis of microtubule-related proteins. Modulation of the elongation phase of protein synthesis is important for numerous physiological processes in neurons. Here, using labeling of new proteins coupled with proteomic techniques in primary cortical neurons, we find that the synthesis of microtubule-related proteins is up-regulated by inhibition of elongation. This suggests that translation elongation is a key regulator of cytoskeletal dynamics in neurons. © 2015 The Authors. Journal of Neurochemistry published by John Wiley & Sons Ltd on behalf of International Society for Neurochemistry.

Inhibition of dipeptidyl peptidase activity by flavonol glycosides of guava (Psidium guajava L.): a key to the beneficial effects of guava in type II diabetes mellitus.

PubMed

Eidenberger, Thomas; Selg, Manuel; Krennhuber, Klaus

2013-09-01

Based on the traditional use in popular medicine, the effect of extracts from Psidium guajava L. leaves and of the main flavonol-glycoside components on dipeptidyl-peptidase IV (DP-IV), a key enzyme of blood glucose homoeostasis, has been investigated in-vitro. An ethanolic extract was prepared from dried, powdered leaves of guava and was found to contain seven main flavonol-glycosides, which were isolated by semipreparative HPLC and tested individually. The ethanolic guava leave extract was shown to exert a dose-dependent inhibition of DP-IV, with an IC50 of 380 μg/ml test assay solution. Also the individual flavonol-glycosides inhibited DP-IV dose-dependently, with variations of the effects by a factor of 10, and an overall effect accounting for 100% of that observed for the total guava extract. The recovery of individual flavonol-glycosides in CaCo-2 epithelial cells, a model of gastrointestinal tract absorption, amounted to 2.3-5.3% of the amount available for absorption over 60 min at 37°C. Copyright © 2013 Elsevier B.V. All rights reserved.

Total Phenolic, Phenolic Acid, Anthocyanin, Flavan-3-ol, and Flavonol Profiles and Antioxidant Properties of Pinto and Black Beans ( Phaseolus vulgaris L.) as Affected by Thermal Processing.

PubMed

Xu, Baojun; Chang, Sam K C

2009-06-10

The effects of boiling and steaming processes at atmospheric and high pressures on the phenolic components and antioxidant properties of pinto and black beans were investigated. In comparison to the original raw beans, all processing methods caused significant (p < 0.05) decreases in total phenolic content (TPC), total flavonoid content (TFC), condensed tannin content (CTC), monomeric anthocyanin content (MAC), DPPH free-radical scavenging activity (DPPH), ferric-reducing antioxidant power (FRAP), and oxygen radical absorbing capacity (ORAC) values in both pinto and black beans. Steaming processing resulted in a greater retention of TPC, DPPH, FRAP, and ORAC values than the boiling processes in both pinto and black beans. To further investigate how thermal processing affected phenolic compositions and to elucidate the contribution of individual phenolic compounds to antioxidant properties, phenolic acids, anthocyanins, flavan-3-ols, and flavonols were quantitatively analyzed by high-performance liquid chromatography (HPLC). All thermal processing significantly (p < 0.05) affected individual phenolic acids, anthocyanins, flavan-3-ols, and flavonols, significantly (p < 0.05) reduced total phenolic acid contents in both pinto and black beans and total flavonol contents in pinto beans, and dramatically reduced anthocyanin contents in black beans. Phenolic acids and flavonols may play important roles on the overall antioxidant activities of pinto beans, while anthocyanins, flavan-3-ols, and flavonols may play important roles on the overall antioxidant activities of black beans.

Biochemical and Molecular Characterization of a Flavonoid 3-O-glycosyltransferase Responsible for Anthocyanins and Flavonols Biosynthesis in Freesia hybrida

PubMed Central

Sun, Wei; Liang, Lingjie; Meng, Xiangyu; Li, Yueqing; Gao, Fengzhan; Liu, Xingxue; Wang, Shucai; Gao, Xiang; Wang, Li

2016-01-01

The glycosylation of flavonoids increases their solubility and stability in plants. Flowers accumulate anthocyanidin and flavonol glycosides which are synthesized by UDP-sugar flavonoid glycosyltransferases (UFGTs). In our previous study, a cDNA clone (Fh3GT1) encoding UFGT was isolated from Freesia hybrida, which was preliminarily proved to be invovled in cyanidin 3-O-glucoside biosynthesis. Here, a variety of anthocyanin and flavonol glycosides were detected in flowers and other tissues of F. hybrida, implying the versatile roles of Fh3GT1 in flavonoids biosynthesis. To further unravel its multi-functional roles, integrative analysis between gene expression and metabolites was investigated. The results showed expression of Fh3GT1 was positively related to the accumulation of anthocyanins and flavonol glycosides, suggesting its potential roles in the biosynthesis of both flavonoid glycosides. Subsequently, biochemical analysis results revealed that a broad range of flavonoid substrates including flavonoid not naturally occurred in F. hybrida could be recognized by the recombinant Fh3GT1. Both UDP-glucose and UDP-galactose could be used as sugar donors by recombinant Fh3GT1, although UDP-galactose was transferred with relatively low activity. Furthermore, regiospecificity analysis demonstrated that Fh3GT1 was able to glycosylate delphinidin at the 3-, 4-′, and 7- positions in a sugar-dependent manner. And the introduction of Fh3GT1 into Arabidopsis UGT78D2 mutant successfully restored the anthocyanins and flavonols phenotypes caused by lost-of-function of the 3GT, indicating that Fh3GT1 functions as a flavonoid 3-O-glucosyltransferase in vivo. In summary, these results demonstrate that Fh3GT1 is a flavonoid 3-O-glycosyltransferase using UDP-glucose as the preferred sugar donor and may involve in flavonoid glycosylation in F. hybrida. PMID:27064818

Biochemical and Molecular Characterization of a Flavonoid 3-O-glycosyltransferase Responsible for Anthocyanins and Flavonols Biosynthesis in Freesia hybrida.

PubMed

Sun, Wei; Liang, Lingjie; Meng, Xiangyu; Li, Yueqing; Gao, Fengzhan; Liu, Xingxue; Wang, Shucai; Gao, Xiang; Wang, Li

2016-01-01

The glycosylation of flavonoids increases their solubility and stability in plants. Flowers accumulate anthocyanidin and flavonol glycosides which are synthesized by UDP-sugar flavonoid glycosyltransferases (UFGTs). In our previous study, a cDNA clone (Fh3GT1) encoding UFGT was isolated from Freesia hybrida, which was preliminarily proved to be invovled in cyanidin 3-O-glucoside biosynthesis. Here, a variety of anthocyanin and flavonol glycosides were detected in flowers and other tissues of F. hybrida, implying the versatile roles of Fh3GT1 in flavonoids biosynthesis. To further unravel its multi-functional roles, integrative analysis between gene expression and metabolites was investigated. The results showed expression of Fh3GT1 was positively related to the accumulation of anthocyanins and flavonol glycosides, suggesting its potential roles in the biosynthesis of both flavonoid glycosides. Subsequently, biochemical analysis results revealed that a broad range of flavonoid substrates including flavonoid not naturally occurred in F. hybrida could be recognized by the recombinant Fh3GT1. Both UDP-glucose and UDP-galactose could be used as sugar donors by recombinant Fh3GT1, although UDP-galactose was transferred with relatively low activity. Furthermore, regiospecificity analysis demonstrated that Fh3GT1 was able to glycosylate delphinidin at the 3-, 4-', and 7- positions in a sugar-dependent manner. And the introduction of Fh3GT1 into Arabidopsis UGT78D2 mutant successfully restored the anthocyanins and flavonols phenotypes caused by lost-of-function of the 3GT, indicating that Fh3GT1 functions as a flavonoid 3-O-glucosyltransferase in vivo. In summary, these results demonstrate that Fh3GT1 is a flavonoid 3-O-glycosyltransferase using UDP-glucose as the preferred sugar donor and may involve in flavonoid glycosylation in F. hybrida.

The flavonol quercetin-3-glucoside inhibits cyanidin-3-glucoside absorption in vitro.

PubMed

Walton, Michaela C; McGhie, Tony K; Reynolds, Gordon W; Hendriks, Wouter H

2006-06-28

At present, little is known about the mechanisms responsible for intestinal absorption of anthocyanins (ACNs). For example, it has not yet been established if ACNs are absorbed through an active transport mechanism, such as the sodium-dependent glucose transporter (SGLT1), or by passive diffusion. Previously, we found that the absorption of ACNs differs between regions of the digestive tract and is maximal in the jejunum, suggesting that an active transport mechanism is involved. In the present study, we examined the effect of d-glucose (main substrate of SGLT1), phloridzin (inhibitor of SGLT1), and quercetin-3-glucose (Q3G, a flavonol) on the absorption of cyanidin-3-glucoside (C3G; approximately 5 micromol/L) by mouse jejunum mounted in Ussing chambers. We found that the presence of either D-glucose (10, 20, and 40 mmol/L) or phloridzin (50, 100, and 200 micromol/L) resulted in a small but insignificant inhibition of C3G disappearance from the mucosal solution (decrease of disappearance with glucose, 33%; with phloridzin, 18%; NS). However, when the flavonol Q3G (50 micromol/L) was added to the mucosal solution together with the C3G, the disappearance of C3G was significantly decreased (74%; p < 0.001), and Q3G disappeared instead. In addition, we found phloretin and quercetin, the aglycones of phloridzin and Q3G, respectively, present in the mucosal solution and tissue extracts, indicating hydrolysis of these compounds by the enterocytes of the jejunum. In contrast, the aglycone cyanidin was not detected at all. Our results show that in the mouse small intestine, ACN absorption is not solely dependent on the activity of the SGLT1 transporter, as d-glucose and phloridzin had only a slight effect on uptake. Q3G, however, clearly inhibited C3G disappearance. These results suggest that there might be a competitive inhibition between C3G and Q3G absorption. It is possible that an absorption mechanism other than the SGLT1 is involved, which has a structural preference

Identification, quantification and antioxidant activity of acylated flavonol glycosides from sea buckthorn (Hippophae rhamnoides ssp. sinensis).

PubMed

Chen, Chu; Xu, Xue-Min; Chen, Yang; Yu, Meng-Yao; Wen, Fei-Yan; Zhang, Hao

2013-12-01

A novel acylated flavonol glycoside: isorhamnetin (3-O-[(6-O-E-sinapoyl)-β-D-glucopyranosyl-(1→2)]-β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside) (1), together with two known acylated flavonol glycosides: quercetin (3-O-[(6-O-E-sinapoyl)-β-D-glucopyranosyl-(1→2)]-β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside) (2) and kaempferol (3-O-[(6-O-E-sinapoyl)-β-D-glucopyranosyl-(1→2)]-β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside) (3) were isolated from the n-butanol fraction of sea buckthorn (Hippophae rhamnoides ssp. sinensis) berries for the first time by chromatographic methods, and their structures were elucidated using UV, MS, (1)H and (13)C NMR, and 2D NMR. Compounds 1-3 showed good scavenging activities, with respective IC50 values of 8.91, 4.26 and 30.90 μM toward the 2,2'-diphenyl-1-picrylhydrazyl (DPPH) radical; respective Trolox equivalent antioxidant capacities of 2.89, 4.04 and 2.44 μM μM(-1) toward 2,2'-azino-bis-3-ethyl-benzothiazoline-6-sulphonate (ABTS) radical. The quantitative analysis of the isolated acylated flavonol glycosides was performed by HPLC-DAD method. The contents of compounds 1-3 were in the range of 12.2-31.4, 4.0-25.3, 7.5-59.7 mg/100 g dried berries and 9.1-34.5, 75.1-182.1, 29.2-113.4 mg/100 g dried leaves, respectively. Copyright © 2013. Published by Elsevier Ltd.

GC/MS-based metabolomic studies reveal key roles of glycine in regulating silk synthesis in silkworm, Bombyx mori.

PubMed

Chen, Quanmei; Liu, Xinyu; Zhao, Ping; Sun, Yanhui; Zhao, Xinjie; Xiong, Ying; Xu, Guowang; Xia, Qingyou

2015-02-01

Metabolic profiling of silkworm, especially the factors that affect silk synthesis at the metabolic level, is little known. Herein, metabolomic method based on gas chromatography-mass spectrometry was applied to identify key metabolic changes in silk synthesis deficient silkworms. Forty-six differential metabolites were identified in Nd group with the defect of silk synthesis. Significant changes in the levels of glycine and uric acid (up-regulation), carbohydrates and free fatty acids (down-regulation) were observed. The further metabolomics of silk synthesis deficient silkworms by decreasing silk proteins synthesis using knocking out fibroin heavy chain gene or extirpating silk glands operation showed that the changes of the metabolites were almost consistent with those of the Nd group. Furthermore, the increased silk yields by supplying more glycine or its related metabolite confirmed that glycine is a key metabolite to regulate silk synthesis. These findings provide important insights into the regulation between metabolic profiling and silk synthesis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Regulation of phospholipid synthesis in phosphatidylserine synthase-deficient (chol) mutants of Saccharomyces cerevisiae

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

Letts, V.A.; Henry, S.A.

1985-08-01

Saccharomyces cerevisiae mutants, chol, are deficient in the synthesis of the phospholipid phosphatidylserine owing to lowered activity of the membrane-associated enzyme phosphatidylserine synthase. These mutants are auxotrophic for ethanolamine or choline and, in the absence of these supplements, cannot synthesize phosphatidylethanolamine or phosphatidylcholine (PC). The authors exploited these characteristics of the chol mutants to examine the regulation of phospholipid metabolism in S. cerevisiae. Macromolecular synthesis and phospholipid metabolism were examined in chol cells starved for ethanolamine. Coupled to the decline in PC biosynthesis was a simultaneous decrease in the overall rate of phospholipid synthesis. In particular, the rate of synthesismore » of phosphatidylinositol decreased in parallel with the decline in PC biosynthesis. However, under conditions of ethanolamine deprivation in chol cells, the cytoplasmic enzyme inositol-1-phosphate synthase could not be repressed by exogenous inositol, and the endogenous synthesis of the phospholipid precursor inositol appeared to be elevated. The implications of these findings with respect to the coordinated regulation of phospholipid synthesis are discussed.« less

A new flavonol glucoside from the aerial parts of Sida glutinosa.

PubMed

Das, Niranjan; Achari, Basudev; Harigaya, Yoshihiro; Dinda, Biswanath

2011-10-01

Phytochemical investigation on the dried aerial parts of Sida glutinosa has led to the isolation of a new flavonol glucoside, glutinoside (1), along with seven known compounds, 24(28)-dehydromakisterone A (2), 1,2,3,9-tetrahydropyrrolo[2,1-b]-quinazolin-3-amine (3), docosanoic acid, 1-triacontanol, campesterol, stigmasterol, and β-sitosterol. The structures of these compounds were elucidated by means of extensive spectroscopic techniques as well as GC/MS analysis (for sterols) and comparison with the literature data. All these seven known compounds are reported from this plant for the first time.

Lewis lung carcinoma regulation of mechanical stretch-induced protein synthesis in cultured myotubes.

PubMed

Gao, Song; Carson, James A

2016-01-01

Mechanical stretch can activate muscle and myotube protein synthesis through mammalian target of rapamycin complex 1 (mTORC1) signaling. While it has been established that tumor-derived cachectic factors can induce myotube wasting, the effect of this catabolic environment on myotube mechanical signaling has not been determined. We investigated whether media containing cachectic factors derived from Lewis lung carcinoma (LLC) can regulate the stretch induction of myotube protein synthesis. C2C12 myotubes preincubated in control or LLC-derived media were chronically stretched. Protein synthesis regulation by anabolic and catabolic signaling was then examined. In the control condition, stretch increased mTORC1 activity and protein synthesis. The LLC treatment decreased basal mTORC1 activity and protein synthesis and attenuated the stretch induction of protein synthesis. LLC media increased STAT3 and AMP-activated protein kinase phosphorylation in myotubes, independent of stretch. Both stretch and LLC independently increased ERK1/2, p38, and NF-κB phosphorylation. In LLC-treated myotubes, the inhibition of ERK1/2 and p38 rescued the stretch induction of protein synthesis. Interestingly, either leukemia inhibitory factor or glycoprotein 130 antibody administration caused further inhibition of mTORC1 signaling and protein synthesis in stretched myotubes. AMP-activated protein kinase inhibition increased basal mTORC1 signaling activity and protein synthesis in LLC-treated myotubes, but did not restore the stretch induction of protein synthesis. These results demonstrate that LLC-derived cachectic factors can dissociate stretch-induced signaling from protein synthesis through ERK1/2 and p38 signaling, and that glycoprotein 130 signaling is associated with the basal stretch response in myotubes. Copyright © 2016 the American Physiological Society.

Simultaneous separation by reversed-phase high-performance liquid chromatography and mass spectral identification of anthocyanins and flavonols in Shiraz grape skin.

PubMed

Downey, Mark O; Rochfort, Simone

2008-08-01

A limitation of large-scale viticultural trials is the time and cost of comprehensive compositional analysis of the fruit by high-performance liquid chromatography (HPLC). In addition, separate methods have generally been required to identify and quantify different classes of metabolites. To address these shortcomings a reversed-phase HPLC method was developed to simultaneously separate the anthocyanins and flavonols present in grape skins. The method employs a methanol and water gradient acidified with 10% formic acid with a run-time of 48 min including re-equilibration. Identity of anthocyanins and flavonols in Shiraz (Vitis vinifera L.) skin was confirmed by mass spectral analysis.

VCP and ATL1 regulate endoplasmic reticulum and protein synthesis for dendritic spine formation

PubMed Central

Shih, Yu-Tzu; Hsueh, Yi-Ping

2016-01-01

Imbalanced protein homeostasis, such as excessive protein synthesis and protein aggregation, is a pathogenic hallmark of a range of neurological disorders. Here, using expression of mutant proteins, a knockdown approach and disease mutation knockin mice, we show that VCP (valosin-containing protein), together with its cofactor P47 and the endoplasmic reticulum (ER) morphology regulator ATL1 (Atlastin-1), regulates tubular ER formation and influences the efficiency of protein synthesis to control dendritic spine formation in neurons. Strengthening the significance of protein synthesis in dendritic spinogenesis, the translation blocker cyclohexamide and the mTOR inhibitor rapamycin reduce dendritic spine density, while a leucine supplement that increases protein synthesis ameliorates the dendritic spine defects caused by Vcp and Atl1 deficiencies. Because VCP and ATL1 are the causative genes of several neurodegenerative and neurodevelopmental disorders, we suggest that impaired ER formation and inefficient protein synthesis are significant in the pathogenesis of multiple neurological disorders. PMID:26984393

VCP and ATL1 regulate endoplasmic reticulum and protein synthesis for dendritic spine formation.

PubMed

Shih, Yu-Tzu; Hsueh, Yi-Ping

2016-03-17

Imbalanced protein homeostasis, such as excessive protein synthesis and protein aggregation, is a pathogenic hallmark of a range of neurological disorders. Here, using expression of mutant proteins, a knockdown approach and disease mutation knockin mice, we show that VCP (valosin-containing protein), together with its cofactor P47 and the endoplasmic reticulum (ER) morphology regulator ATL1 (Atlastin-1), regulates tubular ER formation and influences the efficiency of protein synthesis to control dendritic spine formation in neurons. Strengthening the significance of protein synthesis in dendritic spinogenesis, the translation blocker cyclohexamide and the mTOR inhibitor rapamycin reduce dendritic spine density, while a leucine supplement that increases protein synthesis ameliorates the dendritic spine defects caused by Vcp and Atl1 deficiencies. Because VCP and ATL1 are the causative genes of several neurodegenerative and neurodevelopmental disorders, we suggest that impaired ER formation and inefficient protein synthesis are significant in the pathogenesis of multiple neurological disorders.

Regulation of peroxisome proliferator-activated receptor gamma on milk fat synthesis in dairy cow mammary epithelial cells.

PubMed

Liu, Lili; Lin, Ye; Liu, Lixin; Wang, Lina; Bian, Yanjie; Gao, Xuejun; Li, Qingzhang

2016-12-01

Peroxisome proliferator-activated receptor gamma (PPARγ) participates in lipogenesis in rats, goats, and humans. However, the exact mechanism of PPARγ regulation on milk fat synthesis in dairy cow mammary epithelial cells (DCMECs) remains largely unexplored. The aim of this study was to investigate the role of PPARγ regarding milk fat synthesis in DCMECs and to ascertain whether milk fat precursor acetic acid and palmitic acid could interact with PPARγ signaling to regulate milk fat synthesis. For this study, we examined the effects of PPARγ overexpression and gene silencing on cell growth, triacylglycerol synthesis, and the messenger RNA (mRNA) and protein expression levels of genes involved in milk fat synthesis in DCMECs. In addition, we investigated the influences of acetic acid and palmitic acid on the mRNA and protein levels of milk lipogenic genes and triacylglycerol synthesis in DCMECs transfected with PPARγ small interfering RNA (siRNA) and PPARγ expression vector. The results showed that when PPARγ was silenced, cell viability, proliferation, and triacylglycerol secretion were obviously reduced. Gene silencing of PPARγ significantly downregulated the expression levels of milk fat synthesis-related genes in DCMECs. PPARγ overexpression improved cell viability, proliferation, and triacylglycerol secretion. The expression levels of milk lipogenic genes were significantly increased when PPARγ was overexpressed. Acetic acid and palmitic acid could markedly improve triacylglycerol synthesis and upregulate the expression levels of PPARγ and other lipogenic genes in DCMECs. These results suggest that PPARγ is a positive regulator of milk fat synthesis in DCMECs and that acetic acid and palmitic acid could partly regulate milk fat synthesis in DCMECs via PPARγ signaling.

Protective effects of red wine flavonols on 4-hydroxynonenal-induced apoptosis in PC12 cells.

PubMed

Jang, Young Jin; Kang, Nam Joo; Lee, Ki Won; Lee, Hyong Joo

2009-08-01

There is accumulating evidence that a moderate consumption of red wine has health benefits, such as the inhibition of neurodegenerative diseases. Although this is generally attributed to resveratrol, the protective mechanisms and the active substance(s) remain unclear. We examined whether and how red wine extract (RWE) and red wine flavonols quercetin and myricetin inhibited 4-hydroxynonenal (HNE)-induced apoptosis of rat pheochromocytoma PC12 cells. RWE attenuated HNE-induced PC12 cell death in a dose-dependent manner. HNE induced cleavage of poly(ADP-ribose) polymerase, which is involved in DNA repair in the nucleus, and this was inhibited by RWE treatment. Treatment with RWE also inhibited HNE-induced nuclear condensation in PC12 cells. Data of 2',7'-dichlorofluorescin diacetate showed that RWE protected against apoptosis of PC12 cells by attenuating intracellular reactive oxygen species. The cytoprotective effects on HNE-induced cell death were stronger for quercetin and myricetin than for resveratrol. HNE-induced nuclear condensation was attenuated by quercetin and myricetin. These results suggest that the neuroprotective potential of red wine is attributable to flavonols rather than to resveratrol.

Phytochrome regulation of cellulose synthesis in Arabidopsis.

PubMed

Bischoff, Volker; Desprez, Thierry; Mouille, Gregory; Vernhettes, Samantha; Gonneau, Martine; Höfte, Herman

2011-11-08

Plant development is highly plastic and dependent on light quantity and quality monitored by specific photoreceptors. Although we have a detailed knowledge of light signaling pathways, little is known about downstream targets involved in growth control. Cell size and shape are in part controlled by cellulose microfibrils extruded from large cellulose synthase complexes (CSCs) that migrate in the plasma membrane along cortical microtubules. Here we show a role for the red/far-red light photoreceptor PHYTOCHROME B (PHYB) in the regulation of cellulose synthesis in the growing Arabidopsis hypocotyl. In this organ, CSCs contains three distinct cellulose synthase (CESA) isoform classes: nonredundant CESA1 and CESA3 and a third class represented by partially redundant CESA2, CESA5, and CESA6. Interestingly, in the dark, depending on which CESA subunits occupy the third position, CSC velocity is more or less inhibited through an interaction with microtubules. Activation of PHYB overrules this inhibition. The analysis of cesa5 mutants shows a role for phosphorylation in the control of CSC velocity. These results, combined with the cesa5 mutant phenotype, suggest that cellulose synthesis is fine tuned through the regulated interaction of CSCs with microtubules and that PHYB signaling impinges on this process to maintain cell wall strength and growth in changing environments. Copyright © 2011 Elsevier Ltd. All rights reserved.

Identification and quantification of glucosinolate and flavonol compounds in rocket salad (Eruca sativa, Eruca vesicaria and Diplotaxis tenuifolia) by LC–MS: Highlighting the potential for improving nutritional value of rocket crops

PubMed Central

Bell, Luke; Oruna-Concha, Maria Jose; Wagstaff, Carol

2015-01-01

Liquid chromatography mass spectrometry (LC–MS) was used to obtain glucosinolate and flavonol content for 35 rocket accessions and commercial varieties. 13 glucosinolates and 11 flavonol compounds were identified. Semi-quantitative methods were used to estimate concentrations of both groups of compounds. Minor glucosinolate composition was found to be different between accessions; concentrations varied significantly. Flavonols showed differentiation between genera, with Diplotaxis accumulating quercetin glucosides and Eruca accumulating kaempferol glucosides. Several compounds were detected in each genus that have only previously been reported in the other. We highlight how knowledge of phytochemical content and concentration can be used to breed new, nutritionally superior varieties. We also demonstrate the effects of controlled environment conditions on the accumulations of glucosinolates and flavonols and explore the reasons for differences with previous studies. We stress the importance of consistent experimental design between research groups to effectively compare and contrast results. PMID:25442630

PERK Regulates Working Memory and Protein Synthesis-Dependent Memory Flexibility

PubMed Central

Zhu, Siying; Henninger, Keely; McGrath, Barbara C.; Cavener, Douglas R.

2016-01-01

PERK (EIF2AK3) is an ER-resident eIF2α kinase required for memory flexibility and metabotropic glutamate receptor-dependent long-term depression, processes known to be dependent on new protein synthesis. Here we investigated PERK’s role in working memory, a cognitive ability that is independent of new protein synthesis, but instead is dependent on cellular Ca2+ dynamics. We found that working memory is impaired in forebrain-specific Perk knockout and pharmacologically PERK-inhibited mice. Moreover, inhibition of PERK in wild-type mice mimics the fear extinction impairment observed in forebrain-specific Perk knockout mice. Our findings reveal a novel role of PERK in cognitive functions and suggest that PERK regulates both Ca2+ -dependent working memory and protein synthesis-dependent memory flexibility. PMID:27627766

Nondestructive Optical Sensing of Flavonols and Chlorophyll in White Head Cabbage (Brassica oleracea L. var. capitata subvar. alba) Grown under Different Nitrogen Regimens.

PubMed

Agati, Giovanni; Tuccio, Lorenza; Kusznierewicz, Barbara; Chmiel, Tomasz; Bartoszek, Agnieszka; Kowalski, Artur; Grzegorzewska, Maria; Kosson, Ryszard; Kaniszewski, Stanislaw

2016-01-13

A multiparametric optical sensor was used to nondestructively estimate phytochemical compounds in white cabbage leaves directly in the field. An experimental site of 1980 white cabbages (Brassica oleracea L. var. capitata subvar. alba), under different nitrogen (N) treatments, was mapped by measuring leaf transmittance and chlorophyll fluorescence screening in one leaf/cabbage head. The provided indices of flavonols (FLAV) and chlorophyll (CHL) displayed the opposite response to applied N rates, decreasing and increasing, respectively. The combined nitrogen balance index (NBI = CHL/FLAV) calculated was able to discriminate all of the plots under four N regimens (0, 100, 200, and 400 kg/ha) and was correlated with the leaf N content determined destructively. CHL and FLAV were properly calibrated against chlorophyll (R(2) = 0.945) and flavonol (R(2) = 0.932) leaf contents, respectively, by using a homographic fit function. The proposed optical sensing of cabbage crops can be used to estimate the N status of plants and perform precision fertilization to maintain acceptable crop yield levels and, additionally, to rapidly detect health-promoting flavonol antioxidants in Brassica plants.

Identification and quantification of glucosinolate and flavonol compounds in rocket salad (Eruca sativa, Eruca vesicaria and Diplotaxis tenuifolia) by LC-MS: highlighting the potential for improving nutritional value of rocket crops.

PubMed

Bell, Luke; Oruna-Concha, Maria Jose; Wagstaff, Carol

2015-04-01

Liquid chromatography mass spectrometry (LC-MS) was used to obtain glucosinolate and flavonol content for 35 rocket accessions and commercial varieties. 13 glucosinolates and 11 flavonol compounds were identified. Semi-quantitative methods were used to estimate concentrations of both groups of compounds. Minor glucosinolate composition was found to be different between accessions; concentrations varied significantly. Flavonols showed differentiation between genera, with Diplotaxis accumulating quercetin glucosides and Eruca accumulating kaempferol glucosides. Several compounds were detected in each genus that have only previously been reported in the other. We highlight how knowledge of phytochemical content and concentration can be used to breed new, nutritionally superior varieties. We also demonstrate the effects of controlled environment conditions on the accumulations of glucosinolates and flavonols and explore the reasons for differences with previous studies. We stress the importance of consistent experimental design between research groups to effectively compare and contrast results. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Higher dietary anthocyanin and flavonol intakes are associated with anti-inflammatory effects in a population of US adults.

PubMed

Cassidy, Aedin; Rogers, Gail; Peterson, Julia J; Dwyer, Johanna T; Lin, Honghuang; Jacques, Paul F

2015-07-01

Although growing evidence from trials and population-based studies has supported a protective role for flavonoids in relation to risk of certain chronic diseases, the underlying mechanisms remain unclear. Several previous studies focused on individual inflammatory biomarkers, but because of the limited specificity of any individual marker, an assessment of a combination of biomarkers may be more informative. We used an inflammation score (IS) that integrated 12 individual inflammatory biomarkers for the examination of associations with intakes of different flavonoid classes. The study was a cross-sectional analysis of 2375 Framingham Heart Study Offspring Cohort participants. Intakes of total flavonoids and their classes (anthocyanins, flavonols, flavanones, flavan-3-ols, polymers, and flavones) were calculated from validated food-frequency questionnaires. Individual inflammatory biomarkers were ranked, standardized, and summed to derive an overall IS and subgroup scores of functionally related biomarkers. In multivariate analyses, an inverse association between higher anthocyanin and flavonol intakes and IS was observed with a mean ± SE difference between quintile categories 5 and 1 of -1.48 ± 0.32 (P-trend ≤ 0.001) and -0.72 ± 0.33 (P-trend = 0.01), respectively. Results remained significant after additional adjustment for physical activity and vitamin C and fruit and vegetable intakes. Higher anthocyanin intake was inversely associated with all biomarker subgroups, whereas higher flavonol intake was associated only with lower cytokine and oxidative stress biomarker concentrations. In food-based analyses, higher intakes of apples and pears, red wine, and strawberries were associated with a lower IS with differences between quintiles 5 and 1 of -1.02 ± 0.43 (P = 0.006), -1.73 ± 0.39 (P < 0.001), and -0.44 ± 0.88 (P = 0.02), respectively. Although intakes of other classes were not associated with a reduction in overall IS, higher intakes of flavan-3-ols and

A general approach to quantification of hydroxycinnamic acid derivatives and flavones, flavonols, and their glycosides by UV spectrophotometry

USDA-ARS?s Scientific Manuscript database

A general method was developed for the quantification of hydroxycinnamic acid derivatives and flavones, flavonols, and their glycosides based on the UV molar relative response factors (MRRF) of the standards. Each of these phenolic compounds contains a cinnamoyl structure and has a maximum absorban...

Calyx diversity of flavonols and fatty acids in Roselle (Hibiscus sabdariffa) for use as a potential nutraceutical crop.

USDA-ARS?s Scientific Manuscript database

Flavonols and fatty acids in plants has potential to be used as an antioxidant, lowering of cholesterol, and for cancer prevention. Roselle is a photoperiod and frost-sensitive species requiring greenhouse production in the Griffin, GA environment. Six accessions of roselle calyces were evaluated fo...

Ikarisoside A inhibits acetylcholine-induced catecholamine secretion and synthesis by suppressing nicotinic acetylcholine receptor-ion channels in cultured bovine adrenal medullary cells.

PubMed

Li, Xiaojia; Toyohira, Yumiko; Horisita, Takafumi; Satoh, Noriaki; Takahashi, Keita; Zhang, Han; Iinuma, Munekazu; Yoshinaga, Yukari; Ueno, Susumu; Tsutsui, Masato; Sata, Takeyoshi; Yanagihara, Nobuyuki

2015-12-01

Ikarisoside A is a natural flavonol glycoside derived from plants of the genus Epimedium, which have been used in Traditional Chinese Medicine as tonics, antirheumatics, and aphrodisiacs. Here, we report the effects of ikarisoside A and three other flavonol glycosides on catecholamine secretion and synthesis in cultured bovine adrenal medullary cells. We found that ikarisoside A (1-100 μM), but not icariin, epimedin C, or epimedoside A, concentration-dependently inhibited the secretion of catecholamines induced by acetylcholine, a physiological secretagogue and agonist of nicotinic acetylcholine receptors. Ikarisoside A had little effect on catecholamine secretion induced by veratridine and 56 mM K(+). Ikarisoside A (1-100 μM) also inhibited (22)Na(+) influx and (45)Ca(2+) influx induced by acetylcholine in a concentration-dependent manner similar to that of catecholamine secretion. In Xenopus oocytes expressing α3β4 nicotinic acetylcholine receptors, ikarisoside A (0.1-100 μM) directly inhibited the current evoked by acetylcholine. It also suppressed (14)C-catecholamine synthesis and tyrosine hydroxylase activity induced by acetylcholine at 1-100 μM and 10-100 μM, respectively. The present findings suggest that ikarisoside A inhibits acetylcholine-induced catecholamine secretion and synthesis by suppression of nicotinic acetylcholine receptor-ion channels in bovine adrenal medullary cells.

Regulation of tyramine oxidase synthesis in Klebsiella aerogenes.

PubMed Central

Okamura, H; Murooka, Y; Harada, T

1976-01-01

Tyramine oxidase in Klebsiella aerogenes is highly specific for tyramine, dopamine, octopamine, and norepinephrine, and its synthesis is induced specifically by these compounds. The enzyme is present in a membrane-bound form. The Km value for tyramine is 9 X 10(-4) M. Tyramine oxidase synthesis was subjected to catabolite repression by glucose in the presence of ammonium salts. Addition of cyclic adenosine 3',5'-monophosphate (cAMP) overcame the catabolite repression. A mutant strain, K711, which can produce a high level of beta-galactosidase in the presence of glucose and ammonium chloride, can also synthesize tyramine oxidase and histidase in the presence of inducer in glucose ammonium medium. Catabolite repression of tyramine oxidase synthesis was relieved when the cells were grown under conditions of nitrogen limitation, whereas beta-galactosidase was strongly repressed under these conditions. A cAMP-requiring mutant, MK54, synthesized tyramine oxidase rapidly when tyramine was used as the sole source of nitrogen in the absence of cAMP. However, a glutamine synthetase-constitutive mutant, MK94, failed to synthesize tyramine oxidase in the presence of glucose and ammonium chloride, although it synthesized histidase rapidly under these conditions. These results suggest that catabolite repression of tyramine oxidase synthesis in K. aerogenes is regulated by the intracellular level of cAMP and an unknown cytoplasmic factor that acts independently of cAMP and is formed under conditions of nitrogen limitation. PMID:179974

Flavonol Glucoside and Antioxidant Enzyme Biosynthesis Affected by Mycorrhizal Fungi in Various Cultivars of Onion (Allium cepa L.).

PubMed

Mollavali, Mohanna; Bolandnazar, Saheb Ali; Schwarz, Dietmar; Rohn, Sascha; Riehle, Peer; Zaare Nahandi, Fariborz

2016-01-13

The objective of this study was to investigate the impact of mycorrhizal symbiosis on qualitative characteristics of onion (Allium cepa L.). For this reason, five onion cultivars with different scale color and three different strains of arbuscular mycorrhizal fungi (Diversispora versiformis, Rhizophagus intraradices, Funneliformis mosseae) were used. Red cultivars, mainly 'Red Azar-shahr', showed the highest content in vitamin C, flavonols, and antioxidant enzymes. Mycorrhizal inoculation increased total phenolic, pyruvic acid, and vitamin C of onion plants. Considerable increase was observed in quercetin-4'-O-monoglucoside and isorhamnetin-4'-O-monoglucoside content in plants inoculated with Diversispora versiformis, but quercetin-3,4'-O-diglucoside was not significantly influenced. Analyses for phenylalanine ammonia-lyase (PAL) and antioxiodant enzyme activities such as polyphenol oxidase (PPO), catalase (CAT), and peroxidase (POD) revealed that all except PPO were enhanced by mycorrhizal inoculation. Overall, these findings suggested that mycorrhizal inoculation influenced biosynthesis of flavonol glucosides and antioxidant enzymes by increasing nutrient uptake or by induction of the plant defense system.

Regulation of early mRNA synthesis after bacteriophage T4 infection of Escherichia coli.

PubMed Central

Linder, C H; Fast, R

1975-01-01

Regulation of T4-specific mRNA synthesis was studied during leucine starvation of a leucine-requiring stringent Escherichia coli B strain. This was done by imposing starvation prior to T4 infection and then letting RNA synthesis proceed for different time periods. Rifampin or streptolydigin was added to stop further RNA synthesis, and protein synthesis was restored by addition of leucine. Samples were withdrawn at different times, and the enzyme-forming capacities found that, during conditions which elicit the stringent response in uninfected bacteria, immediate early mRNA is not stringently regulated. This conclusion contradicts the earlier conclusion of others, obtained by measuring incorporation of radioactive uracil; this is explained by the observation of Edlin and Neuhard (1967), confirmed and extended by us to the T4-infected cell, that the incorporation of uracil into RNA of a stringent strain is virtually blocked by amino acid starvation, whereas that of adenine continues at 30 to 50% of the rate seen in the presence of the required amino acid. PMID:1099229

Regulation of monoclonal immunoglobulin G synthesis by antiidiotypic antibody in a patient with hypogammaglobulinemia.

PubMed Central

Mudawwar, F; Awdeh, Z; Ault, K; Geha, R S

1980-01-01

The regulation of in vitro antibody synthesis by antiidiotypic antibodies was studied in a child with hypogammaglobulinemia and a serum immunoglobulin (Ig)G1 kappa M component. A rabbit antiserum was raised against the purified M component and was rendered idiotype specific by extensive absorption with Cohn fraction II and with IgG derived from the patient's parents. Hemagglutination-inhibition studies demonstrated that less than 1 in 300,000 molecules of pooled human IgG carried M component-related idiotypic determinants. 12% of the patient's B cells, but none of her T cells, expressed idiotypic determinants on their surface. Spontaneous de novo synthesis of the M component by the patient's peripheral blood lymphocytes was demonstrated in vitro and was shown to proceed independently of the polyclonal activator pokeweed mitogen. Antiidiotypic rabbit IgG, but not its F(ab')2, fragments, profoundly inhibited the synthesis of M component by the patient's peripheral blood lymphocytes. We concluded that antiidiotypic antibodies may play a role in the regulation of antibody synthesis in man. PMID:6767740

Behavior of flavonols and carotenoids of minimally processed kale leaves during storage in passive modified atmosphere packaging.

PubMed

Kobori, C N; Huber, L S; Sarantópoulos, C I G L; Rodriguez-Amaya, D B

2011-03-01

Minimally processed kale leaves were packed in passive modified atmosphere and stored at 3 conditions: 1 °C in the dark and 11 °C with or without light exposure. The products were evaluated during storage in terms of headspace gas composition, sensory attributes, flavonol, and carotenoid contents. The sensory quality decreased slightly during 17 d at 1 °C in the dark. At 11 °C, the vegetable shelf life was predicted to be 6 d in the dark and 3 d with light. Quercetin and kaempferol were stable during storage for 15 d at 1 °C in the absence of light. At 11 °C in the dark, quercetin was stable during 10 d, increasing slightly on the 8th day. Kaempferol decreased up to the 5th day but increased on the 8th day, decreasing again on the 10th day. After 5 d at 11 °C under light, the flavonol levels were significantly higher than those of the initial values. Neoxanthin and violaxanthin did not change significantly after 15 d at 1 °C in the dark. Lutein and β-carotene, however, decreased 7.1% and 11.3%, respectively. At 11 °C in the dark, neoxanthin, violaxanthin, lutein, and β-carotene decreased 16.1%, 13.2%, 24.1%, and 23.7% after 10 d, respectively. At 11 °C under light, neoxanthin and lutein had a slight increase while violaxanthin and β-carotene decreased 23.1% and 16.5% after 5 d. Practical Application:  Passive modified atmosphere packaging together with refrigeration can extend the shelf life of minimally processed kale, retaining the health-promoting compounds, flavonols and carotenoids. Quercetin, kaempferol, neoxanthin, and violaxanthin are stable and lutein and β-carotene slightly reduced.

An effective identification and quantification method for Ginkgo biloba flavonol glycosides with targeted evaluation of adulterated products.

PubMed

Ma, Yuan-Chun; Mani, Ana; Cai, Yaling; Thomson, Jaclyn; Ma, Jie; Peudru, Flavie; Chen, Sarah; Luo, Mai; Zhang, Junzeng; Chapman, Robert G; Shi, Zhen-Tuo

2016-04-15

Ginkgo biloba L. (Ginkgoaceae) leaf extract is one of the most popular herbal products on the market, as it contains flavone glycosides (≥ 24%) and terpene lactones (≥ 6%), which are proposed to have significant physiological effects. Unfortunately, the challenging financial climate has resulted in a natural health product market containing adulterated ginkgo products. 42 ginkgo samples were analyzed to establish an HPLC profile for authentic ginkgo and common ginkgo adulterants, and to develop a method capable of easily detecting adulteration in ginkgo commercial products. In this study an efficient and targeted HPLC analysis method was established that is capable of distinguishing flavonol glycosides and aglycones simultaneously for the evaluation of ginkgo powdered extracts (PEs) and finished products in a single, 13 min run. Thirteen ginkgo leaf samples, fifteen standardized powdered extracts, and fourteen commercially available ginkgo products have been analyzed using this new HPLC method. Chromatograms were compared to six standard reference materials: one flavonol glycoside (rutin), three aglycones (quercetin, kaempferol and isorhamnetin), and two isoflavones (genestin and genistein). The quantitative chromatographic data was interpreted by principal component analysis (PCA), which assisted in the detection of unexpected chromatographic features in various adulterated botanical products. Only three of the commercially available ginkgo finished products tested in this study were determined to be authentic, with flavonol glycoside rutin, and aglycones quercetin, kaempferol, and isorhamnetin found to be common adulterants in the ginkgo powdered extract and finished product samples. Despite evidence of adulteration in most of the samples, each of the samples discussed herein met most of the current pharmacopeial standards. It is therefore critical that a preliminary evaluation be utilized to detect adulteration in commercial ginkgo products, prior to the

Direct analysis of 18 flavonol glycosides, aglycones and terpene trilactones in Ginkgo biloba tablets by matrix solid phase dispersion coupled with ultra-high performance liquid chromatography tandem triple quadrupole mass spectrometry.

PubMed

Liu, Xin-Guang; Yang, Hua; Cheng, Xiao-Lan; Liu, Lei; Qin, Yong; Wang, Qi; Qi, Lian-Wen; Li, Ping

2014-08-01

Analysis and quality control of Ginkgo biloba have been comprehensively studied. However, little attention has been devoted to the simultaneous extraction and analysis of flavonols and terpene trilactones, especially for direct quantification of flavonol glycosides. This work described a rapid strategy for one-step extraction and quantification of the components. A matrix solid phase dispersion (MSPD) method was designed for the extraction of ginkgo ingredients and compared with the heat-reflux and ultrasonic extraction methods. An ultra-high performance liquid chromatography (UHPLC)-tandem-triple-quadrupole-mass spectrometry (QQQ-MS) method was developed for detection of the 18 components, including 10 original flavonol glycosides, 3 aglycones, and 5 lactones. Subsequently, the proposed strategy was used for the analysis of 12 G. biloba tablets. Results showed that MSPD produced comparable extraction efficiency but consumed less time and required lower solvent volumes compared with conventional methods. Without hydrolysis, the concentration detected was much closer to the original in the sample. The total flavonol glycoside contents in ginkgo tablets ranged from 3.59 to 125.21μgmg(-1), and the terpene trilactone varied from 3.45 to 57.8μgmg(-1) among different manufacturers. In conclusion, the proposed MSPD and UHPLC-QQQ-MS is rapid and sensitive in providing comprehensive profile of chemical constituents especially the genuine flavonol glycosides for improved quality control of ginkgo products. Copyright © 2014 Elsevier B.V. All rights reserved.

Drosophila TIF-IA is required for ribosome synthesis and cell growth and is regulated by the TOR pathway.

PubMed

Grewal, Savraj S; Evans, Justin R; Edgar, Bruce A

2007-12-17

Synthesis of ribosomal RNA (rRNA) is a key step in ribosome biogenesis and is essential for cell growth. Few studies, however, have investigated rRNA synthesis regulation in vivo in multicellular organisms. Here, we present a genetic analysis of transcription initiation factor IA (TIF-IA), a conserved RNA polymerase I transcription factor. Drosophila melanogaster Tif-IA(-/-) mutants have reduced levels of rRNA synthesis and sustain a developmental arrest caused by a block in cellular growth. We find that the target of rapamycin (TOR) pathway regulates TIF-IA recruitment to rDNA. Furthermore, we show that the TOR pathway regulates rRNA synthesis in vivo and that TIF-IA overexpression can maintain rRNA transcription when TOR activity is reduced in developing larvae. We propose that TIF-IA acts in vivo as a downstream growth-regulatory target of the TOR pathway. Overexpression of TIF-IA also elevates levels of both 5S RNA and messenger RNAs encoding ribosomal proteins. Stimulation of rRNA synthesis by TIF-IA may therefore provide a feed-forward mechanism to coregulate the levels of other ribosome components.

Effect of solar radiation on the functional components of mulberry (Morus alba L.) leaves.

PubMed

Sugiyama, Mari; Katsube, Takuya; Koyama, Akio; Itamura, Hiroyuki

2016-08-01

The functional components of mulberry leaves have attracted the attention of the health food industry, and increasing their concentrations is an industry goal. This study investigated the effects of solar radiation, which may influence the production of flavonol and 1-deoxynojirimycin (DNJ) functional components in mulberry leaves, by comparing a greenhouse (poor solar radiation) and outdoor (rich solar radiation) setting. The level of flavonol in leaves cultivated in the greenhouse was markedly decreased when compared with those cultivated outdoors. In contrast, the DNJ content in greenhouse-cultivated plants was increased only slightly when compared with those cultivated outdoors. Interestingly, the flavonol content was markedly increased in the upper leaves of mulberry trees that were transferred from a greenhouse to the outdoors compared with those cultivated only in the outdoors. Solar radiation conditions influence the synthesis of flavonol and DNJ, the functional components of mulberry leaves. Under high solar radiation, the flavonol level becomes very high but the DNJ level becomes slightly lower, suggesting that the impact of solar radiation is great on flavonol but small on DNJ synthesis. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

The Flavonoid Pathway Regulates the Petal Colors of Cotton Flower

PubMed Central

Tan, Jiafu; Wang, Maojun; Tu, Lili; Nie, Yichun; Lin, Yongjun; Zhang, Xianlong

2013-01-01

Although biochemists and geneticists have studied the cotton flower for more than one century, little is known about the molecular mechanisms underlying the dramatic color change that occurs during its short developmental life following blooming. Through the analysis of world cotton germplasms, we found that all of the flowers underwent color changes post-anthesis, but there is a diverse array of petal colors among cotton species, with cream, yellow and red colors dominating the color scheme. Genetic and biochemical analyses indicated that both the original cream and red colors and the color changes post-anthesis were related to flavonoid content. The anthocyanin content and the expression of biosynthesis genes were both increased from blooming to one day post-anthesis (DPA) when the flower was withering and undergoing abscission. Our results indicated that the color changes and flavonoid biosynthesis of cotton flowers were precisely controlled and genetically regulated. In addition, flavonol synthase (FLS) genes involved in flavonol biosynthesis showed specific expression at 11 am when the flowers were fully opened. The anthocyanidin reductase (ANR) genes, which are responsible for proanthocyanidins biosynthesis, showed the highest expression at 6 pm on 0 DPA, when the flowers were withered. Light showed primary, moderate and little effects on flavonol, anthocyanin and proanthocyanidin biosynthesis, respectively. Flavonol biosynthesis was in response to light exposure, while anthocyanin biosynthesis was involved in flower color changes. Further expression analysis of flavonoid genes in flowers of wild type and a flavanone 3-hydroxylase (F3H) silenced line showed that the development of cotton flower color was controlled by a complex interaction between genes and light. These results present novel information regarding flavonoids metabolism and flower development. PMID:23951318

D-tyrosine negatively regulates melanin synthesis by competitively inhibiting tyrosinase activity.

PubMed

Park, Jisu; Jung, Hyejung; Kim, Kyuri; Lim, Kyung-Min; Kim, Ji-Young; Jho, Eek-Hoon; Oh, Eok-Soo

2018-05-01

Although L-tyrosine is well known for its melanogenic effect, the contribution of D-tyrosine to melanin synthesis was previously unexplored. Here, we reveal that, unlike L-tyrosine, D-tyrosine dose-dependently reduced the melanin contents of human MNT-1 melanoma cells and primary human melanocytes. In addition, 500 μM of D-tyrosine completely inhibited 10 μM L-tyrosine-induced melanogenesis, and both in vitro assays and L-DOPA staining MNT-1 cells showed that tyrosinase activity is reduced by D-tyrosine treatment. Thus, D-tyrosine appears to inhibit L-tyrosine-mediated melanogenesis by competitively inhibiting tyrosinase activity. Furthermore, we found that D-tyrosine inhibited melanogenesis induced by α-MSH treatment or UV irradiation, which are the most common environmental factors responsible for melanin synthesis. Finally, we confirmed that D-tyrosine reduced melanin synthesis in the epidermal basal layer of a 3D human skin model. Taken together, these data suggest that D-tyrosine negatively regulates melanin synthesis by inhibiting tyrosinase activity in melanocyte-derived cells. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Water deficits accelerate ripening and induce changes in gene expression regulating flavonoid biosynthesis in grape berries.

PubMed

Castellarin, Simone D; Matthews, Mark A; Di Gaspero, Gabriele; Gambetta, Gregory A

2007-12-01

Water deficits consistently promote higher concentrations of anthocyanins in red winegrapes and their wines. However, controversy remains as to whether there is any direct effect on berry metabolism other than inhibition of growth. Early (ED) and late (LD) season water deficits, applied before or after the onset of ripening (veraison), were imposed on field grown Vitis vinifera "Cabernet Sauvignon", and the responses of gene expression in the flavonoid pathway and their corresponding metabolites were determined. ED accelerated sugar accumulation and the onset of anthocyanin synthesis. Both ED and LD increased anthocyanin accumulation after veraison. Expression profiling revealed that the increased anthocyanin accumulation resulted from earlier and greater expression of the genes controlling flux through the anthocyanin biosynthetic pathway, including F3H, DFR, UFGT and GST. Increases in total anthocyanins resulted predominantly from an increase of 3'4'5'-hydroxylated forms through the differential regulation of F3'H and F3'5'H. There were limited effects on proanthocyanidin, other flavonols, and on expression of genes committed to their synthesis. These results demonstrate that manipulation of abiotic stress through applied water deficits not only modulates compositional changes during berry ripening, but also alters the timing of particular aspects of the ripening process.

Advanced Knowledge of Three Important Classes of Grape Phenolics: Anthocyanins, Stilbenes and Flavonols

PubMed Central

Flamini, Riccardo; Mattivi, Fulvio; De Rosso, Mirko; Arapitsas, Panagiotis; Bavaresco, Luigi

2013-01-01

Grape is qualitatively and quantitatively very rich in polyphenols. In particular, anthocyanins, flavonols and stilbene derivatives play very important roles in plant metabolism, thanks to their peculiar characteristics. Anthocyanins are responsible for the color of red grapes and wines and confer organoleptic characteristics on the wine. They are used for chemotaxonomic studies and to evaluate the polyphenolic ripening stage of grape. They are natural colorants, have antioxidant, antimicrobial and anticarcinogenic activity, exert protective effects on the human cardiovascular system, and are used in the food and pharmaceutical industries. Stilbenes are vine phytoalexins present in grape berries and associated with the beneficial effects of drinking wine. The principal stilbene, resveratrol, is characterized by anticancer, antioxidant, anti-inflammatory and cardioprotective activity. Resveratrol dimers and oligomers also occur in grape, and are synthetized by the vine as active defenses against exogenous attack, or produced by extracellular enzymes released from pathogens in an attempt to eliminate undesirable toxic compounds. Flavonols are a ubiquitous class of flavonoids with photo-protection and copigmentation (together with anthocyanins) functions. The lack of expression of the enzyme flavonoid 3′,5′-hydroxylase in white grapes restricts the presence of these compounds to quercetin, kaempferol and isorhamnetin derivatives, whereas red grapes usually also contain myricetin, laricitrin and syringetin derivatives. In the last ten years, the technological development of analytical instrumentation, particularly mass spectrometry, has led to great improvements and further knowledge of the chemistry of these compounds. In this review, the biosynthesis and biological role of these grape polyphenols are briefly introduced, together with the latest knowledge of their chemistry. PMID:24084717

Drosophila TIF-IA is required for ribosome synthesis and cell growth and is regulated by the TOR pathway

PubMed Central

Grewal, Savraj S.; Evans, Justin R.; Edgar, Bruce A.

2007-01-01

Synthesis of ribosomal RNA (rRNA) is a key step in ribosome biogenesis and is essential for cell growth. Few studies, however, have investigated rRNA synthesis regulation in vivo in multicellular organisms. Here, we present a genetic analysis of transcription initiation factor IA (TIF-IA), a conserved RNA polymerase I transcription factor. Drosophila melanogaster Tif-IA −/− mutants have reduced levels of rRNA synthesis and sustain a developmental arrest caused by a block in cellular growth. We find that the target of rapamycin (TOR) pathway regulates TIF-IA recruitment to rDNA. Furthermore, we show that the TOR pathway regulates rRNA synthesis in vivo and that TIF-IA overexpression can maintain rRNA transcription when TOR activity is reduced in developing larvae. We propose that TIF-IA acts in vivo as a downstream growth–regulatory target of the TOR pathway. Overexpression of TIF-IA also elevates levels of both 5S RNA and messenger RNAs encoding ribosomal proteins. Stimulation of rRNA synthesis by TIF-IA may therefore provide a feed-forward mechanism to coregulate the levels of other ribosome components. PMID:18086911

Regulation of Pectate Lyase Synthesis in Pseudomonas fluorescens and Erwinia carotovora

PubMed Central

Zucker, Milton; Hankin, Lester

1970-01-01

Inducible synthesis of extracellular pectate lyase occurs in Erwinia carotovora, a bacterial soft-rot pathogen of plants, and, to a lesser extent, in a nonpathogenic isolate of Pseudomonas fluorescens. A combination of pectin and a heat-labile factor in fresh potato tissue or acetone powders of the tissue provided the best carbon source for induction. Yields of inducible pectate lyase were much greater than those usually reported. The pathogen, but not the saprophyte, produced a small amount of constitutive enzyme when grown on glucose. The relatively low level or absence of constitutive synthesis in these bacteria did not result from catabolite repression. Attempts were made to relieve any existing catabolite repression by restricting growth through slow feeding of glucose or by growing the organisms on glycerol. These conditions did not significantly alter the differential rate of lyase synthesis compared with changes observed in the presence of inducers. Previous growth history did not affect induction in the pathogen. However, P. fluorescens previously cultured on glucose required 10 to 20 generations of growth on inducing medium before appreciable lyase synthesis occurred. Differences between the pathogen and nonpathogen suggest that regulation of pectate lyase synthesis is related to pathogenicity of soft-rot bacteria. PMID:5473883

A novel dimeric flavonol glycoside from Cynanchum acutum subsp. sibiricum.

PubMed

Yuan, Si-Wen; Dai, Wei; Pan, Xin-Hui; Lu, Yan; Chen, Dao-Feng; Wang, Qi

2018-06-11

A novel dimeric flavonol glycoside, Cynanflavoside A (1), together with six analogues, kaempferol-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranoside (2), quercetin-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranoside (3), kaempferol-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-xylopyranoside (4), quercetin-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-xylopyranoside (5), kaempferol-3-O-β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside (6), and quercetin-3-O-galactoside (7) were isolated from the n-butyl alcohol extract of Cynanchum acutum subsp. sibiricum. Their structures were determined spectroscopically and compared with previously reported spectral data. All compounds were evaluated for their anti-complementary activity in vitro, and only compound 5 exhibited anti-complement effects with CH 50 value of 0.33 mM.

mTOR signaling regulates myotube hypertrophy by modulating protein synthesis, rDNA transcription, and chromatin remodeling.

PubMed

von Walden, Ferdinand; Liu, Chang; Aurigemma, Nicole; Nader, Gustavo A

2016-10-01

Ribosome production is an early event during skeletal muscle hypertrophy and precedes muscle protein accretion. Signaling via mTOR is crucial for ribosome production and hypertrophy; however, the mechanisms by which it regulates these processes remain to be identified. Herein, we investigated the activation of mTOR signaling in hypertrophying myotubes and determined that mTOR coordinates various aspects of gene expression important for ribosome production. First, inhibition of translation with cycloheximide had a more potent effect on protein synthesis than rapamycin indicating that mTOR function during hypertrophy is not on general, but rather on specific protein synthesis. Second, blocking Pol II transcription had a similar effect as Rapamycin and, unexpectedly, revealed the necessity of Pol II transcription for Pol I transcription, suggesting that mTOR may regulate ribosome production also by controlling Class II genes at the transcriptional level. Third, Pol I activity is essential for rDNA transcription and, surprisingly, for protein synthesis as selective Pol I inhibition blunted rDNA transcription, protein synthesis, and the hypertrophic response of myotubes. Finally, mTOR has nuclear localization in muscle, which is not sensitive to rapamycin. Inhibition of mTOR signaling by rapamycin disrupted mTOR-rDNA promoter interaction and resulted in altered histone marks indicative of repressed transcription and formation of higher-order chromatin structure. Thus mTOR signaling appears to regulate muscle hypertrophy by affecting protein synthesis, Class I and II gene expression, and chromatin remodeling. Copyright © 2016 the American Physiological Society.

Flavonol content in the water extract of the mulberry (Morus alba L.) leaf and their antioxidant capacities.

PubMed

Kim, Gyo-Nam; Jang, Hae-Dong

2011-08-01

The biological activities of the mulberry (Morus alba L.) leaf have been attributed to its flavonoid content. The water extract of the mulberry leaf (WEML) was prepared by autoclaving at 121 °C for 15 min, and the flavonol content of the WEML was determined by HPLC The WEML contained 4 flavonols in the following order: quercetin-3-β-D-glucose (QT-G) > quercetin-3-O-glucose-6″-acetate (QT-GA) > rutin (RT) > quercetin (QT). In the oxygen radical absorbance capacity (ORAC) assay, QT had the highest peroxyl radical-scavenging capacity and a similar hydroxyl radical-scavenging capacity as its glycosides (QT-G, QT-GA, and RT). QT exhibited a stronger cellular antioxidant capacity (CAC) against 2,2'-Azobis(2-amidinopropane) dihydrochloride (AAPH)- and Cu²⁺-induced oxidative stress in HepG2 cells compared to its glycosides, indicating that the intracellular antioxidant capacity of QT and its glycosides may depend upon both the permeability across the cell membrane and the peroxyl or hydroxyl radical-scavenging capacity. The information presented might be used for developing mulberry leaf-based functional foods. © 2011 Institute of Food Technologists®

Amino acid metabolism in dairy cows and their regulation in milk synthesis.

PubMed

Wang, Feiran; Shi, Haitao; Wang, Shuxiang; Wang, Yajing; Cao, Zhijun; Li, Shengli

2018-06-10

Reducing dietary crude protein (CP) and supplementing with certain amino acids (AAs) has been known as a potential solution to improve nitrogen (N) efficiency in dairy production. Thus understanding how AAs are utilized in various sites along the gut is critical. AA flow from the intestine to portal-drained viscera (PDV) and liver then to the mammary gland was elaborated in this article. Recoveries in individual AA in PDV and liver seem to share similar AA pattern with input: output ratio in mammary gland, which subdivides essential AA (EAA) into two groups, lysine (Lys) and branched-chain AA (BCAA) in group 1, input: output ratio > 1; methionine (Met), histidine (His), phenylalanine (Phe) etc. in group 2, input: output ratio close to 1. AAs in the mammary gland are either utilized for milk protein synthesis or retained as body tissue, or catabolized. The fractional removal of AAs and the number and activity of AA transporters together contribute to the ability of AAs going through mammary cells. Mammalian target of rapamycin (mTOR) pathway is closely related to milk protein synthesis and provides alternatives for AA regulation of milk protein synthesis, which connects AA with lactose synthesis via α-lactalbumin (gene: LALBA) and links with milk fat synthesis via sterol regulatory element-binding transcription protein 1 (SREBP1) and peroxisome proliferator-activated receptor (PPAR). Overall, AA flow across various tissues reveal AA metabolism and utilization in dairy cows on one hand. While the function of AA in the biosynthesis of milk protein, fat and lactose at both transcriptional and posttranscriptional level from another angle provides the possibility for us to regulate them for higher efficiency. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

AMPD2 Regulates GTP Synthesis and is Mutated in a Potentially-Treatable Neurodegenerative Brainstem Disorder

PubMed Central

Akizu, Naiara; Cantagrel, Vincent; Schroth, Jana; Cai, Na; Vaux, Keith; McCloskey, Douglas; Naviaux, Robert K.; Vleet, Jeremy Van; Fenstermaker, Ali G.; Silhavy, Jennifer L.; Scheliga, Judith S.; Toyama, Keiko; Morisaki, Hiroko; Sonmez, Fatma Mujgan; Celep, Figen; Oraby, Azza; Zaki, Maha S.; Al-Baradie, Raidah; Faqeih, Eissa; Saleh, Mohammad; Spencer, Emily; Rosti, Rasim Ozgur; Scott, Eric; Nickerson, Elizabeth; Gabriel, Stacey; Morisaki, Takayuki; Holmes, Edward W.; Gleeson, Joseph G.

2013-01-01

Purine biosynthesis and metabolism, conserved in all living organisms, is essential for cellular energy homeostasis and nucleic acids synthesis. The de novo synthesis of purine precursors is under tight negative feedback regulation mediated by adenosine and guanine nucleotides. We describe a new distinct early-onset neurodegenerative condition resulting from mutations in the adenosine monophosphate deaminase 2 gene (AMPD2). Patients have characteristic brain imaging features of pontocerebellar hypoplasia (PCH), due to loss of brainstem and cerebellar parenchyma. We found that AMPD2 plays an evolutionary conserved role in the maintenance of cellular guanine nucleotide pools by regulating the feedback inhibition of adenosine derivatives on de novo purine synthesis. AMPD2 deficiency results in defective GTP-dependent initiation of protein translation, which can be rescued by administration of purine precursors. These data suggest AMPD2-related PCH as a new, potentially treatable early-onset neurodegenerative disease. PMID:23911318

Flavonols and ellagic acid derivatives in peels of different species of jabuticaba (Plinia spp.) identified by HPLC-DAD-ESI/MSn.

PubMed

Neves, Nathália de Andrade; Stringheta, Paulo César; Gómez-Alonso, Sergio; Hermosín-Gutiérrez, Isidro

2018-06-30

Extracts of jabuticaba peels show complex chromatographic profiles at 360 nm, with some peaks presenting UV-Vis spectra resembling those of flavonol glycosides and others resembling that of ellagic acid. The presence and tentative identification of these phenolic compounds were comprehensively studied in four species of Brazilian jabuticaba fruit - Plinia trunciflora, variety 'jabuticaba de cabinho'; P. caulifora, varieties 'jabuticaba paulista' and 'jabuticaba canaã-açu'; P. jaboticaba, variety 'jabuticaba sabará'; and P. phitrantha, variety 'jabuticaba branca-vinho' - using HPLC-DAD-ESI-MS n . Seventeen flavonols derived from quercetin and three from myricetin and eighteen derivatives of ellagic acid and eleven of methyl ellagic acid were detected. Most of them were newly described and mainly occurred in glycosylated and acylglycosylated forms. Some compounds were missing in one variety, such as the absence of methyl ellagic acid derivatives in 'jabuticaba branca-vinho', and others only appeared in one variety, thus suggesting potential capacity for varietal differentiation. Copyright © 2018 Elsevier Ltd. All rights reserved.

Calcium regulates glutamate dehydrogenase and poly-γ-glutamic acid synthesis in Bacillus natto.

PubMed

Meng, Yonghong; Dong, Guiru; Zhang, Chen; Ren, Yuanyuan; Qu, Yuling; Chen, Weifeng

2016-04-01

To study the effect of Ca(2+) on glutamate dehydrogenase (GDH) and its role in poly-γ-glutamic acid (γ-PGA) synthesis in Bacillus natto HSF 1410. When the concentration of Ca(2+) varied from 0 to 0.1 g/l in the growth medium of B. natto HSF 1410, γ-PGA production increased from 6.8 to 9.7 g/l, while GDH specific activity and NH4Cl consumption improved from 183 to 295 U/mg and from 0.65 to 0.77 g/l, respectively. GDH with α-ketoglutarate as substrate primarily used NADPH as coenzyme with a K m of 0.08 mM. GDH was responsible for the synthesis of endogenous glutamate. The specific activity of GDH remained essentially unchanged in the presence of CaCl2 (0.05-0.2 g/l) in vitro. However, the specific activity of GDH and its expression was significantly increased by CaCl2 in vivo. Therefore, the regulation of GDH and PGA synthesis by Ca(2+) is an intracellular process. Calcium regulation may be an effective approach for producing γ-PGA on an industrial scale.

Determination of catechins and flavonol glycosides in Chinese tea varieties.

PubMed

Wu, Chunyan; Xu, Hairong; Héritier, Julien; Andlauer, Wilfried

2012-05-01

A standardised profiling method based on high performance liquid chromatography combined with ultraviolet (UV) and mass spectrometric detection (MS) was established to analyse the phenolic compounds of selected tea varieties used for manufacturing of green, black and oolong teas. The composition and content of 24 tea constituents were analysed, including catechins, flavonol and flavones glycosides, phenolic acids and purine alkaloids. Each tea variety had a unique chemical profile. The compositions of catechins were lower in the tea varieties for green tea manufacturing, while the content of myricetin glycosides was the lowest in the tea variety for oolong tea manufacturing. The content of individual phenolic compounds in the selected tea varieties is highly variable. However, the content of total catechins is proposed to be helpful to classify tea according to the future application as non fermented green and fermented oolong or black tea. Copyright © 2011 Elsevier Ltd. All rights reserved.

AMPD2 regulates GTP synthesis and is mutated in a potentially treatable neurodegenerative brainstem disorder.

PubMed

Akizu, Naiara; Cantagrel, Vincent; Schroth, Jana; Cai, Na; Vaux, Keith; McCloskey, Douglas; Naviaux, Robert K; Van Vleet, Jeremy; Fenstermaker, Ali G; Silhavy, Jennifer L; Scheliga, Judith S; Toyama, Keiko; Morisaki, Hiroko; Sonmez, Fatma M; Celep, Figen; Oraby, Azza; Zaki, Maha S; Al-Baradie, Raidah; Faqeih, Eissa A; Saleh, Mohammed A M; Spencer, Emily; Rosti, Rasim Ozgur; Scott, Eric; Nickerson, Elizabeth; Gabriel, Stacey; Morisaki, Takayuki; Holmes, Edward W; Gleeson, Joseph G

2013-08-01

Purine biosynthesis and metabolism, conserved in all living organisms, is essential for cellular energy homeostasis and nucleic acid synthesis. The de novo synthesis of purine precursors is under tight negative feedback regulation mediated by adenosine and guanine nucleotides. We describe a distinct early-onset neurodegenerative condition resulting from mutations in the adenosine monophosphate deaminase 2 gene (AMPD2). Patients have characteristic brain imaging features of pontocerebellar hypoplasia (PCH) due to loss of brainstem and cerebellar parenchyma. We found that AMPD2 plays an evolutionary conserved role in the maintenance of cellular guanine nucleotide pools by regulating the feedback inhibition of adenosine derivatives on de novo purine synthesis. AMPD2 deficiency results in defective GTP-dependent initiation of protein translation, which can be rescued by administration of purine precursors. These data suggest AMPD2-related PCH as a potentially treatable early-onset neurodegenerative disease. Copyright © 2013 Elsevier Inc. All rights reserved.

Phosphatase control of 4E-BP1 phosphorylation state is central for glycolytic regulation of retinal protein synthesis.

PubMed

Gardner, Thomas W; Abcouwer, Steven F; Losiewicz, Mandy K; Fort, Patrice E

2015-09-15

Control of protein synthesis in insulin-responsive tissues has been well characterized, but relatively little is known about how this process is regulated in nervous tissues. The retina exhibits a relatively high protein synthesis rate, coinciding with high basal Akt and metabolic activities, with the majority of retinal ATP being derived from aerobic glycolysis. We examined the dependency of retinal protein synthesis on the Akt-mTOR signaling and glycolysis using ex vivo rat retinas. Akt inhibitors significantly reduced retinal protein synthesis but did not affect glycolytic lactate production. Surprisingly, the glycolytic inhibitor 2-deoxyglucose (2-DG) markedly inhibited Akt1 and Akt3 activities, as well as protein synthesis. The effects of 2-DG, and 2-fluorodeoxyglucose (2-FDG) on retinal protein synthesis correlated with inhibition of lactate production and diminished ATP content, with all these effects reversed by provision of d-mannose. 2-DG treatment was not associated with increased AMPK, eEF2, or eIF2α phosphorylation; instead, it caused rapid dephosphorylation of 4E-BP1. 2-DG reduced total mTOR activity by 25%, but surprisingly, it did not reduce mTORC1 activity, as indicated by unaltered raptor-associated mTOR autophosphorylation and ribosomal protein S6 phosphorylation. Dephosphorylation of 4E-BP1 was largely prevented by inhibition of PP1/PP2A phosphatases with okadaic acid and calyculin A, and inhibition of PPM1 phosphatases with cadmium. Thus, inhibition of retinal glycolysis diminished Akt and protein synthesis coinciding with accelerated dephosphorylation of 4E-BP1 independently of mTORC1. These results demonstrate a novel mechanism regulating protein synthesis in the retina involving an mTORC1-independent and phosphatase-dependent regulation of 4E-BP1. Copyright © 2015 the American Physiological Society.

Abscisic acid negatively regulates elicitor-induced synthesis of capsidiol in wild tobacco.

PubMed

Mialoundama, Alexis Samba; Heintz, Dimitri; Debayle, Delphine; Rahier, Alain; Camara, Bilal; Bouvier, Florence

2009-07-01

In the Solanaceae, biotic and abiotic elicitors induce de novo synthesis of sesquiterpenoid stress metabolites known as phytoalexins. Because plant hormones play critical roles in the induction of defense-responsive genes, we have explored the effect of abscisic acid (ABA) on the synthesis of capsidiol, the major wild tobacco (Nicotiana plumbaginifolia) sesquiterpenoid phytoalexin, using wild-type plants versus nonallelic mutants Npaba2 and Npaba1 that are deficient in ABA synthesis. Npaba2 and Npaba1 mutants exhibited a 2-fold higher synthesis of capsidiol than wild-type plants when elicited with either cellulase or arachidonic acid or when infected by Botrytis cinerea. The same trend was observed for the expression of the capsidiol biosynthetic genes 5-epi-aristolochene synthase and 5-epi-aristolochene hydroxylase. Treatment of wild-type plants with fluridone, an inhibitor of the upstream ABA pathway, recapitulated the behavior of Npaba2 and Npaba1 mutants, while the application of exogenous ABA reversed the enhanced synthesis of capsidiol in Npaba2 and Npaba1 mutants. Concomitant with the production of capsidiol, we observed the induction of ABA 8'-hydroxylase in elicited plants. In wild-type plants, the induction of ABA 8'-hydroxylase coincided with a decrease in ABA content and with the accumulation of ABA catabolic products such as phaseic acid and dihydrophaseic acid, suggesting a negative regulation exerted by ABA on capsidiol synthesis. Collectively, our data indicate that ABA is not required per se for the induction of capsidiol synthesis but is essentially implicated in a stress-response checkpoint to fine-tune the amplification of capsidiol synthesis in challenged plants.

A Heme-responsive Regulator Controls Synthesis of Staphyloferrin B in Staphylococcus aureus*♦

PubMed Central

Laakso, Holly A.; Marolda, Cristina L.; Pinter, Tyler B.; Stillman, Martin J.; Heinrichs, David E.

2016-01-01

Staphylococcus aureus possesses a multitude of mechanisms by which it can obtain iron during growth under iron starvation conditions. It expresses an effective heme acquisition system (the iron-regulated surface determinant system), it produces two carboxylate-type siderophores staphyloferrin A and staphyloferrin B (SB), and it expresses transporters for many other siderophores that it does not synthesize. The ferric uptake regulator protein regulates expression of genes encoding all of these systems. Mechanisms of fine-tuning expression of iron-regulated genes, beyond simple iron regulation via ferric uptake regulator, have not been uncovered in this organism. Here, we identify the ninth gene of the sbn operon, sbnI, as encoding a ParB/Spo0J-like protein that is required for expression of genes in the sbn operon from sbnD onward. Expression of sbnD–I is drastically decreased in an sbnI mutant, and the mutant does not synthesize detectable SB during early phases of growth. Thus, SB-mediated iron acquisition is impaired in an sbnI mutant strain. We show that the protein forms dimers and tetramers in solution and binds to DNA within the sbnC coding region. Moreover, we show that SbnI binds heme and that heme-bound SbnI does not bind DNA. Finally, we show that providing exogenous heme to S. aureus growing in an iron-free medium results in delayed synthesis of SB. This is the first study in S. aureus that identifies a DNA-binding regulatory protein that senses heme to control gene expression for siderophore synthesis. PMID:26534960

Comparative transcriptomic analysis of key genes involved in flavonoid biosynthetic pathway and identification of a flavonol synthase from Artemisia annua L.

PubMed

Liu, S; Liu, L; Tang, Y; Xiong, S; Long, J; Liu, Z; Tian, N

2017-07-01

The regulatory mechanism of flavonoids, which synergise anti-malarial and anti-cancer compounds in Artemisia annua, is still unclear. In this study, an anthocyanidin-accumulating mutant callus was induced from A. annua and comparative transcriptomic analysis of wild-type and mutant calli performed, based on the next-generation Illumina/Solexa sequencing platform and de novo assembly. A total of 82,393 unigenes were obtained and 34,764 unigenes were annotated in the public database. Among these, 87 unigenes were assigned to 14 structural genes involved in the flavonoid biosynthetic pathway and 37 unigenes were assigned to 17 structural genes related to metabolism of flavonoids. More than 30 unigenes were assigned to regulatory genes, including R2R3-MYB, bHLH and WD40, which might regulate flavonoid biosynthesis. A further 29 unigenes encoding flavonoid biosynthetic enzymes or transcription factors were up-regulated in the mutant, while 19 unigenes were down-regulated, compared with the wild type. Expression levels of nine genes involved in the flavonoid pathway were compared using semi-quantitative RT-PCR, and results were consistent with comparative transcriptomic analysis. Finally, a putative flavonol synthase gene (AaFLS1) was identified from enzyme assay in vitro and in vivo through heterogeneous expression, and confirmed comparative transcriptomic analysis of wild-type and mutant callus. The present work has provided important target genes for the regulation of flavonoid biosynthesis in A. annua. © 2017 German Botanical Society and The Royal Botanical Society of the Netherlands.

TIF-IA and Ebp1 regulate RNA synthesis in T cells.

PubMed

Saudemont, Aurore

2015-04-16

In this issue of Blood, Nguyen et al show that mycophenolic acid (MPA) induces GTP depletion, which inhibits the function of transcription initiation factor I (TIF-IA) and impacts the interaction of TIF-IA with ErbB3-binding protein 1 (Ebp1), a key in regulating proliferating cell nuclear antigen (PCNA) expression and ribosomal RNA (rRNA) synthesis in T cells during activation.

Transcriptional regulation of decreased protein synthesis during skeletal muscle unloading

NASA Technical Reports Server (NTRS)

Howard, G.; Steffen, J. M.; Geoghegan, T. E.

1989-01-01

The regulatory role of transcriptional alterations in unloaded skeletal muscles was investigated by determining levels of total muscle RNA and mRNA fractions in soleus, gastrocnemius, and extensor digitorum longus (EDL) of rats subjected to whole-body suspension for up to 7 days. After 7 days, total RNA and mRNA contents were lower in soleus and gastrocnemius, compared with controls, but the concentrations of both RNAs per g muscle were unaltered. Alpha-actin mRNA (assessed by dot hybridization) was significantly reduced in soleus after 1, 3, and 7 days of suspension and in gastrocnemius after 3 and 7 days, but was unchanged in EDL. Protein synthesis directed by RNA extracted from soleus and EDL indicated marked alteration in mRNAs coding for several small proteins. Results suggest that altered transcription and availability of specific mRNAs contribute significantly to the regulation of protein synthesis during skeletal muscle unloading.

The role of microtubules in the regulation of proteoglycan synthesis in chondrocytes under hydrostatic pressure.

PubMed

Jortikka, M O; Parkkinen, J J; Inkinen, R I; Kärner, J; Järveläinen, H T; Nelimarkka, L O; Tammi, M I; Lammi, M J

2000-02-15

Chondrocytes of the articular cartilage sense mechanical factors associated with joint loading, such as hydrostatic pressure, and maintain the homeostasis of the extracellular matrix by regulating the metabolism of proteoglycans (PGs) and collagens. Intermittent hydrostatic pressure stimulates, while continuous high hydrostatic pressure inhibits, the biosynthesis of PGs. High continuous hydrostatic pressure also changes the structure of cytoskeleton and Golgi complex in cultured chondrocytes. Using microtubule (MT)-affecting drugs nocodazole and taxol as tools we examined whether MTs are involved in the regulation of PG synthesis in pressurized primary chondrocyte monolayer cultures. Disruption of the microtubular array by nocodazole inhibited [(35)S]sulfate incorporation by 39-48%, while MT stabilization by taxol caused maximally a 17% inhibition. Continuous hydrostatic pressure further decreased the synthesis by 34-42% in nocodazole-treated cultures. This suggests that high pressure exerts its inhibitory effect through mechanisms independent of MTs. On the other hand, nocodazole and taxol both prevented the stimulation of PG synthesis by cyclic 0. 5 Hz, 5 MPa hydrostatic pressure. The drugs did not affect the structural and functional properties of the PGs, and none of the treatments significantly affected cell viability, as indicated by the high level of PG synthesis 24-48 h after the release of drugs and/or high hydrostatic pressure. Our data on two-dimensional chondrocyte cultures indicate that inhibition of PG synthesis by continuous high hydrostatic pressure does not interfere with the MT-dependent vesicle traffic, while the stimulation of synthesis by cyclic pressure does not occur if the dynamic nature of MTs is disturbed by nocodazole. Similar phenomena may operate in cartilage matrix embedded chondrocytes. Copyright 2000 Academic Press.

Biotin and Lipoic Acid: Synthesis, Attachment and Regulation

PubMed Central

Cronan, John E.

2014-01-01

Summary Two vitamins, biotin and lipoic acid, are essential in all three domains of life. Both coenzymes function only when covalently attached to key metabolic enzymes. There they act as “swinging arms” that shuttle intermediates between two active sites (= covalent substrate channeling) of key metabolic enzymes. Although biotin was discovered over 100 years ago and lipoic acid 60 years ago, it was not known how either coenzyme is made until recently. In Escherichia coli the synthetic pathways for both coenzymes have now been worked out for the first time. The late steps of biotin synthesis, those involved in assembling the fused rings, were well-described biochemically years ago, although recent progress has been made on the BioB reaction, the last step of the pathway in which the biotin sulfur moiety is inserted. In contrast, the early steps of biotin synthesis, assembly of the fatty acid-like “arm” of biotin were unknown. It has now been demonstrated that the arm is made by using disguised substrates to gain entry into the fatty acid synthesis pathway followed by removal of the disguise when the proper chain length is attained. The BioC methyltransferase is responsible for introducing the disguise and the BioH esterase for its removal. In contrast to biotin, which is attached to its cognate proteins as a finished molecule, lipoic acid is assembled on its cognate proteins. An octanoyl moiety is transferred from the octanoyl-ACP of fatty acid synthesis to a specific lysine residue of a cognate protein by the LipB octanoyl transferase followed by sulfur insertion at carbons C6 and C8 by the LipA lipoyl synthetase. Assembly on the cognate proteins regulates the amount of lipoic acid synthesized and thus there is no transcriptional control of the synthetic genes. In contrast transcriptional control of the biotin synthetic genes is wielded by a remarkably sophisticated, yet simple, system, exerted through BirA a dual function protein that both represses

Differential regulation of protein synthesis by amino acids and insulin in peripheral and visceral tissues of neonatal pigs

PubMed Central

Suryawan, Agus; O’Connor, Pamela M. J.; Bush, Jill A.; Nguyen, Hanh V.

2009-01-01

The high efficiency of protein deposition during the neonatal period is driven by high rates of protein synthesis, which are maximally stimulated after feeding. In the current study, we examined the individual roles of amino acids and insulin in the regulation of protein synthesis in peripheral and visceral tissues of the neonate by performing pancreatic glucose–amino acid clamps in overnight-fasted 7-day-old pigs. We infused pigs (n = 8–12/group) with insulin at 0, 10, 22, and 110 ng kg−0.66 min−1 to achieve ~0, 2, 6 and 30 μU ml−1 insulin so as to simulate below fasting, fasting, intermediate, and fed insulin levels, respectively. At each insulin dose, amino acids were maintained at the fasting or fed level. In conjunction with the highest insulin dose, amino acids were also allowed to fall below the fasting level. Tissue protein synthesis was measured using a flooding dose of L-[4-3H] phenylalanine. Both insulin and amino acids increased fractional rates of protein synthesis in longissimus dorsi, gastrocnemius, masseter, and diaphragm muscles. Insulin, but not amino acids, increased protein synthesis in the skin. Amino acids, but not insulin, increased protein synthesis in the liver, pancreas, spleen, and lung and tended to increase protein synthesis in the jejunum and kidney. Neither insulin nor amino acids altered protein synthesis in the stomach. The results suggest that the stimulation of protein synthesis by feeding in most tissues of the neonate is regulated by the post-prandial rise in amino acids. However, the feeding-induced stimulation of protein synthesis in skeletal muscles is independently mediated by insulin as well as amino acids. PMID:18683020

Aspergillus Volatiles Regulate Aflatoxin Synthesis and Asexual Sporulation in Aspergillus parasiticus▿

PubMed Central

Roze, Ludmila V.; Beaudry, Randolph M.; Arthur, Anna E.; Calvo, Ana M.; Linz, John E.

2007-01-01

Aspergillus parasiticus is one primary source of aflatoxin contamination in economically important crops. To prevent the potential health and economic impacts of aflatoxin contamination, our goal is to develop practical strategies to reduce aflatoxin synthesis on susceptible crops. One focus is to identify biological and environmental factors that regulate aflatoxin synthesis and to manipulate these factors to control aflatoxin biosynthesis in the field or during crop storage. In the current study, we analyzed the effects of aspergillus volatiles on growth, development, aflatoxin biosynthesis, and promoter activity in the filamentous fungus A. parasiticus. When colonies of Aspergillus nidulans and A. parasiticus were incubated in the same growth chamber, we observed a significant reduction in aflatoxin synthesis and asexual sporulation by A. parasiticus. Analysis of the headspace gases demonstrated that A. nidulans produced much larger quantities of 2-buten-1-ol (CA) and 2-ethyl-1-hexanol (EH) than A. parasiticus. In its pure form, EH inhibited growth and increased aflatoxin accumulation in A. parasiticus at all doses tested; EH also stimulated aflatoxin transcript accumulation. In contrast, CA exerted dose-dependent up-regulatory or down-regulatory effects on aflatoxin accumulation, conidiation, and aflatoxin transcript accumulation. Experiments with reporter strains carrying nor-1 promoter deletions and mutations suggested that the differential effects of CA were mediated through separate regulatory regions in the nor-1 promoter. The potential efficacy of CA as a tool for analysis of transcriptional regulation of aflatoxin biosynthesis is discussed. We also identify a novel, rapid, and reliable method to assess norsolorinic acid accumulation in solid culture using a Chroma Meter CR-300 apparatus. PMID:17890344

UHPLC-PDA-ESI/HRMS/MSn Analysis of Anthocyanins, Flavonol Glycosides, and Hydroxycinnamic Acid Derivatives in Red Mustard Greens (Brassica juncea Coss Variety)

PubMed Central

Lin, Long-Ze; Sun, Jianghao; Chen, Pei; Harnly, James

2013-01-01

An UHPLC-PDA-ESI/HRMS/MSn profiling method was used for a comprehensive study of the phenolic components of red mustard greens (Brassica juncea Coss variety) and identified 67 anthocyanins, 102 flavonol glycosides, and 40 hydroxycinnamic acid derivatives. The glycosylation patterns of the flavonoids were assigned on the basis of direct comparison of the parent flavonoid glycosides with reference compounds. The putative identifications were obtained from tandem mass data analysis and confirmed by the retention time, elution order, and UV–vis and high-resolution mass spectra. Further identifications were made by comparing the UHPLC-PDA-ESI/HRMS/MSn data with those of reference compounds in the polyphenol database and in the literature. Twenty-seven acylated cyanidin 3-sophoroside-5-diglucosides, 24 acylated cyanidin 3-sophoroside-5- glucosides, 3 acylated cyanidin triglucoside-5-glucosides, 37 flavonol glycosides, and 10 hydroxycinnamic acid derivatives were detected for the first time in brassica vegetables. At least 50 of them are reported for the first time in any plant materials. PMID:21970730

Flavonol dimers from callus cultures of Dysosma versipellis and their in vitro neuraminidase inhibitory activities.

PubMed

Chen, Ridao; Duan, Ruigang; Wei, Yannan; Zou, Jianhua; Li, Junwei; Liu, Xiaoyue; Wang, Haiyan; Guo, Ying; Li, Qiuhong; Dai, Jungui

2015-12-01

A chemical investigation of callus cultures of Dysosma versipellis led to the isolation of five new flavonol dimers, dysoverines A-E (1-5), together with 12 known compounds (6-17). The structures of new compounds were determined by the extensive spectroscopic data analyses. The biosynthetic pathway of the new compounds was proposed to involve O-methylation, prenylation, and Diels-Alder cycloaddition, which successively occurred in cultured plant cells. Compounds 1-17 exhibited in vitro neuraminidase inhibitory activities with the IC50 values of 31.0-93.9μM. Copyright © 2015 Elsevier B.V. All rights reserved.

UHPLC-PDA-ESI/HRMS/MSn analysis of anthocyanins, flavonol glycosides, and hydroxycinnamic acid derivatives in red mustard green (Brassica juncea (L) Coss variety)

USDA-ARS?s Scientific Manuscript database

An UHPLC-PDA-ESI/HRMS/MSn profiling method was used for a comprehensive study of the polyphenols in red mustard greens and identified 209 phenolic compounds: 67 anthocyanin, 102 flavonol glycosides, and 40 hydroxycinnamic acid derivatives. The glycosylation patterns of the flavonoids were assigned ...

Priming of seeds with methyl jasmonate induced resistance to hemi-biotroph Fusarium oxysporum f.sp. lycopersici in tomato via 12-oxo-phytodienoic acid, salicylic acid, and flavonol accumulation.

PubMed

Król, P; Igielski, R; Pollmann, S; Kępczyńska, E

2015-05-01

Methyl jasmonate (MeJA) was tested by seed treatment for its ability to protect tomato seedlings against fusarium wilt caused by the soil-borne fungal pathogen Fusarium oxysporum f.sp. lycopersici. Isolated from Solanum lycopersicon L. seeds, cv. Beta fungus was identified as F. oxysporum f.sp. lycopersici Race 3 fungus by using phytopathological and molecular methods. MeJA applied at 0.01, 0.1 and 1 mM reduced spore germination and mycelial growth in vitro. Soaking of tomato seeds in MeJA solution at 0.1 mM for 1 h significantly enhanced the resistance level against the tested fungus in tomato seedlings 4 weeks after inoculation. The extracts from leaves of 15-day-old seedlings obtained from previously MeJA soaked seeds had the ability to inhibit in vitro spore germination of tested fungus. In these seedlings a significant increase in the levels phenolic compounds such as salicylic acid (SA), kaempferol and quercetin was observed. Up-regulation of phenylalanine ammonia-lyase (PAL5) and benzoic acid/salicylic acid carboxyl methyltransferase (BSMT) genes and down-regulation of the isochorysmate synthase (ICS) gene in response to exogenous MeJA application indicate that the phenylalanine ammonia-lyase (PAL), not the isochorismate (IC) pathway, is the primary route for SA production in tomato. Moreover, the increased accumulation of the flavonols quercetin and kaempferol appears closely related to the increase of PAL5, chalcone synthase (CHS) and flavonol synthase/flavanone 3-hydroxylase-like (FLS) genes. Elevated levels of salicylic acid in seedlings raised from MeJA-soaked seeds were simultaneously accompanied by a decrease of jasmonic acid, the precursor of MeJA, and an increase of 12-oxo-phytodienoic acid (OPDA), the precursor of jasmonic acid. The present results indicate that the priming of tomato seeds with 0.1mM MeJA before sowing enables the seedlings grown from these seeds to reduce the attack of the soil-borne fungal pathogen F. oxysporum f.sp. lycopersici

Choline Transport Activity Regulates Phosphatidylcholine Synthesis through Choline Transporter Hnm1 Stability*

PubMed Central

Fernández-Murray, J. Pedro; Ngo, Michael H.; McMaster, Christopher R.

2013-01-01

Choline is a precursor for the synthesis of phosphatidylcholine through the CDP-choline pathway. Saccharomyces cerevisiae expresses a single high affinity choline transporter at the plasma membrane, encoded by the HNM1 gene. We show that exposing cells to increasing levels of choline results in two different regulatory mechanisms impacting Hnm1 activity. Initial exposure to choline results in a rapid decrease in Hnm1-mediated transport at the level of transporter activity, whereas chronic exposure results in Hnm1 degradation through an endocytic mechanism that depends on the ubiquitin ligase Rsp5 and the casein kinase 1 redundant pair Yck1/Yck2. We present details of how the choline transporter is a major regulator of phosphatidylcholine synthesis. PMID:24187140

Ethylene-Regulated Floral Volatile Synthesis in Petunia Corollas1[w

PubMed Central

Underwood, Beverly A.; Tieman, Denise M.; Shibuya, Kenichi; Dexter, Richard J.; Loucas, Holly M.; Simkin, Andrew J.; Sims, Charles A.; Schmelz, Eric A.; Klee, Harry J.; Clark, David G.

2005-01-01

In many flowering plants, such as petunia (Petunia × hybrida), ethylene produced in floral organs after pollination elicits a series of physiological and biochemical events, ultimately leading to senescence of petals and successful fertilization. Here, we demonstrate, using transgenic ethylene insensitive (44568) and Mitchell Diploid petunias, that multiple components of emission of volatile organic compounds (VOCs) are regulated by ethylene. Expression of benzoic acid/salicylic acid carboxyl methyltransferase (PhBSMT1 and 2) mRNA is temporally and spatially down-regulated in floral organs in a manner consistent with current models for postpollination ethylene synthesis in petunia corollas. Emission of methylbenzoate and other VOCs after pollination and exogenous ethylene treatment parallels a reduction in PhBSMT1 and 2 mRNA levels. Under cyclic light conditions (day/night), PhBSMT mRNA levels are rhythmic and precede emission of methylbenzoate by approximately 6 h. When shifted into constant dark or light conditions, PhBSMT mRNA levels and subsequent methylbenzoate emission correspondingly decrease or increase to minimum or maximum levels observed during normal conditions, thus suggesting that light may be a more critical influence on cyclic emission of methylbenzoate than a circadian clock. Transgenic PhBSMT RNAi flowers with reduced PhBSMT mRNA levels show a 75% to 99% decrease in methylbenzoate emission, with minimal changes in other petunia VOCs. These results implicate PhBSMT1 and 2 as genes responsible for synthesis of methylbenzoate in petunia. PMID:15849311

Cyclic phosphatidic acid and lysophosphatidic acid induce hyaluronic acid synthesis via CREB transcription factor regulation in human skin fibroblasts.

PubMed

Maeda-Sano, Katsura; Gotoh, Mari; Morohoshi, Toshiro; Someya, Takao; Murofushi, Hiromu; Murakami-Murofushi, Kimiko

2014-09-01

Cyclic phosphatidic acid (cPA) is a naturally occurring phospholipid mediator and an analog of the growth factor-like phospholipid lysophosphatidic acid (LPA). cPA has a unique cyclic phosphate ring at the sn-2 and sn-3 positions of its glycerol backbone. We showed before that a metabolically stabilized cPA derivative, 2-carba-cPA, relieved osteoarthritis pathogenesis in vivo and induced hyaluronic acid synthesis in human osteoarthritis synoviocytes in vitro. This study focused on hyaluronic acid synthesis in human fibroblasts, which retain moisture and maintain health in the dermis. We investigated the effects of cPA and LPA on hyaluronic acid synthesis in human fibroblasts (NB1RGB cells). Using particle exclusion and enzyme-linked immunosorbent assays, we found that both cPA and LPA dose-dependently induced hyaluronic acid synthesis. We revealed that the expression of hyaluronan synthase 2 messenger RNA and protein is up-regulated by cPA and LPA treatment time dependently. We then characterized the signaling pathways up-regulating hyaluronic acid synthesis mediated by cPA and LPA in NB1RGB cells. Pharmacological inhibition and reporter gene assays revealed that the activation of the LPA receptor LPAR1, Gi/o protein, phosphatidylinositol-3 kinase (PI3K), extracellular-signal-regulated kinase (ERK), and cyclic adenosine monophosphate response element-binding protein (CREB) but not nuclear factor κB induced hyaluronic acid synthesis by the treatment with cPA and LPA in NB1RGB cells. These results demonstrate for the first time that cPA and LPA induce hyaluronic acid synthesis in human skin fibroblasts mainly through the activation of LPAR1-Gi/o followed by the PI3K, ERK, and CREB signaling pathway. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

Genetic resources of the functional food, teramnus labialis (L.f.) spreng for improving seed number, flavonol content, oil %, and fatty acid compositions

USDA-ARS?s Scientific Manuscript database

Teramnus labialis is used as food in India and has potential to be used as a functional food vegetable in the U.S.A. Photoperiod-sensitive T. labialis accessions were grown in the greenhouse from 2010 to 2011 and evaluated for flavonol content, oil %, and fatty acid compositions. Significant variati...

Oxygen-dependent regulation of c-di-GMP synthesis by SadC controls alginate production in Pseudomonas aeruginosa.

PubMed

Schmidt, Annika; Hammerbacher, Anna Silke; Bastian, Mike; Nieken, Karen Jule; Klockgether, Jens; Merighi, Massimo; Lapouge, Karine; Poschgan, Claudia; Kölle, Julia; Acharya, K Ravi; Ulrich, Martina; Tümmler, Burkhard; Unden, Gottfried; Kaever, Volkhard; Lory, Stephen; Haas, Dieter; Schwarz, Sandra; Döring, Gerd

2016-10-01

Pseudomonas aeruginosa produces increased levels of alginate in response to oxygen-deprived conditions. The regulatory pathway(s) that links oxygen limitation to increased synthesis of alginate has remained elusive. In the present study, using immunofluorescence microscopy, we show that anaerobiosis-induced alginate production by planktonic PAO1 requires the diguanylate cyclase (DGC) SadC, previously identified as a regulator of surface-associated lifestyles. Furthermore, we found that the gene products of PA4330 and PA4331, located in a predicted operon with sadC, have a major impact on alginate production: deletion of PA4330 (odaA, for oxygen-dependent alginate synthesis activator) caused an alginate production defect under anaerobic conditions, whereas a PA4331 (odaI, for oxygen-dependent alginate synthesis inhibitor) deletion mutant produced alginate also in the presence of oxygen, which would normally inhibit alginate synthesis. Based on their sequence, OdaA and OdaI have predicted hydratase and dioxygenase reductase activities, respectively. Enzymatic assays using purified protein showed that unlike OdaA, which did not significantly affect DGC activity of SadC, OdaI inhibited c-di-GMP production by SadC. Our data indicate that SadC, OdaA and OdaI are components of a novel response pathway of P. aeruginosa that regulates alginate synthesis in an oxygen-dependent manner. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

One-step separation and purification of three lignans and one flavonol from Sinopodophyllum emodi by medium-pressure liquid chromatography and high-speed counter-current chromatography.

PubMed

Wang, Ping; Liu, Yongling; Chen, Tao; Xu, Wenhua; You, Jinmao; Liu, Yongjun; Li, Yulin

2013-01-01

Lignans and flavonols are the primary constituents of Sinopodophyllum emodi and have been used as cathartic, anthelmintic, chemotherapeutic and anti-hypertensive compounds. Although these compounds have been isolated, there have been no reports on the separation of 4'-demethyl podophyllotoxin, podophyllotoxin, deoxypodophyllotoxin and kaempferol in one step by medium-pressure liquid chromatography (MPLC) and high-speed counter-current chromatography (HSCCC). Development of an efficient method for the preparative separation and purification of three lignans and one flavonol from S. emodi. The precipitate of crude extracts was first separated by MPLC into four parts, numbered GJ-1, GJ-2, GJ-3 and GJ-4. GJ-1 was separated and purified by HSCCC using a solvent system composed of n-hexane:ethyl acetate:methanol:water (1.75:1.5:1:0.75, v/v/v/v). The purities of the target compounds were assessed using high-performance liquid chromatography (HPLC) and chemical structures were identified by (1) H-NMR and (13) C-NMR. The HSCCC and MPLC methods were successfully used for the preparative separation and purification of 4'-demethyl podophyllotoxin (8.5 mg, 92.4%), podophyllotoxin (40.1 mg, 92.1%), deoxypodophyllotoxin (4.6 mg, 98.1%), and kaempferol (1.6 mg, 96.7%) from a 100 mg sample. Three lignans (4'-demethyl podophyllotoxin, podophyllotoxin, deoxypodophyllotoxin) and one flavonol (kaempferol) were successfully isolated by HSCCC and MPLC in one step. Copyright © 2013 John Wiley & Sons, Ltd.

New phenylpropanoid-substituted flavan-3-ols and flavonols from the leaves of Uncaria rhynchophylla.

PubMed

Li, Ruxin; Cheng, Jintang; Jiao, Mengjiao; Li, Li; Guo, Cong; Chen, Sha; Liu, An

2017-01-01

Uncariols A (1) and B (2), two new phenylpropanoid-substituted flavan-3-ols, and (±)-uncariols C (3a/3b) and D (4a/4b), two pairs of new phenylpropanoid-substituted flavonol enantiomers, together with nine known compounds (5-13), were isolated from the leaves of Uncaria rhynchophylla. The structures of 1-4 were established primarily by NMR and HRESIMS experiments. The absolute configurations of the new ones were deduced via the circular dichroism (CD) and quantum chemical calculations of the electronic circular dichroic (ECD) spectra. In addition, all of the isolated compounds showed potent antioxidant activity in the DPPH radical scavenging test. Copyright © 2016. Published by Elsevier B.V.

Separation and purification of four flavonol diglucosides from the flower of Meconopsis integrifolia by high-speed counter-current chromatography.

PubMed

Huang, Yanfei; Han, Yatao; Chen, Keli; Huang, Bisheng; Liu, Yuan

2015-12-01

Flavonoids are the main components of Meconopsis integrifolia (Maxim.) Franch, which is a traditional Tibetan medicine. However, traditional chromatography separation requires a large quantity of raw M. integrifolia and is very time consuming. Herein, we applied high-speed counter-current chromatography in the separation and purification of flavonoids from the ethanol extracts of M. integrifolia flower. Ethyl acetate/n-butanol/water (2:3:5, v/v/v) was selected as the optimum solvent system to purify the four components, namely quercetin-3-O-β-d-glucopyrannosy-(1→6)-β-d-glucopyranoside (compound 1, 60 mg), quercetin 3-O-[2'''-O-acetyl-β-d-glucopyranosyl-(1→6)-β-d-glucopyranoside (compound 2, 40 mg), quercetin 3-O-[3'''-O-acetyl-β-d-glucopyranosyl-(1→6)-β-d-glucopyranoside (compound 3, 11 mg), and quercetin 3-O-[6'''-O-acetyl-β-d-glucopyranosyl-(1→6)-β-d-glucopyranoside (compound 4, 16 mg). Among the four compounds, 3 and 4 were new acetylated flavonol diglucosides. After the high-speed counter-current chromatography separation, the purities of the four flavonol diglucosides were 98, 95, 90, and 92%, respectively. The structures of these compounds were identified by mass spectrometry and NMR spectroscopy. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

TOR Pathway-Mediated Juvenile Hormone Synthesis Regulates Nutrient-Dependent Female Reproduction in Nilaparvata lugens (Stål)

PubMed Central

Lu, Kai; Chen, Xia; Liu, Wen-Ting; Zhou, Qiang

2016-01-01

The “target of rapamycin” (TOR) nutritional signaling pathway and juvenile hormone (JH) regulation of vitellogenesis has been known for a long time. However, the interplay between these two pathways regulating vitellogenin (Vg) expression remains obscure. Here, we first demonstrated the key role of amino acids (AAs) in activation of Vg synthesis and egg development in Nilaparvata lugens using chemically defined artificial diets. AAs induced the expression of TOR and S6K (S6 kinase), whereas RNAi-mediated silencing of these two TOR pathway genes and rapamycin application strongly inhibited the AAs-induced Vg synthesis. Furthermore, knockdown of Rheb (Ras homologue enriched in brain), TOR, S6K and application of rapamycin resulted in a dramatic reduction in the mRNA levels of jmtN (juvenile hormone acid methyltransferase, JHAMT). Application of JH III on the RNAi (Rheb and TOR) and rapamycin-treated females partially rescued the Vg expression. Conversely, knockdown of either jmtN or met (methoprene-tolerant, JH receptor) and application of JH III had no effects on mRNA levels of Rheb, TOR and S6K and phosphorylation of S6K. In summary, our results demonstrate that the TOR pathway induces JH biosynthesis that in turn regulates AAs-mediated Vg synthesis in N. lugens. PMID:27043527

TOR Pathway-Mediated Juvenile Hormone Synthesis Regulates Nutrient-Dependent Female Reproduction in Nilaparvata lugens (Stål).

PubMed

Lu, Kai; Chen, Xia; Liu, Wen-Ting; Zhou, Qiang

2016-03-28

The "target of rapamycin" (TOR) nutritional signaling pathway and juvenile hormone (JH) regulation of vitellogenesis has been known for a long time. However, the interplay between these two pathways regulating vitellogenin (Vg) expression remains obscure. Here, we first demonstrated the key role of amino acids (AAs) in activation of Vg synthesis and egg development in Nilaparvata lugens using chemically defined artificial diets. AAs induced the expression of TOR and S6K (S6 kinase), whereas RNAi-mediated silencing of these two TOR pathway genes and rapamycin application strongly inhibited the AAs-induced Vg synthesis. Furthermore, knockdown of Rheb (Ras homologue enriched in brain), TOR, S6K and application of rapamycin resulted in a dramatic reduction in the mRNA levels of jmtN (juvenile hormone acid methyltransferase, JHAMT). Application of JH III on the RNAi (Rheb and TOR) and rapamycin-treated females partially rescued the Vg expression. Conversely, knockdown of either jmtN or met (methoprene-tolerant, JH receptor) and application of JH III had no effects on mRNA levels of Rheb, TOR and S6K and phosphorylation of S6K. In summary, our results demonstrate that the TOR pathway induces JH biosynthesis that in turn regulates AAs-mediated Vg synthesis in N. lugens.

Multi-substrate flavonol O-glucosyltransferases from strawberry (Fragaria×ananassa) achene and receptacle

PubMed Central

Griesser, Markus; Vitzthum, Florian; Fink, Barbara; Bellido, Mari Luz; Raasch, Constanze; Munoz-Blanco, Juan; Schwab, Wilfried

2008-01-01

In an effort to characterize fruit ripening-related genes functionally, two glucosyltransferases, FaGT6 and FaGT7, were cloned from a strawberry (Fragaria×ananassa) cDNA library and the full-length open reading frames were amplified by rapid amplification of cDNA ends. FaGT6 and FaGT7 were expressed heterologously as fusion proteins in Escherichia coli and target protein was purified using affinity chromatography. Both recombinant enzymes exhibited a broad substrate tolerance in vitro, accepting numerous flavonoids, hydroxycoumarins, and naphthols. FaGT6 formed 3-O-glucosides and minor amounts of 7-O-, 4′-O-, and 3′-O-monoglucosides and one diglucoside from flavonols such as quercetin. FaGT7 converted quercetin to the 3-O-glucoside and 4′-O-glucoside and minor levels of the 7- and 3′-isomers but formed no diglucoside. Gene expression studies showed that both genes are strongly expressed in achenes of small-sized green fruits, while the expression levels were generally lower in the receptacle. Significant levels of quercetin 3-O-, 7-O-, and 4′-O-glucosides, kaempferol 3-O- and 7-O-glucosides, as well as isorhamnetin 7-O-glucoside, were identified in achenes and the receptacle. In the receptacle, the expression of both genes is negatively controlled by auxin which correlates with the ripening-related gene expression in this tissue. Salicylic acid, a known signal molecule in plant defence, induces the expression of both genes. Thus, it appears that FaGT6 and FaGT7 are involved in the glucosylation of flavonols and may also participate in xenobiotic metabolism. The latter function is supported by the proven ability of strawberries to glucosylate selected unnatural substrates injected in ripe fruits. This report presents the first biochemical characterization of enzymes mainly expressed in strawberry achenes and provides the foundation of flavonoid metabolism in the seeds. PMID:18487633

The hemostatic effect study of Cirsium setosum on regulating α1-ARs via mediating norepinephrine synthesis by enzyme catalysis.

PubMed

Chang, Nianwei; Li, Yanmei; Zhou, Mengge; Gao, Jie; Hou, Yuanyuan; Jiang, Min; Bai, Gang

2017-03-01

Cirsium setosum (CS) is the aboveground part of Cephalanoplos segetum Kitam. Although it has been used as a hemostatic treatment for thousands of years and is still in use today, the mechanism of CS on regulating ARs is still not clear. In this study, we aimed to clarify the mechanism of CS on regulating ARs. We developed a simple method based on UPLC/Q-TOF MS combined adrenergic receptor dual-luciferase reporter assay systems for the rapid determination of active constituents in CS. The mechanism of tyramine, the main active component for regulating ARs, was further investigated by an in vitro norepinephrine biotransformation test and in vivo vaso activity tests. Two phenethylamine ARs regulators (tyramine and N-methyltyramine) in CS were characterized, and it was found that tyramine could induce vasoconstriction via regulation of α1-ARs by mediating norepinephrine synthesis. The hemostatic effect of CS is associated with tyramine and N-methyltyramine, via regulation of α1-ARs, and the mechanism of tyramine is related to mediating norepinephrine synthesis by enzyme catalysis. Copyright © 2017. Published by Elsevier Masson SAS.

Abscisic Acid Negatively Regulates Elicitor-Induced Synthesis of Capsidiol in Wild Tobacco1[W

PubMed Central

Mialoundama, Alexis Samba; Heintz, Dimitri; Debayle, Delphine; Rahier, Alain; Camara, Bilal; Bouvier, Florence

2009-01-01

In the Solanaceae, biotic and abiotic elicitors induce de novo synthesis of sesquiterpenoid stress metabolites known as phytoalexins. Because plant hormones play critical roles in the induction of defense-responsive genes, we have explored the effect of abscisic acid (ABA) on the synthesis of capsidiol, the major wild tobacco (Nicotiana plumbaginifolia) sesquiterpenoid phytoalexin, using wild-type plants versus nonallelic mutants Npaba2 and Npaba1 that are deficient in ABA synthesis. Npaba2 and Npaba1 mutants exhibited a 2-fold higher synthesis of capsidiol than wild-type plants when elicited with either cellulase or arachidonic acid or when infected by Botrytis cinerea. The same trend was observed for the expression of the capsidiol biosynthetic genes 5-epi-aristolochene synthase and 5-epi-aristolochene hydroxylase. Treatment of wild-type plants with fluridone, an inhibitor of the upstream ABA pathway, recapitulated the behavior of Npaba2 and Npaba1 mutants, while the application of exogenous ABA reversed the enhanced synthesis of capsidiol in Npaba2 and Npaba1 mutants. Concomitant with the production of capsidiol, we observed the induction of ABA 8′-hydroxylase in elicited plants. In wild-type plants, the induction of ABA 8′-hydroxylase coincided with a decrease in ABA content and with the accumulation of ABA catabolic products such as phaseic acid and dihydrophaseic acid, suggesting a negative regulation exerted by ABA on capsidiol synthesis. Collectively, our data indicate that ABA is not required per se for the induction of capsidiol synthesis but is essentially implicated in a stress-response checkpoint to fine-tune the amplification of capsidiol synthesis in challenged plants. PMID:19420326

Enzymological mechanism for the regulation of lanthanum chloride on flavonoid synthesis of soybean seedlings under enhanced ultraviolet-B radiation.

PubMed

Fan, Caixia; Hu, Huiqing; Wang, Lihong; Zhou, Qing; Huang, Xiaohua

2014-01-01

In order to probe into the enzymological mechanism for the regulation of lanthanum chloride (LaCl3) on flavonoid synthesis in plants under enhanced ultraviolet-B (UV-B) radiation, the effects of LaCl₃ (20 and 60 mg l(-1)) on the content of flavonoids as well as the activities of phenylalanine ammonia-lyase (PAL), cinnamate-4-hydroxylase (C4H), 4-coumarate : coenzyme A ligase (4CL), and chalcone synthase (CHS) in soybean seedlings under enhanced UV-B radiation (2.6 and 6.2 kJ m(-2) day(-1)) were investigated. Enhanced UV-B radiation (2.6 and 6.2 kJ m(-2) day(-1)) caused the increase in the content of flavonoids as well as the activities of PAL, C4H, 4CL, and CHS in soybean seedlings. The treatment of 20 mg l(-1) LaCl₃ also efficiently increased these indices, which promoted the flavonoid synthesis and provided protective effects for resisting enhanced UV-B radiation. On the contrary, the treatment of 60 mg l(-1) LaCl₃ decreased the content of flavonoids as well as the activities of C4H, 4CL, and CHS in soybean seedlings except increasing the activity of PAL, which were not beneficial to the flavonoid synthesis and provided negative effects for resisting enhanced UV-B radiation. In conclusion, enhanced UV-B radiation caused the increase in the flavonoid synthesis by promoting the activities of PAL, C4H, 4CL, and CHS in soybean seedlings. The treatment of LaCl₃ could change flavonoid synthesis in soybean seedlings under enhanced UV-B radiation by regulating the activities of PAL, C4H, 4CL, and CHS, which is an enzymological mechanism for the regulation of LaCl₃ on flavonoid synthesis in plants under enhanced UV-B radiation.

Endoplasmic reticulum stress inhibits expression of genes involved in thyroid hormone synthesis and their key transcriptional regulators in FRTL-5 thyrocytes

PubMed Central

Wen, Gaiping; Eder, Klaus

2017-01-01

Endoplasmic reticulum (ER) stress is characterized by the accumulation of misfolded proteins due to an impairment of ER quality control pathways leading to the activation of a defense system, called unfolded protein response (UPR). While thyrocytes are supposed to be highly susceptible to environmental conditions that cause ER stress due to the synthesis of large amounts of secretory proteins required for thyroid hormone synthesis, systematic investigations on the effect of ER stress on expression of key genes of thyroid hormone synthesis and their transcriptional regulators are lacking. Since the aim of the ER stress-induced UPR is to restore ER homeostasis and to facilitate cell survival through transient shutdown of ribosomal protein translation, we hypothesized that the expression of genes involved in thyroid hormone synthesis and their transcriptional regulators, all of which are not essential for cell survival, are down-regulated in thyrocytes during ER stress, while sterol regulatory element-binding proteins (SREBPs) are activated during ER stress in thyrocytes. Treatment of FRTL-5 thyrocytes with the ER stress inducer tunicamycin (TM) dose-dependently increased the mRNA and/or protein levels of known UPR target genes, stimulated phosphorylation of the ER stress sensor protein kinase RNA-like ER kinase (PERK) and of the PERK target protein eukaryotic initiation factor 2α (eIF2α) and caused splicing of the ER stress-sensitive transcription factor X-box binding protein (XBP-1) (P < 0.05). The mRNA levels and/or protein levels of genes involved in thyroid hormone synthesis, sodium/iodide symporter (NIS), thyroid peroxidase (TPO) and thyroglobulin (TG), their transcriptional regulators and thyrotropin (TSH) receptor and the uptake of Na125I were reduced at the highest concentration of TM tested (0.1 μg/mL; P < 0.05). Proteolytic activation of the SREBP-1c pathway was not observed in FRTL-5 cells treated with TM, whereas TM reduced proteolytic activation of

Endoplasmic reticulum stress inhibits expression of genes involved in thyroid hormone synthesis and their key transcriptional regulators in FRTL-5 thyrocytes.

PubMed

Wen, Gaiping; Ringseis, Robert; Eder, Klaus

2017-01-01

Endoplasmic reticulum (ER) stress is characterized by the accumulation of misfolded proteins due to an impairment of ER quality control pathways leading to the activation of a defense system, called unfolded protein response (UPR). While thyrocytes are supposed to be highly susceptible to environmental conditions that cause ER stress due to the synthesis of large amounts of secretory proteins required for thyroid hormone synthesis, systematic investigations on the effect of ER stress on expression of key genes of thyroid hormone synthesis and their transcriptional regulators are lacking. Since the aim of the ER stress-induced UPR is to restore ER homeostasis and to facilitate cell survival through transient shutdown of ribosomal protein translation, we hypothesized that the expression of genes involved in thyroid hormone synthesis and their transcriptional regulators, all of which are not essential for cell survival, are down-regulated in thyrocytes during ER stress, while sterol regulatory element-binding proteins (SREBPs) are activated during ER stress in thyrocytes. Treatment of FRTL-5 thyrocytes with the ER stress inducer tunicamycin (TM) dose-dependently increased the mRNA and/or protein levels of known UPR target genes, stimulated phosphorylation of the ER stress sensor protein kinase RNA-like ER kinase (PERK) and of the PERK target protein eukaryotic initiation factor 2α (eIF2α) and caused splicing of the ER stress-sensitive transcription factor X-box binding protein (XBP-1) (P < 0.05). The mRNA levels and/or protein levels of genes involved in thyroid hormone synthesis, sodium/iodide symporter (NIS), thyroid peroxidase (TPO) and thyroglobulin (TG), their transcriptional regulators and thyrotropin (TSH) receptor and the uptake of Na125I were reduced at the highest concentration of TM tested (0.1 μg/mL; P < 0.05). Proteolytic activation of the SREBP-1c pathway was not observed in FRTL-5 cells treated with TM, whereas TM reduced proteolytic activation of

Metabolite profiling of flavonols and in vitro antioxidant activity of young shoots of wild Humulus lupulus L. (hop).

PubMed

Maietti, Annalisa; Brighenti, Virginia; Bonetti, Gianpiero; Tedeschi, Paola; Prencipe, Francesco Pio; Benvenuti, Stefania; Brandolini, Vincenzo; Pellati, Federica

2017-08-05

Humulus lupulus L., commonly named hop, is well-known for its sedative and estrogenic activity. While hop cones are widely characterized, only few works have been carried out on the young shoots of this plant. In the light of this, the aim of this study was to identify for the first time the flavonoids present in young hop shoots and to compare the composition of samples harvested from different locations in Northern Italy with their antioxidant activity. The samples were extracted by means of dynamic maceration with methanol. The HPLC-UV/DAD, HPLC-ESI-MS and MS 2 analysis were carried out by using an Ascentis C 18 column (250×4.6mm I.D., 5μm), with a mobile phase composed of 0.1M formic acid in both water and acetonitrile, under gradient elution. Quercetin and kaempferol glycosides were the main compounds identified and quantified in hop shoot extracts. Total flavonols ranged from 2698±185 to 517±48μg/g (fresh weight). The antioxidant activity was determined by means of the radical scavenging activity assay against diphenylpicrylhydrazyl (DPPH) and by using a photochemiluscence assay with a Photochem ® apparatus. The results showed that hop shoots represent a new source of flavonols; therefore, they can be useful for a possible incorporation in the diet as a functional food or applied in the nutraceutical ambit. Copyright © 2017 Elsevier B.V. All rights reserved.

Keratinocyte-derived Laminin-332 Protein Promotes Melanin Synthesis via Regulation of Tyrosine Uptake*

PubMed Central

Chung, Heesung; Jung, Hyejung; Lee, Jung-hyun; Oh, Hye Yun; Kim, Ok Bin; Han, Inn-Oc; Oh, Eok-Soo

2014-01-01

Melanocytes, which produce the pigment melanin, are known to be closely regulated by neighboring keratinocytes. However, how keratinocytes regulate melanin production is unclear. Here we report that melanin production in melanoma cells (B16F10 and MNT-1) was increased markedly on a keratinocyte-derived extracellular matrix compared with a melanoma cell-derived extracellular matrix. siRNA-mediated reduction of keratinocyte-derived laminin-332 expression decreased melanin synthesis in melanoma cells, and laminin-332, but not fibronectin, enhanced melanin content and α-melanocyte-stimulating hormone-regulated melanin production in melanoma cells. Similar effects were observed in human melanocytes. Interestingly, however, laminin-332 did not affect the expression or activity of tyrosinase. Instead, laminin-332 promoted the uptake of extracellular tyrosine and, subsequently, increased intracellular levels of tyrosine in both melanocytes and melanoma cells. Taken together, these data strongly suggest that keratinocyte-derived laminin-332 contributes to melanin production by regulating tyrosine uptake. PMID:24951591

Keratinocyte-derived laminin-332 protein promotes melanin synthesis via regulation of tyrosine uptake.

PubMed

Chung, Heesung; Jung, Hyejung; Lee, Jung-Hyun; Oh, Hye Yun; Kim, Ok Bin; Han, Inn-Oc; Oh, Eok-Soo

2014-08-01

Melanocytes, which produce the pigment melanin, are known to be closely regulated by neighboring keratinocytes. However, how keratinocytes regulate melanin production is unclear. Here we report that melanin production in melanoma cells (B16F10 and MNT-1) was increased markedly on a keratinocyte-derived extracellular matrix compared with a melanoma cell-derived extracellular matrix. siRNA-mediated reduction of keratinocyte-derived laminin-332 expression decreased melanin synthesis in melanoma cells, and laminin-332, but not fibronectin, enhanced melanin content and α-melanocyte-stimulating hormone-regulated melanin production in melanoma cells. Similar effects were observed in human melanocytes. Interestingly, however, laminin-332 did not affect the expression or activity of tyrosinase. Instead, laminin-332 promoted the uptake of extracellular tyrosine and, subsequently, increased intracellular levels of tyrosine in both melanocytes and melanoma cells. Taken together, these data strongly suggest that keratinocyte-derived laminin-332 contributes to melanin production by regulating tyrosine uptake. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Regulation of Viral RNA Synthesis by the V Protein of Parainfluenza Virus 5

PubMed Central

Yang, Yang; Zengel, James; Sun, Minghao; Sleeman, Katrina; Timani, Khalid Amine; Aligo, Jason; Rota, Paul

2015-01-01

ABSTRACT Paramyxoviruses include many important animal and human pathogens. The genome of parainfluenza virus 5 (PIV5), a prototypical paramyxovirus, encodes a V protein that inhibits viral RNA synthesis. In this work, the mechanism of inhibition was investigated. Using mutational analysis and a minigenome system, we identified regions in the N and C termini of the V protein that inhibit viral RNA synthesis: one at the very N terminus of V and the second at the C terminus of V. Furthermore, we determined that residues L16 and I17 are critical for the inhibitory function of the N-terminal region of the V protein. Both regions interact with the nucleocapsid protein (NP), an essential component of the viral RNA genome complex (RNP). Mutations at L16 and I17 abolished the interaction between NP and the N-terminal domain of V. This suggests that the interaction between NP and the N-terminal domain plays a critical role in V inhibition of viral RNA synthesis by the N-terminal domain. Both the N- and C-terminal regions inhibited viral RNA replication. The C terminus inhibited viral RNA transcription, while the N-terminal domain enhanced viral RNA transcription, suggesting that the two domains affect viral RNA through different mechanisms. Interestingly, V also inhibited the synthesis of the RNA of other paramyxoviruses, such as Nipah virus (NiV), human parainfluenza virus 3 (HPIV3), measles virus (MeV), mumps virus (MuV), and respiratory syncytial virus (RSV). This suggests that a common host factor may be involved in the replication of these paramyxoviruses. IMPORTANCE We identified two regions of the V protein that interact with NP and determined that one of these regions enhances viral RNA transcription via its interaction with NP. Our data suggest that a common host factor may be involved in the regulation of paramyxovirus replication and could be a target for broad antiviral drug development. Understanding the regulation of paramyxovirus replication will enable the

MicroRNAs in the pineal gland: miR-483 regulates melatonin synthesis by targeting arylalkylamine N-acetyltransferase.

PubMed

Clokie, Samuel J H; Lau, Pierre; Kim, Hyun Hee; Coon, Steven L; Klein, David C

2012-07-20

MicroRNAs (miRNAs) play a broad range of roles in biological regulation. In this study, rat pineal miRNAs were profiled for the first time, and their importance was evaluated by focusing on the main function of the pineal gland, melatonin synthesis. Massively parallel sequencing and related methods revealed the miRNA population is dominated by a small group of miRNAs as follows: ~75% is accounted for by 15 miRNAs; miR-182 represents 28%. In addition to miR-182, miR-183 and miR-96 are also highly enriched in the pineal gland, a distinctive pattern also found in the retina. This effort also identified previously unrecognized miRNAs and other small noncoding RNAs. Pineal miRNAs do not exhibit a marked night/day difference in abundance with few exceptions (e.g. 2-fold night/day differences in the abundance of miR-96 and miR-182); this contrasts sharply with the dynamic 24-h pattern that characterizes the pineal transcriptome. During development, the abundance of most pineal gland-enriched miRNAs increases; however, there is a marked decrease in at least one, miR-483. miR-483 is a likely regulator of melatonin synthesis, based on the following. It inhibits melatonin synthesis by pinealocytes in culture; it acts via predicted binding sites in the 3"-UTR of arylalkylamine N-acetyltransferase (Aanat) mRNA, the penultimate enzyme in melatonin synthesis, and it exhibits a developmental profile opposite to that of Aanat transcripts. Additionally, a miR-483 targeted antagonist increased melatonin synthesis in neonatal pinealocytes. These observations support the hypothesis that miR-483 suppresses Aanat mRNA levels during development and that the developmental decrease in miR-483 abundance promotes melatonin synthesis.

DNA polymerase-α regulates type I interferon activation through cytosolic RNA:DNA synthesis

PubMed Central

Starokadomskyy, Petro; Gemelli, Terry; Rios, Jonathan J.; Xing, Chao; Wang, Richard C.; Li, Haiying; Pokatayev, Vladislav; Dozmorov, Igor; Khan, Shaheen; Miyata, Naoteru; Fraile, Guadalupe; Raj, Prithvi; Xu, Zhe; Xu, Zigang; Ma, Lin; Lin, Zhimiao; Wang, Huijun; Yang, Yong; Ben-Amitai, Dan; Orenstein, Naama; Mussaffi, Huda; Baselga, Eulalia; Tadini, Gianluca; Grunebaum, Eyal; Sarajlija, Adrijan; Krzewski, Konrad; Wakeland, Edward K.; Yan, Nan; de la Morena, Maria Teresa; Zinn, Andrew R.; Burstein, Ezra

2016-01-01

Aberrant nucleic acids generated during viral replication are the main trigger for antiviral immunity, and mutations disrupting nucleic acid metabolism can lead to autoinflammatory disorders. Here we investigated the etiology of X-linked reticulate pigmentary disorder (XLPDR), a primary immunodeficiency with autoinflammatory features. We discovered that XLPDR is caused by an intronic mutation that disrupts expression of POLA1, the gene encoding the catalytic subunit of DNA polymerase-α. Unexpectedly, POLA1 deficiency results in increased type I interferon production. This enzyme is necessary for RNA:DNA primer synthesis during DNA replication and strikingly, POLA1 is also required for the synthesis of cytosolic RNA:DNA, which directly modulates interferon activation. Altogether, this work identified POLA1 as a critical regulator of the type I interferon response. PMID:27019227

Naringenin Regulates Expression of Genes Involved in Cell Wall Synthesis in Herbaspirillum seropedicae▿

PubMed Central

Tadra-Sfeir, M. Z.; Souza, E. M.; Faoro, H.; Müller-Santos, M.; Baura, V. A.; Tuleski, T. R.; Rigo, L. U.; Yates, M. G.; Wassem, R.; Pedrosa, F. O.; Monteiro, R. A.

2011-01-01

Five thousand mutants of Herbaspirillum seropedicae SmR1 carrying random insertions of transposon pTnMod-OGmKmlacZ were screened for differential expression of LacZ in the presence of naringenin. Among the 16 mutants whose expression was regulated by naringenin were genes predicted to be involved in the synthesis of exopolysaccharides, lipopolysaccharides, and auxin. These loci are probably involved in establishing interactions with host plants. PMID:21257805

Naringenin regulates expression of genes involved in cell wall synthesis in Herbaspirillum seropedicae.

PubMed

Tadra-Sfeir, M Z; Souza, E M; Faoro, H; Müller-Santos, M; Baura, V A; Tuleski, T R; Rigo, L U; Yates, M G; Wassem, R; Pedrosa, F O; Monteiro, R A

2011-03-01

Five thousand mutants of Herbaspirillum seropedicae SmR1 carrying random insertions of transposon pTnMod-OGmKmlacZ were screened for differential expression of LacZ in the presence of naringenin. Among the 16 mutants whose expression was regulated by naringenin were genes predicted to be involved in the synthesis of exopolysaccharides, lipopolysaccharides, and auxin. These loci are probably involved in establishing interactions with host plants.

[Regulation of the expression of coenzyme Q-synthesis complex during ageing].

PubMed

Campos-Silva, Carmen; Reyes-Torres, Iván; Rivera, Maximiliano; Meza-Torres, Catherine; Hernández-Camacho, Juan Diego; Rodríguez-Bies, Elisabet; Navas, Plácido; López-Lluch, Guillermo

Coenzyme Q is an essential component in the activity of the mitochondrial electron transport chain. Its synthesis involves, at least, a complex of ten different proteins. In this study, an attempt is made to determine the evolution of the expression of the genes involved in coenzyme Q synthesis during mouse ageing. The messenger RNA (mRNA) of different organs, such as brain, liver, kidney and skeletal muscle from young (8 months), mature (18 months), and old (24 months) mice was extracted by using Trizol and was then analysed by real time PCR (qPCR) using specific primers for all the known components of the coenzyme Q-synthesis complex (COQ genes). Liver showed the highest age-dependent changes in mRNA levels of the different components of Q-synthesis complex, affecting the extent of the variation as well as the significance of the change. In most of the cases, mRNA levels of the different components were higher in mature animals compared to young and old animals. When mRNAs of young and old animals were compared, only minor reductions of mRNA levels were found. Kidney showed a pattern similar to that found in liver as regards the changes in expression, although with lower increases in mature animals than those observed in the liver. Brain and skeletal muscle showed low variations, with muscle being the tissue with less changes, although a pattern similar to that found in liver and kidney was found, with slight increases in mature animals. The results of this study indicate that ageing is an important factor affecting COQ gene expression, but its effect depends on the organ, and that mature animals show higher levels of mRNA than young and old animals. Taken into consideration the importance of coenzyme Q in cell metabolism and ageing, a more detailed study is needed to understand the gene regulation of the coenzyme Q-synthesis mechanisms during ageing. Copyright © 2017 SEGG. Publicado por Elsevier España, S.L.U. All rights reserved.

Cynaropicrin is dual regulator for both degradation factors and synthesis factors in the cartilage metabolism.

PubMed

Masutani, Teruaki; Tanaka, Yuka Tsuda; Kojima, Hiroyuki; Tsuboi, Makoto; Hara, Akira; Niwa, Masayuki

2016-08-01

The molecular mechanism of osteoarthritis (OA) has never been understood clearly, but it has been suggested that imbalance of degradation and synthesis in cartilage contribute to the underlying mechanisms of OA. In this study, we investigated the effectiveness in the cartilage metabolism of the artichoke extract that includes the compound cynaropicrin. We evaluated the efficacy of the artichoke extract or cynaropicrin in the cartilage metabolism factors and NF-κB signaling activity stimulated by inflammatory cytokine in chondrogenic cell lines, OUMS-27 and SW1353, using qRT-PCR, immunofluorescence and immunoblotting. We initially found that an artichoke extract and cynaropicrin both inhibited the increase of cartilage degradation factor MMP13 and further decreased the synthesis factor aggrecan induced by TNF-α in OUMS-27. In addition, cynaropicrin suppressed the enhancement of master regulator HIF-2α on cartilage degradation and further reduced the master regulator Sox9 on cartilage synthesis induced by TNF-α. We observed that cynaropicrin suppresses NF-κB signaling, which controls HIF-2α and Sox9. Since, HIF-2α is induced by p65 (RelA), we evaluated the effect of cynaropicrin and observed that it suppressed the nuclear transport of p65 (RelA) by inhibiting phosphorylation of IκBα. Moreover, cynaropicrin not only suppressed TNF-α stimulation, it had a similar effect on IL-1β stimulation. No significant cytotoxicity with cynaropicrin was observed. These finding suggest that cynaropicrin is an effective substance that can improve the balance of cartilage metabolism, by altering the equilibrium of cartilage degradation and synthesis induced by multiple mediators know to contribute to OA. Copyright © 2016 Elsevier Inc. All rights reserved.

Antiaging Gene Klotho Regulates Adrenal CYP11B2 Expression and Aldosterone Synthesis

PubMed Central

Zhou, Xiaoli; Chen, Kai; Wang, Yongjun; Schuman, Mariano; Lei, Han

2016-01-01

Deficiency of the antiaging gene Klotho (KL) induces renal damage and hypertension through unknown mechanisms. In this study, we assessed whether KL regulates expression of CYP11B2, a key rate–limiting enzyme in aldosterone synthesis, in adrenal glands. We found that haplodeficiency of KL(+/−) in mice increased the plasma level of aldosterone by 16 weeks of age, which coincided with spontaneous and persistent elevation of BP. Blockade of aldosterone actions by eplerenone reversed KL deficiency–induced hypertension and attenuated the kidney damage. Protein expression of CYP11B2 was upregulated in adrenal cortex of KL(+/−) mice. KL and CYP11B2 proteins colocalized in adrenal zona glomerulosa cells. Silencing of KL upregulated and overexpression of KL downregulated CYP11B2 expression in human adrenocortical cells. Notably, silencing of KL decreased expression of SF-1, a negative transcription factor of CYP11B2, but increased phosphorylation of ATF2, a positive transcription factor of CYP11B2, which may contribute to upregulation of CYP11B2 expression. Therefore, these results show that KL regulates adrenal CYP11B2 expression. KL deficiency–induced spontaneous hypertension and kidney damage may be partially attributed to the upregulation of CYP11B2 expression and aldosterone synthesis. PMID:26471128

Nutrient-dependent phosphorylation channels lipid synthesis to regulate PPARα

PubMed Central

Jensen-Urstad, Anne P. L.; Song, Haowei; Lodhi, Irfan J.; Funai, Katsuhiko; Yin, Li; Coleman, Trey; Semenkovich, Clay F.

2013-01-01

Peroxisome proliferator-activated receptor (PPAR)α is a nuclear receptor that coordinates liver metabolism during fasting. Fatty acid synthase (FAS) is an enzyme that stores excess calories as fat during feeding, but it also activates hepatic PPARα by promoting synthesis of an endogenous ligand. Here we show that the mechanism underlying this paradoxical relationship involves the differential regulation of FAS in at least two distinct subcellular pools: cytoplasmic and membrane-associated. In mouse liver and cultured hepatoma cells, the ratio of cytoplasmic to membrane FAS-specific activity was increased with fasting, indicating higher cytoplasmic FAS activity under conditions associated with PPARα activation. This effect was due to a nutrient-dependent and compartment-selective covalent modification of FAS. Cytoplasmic FAS was preferentially phosphorylated during feeding or insulin treatment at Thr-1029 and Thr-1033, which flank a dehydratase domain catalytic residue. Mutating these sites to alanines promoted PPARα target gene expression. Rapamycin-induced inhibition of mammalian/mechanistic target of rapamycin complex 1 (mTORC1), a mediator of the feeding/insulin signal to induce lipogenesis, reduced FAS phosphorylation, increased cytoplasmic FAS enzyme activity, and increased PPARα target gene expression. Rapamycin-mediated induction of the same gene was abrogated with FAS knockdown. These findings suggest that hepatic FAS channels lipid synthesis through specific subcellular compartments that allow differential gene expression based on nutritional status. PMID:23585690

Cellulose Synthesis and Its Regulation

PubMed Central

Li, Shundai; Bashline, Logan; Lei, Lei; Gu, Ying

2014-01-01

Cellulose, the most abundant biopolymer synthesized on land, is made of linear chains of ß (1–4) linked D-glucose. As a major structural component of the cell wall, cellulose is important not only for industrial use but also for plant growth and development. Cellulose microfibrils are tethered by other cell wall polysaccharides such as hemicellulose, pectin, and lignin. In higher plants, cellulose is synthesized by plasma membrane-localized rosette cellulose synthase complexes. Despite the recent advances using a combination of molecular genetics, live cell imaging, and spectroscopic tools, many aspects of the cellulose synthesis remain a mystery. In this chapter, we highlight recent research progress towards understanding the mechanism of cellulose synthesis in Arabidopsis. PMID:24465174

Linkage mapping, molecular cloning and functional analysis of soybean gene Fg3 encoding flavonol 3-O-glucoside/galactoside (1 → 2) glucosyltransferase.

PubMed

Di, Shaokang; Yan, Fan; Rodas, Felipe Rojas; Rodriguez, Tito O; Murai, Yoshinori; Iwashina, Tsukasa; Sugawara, Satoko; Mori, Tetsuya; Nakabayashi, Ryo; Yonekura-Sakakibara, Keiko; Saito, Kazuki; Takahashi, Ryoji

2015-05-23

Flavonol glycosides (FGs) are major components of soybean leaves and there are substantial differences in FG composition among genotypes. The first objective of this study was to identify genes responsible for FG biosynthesis and to locate them in the soybean genome. The second objective was to clone the candidate genes and to verify their function. Recombinant inbred lines (RILs) were developed from a cross between cultivars Nezumisaya and Harosoy. HPLC comparison with authentic samples suggested that FGs having glucose at the 2″-position of glucose or galactose that is bound to the 3-position of kaempferol were present in Nezumisaya, whereas FGs of Harosoy were devoid of 2″-glucose. Conversely, FGs having glucose at the 6″-position of glucose or galactose that is bound to the 3-position of kaempferol were present in Harosoy, whereas these FGs were absent in Nezumisaya. Genetic analysis suggested that two genes control the pattern of attachment of these sugar moieties in FGs. One of the genes may be responsible for attachment of glucose to the 2″-position, probably encoding for a flavonol 3-O-glucoside/galactoside (1 → 2) glucosyltransferase. Nezumisaya may have a dominant whereas Harosoy may have a recessive allele of the gene. Based on SSR analysis, linkage mapping and genome database survey, we cloned a candidate gene designated as GmF3G2″Gt in the molecular linkage group C2 (chromosome 6). The open reading frame of GmF3G2″Gt is 1380 bp long encoding 459 amino acids with four amino acid substitutions among the cultivars. The GmF3G2″Gt recombinant protein converted kaempferol 3-O-glucoside to kaempferol 3-O-sophoroside. GmF3G2″Gt of Nezumisaya showed a broad activity for kaempferol/quercetin 3-O-glucoside/galactoside derivatives but it did not glucosylate kaempferol 3-O-rhamnosyl-(1 → 4)-[rhamnosyl-(1 → 6)-glucoside] and 3-O-rhamnosyl-(1 → 4)-[glucosyl-(1 → 6)-glucoside]. GmF3G2″Gt encodes a flavonol 3-O

cAMP dependent and independent regulation of thyroglobulin synthesis by two clones of the OVNIS 6H thyroid cell line.

PubMed

Aouani, A; Hovsépian, S; Fayet, G

1987-07-01

The hormonal regulation of thyroglobulin synthesis has been studied using two independent clones of the OVNIS 6H cell line. Insulin, hydrocortisone and TSH were able to stimulate thyroglobulin synthesis, whereas transferrin, somatostatin and glycyl-histidyl-lysine were without effect. Insulin stimulated thyroglobulin synthesis without affecting cAMP production. Hydrocortisone, when combined with insulin was a stimulator too; this stimulation was not accompanied by an increase in cAMP. TSH alone was unable to stimulate either cAMP or thyroglobulin synthesis. The stimulatory effect of TSH on thyroglobulin synthesis took place only when combined with insulin or insulin plus hydrocortisone, and was mediated by cAMP. Consequently, insulin and hydrocortisone stimulated thyroglobulin synthesis by cAMP-independent mechanisms, whereas TSH acted via the cAMP system. Forskolin mimicked TSH effects on cAMP and thyroglobulin synthesis. Calf serum inhibited cAMP and thyroglobulin production. Optimal cAMP and thyroglobulin synthesis as well as TSH responsiveness were obtained in serum-free medium supplemented with 5 micrograms/ml insulin, 100 nM hydrocortisone and 1 mU/ml TSH.

Synthesis, Delivery and Regulation of Eukaryotic Heme and Fe-S Cluster Cofactors

PubMed Central

Barupala, Dulmini P.; Dzul, Stephen P.; Riggs-Gelasco, Pamela Jo; Stemmler, Timothy L.

2016-01-01

In humans, the bulk of iron in the body (over 75%) is directed towards heme- or Fe-S cluster cofactor synthesis, and the complex, highly regulated pathways in place to accomplish biosynthesis have evolved to safely assemble and load these cofactors into apoprotein partners. In eukaryotes, heme biosynthesis is both initiated and finalized within the mitochondria, while cellular Fe-S cluster assembly is controlled by correlated pathways both within the mitochondria and within the cytosol. Iron plays a vital role in a wide array of metabolic processes and defects in iron cofactor assembly leads to human diseases. This review describes progress towards our molecular-level understanding of cellular heme and Fe-S cluster biosynthesis, focusing on the regulation and mechanistic details that are essential for understanding human disorders related to the breakdown in these essential pathways. PMID:26785297

Higher Antioxidant Activity, Total Flavonols, and Specific Quercetin Glucosides in Two Different Onion (Allium cepa L.) Varieties Grown under Organic Production: Results from a 6-Year Field Study.

PubMed

Ren, Feiyue; Reilly, Kim; Kerry, Joseph P; Gaffney, Michael; Hossain, Mohammad; Rai, Dilip K

2017-06-28

We carried out a 6-year study to assess the effect of conventional, organic, and mixed cultivation practices on bioactive compounds (flavonoids, anthocyanins) and antioxidant capacity in onion. Total flavonoids, total anthocyanins, individual flavonols, individual anthocyanins, and antioxidant activity were measured in two varieties ('Hyskin' and 'Red Baron') grown in a long-term split-plot factorial systems comparison trial. This is the first report of repeated measurements of bioactive content over an extensive time period in a single crop type within the same trial. Antioxidant activity (DPPH and FRAP), total flavonol content, and levels of Q 3,4' D and Q 3 G were higher in both varieties under fully organic compared to fully conventional management. Total flavonoids were higher in 'Red Baron' and when onions were grown under organic soil treatment. Differences were primarily due to different soil management practices used in organic agriculture rather than pesticide/ herbicide application.

Upregulation of capacity for glutathione synthesis in response to amino acid deprivation: regulation of glutamate-cysteine ligase subunits

PubMed Central

Sikalidis, Angelos K.; Mazor, Kevin M.; Lee, Jeong-In; Roman, Heather B.; Hirschberger, Lawrence L.; Stipanuk, Martha H.

2014-01-01

Using HepG2/C3A cells and MEFs, we investigated whether induction of GSH synthesis in response to sulfur amino acid deficiency is mediated by the decrease in cysteine levels or whether it requires a decrease in GSH levels per se. Both the glutamate-cysteine ligase catalytic (GCLC) and modifier (GCLM) subunit mRNA levels were upregulated in response to a lack of cysteine or other essential amino acids, independent of GSH levels. This upregulation did not occur in MEFs lacking GCN2 (general control non-derepressible 2, also known as eIF2α kinase 4) or in cells expressing mutant eIF2α lacking the eIF2α kinase Ser51 phosphorylation site, indicating that expression of both GCLC and GCLM was mediated by the GCN2/ATF4 stress response pathway. Only the increase in GCLM mRNA level, however, was accompanied by a parallel increase in protein expression, suggesting that the enhanced capacity for GSH synthesis depended largely on increased association of GCLC with its regulatory subunit. Upregulation of both GCLC and GLCM mRNA levels in response to cysteine deprivation was dependent on new protein synthesis, which is consistent with expression of GCLC and GCLM being mediated by proteins whose synthesis depends on activation of the GCN2/ATF4 pathway. Our data suggest that the regulation of GCLC expression may be mediated by changes in the abundance of transcriptional regulators, whereas the regulation of GCLM expression may be mediated by changes in the abundance of mRNA stabilizing or destabilizing proteins. Upregulation of GCLM levels in response to low cysteine levels may serve to protect the cell in the face of a future stress requiring GSH as an antioxidant or conjugating/detoxifying agent. PMID:24557597

The role of mTOR signaling in the regulation of protein synthesis and muscle mass during immobilization in mice

PubMed Central

You, Jae-Sung; Anderson, Garrett B.; Dooley, Matthew S.; Hornberger, Troy A.

2015-01-01

ABSTRACT The maintenance of skeletal muscle mass contributes substantially to health and to issues associated with the quality of life. It has been well recognized that skeletal muscle mass is regulated by mechanically induced changes in protein synthesis, and that signaling by mTOR is necessary for an increase in protein synthesis and the hypertrophy that occurs in response to increased mechanical loading. However, the role of mTOR signaling in the regulation of protein synthesis and muscle mass during decreased mechanical loading remains largely undefined. In order to define the role of mTOR signaling, we employed a mouse model of hindlimb immobilization along with pharmacological, mechanical and genetic means to modulate mTOR signaling. The results first showed that immobilization induced a decrease in the global rates of protein synthesis and muscle mass. Interestingly, immobilization also induced an increase in mTOR signaling, eIF4F complex formation and cap-dependent translation. Blocking mTOR signaling during immobilization with rapamycin not only impaired the increase in eIF4F complex formation, but also augmented the decreases in global protein synthesis and muscle mass. On the other hand, stimulating immobilized muscles with isometric contractions enhanced mTOR signaling and rescued the immobilization-induced decrease in global protein synthesis through a rapamycin-sensitive mechanism that was independent of ribosome biogenesis. Unexpectedly, the effects of isometric contractions were also independent of eIF4F complex formation. Similar to isometric contractions, overexpression of Rheb in immobilized muscles enhanced mTOR signaling, cap-dependent translation and global protein synthesis, and prevented the reduction in fiber size. Therefore, we conclude that the activation of mTOR signaling is both necessary and sufficient to alleviate the decreases in protein synthesis and muscle mass that occur during immobilization. Furthermore, these results indicate

Comparative Characterization of Total Flavonol Glycosides and Terpene Lactones at Different Ages, from Different Cultivation Sources and Genders of Ginkgo biloba Leaves

PubMed Central

Yao, Xin; Shang, Erxin; Zhou, Guisheng; Tang, Yuping; Guo, Sheng; Su, Shulan; Jin, Chun; Qian, Dawei; Qin, Yong; Duan, Jin-Ao

2012-01-01

The extract from Ginkgo biloba leaves has become a very popular plant medicine and herbal supplement for its potential benefit in alleviating symptoms associated with peripheral vascular disease, dementia, asthma and tinnitus. Most research on G. biloba leaves focus on the leaves collected in July and August from four to seven year-old trees, however a large number of leaves from fruit cultivars (trees older than 10 years) are ignored and become obsolete after fruit harvest season (November). In this paper, we expand the tree age range (from one to 300 years) and first comparatively analyze the total flavonol glycosides and terpene lactones at different ages, from different cultivation sources and genders of G. biloba leaves collected in November by using the validated HPLC-ELSD and HPLC-PDA methods. The results show that the contents of total terpene lactones and flavonol glycosides in the leaves of young ginkgo trees are higher than those in old trees, and they are higher in male trees than in female trees. Geographical factors appear to have a significant influence on the contents as well. These results will provide a good basis for the comprehensive utilization of G. biloba leaves, especially the leaves from fruit cultivars. PMID:22949862

Antiaging Gene Klotho Regulates Adrenal CYP11B2 Expression and Aldosterone Synthesis.

PubMed

Zhou, Xiaoli; Chen, Kai; Wang, Yongjun; Schuman, Mariano; Lei, Han; Sun, Zhongjie

2016-06-01

Deficiency of the antiaging gene Klotho (KL) induces renal damage and hypertension through unknown mechanisms. In this study, we assessed whether KL regulates expression of CYP11B2, a key rate-limiting enzyme in aldosterone synthesis, in adrenal glands. We found that haplodeficiency of KL(+/-) in mice increased the plasma level of aldosterone by 16 weeks of age, which coincided with spontaneous and persistent elevation of BP. Blockade of aldosterone actions by eplerenone reversed KL deficiency-induced hypertension and attenuated the kidney damage. Protein expression of CYP11B2 was upregulated in adrenal cortex of KL(+/-) mice. KL and CYP11B2 proteins colocalized in adrenal zona glomerulosa cells. Silencing of KL upregulated and overexpression of KL downregulated CYP11B2 expression in human adrenocortical cells. Notably, silencing of KL decreased expression of SF-1, a negative transcription factor of CYP11B2, but increased phosphorylation of ATF2, a positive transcription factor of CYP11B2, which may contribute to upregulation of CYP11B2 expression. Therefore, these results show that KL regulates adrenal CYP11B2 expression. KL deficiency-induced spontaneous hypertension and kidney damage may be partially attributed to the upregulation of CYP11B2 expression and aldosterone synthesis. Copyright © 2016 by the American Society of Nephrology.

A role for PPARα in the regulation of arginine metabolism and nitric oxide synthesis.

PubMed

Guelzim, Najoua; Mariotti, François; Martin, Pascal G P; Lasserre, Frédéric; Pineau, Thierry; Hermier, Dominique

2011-10-01

The pleiotropic effects of PPARα may include the regulation of amino acid metabolism. Nitric oxide (NO) is a key player in vascular homeostasis. NO synthesis may be jeopardized by a differential channeling of arginine toward urea (via arginase) versus NO (via NO synthase, NOS). This was studied in wild-type (WT) and PPARα-null (KO) mice fed diets containing either saturated fatty acids (COCO diet) or 18:3 n-3 (LIN diet). Metabolic markers of arginine metabolism were assayed in urine and plasma. mRNA levels of arginases and NOS were determined in liver. Whole-body NO synthesis and the conversion of systemic arginine into urea were assessed by using (15)N(2)-guanido-arginine and measuring urinary (15)NO(3) and [(15)N]-urea. PPARα deficiency resulted in a markedly lower whole-body NO synthesis, whereas the conversion of systemic arginine into urea remained unaffected. PPARα deficiency also increased plasma arginine and decreased citrulline concentration in plasma. These changes could not be ascribed to a direct effect on hepatic target genes, since NOS mRNA levels were unaffected, and arginase mRNA levels decreased in KO mice. Despite the low level in the diet, the nature of the fatty acids modulated some effects of PPARα deficiency, including plasma arginine and urea, which increased more in KO mice fed the LIN diet than in those fed the COCO diet. In conclusion, PPARα is largely involved in normal whole-body NO synthesis. This warrants further study on the potential of PPARα activation to maintain NO synthesis in the initiation of the metabolic syndrome.

Regulation of vascular prostaglandin synthesis by metabolites of arachidonic acid in perfused rabbit aorta.

PubMed Central

Kent, R S; Diedrich, S L; Whorton, A R

1983-01-01

To address the hypothesis that metabolites of arachidonic acid are important regulators of prostaglandin (PG) synthesis in intact vascular tissue, we studied arachidonate metabolism in rabbit aortas in response to a continuous infusion of arachidonic acid, 10 micrograms/ml. Prostacyclin (PGI2; measured as 6-keto-PGF1 alpha) production rate accelerated during the first 2 min, reached peak velocity at 2 min, and then progressively decelerated. The velocity profile of PGI2 production was similar to that previously reported for cyclooxygenase holoenzyme assayed in vitro, and was consistent with progressive inactivation of the enzymes leading to PGI2 synthesis. We determined the specific inhibition of cyclooxygenase and prostacyclin synthetase by measuring PGI2 and PGE2 production rates and by infusing cyclic endoperoxides. Our results indicate preferential inactivation of cyclooxygenase during arachidonate metabolism, most likely due to cyclooxygenase-derived oxidative intermediates. This was a dose-dependent response and resulted in a progressive decrease in the 6-keto-PGF1 alpha/PGE2 ratio. Exogenously added 15-hydroperoxy eicosatetraenoic acid, on the other hand, actually stimulated cyclooxygenase activity at low doses, while markedly inhibiting prostacyclin synthetase. This finding, along with the accelerating nature of arachidonate metabolism, is consistent with the concept of "peroxide tone" as a mediator of cyclooxygenase activity in this system. These results demonstrate that arachidonate metabolites regulate PG synthesis in intact blood vessels. The progressive enzymatic inhibition intrinsic to arachidonate metabolism may be a model for similar changes occurring in states of enhanced lipid peroxidation. These metabolic alterations might greatly influence the numerous vascular functions known to involve arachidonic acid metabolism. PMID:6409932

Phenolic antioxidants (hydrolyzable tannins, flavonols, and anthocyanins) identified by LC-ESI-MS and MALDI-QIT-TOF MS from Rosa chinensis flowers.

PubMed

Cai, Yi-Zhong; Xing, Jie; Sun, Mei; Zhan, Zhao-Qi; Corke, Harold

2005-12-28

Rosa chinensis (Yuejihua) is a well-known ornamental plant, and its flowers are commonly used in traditional Chinese medicine. Methanolic crude extracts of dried R. chinensis flowers were used for simultaneous determination of phenolic constituents by liquid chromatography-mass spectrometry (LC-MS) and matrix-assisted laser desorption/ionization quadrupole ion trap time-of-flight mass spectrometry (MALDI-QIT-TOF MS). A total of 36 known and unknown phenolics were identified as hydrolyzable tannins, flavonols, and anthocyanins, mainly including gallotannins (mono-, di-, or trigalloylglucopyranosides), ellagitannins, quercetin, quercetin/kaempferol mono- and diglycosides, and cyanidin/pelargonidin diglycosides. MALDI-QIT-TOF MS was applied not only to verify most phenolics isolated and identified by LC-MS but also to tentatively identify two ellagitannins (rugosins B and C) not isolated and unidentified by LC-MS. This study is the first to demonstrate the rapid and successful use of MALDI-QIT-TOF MS and LC-MS to directly and simultaneously identify phenolics in the crude extracts of R. chinensis flowers without any purification. The antioxidant activity of the crude extracts from R. chinensis flowers was also measured with three assay methods. The results showed that the phenolic antioxidants from R. chinensis flowers exhibited very strong radical scavenging effect and antioxidant power. High levels of flavonols and hydrolyzable tannins might be important bioactive principles in the dried R. chinensis flowers.

Complementary cereals and legumes for health: Synergistic interaction of sorghum flavones and cowpea flavonols against LPS-induced inflammation in colonic myofibroblasts.

PubMed

Agah, Shima; Kim, Hyemee; Mertens-Talcott, Susanne U; Awika, Joseph M

2017-07-01

Cereals and legumes are traditionally consumed together in many cultures, and may provide complementary health benefits beyond what is known about improved indispensable amino acid intake. Here, we use an in vitro model of inflammatory pathways to investigate whether the different flavonoids in sorghum and cowpea could synergistically reduce inflammation. Interactive effect of combining apigenin and quercetin, as well as extracts (70% acetone, v/v) from a flavone-dominated white sorghum and flavonol-dominated white cowpea, against LPS-induced NF-κB and downstream cytokines (TNF-α, IL-6, IL-8) gene and protein expression was evaluated using the CCD18Co colon myofibroblasts. Combination of apigenin and quercetin, and sorghum and cowpea extracts synergistically downregulated LPS-induced NF-κB gene and protein expression in a dose-dependent manner, with additive effect producing IC 50 values that were 14.6 and 14.0 times, respectively, higher than 1:1 combined treatments. Similar strong synergistic interactions were observed for the downstream cytokines (IC 50 values for additive effect 8.3-21 times higher than combined treatments). Furthermore, the ratios of the different combined treatments significantly affected the magnitude of synergy. Combining the structurally related cereal flavones and legume flavonols elicit strong synergistic anti-inflammatory response in LPS-stimulated nonmalignant colonocytes, likely by targeting interdependent mechanisms. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Diversity‐Oriented Synthesis of Natural‐Product‐like Libraries Containing a 3‐Methylbenzofuran Moiety for the Discovery of New Chemical Elicitors

PubMed Central

He, Xingrui; Chen, Xia; Lin, Songbo; Mo, Xiaochang; Zhou, Pengyong; Zhang, Zhihao; Lu, Yaoyao; Yang, Yu; Gu, Haining

2016-01-01

Abstract Natural products are a major source of biological molecules. The 3‐methylfuran scaffold is found in a variety of plant secondary metabolite chemical elicitors that confer host‐plant resistance against insect pests. Herein, the diversity‐oriented synthesis of a natural‐product‐like library is reported, in which the 3‐methylfuran core is fused in an angular attachment to six common natural product scaffolds—coumarin, chalcone, flavone, flavonol, isoflavone and isoquinolinone. The structural diversity of this library is assessed computationally using cheminformatic analysis. Phenotypic high‐throughput screening of β‐glucuronidase activity uncovers several hits. Further in vivo screening confirms that these hits can induce resistance in rice to nymphs of the brown planthopper Nilaparvata lugens. This work validates the combination of diversity‐oriented synthesis and high‐throughput screening of β‐glucuronidase activity as a strategy for discovering new chemical elicitors. PMID:28168155

Evidence for a systemic regulation of neurotrophin synthesis in response to peripheral nerve injury.

PubMed

Shakhbazau, Antos; Martinez, Jose A; Xu, Qing-Gui; Kawasoe, Jean; van Minnen, Jan; Midha, Rajiv

2012-08-01

Up-regulation of neurotrophin synthesis is an important mechanism of peripheral nerve regeneration after injury. Neurotrophin expression is regulated by a complex series of events including cell interactions and multiple molecular stimuli. We have studied neurotrophin synthesis at 2 weeks time-point in a transvertebral model of unilateral or bilateral transection of sciatic nerve in rats. We have found that unilateral sciatic nerve transection results in the elevation of nerve growth factor (NGF) and NT-3, but not glial cell-line derived neurotrophic factor or brain-derived neural factor, in the uninjured nerve on the contralateral side, commonly considered as a control. Bilateral transection further increased NGF but not other neurotrophins in the nerve segment distal to the transection site, as compared to the unilateral injury. To further investigate the distinct role of NGF in regeneration and its potential for peripheral nerve repair, we transduced isogeneic Schwann cells with NGF-encoding lentivirus and transplanted the over-expressing cells into the distal segment of a transected nerve. Axonal regeneration was studied at 2 weeks time-point using pan-neuronal marker NF-200 and found to directly correlate with NGF levels in the regenerating nerve. © 2012 The Authors. Journal of Neurochemistry © 2012 International Society for Neurochemistry.

Pretranslational regulation of the synthesis of the third component of complement in human mononuclear phagocytes by the lipid A portion of lipopolysaccharide.

PubMed Central

Strunk, R C; Whitehead, A S; Cole, F S

1985-01-01

The third component of complement (C3) is a plasma glycoprotein with a variety of biologic functions in the initiation and maintenance of host response to infectious agents. While the hepatocyte is the primary source of plasma C3, mononuclear phagocytes contribute to the regulation of tissue availability of C3. Lipopolysaccharide (LPS), a constituent of cell walls of gram-negative bacteria, consists of a polysaccharide moiety (core polysaccharide and O antigen) covalently linked to a lipid portion (lipid A). Using metabolic labeling with [35S]methionine, immunoprecipitation, and SDS-polyacrylamide gel electrophoresis, we examined the effects of LPS on synthesis of C3 by human mononuclear phagocytes as well as synthesis of the second component of complement (C2), factor B, lysozyme, and total protein. LPS increased C3 synthesis 5-30-fold without affecting the kinetics of secretion of C3 or the synthesis of C2, lysozyme, or total protein. Factor B synthesis was consistently increased by LPS. Experiments with lipid A-inactivated LPS (alkaline treated), LPS from a polysaccharide mutant strain, and lipid X (a lipid A precursor) indicated that the lipid A portion is the structural element required for this effect. Northern blot analysis demonstrated at least a fivefold increase in C3 mRNA in LPS-treated monolayers, which suggests that the regulation of the increase in C3 synthesis is pretranslational. C2 mRNA and factor B mRNA were increased approximately twofold. The availability of specific gene products in human mononuclear phagocytes that respond to LPS should permit understanding of the molecular regulation of more complex functions of these cells elicited by LPS in which multiple gene products are coordinately expressed. Images PMID:3900137

A sensitive microextraction by packed sorbent-based methodology combined with ultra-high pressure liquid chromatography as a powerful technique for analysis of biologically active flavonols in wines.

PubMed

Silva, Catarina L; Gonçalves, João L; Câmara, José S

2012-08-20

A new approach based on microextraction by packed sorbent (MEPS) and reversed-phase high-throughput ultra high pressure liquid chromatography (UHPLC) method that uses a gradient elution and diode array detection to quantitate three biologically active flavonols in wines, myricetin, quercetin, and kaempferol, is described. In addition to performing routine experiments to establish the validity of the assay to internationally accepted criteria (selectivity, linearity, sensitivity, precision, accuracy), experiments are included to assess the effect of the important experimental parameters such as the type of sorbent material (C2, C8, C18, SIL, and C8/SCX), number of extraction cycles (extract-discard), elution volume, sample volume, and ethanol content, on the MEPS performance. The optimal conditions of MEPS extraction were obtained using C8 sorbent and small sample volumes (250μL) in five extraction cycle and in a short time period (about 5min for the entire sample preparation step). Under optimized conditions, excellent linearity (R(values)(2)>0.9963), limits of detection of 0.006μgmL(-1) (quercetin) to 0.013μgmL(-1) (myricetin) and precision within 0.5-3.1% were observed for the target flavonols. The average recoveries of myricetin, quercetin and kaempferol for real samples were 83.0-97.7% with relative standard deviation (RSD, %) lower than 1.6%. The results obtained showed that the most abundant flavonol in the analyzed samples was myricetin (5.8±3.7μgmL(-1)). Quercetin (0.97±0.41μgmL(-1)) and kaempferol (0.66±0.24μgmL(-1)) were found in a lower concentration. The optimized MEPS(C8) method was compared with a reverse-phase solid-phase extraction (SPE) procedure using as sorbent a macroporous copolymer made from a balanced ratio of two monomers, the lipophilic divinylbenzene and the hydrophilic N-vinylpyrrolidone (Oasis HLB) were used as reference. MEPS(C8) approach offers an attractive alternative for analysis of flavonols in wines, providing a number of

Vitamin B12–dependent taurine synthesis regulates growth and bone mass

PubMed Central

Roman-Garcia, Pablo; Quiros-Gonzalez, Isabel; Mottram, Lynda; Lieben, Liesbet; Sharan, Kunal; Wangwiwatsin, Arporn; Tubio, Jose; Lewis, Kirsty; Wilkinson, Debbie; Santhanam, Balaji; Sarper, Nazan; Clare, Simon; Vassiliou, George S.; Velagapudi, Vidya R.; Dougan, Gordon; Yadav, Vijay K.

2014-01-01

Both maternal and offspring-derived factors contribute to lifelong growth and bone mass accrual, although the specific role of maternal deficiencies in the growth and bone mass of offspring is poorly understood. In the present study, we have shown that vitamin B12 (B12) deficiency in a murine genetic model results in severe postweaning growth retardation and osteoporosis, and the severity and time of onset of this phenotype in the offspring depends on the maternal genotype. Using integrated physiological and metabolomic analysis, we determined that B12 deficiency in the offspring decreases liver taurine production and associates with abrogation of a growth hormone/insulin-like growth factor 1 (GH/IGF1) axis. Taurine increased GH-dependent IGF1 synthesis in the liver, which subsequently enhanced osteoblast function, and in B12-deficient offspring, oral administration of taurine rescued their growth retardation and osteoporosis phenotypes. These results identify B12 as an essential vitamin that positively regulates postweaning growth and bone formation through taurine synthesis and suggests potential therapies to increase bone mass. PMID:24911144

Synthesis, delivery and regulation of eukaryotic heme and Fe-S cluster cofactors.

PubMed

Barupala, Dulmini P; Dzul, Stephen P; Riggs-Gelasco, Pamela Jo; Stemmler, Timothy L

2016-02-15

In humans, the bulk of iron in the body (over 75%) is directed towards heme- or Fe-S cluster cofactor synthesis, and the complex, highly regulated pathways in place to accomplish biosynthesis have evolved to safely assemble and load these cofactors into apoprotein partners. In eukaryotes, heme biosynthesis is both initiated and finalized within the mitochondria, while cellular Fe-S cluster assembly is controlled by correlated pathways both within the mitochondria and within the cytosol. Iron plays a vital role in a wide array of metabolic processes and defects in iron cofactor assembly leads to human diseases. This review describes progress towards our molecular-level understanding of cellular heme and Fe-S cluster biosynthesis, focusing on the regulation and mechanistic details that are essential for understanding human disorders related to the breakdown in these essential pathways. Copyright © 2016 Elsevier Inc. All rights reserved.

Synthesis and regulation of chlorogenic acid in potato: Rerouting phenylpropanoid flux in HQT-silenced lines.

PubMed

Payyavula, Raja S; Shakya, Roshani; Sengoda, Venkatesan G; Munyaneza, Joseph E; Swamy, Prashant; Navarre, Duroy A

2015-05-01

Chlorogenic acid (CGA) is the major phenolic sink in potato tubers and can constitute over 90% of total phenylpropanoids. The regulation of CGA biosynthesis in potato and the role of the CGA biosynthetic gene hydroxycinnamoyl CoA:quinate hydroxycinnamoyl transferase (HQT) was characterized. A sucrose induced accumulation of CGA correlated with the increased expression of phenylalanine ammonia-lyase (PAL) rather than HQT. Transient expression of the potato MYB transcription factor StAN1 (anthocyanin 1) in tobacco increased CGA. RNAi suppression of HQT resulted in over a 90% reduction in CGA and resulted in early flowering. The reduction in total phenolics and antioxidant capacity was less than the reduction in CGA, suggesting flux was rerouted into other phenylpropanoids. Network analysis showed distinct patterns in different organs, with anthocyanins and phenolic acids showing negative correlations in leaves and flowers and positive in tubers. Some flavonols increased in flowers, but not in leaves or tubers. Anthocyanins increased in flowers and showed a trend to increase in leaves, but not tubers. HQT suppression increased biosynthesis of caffeoyl polyamines, some of which are not previously reported in potato. Decreased PAL expression and enzyme activity was observed in HQT suppressed lines, suggesting the existence of a regulatory loop between CGA and PAL. Electrophysiology detected no effect of CGA suppression on potato psyllid feeding. Collectively, this research showed that CGA in potatoes is synthesized through HQT and HQT suppression altered phenotype and redirected phenylpropanoid flux. © 2014 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Interaction of moderate UV-B exposure and temperature on the formation of structurally different flavonol glycosides and hydroxycinnamic acid derivatives in kale (Brassica oleracea var. sabellica).

PubMed

Neugart, Susanne; Fiol, Michaela; Schreiner, Monika; Rohn, Sascha; Zrenner, Rita; Kroh, Lothar W; Krumbein, Angelika

2014-05-07

Kale has a high number of structurally different flavonol glycosides and hydroxycinnamic acid derivatives. In this study we investigated the interaction of moderate UV-B radiation and temperature on these compounds. Kale plants were grown at daily mean temperatures of 5 or 15 °C and were exposed to five subsequent daily doses (each 0.25 kJ m(-2) d(-1)) of moderate UV-B radiation at 1 d intervals. Of 20 phenolic compounds, 11 were influenced by an interaction of UV-B radiation and temperature, e.g., monoacylated quercetin glycosides. Concomitantly, enhanced mRNA expression of flavonol 3'- hydroxylase showed an interaction of UV-B and temperature, highest at 0.75 kJ m(-2) and 15 °C. Kaempferol glycosides responded diversely and dependent on, e.g., the hydroxycinnamic acid residue. Compounds containing a catechol structure seem to be favored in the response to UV-B. Taken together, subsequent exposure to moderate UV-B radiation is a successful tool for enhancing the flavonoid profile of plants, and temperature should be considered.

Skeletal muscle plasticity induced by seasonal acclimatization in carp involves differential expression of rRNA and molecules that epigenetically regulate its synthesis.

PubMed

Fuentes, Eduardo N; Zuloaga, Rodrigo; Nardocci, Gino; Fernandez de la Reguera, Catalina; Simonet, Nicolas; Fumeron, Robinson; Valdes, Juan Antonio; Molina, Alfredo; Alvarez, Marco

2014-01-01

Ribosomal biogenesis controls cellular growth in living organisms, with the rate-limiting step of this process being the transcription of ribosomal DNA (rDNA). Considering that epigenetic mechanisms allow an organism to respond to environmental changes, the expression in muscle of several molecules that regulate epigenetic rRNA synthesis, as well as rDNA transcription, were evaluated during the seasonal acclimatization of the carp. First, the nucleotide sequences encoding the components forming the NoRC (ttf-I, tip5) and eNoSC (sirt1, nml, suv39h1), two chromatin remodeling complexes that silence rRNA synthesis, as well as the sequence of ubf1, a key regulator of rDNA transcription, were obtained. Subsequently the transcriptional regulation of the aforementioned molecules, and other key molecules involved in rRNA synthesis (mh2a1, mh2a2, h2a.z, h2a.z.7, nuc, p80), was assessed. The carp sequences for TTF-I, TIP5, SIRT1, NML, SUV39H1, and UBF1 showed a high conservation of domains and key amino acids in comparison with other fish and higher vertebrates. The mRNA contents in muscle for ttf-I, tip5, sirt1, nml, suv39h1, mh2a1, mh2a.z, and nuc were up-regulated during winter in comparison with summer, whereas the mRNA levels of mh2a2, ubf1, and p80 were down-regulated. Also, the contents of molecules involved in processing the rRNA (snoRNAs) and pRNA, a stabilizer of NoRC complex, were analyzed, finding that these non-coding RNAs were not affected by seasonal acclimatization. These results suggest that variations in the expression of rRNA and the molecules that epigenetically regulate its synthesis are contributing to the muscle plasticity induced by seasonal acclimatization in carp. Copyright © 2014 Elsevier Inc. All rights reserved.

Live Attenuated Salmonella Vaccines Displaying Regulated Delayed Lysis and Delayed Antigen Synthesis To Confer Protection against Mycobacterium tuberculosis

PubMed Central

Juárez-Rodríguez, María Dolores; Yang, Jiseon; Kader, Rebin; Alamuri, Praveen; Curtiss, Roy

2012-01-01

Live recombinant attenuated Salmonella vaccine (RASV) strains have great potential to induce protective immunity against Mycobacterium tuberculosis by delivering M. tuberculosis antigens. Recently, we reported that, in orally immunized mice, RASV strains delivering the M. tuberculosis early secreted antigenic target 6-kDa (ESAT-6) protein and culture filtrate protein 10 (CFP-10) antigens via the Salmonella type III secretion system (SopE amino-terminal region residues 1 to 80 with two copies of ESAT-6 and one copy of CFP-10 [SopENt80-E2C]) afforded protection against aerosol challenge with M. tuberculosis. Here, we constructed and evaluated an improved Salmonella vaccine against M. tuberculosis. We constructed translational fusions for the synthesis of two copies of ESAT-6 plus CFP-10 fused to the OmpC signal sequence (OmpCSS-E2C) and amino acids 44 to 338 of antigen 85A (Ag85A294) flanked by the signal sequence (SS) and C-terminal peptide (CT) of β-lactamase (BlaSS-Ag85A294-BlaCT) to enable delivery via the Salmonella type II secretion system. The genes expressing these proteins were cloned as an operon transcribed from Ptrc into isogenic Asd+/MurA+ pYA3681 lysis vector derivatives with different replication origins (pBR, p15A, pSC101), resulting in pYA4890, pYA4891, and pYA4892 for SopENt80-E2C/Ag85A294 synthesis and pYA4893 and pYA4894 for OmpCSS-E2C/Ag85A294 synthesis. Mice orally immunized with the RASV χ11021 strain engineered to display regulated delayed lysis and regulated delayed antigen synthesis in vivo and harboring pYA4891, pYA4893, or pYA4894 elicited significantly greater humoral and cellular immune responses, and the RASV χ11021 strain afforded a greater degree of protection against M. tuberculosis aerosol challenge in mice than RASVs harboring any other Asd+/MurA+ lysis plasmid and immunization with M. bovis BCG, demonstrating that RASV strains displaying regulated delayed lysis with delayed antigen synthesis resulted in highly immunogenic

Regulation of leptin synthesis in white adipose tissue of the female fruit bat, Cynopterus sphinx: role of melatonin with or without insulin.

PubMed

Banerjee, A; Udin, S; Krishna, A

2011-02-01

Factors regulating leptin synthesis during adipogenesis in wild species are not well known. Studies in the female Cynopterus sphinx bat have shown that it undergoes seasonal changes in its fat deposition and serum leptin and melatonin levels. The aim of the present study was to investigate the hormonal regulation of leptin synthesis by the white adipose tissue during the period of fat deposition in female C. sphinx. This study showed a significant correlation between the seasonal changes in serum melatonin level with the circulating leptin level (r = 0.78; P < 0.05) and with the changes in body fat mass (r = 0.88; P < 0.05) in C. sphinx. A significant correlation between circulating insulin and leptin levels (r = 0.65; P < 0.05) was also found in this species. This in vivo finding suggests that melatonin together with insulin may enhance leptin synthesis by increasing adipose tissue accumulation. The in vitro study showed that melatonin interacts synergistically with insulin in stimulating leptin synthesis by adipose tissue in C. sphinx. The study showed MT(2) receptors in adipose tissue and a stimulatory effect of melatonin on leptin synthesis, which was blocked by treatment with an MT(2) receptor antagonist, suggesting that the effect of melatonin on leptin synthesis by adipose tissue is mediated through the MT(2) receptor in C. sphinx. The in vitro study showed that the synthesis of leptin is directly proportional to the amount of glucose uptake by the adipose tissue. It further showed that melatonin together with insulin synergistically enhanced the leptin synthesis by adipose tissue through phosphorylation of mitogen-activated protein kinase in C. sphinx.

Interaction of TIF-90 and filamin A in the regulation of rRNA synthesis in leukemic cells.

PubMed

Nguyen, Le Xuan Truong; Chan, Steven M; Ngo, Tri Duc; Raval, Aparna; Kim, Kyeong Kyu; Majeti, Ravindra; Mitchell, Beverly S

2014-07-24

The transcription initiation factor I (TIF-IA) is an important regulator of the synthesis of ribosomal RNA (rRNA) through its facilitation of the recruitment of RNA polymerase I (Pol I) to the ribosomal DNA promoter. Activation of the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway, which occurs commonly in acute myelogenous leukemia, enhances rRNA synthesis through TIF-IA stabilization and phosphorylation. We have discovered that TIF-IA coexists with a splicing isoform, TIF-90, which is expressed preferentially in the nucleolus and at higher levels in proliferating and transformed hematopoietic cells. TIF-90 interacts directly with Pol I to increase rRNA synthesis as a consequence of Akt activation. Furthermore, TIF-90 binds preferentially to a 90-kDa cleavage product of the actin binding protein filamin A (FLNA) that inhibits rRNA synthesis. Increased expression of TIF-90 overcomes the inhibitory effect of this cleavage product and stimulates rRNA synthesis. Because activated Akt also reduces FLNA cleavage, these results indicate that activated Akt and TIF-90 function in parallel to increase rRNA synthesis and, as a consequence, cell proliferation in leukemic cells. These results provide evidence that the direct targeting of Akt would be an effective therapy in acute leukemias in which Akt is activated. © 2014 by The American Society of Hematology.

Catalase anabolism in yeast: loss of regulation by oxygen of catalase apoprotein synthesis after mutation.

PubMed

Berte, C; Sels, A

1979-04-17

A mutant of Saccharomyces cerevisiae which displays catalase activity when grown under strictly anaerobic conditions has been selected on solid media. Although some preformed holoenzyme has accumulated in anaerobic cells, a sharp increase of activity is still measured during adaptation to oxygen in glucose-buffer; however, a striking difference with the wild-type strain is that in the mutant, catalase formation is observed in the presence of cycloheximide that totally inhibits cytoplasmic translation. It is concluded that kat 80 mutant has lost the regulatory control by oxygen of apocatalase synthesis; the later precursor, characterized as apocatalase synthesis; the latter precursor, characterized as apocatalase T, is thought to be activated in vivo, under aerobic conditions, by inclusion of prosthetic group. Regulation of enzyme synthesis by catabolite repression (glucose erfect) persists, unmodified by reference to the wild-type parental strain. Mutation kat 80 specifically hits catalase anabolism, as no significant variations were observed for the edification of the respiratory system and (apo)cytochrome c peroxidase production. Genetic analysis shows that kat 80 phenotype, recessive in heterozygotes, results from a single nuclear mutation.

Increased chalcone synthase (CHS) expression is associated with dicamba resistance in Kochia scoparia.

PubMed

Pettinga, Dean J; Ou, Junjun; Patterson, Eric L; Jugulam, Mithila; Westra, Philip; Gaines, Todd A

2017-10-30

Resistance to the synthetic auxin herbicide dicamba is increasingly problematic in Kochia scoparia. The resistance mechanism in an inbred dicamba-resistant K. scoparia line (9425R) was investigated using physiological and transcriptomics (RNA-Seq) approaches. No differences were found in dicamba absorption or metabolism between 9425R and a dicamba-susceptible line, but 9425R was found to have significantly reduced dicamba translocation. Known auxin-responsive genes ACC synthase (ACS) and indole-3-acetic acid amino synthetase (GH3) were transcriptionally induced following dicamba treatment in dicamba-susceptible K. scoparia but not in 9425R. Chalcone synthase (CHS), the gene regulating synthesis of the flavonols quertecin and kaemperfol, was found to have twofold higher transcription in 9425R both without and 12 h after dicamba treatment. Increased CHS transcription co-segregated with dicamba resistance in a forward genetics screen using an F 2 population. Prior work has shown that the flavonols quertecin and kaemperfol compete with auxin for intercellular movement and vascular loading via ATP-binding cassette subfamily B (ABCB) membrane transporters. The results of this study support a model in which constitutively increased CHS expression in the meristem produces more flavonols that would compete with dicamba for intercellular transport by ABCB transporters, resulting in reduced dicamba translocation. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Sumoylation of the Tumor Suppressor Promyelocytic Leukemia Protein Regulates Arsenic Trioxide-Induced Collagen Synthesis in Osteoblasts.

PubMed

Xu, Wen-Xiao; Liu, Sheng-Zhi; Wu, Di; Qiao, Guo-Fen; Yan, Jinglong

2015-01-01

Promyelocytic leukemia (PML) protein is a tumor suppressor that fuses with retinoic acid receptor-α (PML-RARα) to contribute to the initiation of acute promyelocytic leukemia (APL). Arsenic trioxide (ATO) upregulates expression of TGF-β1, promoting collagen synthesis in osteoblasts, and ATO binds directly to PML to induce oligomerization, sumoylation, and ubiquitination. However, how ATO upregulates TGF-β1 expression is uncertain. Thus, we suggested that PML sumoylation is responsible for regulation of TGF-β1 protein expression. Kunming mice were treated with ATO, and osteoblasts were counted under scanning electron microscopy. Masson's staining was used to quantify collagen content. hFOB1.19 cells were transfected with siRNA against UBC9 or RNF4, and then treated with ATO or FBS. TGF-β1, PML expression, and sumoylation were quantified with Western blot, and collagen quantified via immunocytochemistry. ATO enhanced osteoblast accumulation, collagen synthesis, and PML-NB formation in vivo. Knocking down UBC9 in hFOB1.19 cells inhibited ATO- and FBS-induced PML sumoylation, TGF-β1 expression, and collagen synthesis. Conversely, knocking down RNF4 enhanced ATO- and FBS-induced PML sumoylation, TGF-β1 expression, and collagen synthesis. These data suggest that PML sumoylation is required for ATO-induced collagen synthesis in osteoblasts. © 2015 S. Karger AG, Basel.

PECULIAIRITIES OF MELATONIN EFFECT ON CHONORHYTMIC ORGANIZATION OF KIDNEY ACID-REGULATING FUNCTION INFLUENCED BY NITROGEN MONOXIDE SYNTHESIS BLOCKADE UNDER CONDITIONS OF PINEAL GLAND HYPOFUNCTION.

PubMed

Semenenko, S; Tymofiychuk, I; Boreyko, L; Karatieieva, S; Slobodian, K

2017-10-01

The objective of research is to study the peculiarities of melatonin effect on chronorhythmic organization of the kidney acid-regulating function influenced by nitrogen monoxide (NO) synthesis blockade under conditions of pineal gland (PG) hypofunction. The experiments were conducted on 72 mature non-linear albino male rats with their body mass 0,15-0,18 kg. The animals were kept under vivarium conditions at a stable temperature and air humidity fed on a standard dietary intake. The control group included animals (n=36) kept under conditions of usual light regimen (12.00С:12.00Т) during 7 days. The experimental group included animals (n=36) injected with N-nitro-L-arginine (L-NNA) in the dose of 20 mg/kg during 7 days under conditions of continuous light (12.00С:12.00С) and melatonin in the dose of 0,5 mg/kg during 7 days simultaneously. On the 8th day the animals were exposed to 5% water load with heated to room temperature water supplied and the parameters of the kidney acid-regulating function under conditions of forced diuresis were investigated. Kidney functions in the control animals are subordinated to accurate circadian organization. Daily rhythms of the parameters of kidney acid-regulating functions reflect similar changes of the renal processes. Chronorhythmic transformations of the kidney acid-regulating functions in animals with blocked NO synthesis against continuous light and parallel injection of melatonin enable to suggest that NO synthesis blockade under conditions of melatonin correction reduces daily mean pH level as compared to the control. Although, it was higher than that in the animals with blocked NO synthesis against the ground of physiological function of the pineal gland, and animals with PG hypofunction under conditions of NO synthesis blockadeю Therefore, under conditions of L-NNA blockade of NO synthesis and injection of melatonin influenced by PG hypofunction chronorhythmic transformations of architectonics and phase structure of

Characterization and content of flavonol derivatives of Allium ursinum L. plant.

PubMed

Oszmiański, J; Kolniak-Ostek, J; Wojdyło, A

2013-01-09

The phenolic compounds were extracted from green and yellow leaves, stalks, and seeds of garlic ( Allium ursinum L.). The extracts were analyzed by liquid chromatography-photodiode array detector-electrospray ionization-tandem mass spectrometry (LC-PDA-ESI-MS/MS). In total, 21 compounds were detected. The flavonol derivatives were identified on the basis of their ultraviolet (UV) spectra and fragmentation patterns in collision-induced dissociation experiments. On the basis of accurate MS and MS/MS data, six compounds were newly identified in bear's garlic, mainly the kaempferol derivatives. As far as the investigated parts of garlic are concerned, the kaempferol derivatives were found to be predominant in yellow leaves [2362.96 mg/100 g of dry matter (dm)], followed by green leaves (1856.31 mg/100 g of dm). Seeds contained the minimal phenolic compounds, less than stalks. The yellow leaves of A. ursinum possessed a much larger content of compounds acylated with p-coumaric acid than green leaves (1299.97 versus 855.67 mg/100 g of dm, respectively). The stalks and seeds contained much more non-acetylated than acetylated flavonoid glycosides with p-coumaric acid compounds (162.4 versus 62.82 mg/100 g of dm and 105.49 versus 24.18 mg/100 g of dm, respectively).

Regulation of error-prone translesion synthesis by Spartan/C1orf124

PubMed Central

Kim, Myoung Shin; Machida, Yuka; Vashisht, Ajay A.; Wohlschlegel, James A.; Pang, Yuan-Ping; Machida, Yuichi J.

2013-01-01

Translesion synthesis (TLS) employs low fidelity polymerases to replicate past damaged DNA in a potentially error-prone process. Regulatory mechanisms that prevent TLS-associated mutagenesis are unknown; however, our recent studies suggest that the PCNA-binding protein Spartan plays a role in suppression of damage-induced mutagenesis. Here, we show that Spartan negatively regulates error-prone TLS that is dependent on POLD3, the accessory subunit of the replicative DNA polymerase Pol δ. We demonstrate that the putative zinc metalloprotease domain SprT in Spartan directly interacts with POLD3 and contributes to suppression of damage-induced mutagenesis. Depletion of Spartan induces complex formation of POLD3 with Rev1 and the error-prone TLS polymerase Pol ζ, and elevates mutagenesis that relies on POLD3, Rev1 and Pol ζ. These results suggest that Spartan negatively regulates POLD3 function in Rev1/Pol ζ-dependent TLS, revealing a previously unrecognized regulatory step in error-prone TLS. PMID:23254330

An Unconventional Diacylglycerol Kinase That Regulates Phospholipid Synthesis and Nuclear Membrane Growth*♦

PubMed Central

Han, Gil-Soo; O'Hara, Laura; Carman, George M.; Siniossoglou, Symeon

2008-01-01

Changes in nuclear size and shape during the cell cycle or during development require coordinated nuclear membrane remodeling, but the underlying molecular events are largely unknown. We have shown previously that the activity of the conserved phosphatidate phosphatase Pah1p/Smp2p regulates nuclear structure in yeast by controlling phospholipid synthesis and membrane biogenesis at the nuclear envelope. Two screens for novel regulators of phosphatidate led to the identification of DGK1. We show that Dgk1p is a unique diacylglycerol kinase that uses CTP, instead of ATP, to generate phosphatidate. DGK1 counteracts the activity of PAH1 at the nuclear envelope by controlling phosphatidate levels. Overexpression of DGK1 causes the appearance of phosphatidate-enriched membranes around the nucleus and leads to its expansion, without proliferating the cortical endoplasmic reticulum membrane. Mutations that decrease phosphatidate levels decrease nuclear membrane growth in pah1Δ cells. We propose that phosphatidate metabolism is a critical factor determining nuclear structure by regulating nuclear membrane biogenesis. PMID:18458075

The Alternative Sigma Factor SigX Controls Bacteriocin Synthesis and Competence, the Two Quorum Sensing Regulated Traits in Streptococcus mutans

PubMed Central

Reck, Michael; Tomasch, Jürgen; Wagner-Döbler, Irene

2015-01-01

Two small quorum sensing (QS) peptides regulate competence in S. mutans in a cell density dependent manner: XIP (sigX inducing peptide) and CSP (competence stimulating peptide). Depending on the environmental conditions isogenic S. mutans cells can split into a competent and non-competent subpopulation. The origin of this population heterogeneity has not been experimentally determined and it is unknown how the two QS systems are connected. We developed a toolbox of single and dual fluorescent reporter strains and systematically knocked out key genes of the competence signaling cascade in the reporter strain backgrounds. By following signal propagation on the single cell level we discovered that the master regulator of competence, the alternative sigma factor SigX, directly controls expression of the response regulator for bacteriocin synthesis ComE. Consequently, a SigX binding motif (cin-box) was identified in the promoter region of comE. Overexpressing the genetic components involved in competence development demonstrated that ComRS represents the origin of bimodality and determines the modality of the downstream regulators SigX and ComE. Moreover these analysis showed that there is no direct regulatory link between the two QS signaling cascades. Competence is induced through a hierarchical XIP signaling cascade, which has no regulatory input from the CSP cascade. CSP exclusively regulates bacteriocin synthesis. We suggest renaming it mutacin inducing peptide (MIP). Finally, using phosphomimetic comE mutants we show that unimodal bacteriocin production is controlled posttranslationally, thus solving the puzzling observation that in complex media competence is observed in a subpopulation only, while at the same time all cells produce bacteriocins. The control of both bacteriocin synthesis and competence through the alternative sigma-factor SigX suggests that S. mutans increases its genetic repertoire via QS controlled predation on neighboring species in its

Early steps in protein synthesis and their regulation: a background study related to the biological effects of radiation. Progress report, July 1, 1975--June 30, 1976

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

Zamecnik, P.C.

This is a continuing study of protein synthesis, involving a search for the role of Ap/sub 4/A and other unusual nucleotides in growth regulation; studies of the mechanism of action of aminoacyl-tRNA ligases and the effect thereof on protein synthesis; a search for new regulators of the translation step, in cell-free systems; and an effort to improve the sensitivity and quantitation of chemical sequencing at the 3'-end of messenger RNA.

Gene expression and metabolite profiling of developing highbush blueberry fruit indicates transcriptional regulation of flavonoid metabolism and activation of abscisic acid metabolism.

PubMed

Zifkin, Michael; Jin, Alena; Ozga, Jocelyn A; Zaharia, L Irina; Schernthaner, Johann P; Gesell, Andreas; Abrams, Suzanne R; Kennedy, James A; Constabel, C Peter

2012-01-01

Highbush blueberry (Vaccinium corymbosum) fruits contain substantial quantities of flavonoids, which are implicated in a wide range of health benefits. Although the flavonoid constituents of ripe blueberries are known, the molecular genetics underlying their biosynthesis, localization, and changes that occur during development have not been investigated. Two expressed sequence tag libraries from ripening blueberry fruit were constructed as a resource for gene identification and quantitative real-time reverse transcription-polymerase chain reaction primer design. Gene expression profiling by quantitative real-time reverse transcription-polymerase chain reaction showed that flavonoid biosynthetic transcript abundance followed a tightly regulated biphasic pattern, and transcript profiles were consistent with the abundance of the three major classes of flavonoids. Proanthocyanidins (PAs) and corresponding biosynthetic transcripts encoding anthocyanidin reductase and leucoanthocyanidin reductase were most concentrated in young fruit and localized predominantly to the inner fruit tissue containing the seeds and placentae. Mean PA polymer length was seven to 8.5 subunits, linked predominantly via B-type linkages, and was relatively constant throughout development. Flavonol accumulation and localization patterns were similar to those of the PAs, and the B-ring hydroxylation pattern of both was correlated with flavonoid-3'-hydroxylase transcript abundance. By contrast, anthocyanins accumulated late in maturation, which coincided with a peak in flavonoid-3-O-glycosyltransferase and flavonoid-3'5'-hydroxylase transcripts. Transcripts of VcMYBPA1, which likely encodes an R2R3-MYB transcriptional regulator of PA synthesis, were prominent in both phases of development. Furthermore, the initiation of ripening was accompanied by a substantial rise in abscisic acid, a growth regulator that may be an important component of the ripening process and contribute to the regulation of

Identification of flavonols in leaves of Maytenus ilicifolia and M. aquifolium (Celastraceae) by LC/UV/MS analysis.

PubMed

Tiberti, Luciana A; Yariwake, Janete H; Ndjoko, Karine; Hostettmann, Kurt

2007-02-01

A comparative analysis of the flavonoid components of the leaves of two medicinal plants known in Brazil as "espinheira santa", namely, Maytenus ilicifolia Mart. ex Reiss. and M. aquifolium Mart. (Celastraceae), and a hybrid plant, M. aquifoliumxM. ilicifolia, has been carried out using high-performance liquid chromatography coupled with photodiode array UV detection and mass spectrometry. One methoxyflavonoid glycoside and 18 flavonol-3-O-glycosides were identified in the extracts on the basis of their on-line UV spectra (measured in the absence and presence of shift reagents) and multiple stage mass spectral data. Fingerprint analysis of the flavonoid extracts revealed significant differences in the profiles of the two Maytenus species, while the hybrid plant contained flavonoids found in both parent species.

Tc-MYBPA an Arabidopsis TT2-like transcription factor and functions in the regulation of proanthocyanidin synthesis in Theobroma cacao.

PubMed

Liu, Yi; Shi, Zi; Maximova, Siela N; Payne, Mark J; Guiltinan, Mark J

2015-06-25

The flavan-3-ols catechin and epicatechin, and their polymerized oligomers, the proanthocyanidins (PAs, also called condensed tannins), accumulate to levels of up to 15 % of the total weight of dry seeds of Theobroma cacao L. These compounds have been associated with several health benefits in humans. They also play important roles in pest and disease defense throughout the plant. In Arabidopsis, the R2R3 type MYB transcription factor TT2 regulates the major genes leading to the synthesis of PA. To explore the transcriptional regulation of the PA synthesis pathway in cacao, we isolated and characterized an R2R3 type MYB transcription factor MYBPA from cacao. We examined the spatial and temporal gene expression patterns of the Tc-MYBPA gene and found it to be developmentally expressed in a manner consistent with its involvement in PAs and anthocyanin synthesis. Functional complementation of an Arabidopsis tt2 mutant with Tc-MYBPA suggested that it can functionally substitute the Arabidopsis TT2 gene. Interestingly, in addition to PA accumulation in seeds of the Tc-MYBPA expressing plants, we also observed an obvious increase of anthocyanidin accumulation in hypocotyls. We observed that overexpression of the Tc-MYBPA gene resulted in increased expression of several key genes encoding the major structural enzymes of the PA and anthocyanidin pathway, including DFR (dihydroflavanol reductase), LDOX (leucoanthocyanidin dioxygenase) and BAN (ANR, anthocyanidin reductase). We conclude that the Tc-MYBPA gene that encodes an R2R3 type MYB transcription factor is an Arabidopsis TT2 like transcription factor, and may be involved in the regulation of both anthocyanin and PA synthesis in cacao. This research may provide molecular tools for breeding of cacao varieties with improved disease resistance and enhanced flavonoid profiles for nutritional and pharmaceutical applications.

Effects of Parsley (Petroselinum crispum) and its Flavonol Constituents, Kaempferol and Quercetin, on Serum Uric Acid Levels, Biomarkers of Oxidative Stress and Liver Xanthine Oxidoreductase Aactivity inOxonate-Induced Hyperuricemic Rats.

PubMed

Haidari, Fatemeh; Keshavarz, Seid Ali; Mohammad Shahi, Majid; Mahboob, Soltan-Ali; Rashidi, Mohammad-Reza

2011-01-01

Increased serum uric acid is known to be a major risk related to the development of several oxidative stress diseases. The aim of this study was to investigate the effect of parsley, quercetin and kaempferol on serum uric acid levels, liver xanthine oxidoreductase activity and two non-invasive biomarkers of oxidative stress (total antioxidant capacity and malondialdehyde concentration) in normal and oxonate-induced hyperuricemic rats. A total of 60 male Wistar rats were randomly divided into ten equal groups; including 5 normal groups (vehicle, parsley, quercetin, kaempferol and allopurinol) and 5 hyperuricemic groups (vehicle, parsley, quercetin, kaempferol and allopurinol). Parsley (5 g/Kg), quercetin (5 mg/Kg), kaempferol (5 mg/Kg) and allopurinol (5 mg/Kg) were administrated to the corresponding groups by oral gavage once a day for 2 weeks. The results showed that parsley and its flavonol did not cause any significant reduction in the serum uric acid levels in normal rats, but significantly reduced the serum uric acid levels of hyperuricemic rats in a time-dependent manner. All treatments significantly inhibited liver xanthine oxidoreductase activity. Parsley, kaempferol and quercetin treatment led also to a significant increase in total antioxidant capacity and decrease in malondialdehyde concentration in hyperuricemic rats. Although the hypouricemic effect of allopurinol was much higher than that of parsley and its flavonol constituents, it could not significantly change oxidative stress biomarkers. These features of parsley and its flavonols make them as a possible alternative for allopurinol, or at least in combination therapy to minimize the side effects of allopurinol to treat hyperuricemia and oxidative stress diseases.

Effects of Parsley (Petroselinum crispum) and its Flavonol Constituents, Kaempferol and Quercetin, on Serum Uric Acid Levels, Biomarkers of Oxidative Stress and Liver Xanthine Oxidoreductase Aactivity inOxonate-Induced Hyperuricemic Rats

PubMed Central

Haidari, Fatemeh; Keshavarz, Seid Ali; Mohammad Shahi, Majid; Mahboob, Soltan-Ali; Rashidi, Mohammad-Reza

2011-01-01

Increased serum uric acid is known to be a major risk related to the development of several oxidative stress diseases. The aim of this study was to investigate the effect of parsley, quercetin and kaempferol on serum uric acid levels, liver xanthine oxidoreductase activity and two non-invasive biomarkers of oxidative stress (total antioxidant capacity and malondialdehyde concentration) in normal and oxonate-induced hyperuricemic rats. A total of 60 male Wistar rats were randomly divided into ten equal groups; including 5 normal groups (vehicle, parsley, quercetin, kaempferol and allopurinol) and 5 hyperuricemic groups (vehicle, parsley, quercetin, kaempferol and allopurinol). Parsley (5 g/Kg), quercetin (5 mg/Kg), kaempferol (5 mg/Kg) and allopurinol (5 mg/Kg) were administrated to the corresponding groups by oral gavage once a day for 2 weeks. The results showed that parsley and its flavonol did not cause any significant reduction in the serum uric acid levels in normal rats, but significantly reduced the serum uric acid levels of hyperuricemic rats in a time-dependent manner. All treatments significantly inhibited liver xanthine oxidoreductase activity. Parsley, kaempferol and quercetin treatment led also to a significant increase in total antioxidant capacity and decrease in malondialdehyde concentration in hyperuricemic rats. Although the hypouricemic effect of allopurinol was much higher than that of parsley and its flavonol constituents, it could not significantly change oxidative stress biomarkers. These features of parsley and its flavonols make them as a possible alternative for allopurinol, or at least in combination therapy to minimize the side effects of allopurinol to treat hyperuricemia and oxidative stress diseases. PMID:24250417

FLOURY ENDOSPERM7 encodes a regulator of starch synthesis and amyloplast development essential for peripheral endosperm development in rice

PubMed Central

Zhang, Long; Ren, Yulong; Lu, Bingyue; Yang, Chunyan; Feng, Zhiming; Liu, Zhou; Chen, Jun; Ma, Weiwei; Wang, Ying; Yu, Xiaowen; Wang, Yunlong; Zhang, Wenwei; Wang, Yihua; Liu, Shijia; Wu, Fuqing; Zhang, Xin; Guo, Xiuping; Bao, Yiqun; Jiang, Ling; Wan, Jianmin

2016-01-01

In cereal crops, starch synthesis and storage depend mainly on a specialized class of plastids, termed amyloplasts. Despite the importance of starch, the molecular machinery regulating starch synthesis and amyloplast development remains largely unknown. Here, we report the characterization of the rice (Oryza sativa) floury endosperm7 (flo7) mutant, which develops a floury-white endosperm only in the periphery and not in the inner portion. Consistent with the phenotypic alternation in flo7 endosperm, the flo7 mutant had reduced amylose content and seriously disrupted amylopectin structure only in the peripheral endosperm. Notably, flo7 peripheral endosperm cells showed obvious defects in compound starch grain development. Map-based cloning of FLO7 revealed that it encodes a protein of unknown function. FLO7 harbors an N-terminal transit peptide capable of targeting functional FLO7 fused to green fluorescent protein to amyloplast stroma in developing endosperm cells, and a domain of unknown function 1338 (DUF1338) that is highly conserved in green plants. Furthermore, our combined β-glucuronidase activity and RNA in situ hybridization assays showed that the FLO7 gene was expressed ubiquitously but exhibited a specific expression in the endosperm periphery. Moreover, a set of in vivo experiments demonstrated that the missing 32 aa in the flo7 mutant protein are essential for the stable accumulation of FLO7 in the endosperm. Together, our findings identify FLO7 as a unique plant regulator required for starch synthesis and amyloplast development within the peripheral endosperm and provide new insights into the spatial regulation of endosperm development in rice. PMID:26608643

Infrared spectroscopy of flavones and flavonols. Reexamination of the hydroxyl and carbonyl vibrations in relation to the interactions of flavonoids with membrane lipids

NASA Astrophysics Data System (ADS)

Baranović, Goran; Šegota, Suzana

2018-03-01

Detailed vibrational assignments for twelve flavonoids (seven flavones (flavone, 3- and 5-hydroxyflavone, chrysin, apigenin, fisetin and luteolin) and five flavonols (galangin, kaempferol, quercetin, morin and myricetin)) have been made based on own and reported experimental data and calculations at the B3LYP/6-31 + G(d,p) level of theory. All the molecules are treated in a uniform way by using the same set of redundancy-free set of internal coordinates. A generalized harmonic mode mixing is used to corroborate the vibrational characteristics of this important class of molecules. Each flavonoid molecule can be treated from the vibrational point of view as made of relatively weakly coupled chromone and phenyl part. It has been shown that the strongest band around 1600 cm- 1 need not be attributable to the Cdbnd O stretching. The way the vibrations of any of the hydroxyl groups are mixed with ring vibrations and vibrations of other neighboring hydroxyl groups is rather involved. This imposes severe limitations on any attempt to describe normal modes of a flavonol in terms of hydroxyl or carbonyl group vibrations. The role of water molecules in the appearance of flavonoid IR spectra is emphasized. Knowing for the great affinity of phosphate groups in lipids towards water, the immediate consequence is a reasonable assumption that flavonoid lipid interactions is mediated by water.

Regulation of aortic extracellular matrix synthesis via noradrenergic system and angiotensin II in juvenile rats.

PubMed

Dab, Houcine; Hachani, Rafik; Dhaouadi, Nedra; Sakly, Mohsen; Hodroj, Wassim; Randon, Jacques; Bricca, Giampiero; Kacem, Kamel

2012-10-01

Extracellular matrix (ECM) synthesis regulation by sympathetic nervous system (SNS) or angiotensin II (ANG II) was widely reported, but interaction between the two systems on ECM synthesis needs further investigation. We tested implication of SNS and ANG II on ECM synthesis in juvenile rat aorta. Sympathectomy with guanethidine (50 mg/kg, subcutaneous) and blockade of the ANG II AT1 receptors (AT1R) blocker with losartan (20 mg/kg/day in drinking water) were performed alone or in combination in rats. mRNA and protein synthesis of collagen and elastin were examined by Q-RT-PCR and immunoblotting. Collagen type I and III mRNA were increased respectively by 62 and 43% after sympathectomy and decreased respectively by 31 and 60% after AT1R blockade. Combined treatment increased collagen type III by 36% but not collagen type I. The same tendency of collagen expression was observed at mRNA and protein levels after the three treatments. mRNA and protein level of elastin was decreased respectively by 63 and 39% and increased by 158 and 15% after losartan treatment. Combined treatment abrogates changes induced by single treatments. The two systems act as antagonists on ECM expression in the aorta and combined inhibition of the two systems prevents imbalance of mRNA and protein level of collagen I and elastin induced by single treatment. Combined inhibition of the two systems prevents deposit or excessive reduction of ECM and can more prevent cardiovascular disorders.

Matrix Rigidity Regulates Cancer Cell Growth by Modulating Cellular Metabolism and Protein Synthesis

PubMed Central

Tilghman, Robert W.; Blais, Edik M.; Cowan, Catharine R.; Sherman, Nicholas E.; Grigera, Pablo R.; Jeffery, Erin D.; Fox, Jay W.; Blackman, Brett R.; Tschumperlin, Daniel J.; Papin, Jason A.; Parsons, J. Thomas

2012-01-01

Background Tumor cells in vivo encounter diverse types of microenvironments both at the site of the primary tumor and at sites of distant metastases. Understanding how the various mechanical properties of these microenvironments affect the biology of tumor cells during disease progression is critical in identifying molecular targets for cancer therapy. Methodology/Principal Findings This study uses flexible polyacrylamide gels as substrates for cell growth in conjunction with a novel proteomic approach to identify the properties of rigidity-dependent cancer cell lines that contribute to their differential growth on soft and rigid substrates. Compared to cells growing on more rigid/stiff substrates (>10,000 Pa), cells on soft substrates (150–300 Pa) exhibited a longer cell cycle, due predominantly to an extension of the G1 phase of the cell cycle, and were metabolically less active, showing decreased levels of intracellular ATP and a marked reduction in protein synthesis. Using stable isotope labeling of amino acids in culture (SILAC) and mass spectrometry, we measured the rates of protein synthesis of over 1200 cellular proteins under growth conditions on soft and rigid/stiff substrates. We identified cellular proteins whose syntheses were either preferentially inhibited or preserved on soft matrices. The former category included proteins that regulate cytoskeletal structures (e.g., tubulins) and glycolysis (e.g., phosphofructokinase-1), whereas the latter category included proteins that regulate key metabolic pathways required for survival, e.g., nicotinamide phosphoribosyltransferase, a regulator of the NAD salvage pathway. Conclusions/Significance The cellular properties of rigidity-dependent cancer cells growing on soft matrices are reminiscent of the properties of dormant cancer cells, e.g., slow growth rate and reduced metabolism. We suggest that the use of relatively soft gels as cell culture substrates would allow molecular pathways to be studied under

Hydrocortisone and triiodothyronine regulate hyaluronate synthesis in a tissue-engineered human dermal equivalent through independent pathways.

PubMed

Deshpande, Madhura; Papp, Suzanne; Schaffer, Lana; Pouyani, Tara

2015-02-01

Hydrocortisone (HC) and triiodothyronine (T3) have both been shown to be capable of independently inhibiting hyaluronate (HA, hyaluronic acid) synthesis in a self-assembled human dermal equivalent (human dermal matrix). We sought to investigate the action of these two hormones in concert on extracellular matrix formation and HA inhibition in the tissue engineered human dermal matrix. To this end, neonatal human dermal fibroblasts were cultured in defined serum-free medium for 21 days in the presence of each hormone alone, or in combination, in varying concentrations. Through a process of self-assembly, a substantial dermal extracellular matrix formed that was characterized. The results of these studies demonstrate that combinations of the hormones T3 and hydrocortisone showed significantly higher levels of hyaluronate inhibition as compared to each hormone alone in the human dermal matrix. In order to gain preliminary insight into the genes regulating HA synthesis in this system, a differential gene array analysis was conducted in which the construct prepared in the presence of 200 μg/mL HC and 0.2 nM T3 was compared to the normal construct (0.4 μg/mL HC and 20 pM T3). Using a GLYCOv4 gene chip containing approximately 1260 human genes, we observed differential expression of 131 genes. These data suggest that when these two hormones are used in concert a different mechanism of inhibition prevails and a combination of degradation and inhibition of HA synthesis may be responsible for HA regulation in the human dermal matrix. Copyright © 2014 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

A novel MYB Transcription Factor regulates AsA synthesis and effects cold tolerance.

PubMed

Xing, Caihua; Liu, Yue; Zhao, Liangyi; Zhang, Shaoling; Huang, Xiaosan

2018-06-21

Dehydroascorbate reductase (DHAR) plays an important role in stress responses, but the transcriptional regulation of DHAR in response to abiotic stress is still poorly understood. In this study, we isolated a novel R2R3-type MYB transcription factor from Pyrus betulaefolia by yeast one-hybrid screening, designated as PbrMYB5. PbrMYB5 was localized in the nucleus and could bind specifically to the promoter of PbrDHAR2. PbrMYB5 was greatly induced by cold and salt, but slightly by dehydration. Overexpression of PbrMYB5 in tobacco conferred enhanced tolerance to chilling stresses, whereas down-regulation of PbrMYB5 in Pyrus betulaefolia by virus-induced gene silencing (VIGS) resulted in elevated chilling sensitivity. Transgenic tobacco exhibited higher expression levels of DHAR2 and accumulated larger amount of AsA than the WT plants. VIGS of PbrMYB5 in Pyrus betulaefolia down-regulated PbrDHAR2 abundance and decreased AsA level, accompanied by an increased sensitivity to the chilling stress. Taken together, these results demonstrated that PbrMYB5 is an activator of AsA biosynthesis and may play a positive role in chilling tolerance, at least in part, due to the modulation of AsA synthesis by regulating the PbrDHAR2 expression. This article is protected by copyright. All rights reserved.

Regulation of yeast DNA polymerase δ-mediated strand displacement synthesis by 5′-flaps

PubMed Central

Koc, Katrina N.; Stodola, Joseph L.; Burgers, Peter M.; Galletto, Roberto

2015-01-01

The strand displacement activity of DNA polymerase δ is strongly stimulated by its interaction with proliferating cell nuclear antigen (PCNA). However, inactivation of the 3′–5′ exonuclease activity is sufficient to allow the polymerase to carry out strand displacement even in the absence of PCNA. We have examined in vitro the basic biochemical properties that allow Pol δ-exo− to carry out strand displacement synthesis and discovered that it is regulated by the 5′-flaps in the DNA strand to be displaced. Under conditions where Pol δ carries out strand displacement synthesis, the presence of long 5′-flaps or addition in trans of ssDNA suppress this activity. This suggests the presence of a secondary DNA binding site on the enzyme that is responsible for modulation of strand displacement activity. The inhibitory effect of a long 5′-flap can be suppressed by its interaction with single-stranded DNA binding proteins. However, this relief of flap-inhibition does not simply originate from binding of Replication Protein A to the flap and sequestering it. Interaction of Pol δ with PCNA eliminates flap-mediated inhibition of strand displacement synthesis by masking the secondary DNA site on the polymerase. These data suggest that in addition to enhancing the processivity of the polymerase PCNA is an allosteric modulator of other Pol δ activities. PMID:25813050

Hydrolyzable Tannins, Flavonol Glycosides, and Phenolic Acids Show Seasonal and Ontogenic Variation in Geranium sylvaticum.

PubMed

Tuominen, Anu; Salminen, Juha-Pekka

2017-08-09

The seasonal variation of polyphenols in the aboveground organs and roots of Geranium sylvaticum in four populations was studied using UPLC-DAD-ESI-QqQ-MS/MS. The content of the main compound, geraniin, was highest (16% of dry weight) in the basal leaves after the flowering period but stayed rather constant throughout the growing season. Compound-specific mass spectrometric methods revealed the different seasonal patterns in minor polyphenols. Maximum contents of galloylglucoses and flavonol glycosides were detected in the small leaves in May, whereas the contents of further modified ellagitannins, such as ascorgeraniin and chebulagic acid, increased during the growing season. In flower organs, the polyphenol contents differed significantly between ontogenic phases so that maximum amounts were typically found in the bud phase, except in pistils the amount of gallotannins increased significantly in the fruit phase. These results can be used in evaluating the role of polyphenols in plant-herbivore interactions or in planning the best collection times of G. sylvaticum for compound isolation purposes.

Poly-L-glutamate/glutamine synthesis in the cell wall of Mycobacterium bovis is regulated in response to nitrogen availability

PubMed Central

2013-01-01

Background The cell wall of pathogenic mycobacteria is known to possess poly-L-glutamine (PLG) layer. PLG synthesis has been directly linked to glutamine synthetase (GS) enzyme. glnA1 gene encodes for GS enzyme in mycobacteria. PLG layer is absent in cell wall of avirulent Mycobacterium smegmatis, although M. smegmatis strain expressing GS enzyme of pathogenic mycobacteria can synthesize PLG layer in the cell wall. The role of GS enzyme has been extensively studied in Mycobacterium tuberculosis, however, little is known about GS enzyme in other mycobacterial species. Mycobacterium bovis, as an intracellular pathogen encounters nitrogen stress inside macrophages, thus it has developed nitrogen assimilatory pathways to survive in adverse conditions. We have investigated the expression and activity of M. bovis GS in response to nitrogen availability and effect on synthesis of PLG layer in the cell wall. M. smegmatis was used as a model to study the behaviour of glnA1 locus of M. bovis. Results We observed that GS expression and activity decreased significantly in high nitrogen grown conditions. In high nitrogen conditions, the amount of PLG in cell wall was drastically reduced (below detectable limits) as compared to low nitrogen condition in M. bovis and in M. smegmatis strain complemented with M. bovis glnA1. Additionally, biofilm formation by M. smegmatis strain complemented with M. bovis glnA1 was increased than the wild type M. smegmatis strain. Conclusions The physiological regulation of GS in M. bovis was found to be similar to that reported in other mycobacteria but this data revealed that PLG synthesis in the cell wall of pathogenic mycobacteria occurs only in nitrogen limiting conditions and on the contrary high nitrogen conditions inhibit PLG synthesis. This indicates that PLG synthesis may be a form of nitrogen assimilatory pathway during ammonium starvation in virulent mycobacteria. Also, we have found that M. smegmatis complemented with M. bovis glnA1

Parkin regulates translesion DNA synthesis in response to UV radiation.

PubMed

Zhu, Xuefei; Ma, Xiaolu; Tu, Yingfeng; Huang, Min; Liu, Hongmei; Wang, Fengli; Gong, Juanjuan; Wang, Jiuqiang; Li, Xiaoling; Chen, Qian; Shen, Hongyan; Zhu, Shu; Wang, Yun; Liu, Yang; Guo, Caixia; Tang, Tie-Shan

2017-05-30

Deficiency of Parkin is a major cause of early-onset Parkinson's disease (PD). Notably, PD patients also exhibit a significantly higher risk in melanoma and other skin tumors, while the mechanism remains largely unknown. In this study, we show that depletion of Parkin causes compromised cell viability and genome stability after ultraviolet (UV) radiation. We demonstrate that Parkin promotes efficient Rad18-dependent proliferating cell nuclear antigen (PCNA) monoubiquitination by facilitating the formation of Replication protein A (RPA)-coated ssDNA upon UV radiation. Furthermore, Parkin is found to physically interact with NBS1 (Nijmegen breakage syndrome 1), and to be required for optimal recruitment of NBS1 and DNA polymerase eta (Polη) to UV-induced damage sites. Consequently, depletion of Parkin leads to increased UV-induced mutagenesis. These findings unveil an important role of Parkin in protecting genome stability through positively regulating translesion DNA synthesis (TLS) upon UV damage, providing a novel mechanistic link between Parkin deficiency and predisposition to skin cancers in PD patients.

Increasing Malonyl-CoA Derived Product through Controlling the Transcription Regulators of Phospholipid Synthesis in Saccharomyces cerevisiae.

PubMed

Chen, Xiaoxu; Yang, Xiaoyu; Shen, Yu; Hou, Jin; Bao, Xiaoming

2017-05-19

Malonyl-CoA is a precursor of a variety of compounds such as polyketides and flavonoids. In Saccharomyces cerevisiae, malonyl-CoA concentration is tightly regulated and therefore maintained at a very low level, limiting the production of malonyl-CoA-derived chemicals. Here we manipulated the phospholipid synthesis transcriptional regulators to control the malonyl-CoA levels and increase the downstream product. Through manipulating different regulators including Ino2p, Ino4p, Opi1p, and a series of synthetic Ino2p variants, combining with studying the inositol and choline effect, the engineered strain achieved a 9-fold increase of the titer of malonyl-CoA-derived product 3-hydroxypropionic acid, which is among the highest improvement relative to previously reported strategies. Our study provides a new strategy to regulate malonyl-CoA availability and will contribute to the production of other highly valued malonyl-CoA-derived chemicals.

Role for tryptophan in regulation of protein synthesis in porcine muscle

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

Lin, F.D.; Smith, T.K.; Bayley, H.S.

1988-04-01

Experiments were conducted to determine the effect of varying concentrations of dietary tryptophan on growth rate and protein synthesis in edible muscle tissues of growing swine. A total of 45 immature swine (initial weight approximately 24 kg) were fed corn-gelatin diets containing 0.5 (n = 8), 0.8 (n = 10), 1.3 (n = 10), 1.5 (n = 7) or 2.0 (n = 10) g tryptophan/kg diet for 35 d. Animals fed 0.5 and 0.8 g tryptophan/kg grew more slowly, consumed less feed and had a lower efficiency of feed utilization than animals fed higher concentrations of tryptophan. Thirty similar animalsmore » were used in a second experiment. Diets containing 0.5, 0.8, 1.0, 1.5 or 2.0 g tryptophan/kg diet (n = 6) were fed for 14 d, after which all animals were killed and samples were taken of longissimus dorsi, triceps brachii and biceps femoris. Protein synthetic activity was determined by monitoring the incorporation of (/sup 14/C)phenylalanine into protein in vitro. There was no significant difference in synthetic activity between different muscle types. There was no effect of diet on the activity of the muscle soluble protein fraction. The activity of the muscle ribosomal fraction, however, was positively correlated with increasing concentrations of dietary tryptophan. It was concluded that tryptophan has the potential to regulate muscle protein synthesis in a manner beyond serving simply as a component of protein.« less

Chromatin-associated regulation of sorbitol synthesis in flower buds of peach.

PubMed

Lloret, Alba; Martínez-Fuentes, Amparo; Agustí, Manuel; Badenes, María Luisa; Ríos, Gabino

2017-11-01

PpeS6PDH gene is postulated to mediate sorbitol synthesis in flower buds of peach concomitantly with specific chromatin modifications. Perennial plants have evolved an adaptive mechanism involving protection of meristems within specialized structures named buds in order to survive low temperatures and water deprivation during winter. A seasonal period of dormancy further improves tolerance of buds to environmental stresses through specific mechanisms poorly known at the molecular level. We have shown that peach PpeS6PDH gene is down-regulated in flower buds after dormancy release, concomitantly with changes in the methylation level at specific lysine residues of histone H3 (H3K27 and H3K4) in the chromatin around the translation start site of the gene. PpeS6PDH encodes a NADPH-dependent sorbitol-6-phosphate dehydrogenase, the key enzyme for biosynthesis of sorbitol. Consistently, sorbitol accumulates in dormant buds showing higher PpeS6PDH expression. Moreover, PpeS6PDH gene expression is affected by cold and water deficit stress. Particularly, its expression is up-regulated by low temperature in buds and leaves, whereas desiccation treatment induces PpeS6PDH in buds and represses the gene in leaves. These data reveal the concurrent participation of chromatin modification mechanisms, transcriptional regulation of PpeS6PDH and sorbitol accumulation in flower buds of peach. In addition to its role as a major translocatable photosynthate in Rosaceae species, sorbitol is a widespread compatible solute and cryoprotectant, which suggests its participation in tolerance to environmental stresses in flower buds of peach.

Multi-Drug Resistance Transporter 2 Regulates Mucosal Inflammation by Facilitating the Synthesis of Hepoxilin A3

PubMed Central

Pazos, Michael; Siccardi, Dario; Mumy, Karen L.; Bien, Jeffrey D.; Louie, Steve; Shi, Hai Ning; Gronert, Karsten; Mrsny, Randall J.; McCormick, Beth A.

2008-01-01

Neutrophil transmigration across mucosal surfaces contributes to dysfunction of epithelial barrier properties, a characteristic underlying many mucosal inflammatory diseases. Thus, insight into the directional movement of neutrophils across epithelial barriers will provide important information relating to the mechanisms of such inflammatory disorders. The eicosanoid hepoxilin A3, an endogenous product of 12-lipoxygenase activity, is secreted from the apical surface of the epithelial barrier and establishes a chemotatic gradient to guide neutrophils from the submucosa, across epithelia to the luminal site of an inflammatory stimulus - the final step in neutrophil recruitment. Currently, little is known regarding how hepoxilin A3 is secreted from the intestinal epithelium during an inflammatory insult. In this study we reveal that hepoxilin A3 is a substrate for the apical efflux ABC transporter, multi-drug resistance protein 2 (MRP2). Moreover, using multiple in vitro and in vivo models we show that induction of intestinal inflammation profoundly up-regulates apical expression of MRP2, and that interfering with hepoxilin A3 synthesis and/or inhibition of MRP2 function results in a marked reduction in inflammation and severity of disease. Lastly, examination of inflamed intestinal epithelia in human biopsies revealed up-regulation of MRP2. Thus, blocking hepoxilin A3 synthesis and/or inhibiting MRP2 may lead to the development of new therapeutic strategies for the treatment of epithelial-associated inflammatory conditions. PMID:19017997

Quantitative Analysis and Comparison of Four Major Flavonol Glycosides in the Leaves of Toona sinensis (A. Juss.) Roemer (Chinese Toon) from Various Origins by High-Performance Liquid Chromatography-Diode Array Detector and Hierarchical Clustering Analysis

PubMed Central

Sun, Xiaoxiang; Zhang, Liting; Cao, Yaqi; Gu, Qinying; Yang, Huan; Tam, James P.

2016-01-01

Background: Toona sinensis (A. Juss.) Roemer is an endemic species of Toona genus native to Asian area. Its dried leaves are applied in the treatment of many diseases; however, few investigations have been reported for the quantitative analysis and comparison of major bioactive flavonol glycosides in the leaves harvested from various origins. Objective: To quantitatively analyze four major flavonol glycosides including rutinoside, quercetin-3-O-β-D-glucoside, quercetin-3-O-α-L-rhamnoside, and kaempferol-3-O-α-L-rhamnoside in the leaves from different production sites and classify them according to the content of these glycosides. Materials and Methods: A high-performance liquid chromatography-diode array detector (HPLC-DAD) method for their simultaneous determination was developed and validated for linearity, precision, accuracy, stability, and repeatability. Moreover, the method established was then employed to explore the difference in the content of these four glycosides in raw materials. Finally, a hierarchical clustering analysis was performed to classify 11 voucher specimens. Results: The separation was performed on a Waters XBridge Shield RP18 column (150 mm × 4.6 mm, 3.5 μm) kept at 35°C, and acetonitrile and H2O containing 0.30% trifluoroacetic acid as mobile phase was driven at 1.0 mL/min during the analysis. Ten microliters of solution were injected and 254 nm was selected to monitor the separation. A strong linear relationship between the peak area and concentration of four analytes was observed. And, the method was also validated to be repeatable, stable, precise, and accurate. Conclusion: An efficient and reliable HPLC-DAD method was established and applied in the assays for the samples from 11 origins successfully. Moreover, the content of those flavonol glycosides varied much among different batches, and the flavonoids could be considered as biomarkers to control the quality of Chinese Toon. SUMMARY Four major flavonol glycosides in the leaves

Urea Synthesis and Excretion in Aedes aegypti Mosquitoes Are Regulated by a Unique Cross-Talk Mechanism

PubMed Central

Isoe, Jun; Scaraffia, Patricia Y.

2013-01-01

Aedes aegypti mosquitoes do not have a typical functional urea cycle for ammonia disposal such as the one present in most terrestrial vertebrates. However, they can synthesize urea by two different pathways, argininolysis and uricolysis. We investigated how formation of urea by these two pathways is regulated in females of A. aegypti. The expression of arginase (AR) and urate oxidase (UO), either separately or simultaneously (ARUO) was silenced by RNAi. The amounts of several nitrogen compounds were quantified in excreta using mass spectrometry. Injection of mosquitoes with either dsRNA-AR or dsRNA-UO significantly decreased the expressions of AR or UO in the fat body (FB) and Malpighian tubules (MT). Surprisingly, the expression level of AR was increased when UO was silenced and vice versa, suggesting a cross-talk regulation between pathways. In agreement with these data, the amount of urea measured 48 h after blood feeding remained unchanged in those mosquitoes injected with dsRNA-AR or dsRNA-UO. However, allantoin significantly increased in the excreta of dsRNA-AR-injected females. The knockdown of ARUO mainly led to a decrease in urea and allantoin excretion, and an increase in arginine excretion. In addition, dsRNA-AR-injected mosquitoes treated with a specific nitric oxide synthase inhibitor showed an increase of UO expression in FB and MT and a significant increase in the excretion of nitrogen compounds. Interestingly, both a temporary delay in the digestion of a blood meal and a significant reduction in the expression of several genes involved in ammonia metabolism were observed in dsRNA-AR, UO or ARUO-injected females. These results reveal that urea synthesis and excretion in A. aegypti are tightly regulated by a unique cross-talk signaling mechanism. This process allows blood-fed mosquitoes to regulate the synthesis and/or excretion of nitrogen waste products, and avoid toxic effects that could result from a lethal concentration of ammonia in their

Comparative Analysis of Two Flavonol Synthases from Different-Colored Onions Provides Insight into Flavonoid Biosynthesis.

PubMed

Park, Sangkyu; Kim, Da-Hye; Lee, Jong-Yeol; Ha, Sun-Hwa; Lim, Sun-Hyung

2017-07-05

We isolated cDNAs encoding flavonol synthase (FLS) from the red onion "H6" (AcFLS-H6) and the yellow onion "Hwangryongball" (AcFLS-HRB). We found three amino acid variations between the two sequences. Kinetic analysis with recombinant proteins revealed that AcFLS-HRB exhibited approximately 2-fold higher catalytic efficiencies than AcFLS-H6 for dihydroflavonol substrates and that both proteins preferred dihydroquercetin to dihydrokaempferol. The expression patterns of flavonoid biosynthesis genes corresponded to the accumulation patterns of flavonoid aglycones in both onions. Whereas the other flavonoid biosynthesis genes were weakly expressed in the HRB sheath compared to that of H6, the expression of FLS was similar in both onions. This relatively enhanced FLS expression, along with the higher activity of AcFLS-HRB, could increase the quercetin production in the HRB sheath. The quercetin content was approximately 12-fold higher than the cyanidin content in the H6 sheath, suggesting that FLS has priority in the competition between FLS and dihydroflavonol 4-reductase (DFR) for their substrate dihydroquercetin.

Role of Flavonols and Proanthocyanidins in the Sensory Quality of Sea Buckthorn (Hippophaë rhamnoides L.) Berries.

PubMed

Ma, Xueying; Yang, Wei; Laaksonen, Oskar; Nylander, Merja; Kallio, Heikki; Yang, Baoru

2017-11-15

Sensory profile, flavonols, proanthocyanidins, sugars, and organic acids were investigated in purees of six sea buckthorn (Hippophaë rhamnoides) cultivars. The sensory profiles of the purees were dominated by intense sourness followed by astringency and bitterness due to the high content of malic acid. Malic acid and isorhamnetin glycosides, especially isorhamnetin-3-O-sophoroside-7-O-rhamnoside, had close association with the astringent attributes in the different purees, whereas some of the known astringent compounds such as proanthocyanidin dimers and trimers or quercetin glycosides, had less impact. Moreover, the ratios between contents of acids and phenolic compounds were more important predictors of bitterness than the individual variables alone. Astringency and bitterness are important sensory factors for the consumer acceptance of sea buckthorn products. The current study provides new knowledge on the correlations between sensory properties and composition and supports industrial utilization of the sea buckthorn berries.

Chronological protein synthesis in regenerating rat liver.

PubMed

He, Jinjun; Hao, Shuai; Zhang, Hao; Guo, Fuzheng; Huang, Lingyun; Xiao, Xueyuan; He, Dacheng

2015-07-01

Liver regeneration has been studied for decades; however, its regulation remains unclear. In this study, we report a dynamic tracing of protein synthesis in rat regenerating liver with a new proteomic technique, (35) S in vivo labeling analysis for dynamic proteomics (SiLAD). Conventional proteomic techniques typically measure protein alteration in accumulated amounts. The SiLAD technique specifically detects protein synthesis velocity instead of accumulated amounts of protein through (35) S pulse labeling of newly synthesized proteins, providing a direct way for analyzing protein synthesis variations. Consequently, protein synthesis within short as 30 min was visualized and protein regulations in the first 8 h of regenerating liver were dynamically traced. Further, the 3.5-5 h post partial hepatectomy (PHx) was shown to be an important regulatory turning point by acute regulation of many proteins in the initiation of liver regeneration. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Discrimination and Measurements of Three Flavonols with Similar Structure Using Terahertz Spectroscopy and Chemometrics

NASA Astrophysics Data System (ADS)

Yan, Ling; Liu, Changhong; Qu, Hao; Liu, Wei; Zhang, Yan; Yang, Jianbo; Zheng, Lei

2018-03-01

Terahertz (THz) technique, a recently developed spectral method, has been researched and used for the rapid discrimination and measurements of food compositions due to its low-energy and non-ionizing characteristics. In this study, THz spectroscopy combined with chemometrics has been utilized for qualitative and quantitative analysis of myricetin, quercetin, and kaempferol with concentrations of 0.025, 0.05, and 0.1 mg/mL. The qualitative discrimination was achieved by KNN, ELM, and RF models with the spectra pre-treatments. An excellent discrimination (100% CCR in the prediction set) could be achieved using the RF model. Furthermore, the quantitative analyses were performed by partial least square regression (PLSR) and least squares support vector machine (LS-SVM). Comparing to the PLSR models, the LS-SVM yielded better results with low RMSEP (0.0044, 0.0039, and 0.0048), higher Rp (0.9601, 0.9688, and 0.9359), and higher RPD (8.6272, 9.6333, and 7.9083) for myricetin, quercetin, and kaempferol, respectively. Our results demonstrate that THz spectroscopy technique is a powerful tool for identification of three flavonols with similar chemical structures and quantitative determination of their concentrations.

A new flavonol glycoside and other flavonoids from the aerial parts of Taverniera aegyptiaca.

PubMed

Hassan, Ahmed R; Amer, Khadiga F; El-Toumy, Sayed A; Nielsen, John; Christensen, Søren Brøgger

2018-04-17

Isolation of flavonoids from the aerial parts of Taverniera aegyptiaca Bioss. (Fabaceae) led to identification of one new flavonol glycoside, isorhamnetin-3-O-α-l-rhamnopyranosyl-(1→2)-α-l-arabinopyranoside (1), along with eleven compounds, which previously have not been isolated from this plant quercetin-3-O-α-l-rhamnopyranosyl-(1→2)-[α-l-rhamnopyranosyl-(1→6)-β-d-galactopyranoside] (2), isorhamnetin-3-O-α-l-arabinopyranoside (3), quercetin-3-O-α-l-rhamnopyranosyl-(1→6)-β-d-glucopyranoside (4), isorhamnetin-3-O-α-l-rhamnopyranosyl-(1→6)-β-d-glucopyranoside (7), isorhamnetin 3-O-α-l-rhamnopyranosyl-(1→2)-[α-l-rhamnopyranosyl-(1→6)-β-d-galactopyranoside] (8), isorhamnetin 3-O-α-l-rhamnopyranosyl-(1→2)-[α-l-rhamnopyranosyl-(1→6)-β-d-glucopyranoside] (9), kaempferol 3-O-α-l-rhamnopyranosyl-(1→2)-[α-l-rhamnopyranosyl-(1→6)-β-d-galactopyranoside] (10), isorhamnetin (11), 4,4'-dihydroxy-2'-methoxychalcone (12), formononetin (13) and calycosin (15)] and some compounds already known from this plant [quercetin-3-O-robinobioside (5), isorhamnetin-3-O-robinobioside (6), afrormosin (14) and odoratin (16)].

Inositol synthesis regulates activation of GSK-3α in neuronal cells

PubMed Central

Ye, Cunqi; Greenberg, Miriam L.

2015-01-01

The synthesis of inositol provides precursors of inositol lipids and inositol phosphates that are pivotal for cell signaling. Mood-stabilizers lithium and valproic acid (VPA), used for treating bipolar disorder, cause cellular inositol depletion, which has been proposed as a therapeutic mechanism of action of both drugs. Despite the importance of inositol, the requirement for inositol synthesis in neuronal cells is not well understood. Here, we examined inositol effects on proliferation of SK-N-SH neuroblastoma cells. The essential role of inositol synthesis in proliferation is underscored by the findings that exogenous inositol was dispensable for proliferation, and inhibition of inositol synthesis decreased proliferation. Interestingly, the inhibition of inositol synthesis by knocking down INO1, which encodes inositol-3-phosphate synthase, the rate-limiting enzyme of inositol synthesis, led to inactivation of GSK-3α by increasing the inhibitory phosphorylation of this kinase. Similarly, the mood-stabilizer VPA effected transient decreases in intracellular inositol, leading to inactivation of GSK-3α. As GSK-3 inhibition has been proposed as a likely therapeutic mechanism of action, the finding that inhibition of inositol synthesis results in inactivation of GSK-3α suggests a unifying hypothesis for mechanism of mood-stabilizing drugs. PMID:25345501

Gene Expression and Metabolite Profiling of Developing Highbush Blueberry Fruit Indicates Transcriptional Regulation of Flavonoid Metabolism and Activation of Abscisic Acid Metabolism1[W][OA

PubMed Central

Zifkin, Michael; Jin, Alena; Ozga, Jocelyn A.; Zaharia, L. Irina; Schernthaner, Johann P.; Gesell, Andreas; Abrams, Suzanne R.; Kennedy, James A.; Constabel, C. Peter

2012-01-01

Highbush blueberry (Vaccinium corymbosum) fruits contain substantial quantities of flavonoids, which are implicated in a wide range of health benefits. Although the flavonoid constituents of ripe blueberries are known, the molecular genetics underlying their biosynthesis, localization, and changes that occur during development have not been investigated. Two expressed sequence tag libraries from ripening blueberry fruit were constructed as a resource for gene identification and quantitative real-time reverse transcription-polymerase chain reaction primer design. Gene expression profiling by quantitative real-time reverse transcription-polymerase chain reaction showed that flavonoid biosynthetic transcript abundance followed a tightly regulated biphasic pattern, and transcript profiles were consistent with the abundance of the three major classes of flavonoids. Proanthocyanidins (PAs) and corresponding biosynthetic transcripts encoding anthocyanidin reductase and leucoanthocyanidin reductase were most concentrated in young fruit and localized predominantly to the inner fruit tissue containing the seeds and placentae. Mean PA polymer length was seven to 8.5 subunits, linked predominantly via B-type linkages, and was relatively constant throughout development. Flavonol accumulation and localization patterns were similar to those of the PAs, and the B-ring hydroxylation pattern of both was correlated with flavonoid-3′-hydroxylase transcript abundance. By contrast, anthocyanins accumulated late in maturation, which coincided with a peak in flavonoid-3-O-glycosyltransferase and flavonoid-3′5′-hydroxylase transcripts. Transcripts of VcMYBPA1, which likely encodes an R2R3-MYB transcriptional regulator of PA synthesis, were prominent in both phases of development. Furthermore, the initiation of ripening was accompanied by a substantial rise in abscisic acid, a growth regulator that may be an important component of the ripening process and contribute to the regulation

A novel CARD containing splice-isoform of CIITA regulates nitric oxide synthesis in dendritic cells.

PubMed

Huang, Dachuan; Lim, Sylvia; Chua, Rong Yuan Ray; Shi, Hong; Ng, Mah Lee; Wong, Siew Heng

2010-03-01

MHC class II expression is controlled mainly at transcriptional level by class II transactivator (CIITA), which is a non-DNA binding coactivator and serves as a master control factor for MHC class II genes expression. Here, we describe the function of a novel splice-isoform of CIITA, DC-expressed caspase inhibitory isoform of CIITA (or DC-CASPIC), and we show that the expression of DCCASPIC in DC is upregulated upon lipopolysaccharides (LPS) induction. DC-CASPIC localizes to mitochondria, and protein-protein interaction study demonstrates that DC-CASPIC interacts with caspases and inhibits its activity in DC. Consistently, DC-CASPIC suppresses caspases-induced degradation of nitric oxide synthase-2 (NOS2) and subsequently promotes the synthesis of nitric oxide (NO). NO is an essential regulatory molecule that modulates the capability of DC in stimulating T cell proliferation/activation in vitro; hence, overexpression of DC-CASPIC in DC enhances this stimulation. Collectively, our findings reveal that DC-CASPIC is a key molecule that regulates caspases activity and NO synthesis in DC.

Nicotine-induced retardation of chondrogenesis through down-regulation of IGF-1 signaling pathway to inhibit matrix synthesis of growth plate chondrocytes in fetal rats

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

Deng, Yu; Cao, Hong; Cu, Fenglong

Previous studies have confirmed that maternal tobacco smoking causes intrauterine growth retardation (IUGR) and skeletal growth retardation. Among a multitude of chemicals associated with cigarette smoking, nicotine is one of the leading candidates for causing low birth weights. However, the possible mechanism of delayed chondrogenesis by prenatal nicotine exposure remains unclear. We investigated the effects of nicotine on fetal growth plate chondrocytes in vivo and in vitro. Rats were given 2.0 mg/kg·d of nicotine subcutaneously from gestational days 11 to 20. Prenatal nicotine exposure increased the levels of fetal blood corticosterone and resulted in fetal skeletal growth retardation. Moreover, nicotinemore » exposure induced the inhibition of matrix synthesis and down-regulation of insulin-like growth factor 1 (IGF-1) signaling in fetal growth plates. The effects of nicotine on growth plates were studied in vitro by exposing fetal growth plate chondrocytes to 0, 1, 10, or 100 μM of nicotine for 10 days. Nicotine inhibited matrix synthesis and down-regulated IGF-1 signaling in chondrocytes in a concentration-dependent manner. These results suggest that prenatal nicotine exposure induces delayed chondrogenesis and that the mechanism may involve the down-regulation of IGF-1 signaling and the inhibition of matrix synthesis by growth plate chondrocytes. The present study aids in the characterization of delayed chondrogenesis caused by prenatal nicotine exposure, which might suggest a candidate mechanism for intrauterine origins of osteoporosis and osteoarthritis. - Highlights: ► Prenatal nicotine-exposure could induce delayed chondrogenesis in fetal rats. ► Nicotine inhibits matrix synthesis of fetal growth plate chondrocytes. ► Nicotine inhibits IGF-1 signaling pathway in fetal growth plate chondrocytes.« less

Parkin regulates translesion DNA synthesis in response to UV radiation

PubMed Central

Huang, Min; Liu, Hongmei; Wang, Fengli; Gong, Juanjuan; Wang, Jiuqiang; Li, Xiaoling; Chen, Qian; Shen, Hongyan; Zhu, Shu; Wang, Yun; Liu, Yang; Guo, Caixia; Tang, Tie-Shan

2017-01-01

Deficiency of Parkin is a major cause of early-onset Parkinson's disease (PD). Notably, PD patients also exhibit a significantly higher risk in melanoma and other skin tumors, while the mechanism remains largely unknown. In this study, we show that depletion of Parkin causes compromised cell viability and genome stability after ultraviolet (UV) radiation. We demonstrate that Parkin promotes efficient Rad18-dependent proliferating cell nuclear antigen (PCNA) monoubiquitination by facilitating the formation of Replication protein A (RPA)-coated ssDNA upon UV radiation. Furthermore, Parkin is found to physically interact with NBS1 (Nijmegen breakage syndrome 1), and to be required for optimal recruitment of NBS1 and DNA polymerase eta (Polη) to UV-induced damage sites. Consequently, depletion of Parkin leads to increased UV-induced mutagenesis. These findings unveil an important role of Parkin in protecting genome stability through positively regulating translesion DNA synthesis (TLS) upon UV damage, providing a novel mechanistic link between Parkin deficiency and predisposition to skin cancers in PD patients. PMID:28430587

Regulation of Insulin Synthesis and Secretion and Pancreatic Beta-Cell Dysfunction in Diabetes

PubMed Central

Fu, Zhuo; Gilbert, Elizabeth R.; Liu, Dongmin

2014-01-01

Pancreatic β-cell dysfunction plays an important role in the pathogenesis of both type 1 and type 2 diabetes. Insulin, which is produced in β-cells, is a critical regulator of metabolism. Insulin is synthesized as preproinsulin and processed to proinsulin. Proinsulin is then converted to insulin and C-peptide and stored in secretary granules awaiting release on demand. Insulin synthesis is regulated at both the transcriptional and translational level. The cis-acting sequences within the 5′ flanking region and trans-activators including paired box gene 6 (PAX6), pancreatic and duodenal homeobox-1(PDX-1), MafA, and B-2/Neurogenic differentiation 1 (NeuroD1) regulate insulin transcription, while the stability of preproinsulin mRNA and its untranslated regions control protein translation. Insulin secretion involves a sequence of events in β-cells that lead to fusion of secretory granules with the plasma membrane. Insulin is secreted primarily in response to glucose, while other nutrients such as free fatty acids and amino acids can augment glucose-induced insulin secretion. In addition, various hormones, such as melatonin, estrogen, leptin, growth hormone, and glucagon like peptide-1 also regulate insulin secretion. Thus, the β-cell is a metabolic hub in the body, connecting nutrient metabolism and the endocrine system. Although an increase in intracellular [Ca2+] is the primary insulin secretary signal, cAMP signaling-dependent mechanisms are also critical in the regulation of insulin secretion. This article reviews current knowledge on how β-cells synthesize and secrete insulin. In addition, this review presents evidence that genetic and environmental factors can lead to hyperglycemia, dyslipidemia, inflammation, and autoimmunity, resulting in β-cell dysfunction, thereby triggering the pathogenesis of diabetes. PMID:22974359

MicroRNA and Transcriptomic Profiling Showed miRNA-Dependent Impairment of Systemic Regulation and Synthesis of Biomolecules in Rag2 KO Mice.

PubMed

Reza, Abu Musa Md Talimur; Choi, Yun-Jung; Kim, Jin-Hoi

2018-02-27

The Rag2 knockout (KO) mouse is a well-established immune-compromised animal model for biomedical research. A comparative study identified the deregulated expression of microRNAs (miRNAs) and messenger RNAs (mRNAs) in Rag2 KO mice. However, the interaction between deregulated genes and miRNAs in the alteration of systemic (cardiac, renal, hepatic, nervous, and hematopoietic) regulations and the synthesis of biomolecules (such as l-tryptophan, serotonin, melatonin, dopamine, alcohol, noradrenaline, putrescine, and acetate) are unclear. In this study, we analyzed both miRNA and mRNA expression microarray data from Rag2 KO and wild type mice to investigate the possible role of miRNAs in systemic regulation and biomolecule synthesis. A notable finding obtained from this analysis is that the upregulation of several genes which are target molecules of the downregulated miRNAs in Rag2 KO mice, can potentially trigger the degradation of l-tryptophan, thereby leading to the systemic impairment and alteration of biomolecules synthesis as well as changes in behavioral patterns (such as stress and fear responses, and social recognition memory) in Rag2 gene-depleted mice. These findings were either not observed or not explicitly described in other published Rag2 KO transcriptome analyses. In conclusion, we have provided an indication of miRNA-dependent regulations of clinical and pathological conditions in cardiac, renal, hepatic, nervous, and hematopoietic systems in Rag2 KO mice. These results may significantly contribute to the prediction of clinical disease caused by Rag2 deficiency.

Influence of virgin coconut oil-enriched diet on the transcriptional regulation of fatty acid synthesis and oxidation in rats - a comparative study.

PubMed

Arunima, Sakunthala; Rajamohan, Thankappan

2014-05-28

The present study was carried out to evaluate the effects of virgin coconut oil (VCO) compared with copra oil, olive oil and sunflower-seed oil on the synthesis and oxidation of fatty acids and the molecular regulation of fatty acid metabolism in normal rats. Male Sprague-Dawley rats were fed the test oils at 8 % for 45 d along with a synthetic diet. Dietary supplementation of VCO decreased tissue lipid levels and reduced the activity of the enzymes involved in lipogenesis, namely acyl CoA carboxylase and fatty acid synthase (FAS) (P< 0·05). Moreover, VCO significantly (P< 0·05) reduced the de novo synthesis of fatty acids by down-regulating the mRNA expression of FAS and its transcription factor, sterol regulatory element-binding protein-1c, compared with the other oils. VCO significantly (P< 0·05) increased the mitochondrial and peroxisomal β-oxidation of fatty acids, which was evident from the increased activities of carnitine palmitoyl transferase I, acyl CoA oxidase and the enzymes involved in mitochondrial β-oxidation; this was accomplished by up-regulating the mRNA expression of PPARα and its target genes involved in fatty acid oxidation. In conclusion, the present results confirmed that supplementation of VCO has beneficial effects on lipid parameters by reducing lipogenesis and enhancing the rate of fatty acid catabolism; this effect was mediated at least in part via PPARα-dependent pathways. Thus, dietary VCO reduces the risk for CHD by beneficially modulating the synthesis and degradation of fatty acids.

Piceid (resveratrol glucoside) synthesis in stilbene synthase transgenic apple fruit.

PubMed

Rühmann, Susanne; Treutter, Dieter; Fritsche, Steffi; Briviba, Karlis; Szankowski, Iris

2006-06-28

A stilbene synthase gene along with the selectable marker gene bar for herbicide resistance was transferred via Agrobacterium tumefaciens mediated transformation into apple (Malus domesticaBorkh.) cvs. 'Elstar' and 'Holsteiner Cox'. The stilbene synthase catalyzes the conversion of 1 molecule of p-coumaroyl-CoA and 3 molecules of malonyl-CoA into 3,4',5-trihydroxystilbene, commonly known as resveratrol. This phytoalexin has implications in both phytopathology and human health. Greenhouse-grown transgenic and nontransformed control plants were grafted onto dwarfing rootstock M27. Flowering and fruiting occurred within the following years, offering the opportunity to analyze transgenic apple fruit and fertility of transgenic plants as well as inheritance of the transgenes into the seedling progeny. Molecular analysis revealed that the stilbene synthase is expressed in transgenic plants and in the skin and flesh of transgenic apple fruit. After formation, resveratrol is modified by the addition of a hexose sugar. The resulting component was characterized as piceid. With the aim of characterizing the influence of the novel biosynthetic pathway on the accumulation of other phenolic compounds naturally present in apple fruit, the amounts of flavanols, flavonols, phloretin derivatives and hydroxycinnamic acids in wild type and transgenic fruit were determined by HPLC. In all investigated transformed lines that accumulated piceid, no negative correlation between levels of piceid and the above-mentioned compounds was observed, except for the flavonol contents, which slightly decreased. Inheritance of the transgenes was confirmed in the seedling progeny, which were obtained after pollination of transgenic plants with nontransgenic pollen and vice versa after pollination of nontransgenic plants with pollen obtained from transgenic plants. The fertility of stilbene synthase transgenic plants was demonstrated. To the authors' knowledge this is the first time that data are

Poly-γ-glutamic Acid Synthesis, Gene Regulation, Phylogenetic Relationships, and Role in Fermentation

PubMed Central

Hsueh, Yi-Huang; Huang, Kai-Yao; Kunene, Sikhumbuzo Charles; Lee, Tzong-Yi

2017-01-01

Poly-γ-glutamic acid (γ-PGA) is a biodegradable biopolymer produced by several bacteria, including Bacillus subtilis and other Bacillus species; it has good biocompatibility, is non-toxic, and has various potential biological applications in the food, pharmaceutical, cosmetic, and other industries. In this review, we have described the mechanisms of γ-PGA synthesis and gene regulation, its role in fermentation, and the phylogenetic relationships among various pgsBCAE, a biosynthesis gene cluster of γ-PGA, and pgdS, a degradation gene of γ-PGA. We also discuss potential applications of γ-PGA and highlight the established genetic recombinant bacterial strains that produce high levels of γ-PGA, which can be useful for large-scale γ-PGA production. PMID:29215550

Mlc Is a Transcriptional Activator with a Key Role in Integrating Cyclic AMP Receptor Protein and Integration Host Factor Regulation of Leukotoxin RNA Synthesis in Aggregatibacter actinomycetemcomitans

PubMed Central

Childress, Catherine; Feuerbacher, Leigh A.; Phillips, Linda; Burgum, Alex

2013-01-01

Aggregatibacter actinomycetemcomitans, a periodontal pathogen, synthesizes leukotoxin (LtxA), a protein that helps the bacterium evade the host immune response. Transcription of the ltxA operon is induced during anaerobic growth. The cyclic AMP (cAMP) receptor protein (CRP) indirectly increases ltxA expression, but the intermediary regulator is unknown. Integration host factor (IHF) binds to and represses the leukotoxin promoter, but neither CRP nor IHF is responsible for the anaerobic induction of ltxA RNA synthesis. Thus, we have undertaken studies to identify other regulators of leukotoxin transcription and to demonstrate how these proteins work together to modulate leukotoxin synthesis. First, analyses of ltxA RNA expression from defined leukotoxin promoter mutations in the chromosome identify positions −69 to −35 as the key control region and indicate that an activator protein modulates leukotoxin transcription. We show that Mlc, which is a repressor in Escherichia coli, functions as a direct transcriptional activator in A. actinomycetemcomitans; an mlc deletion mutant reduces leukotoxin RNA synthesis, and recombinant Mlc protein binds specifically at the −68 to −40 region of the leukotoxin promoter. Furthermore, we show that CRP activates ltxA expression indirectly by increasing the levels of Mlc. Analyses of Δmlc, Δihf, and Δihf Δmlc strains demonstrate that Mlc can increase RNA polymerase (RNAP) activity directly and that IHF represses ltxA RNA synthesis mainly by blocking Mlc binding. Finally, a Δihf Δmlc mutant still induces ltxA during anaerobic growth, indicating that there are additional factors involved in leukotoxin transcriptional regulation. A model for the coordinated regulation of leukotoxin transcription is presented. PMID:23475968

Porcine Reproductive and Respiratory Syndrome Virus Nucleocapsid Protein Interacts with Nsp9 and Cellular DHX9 To Regulate Viral RNA Synthesis.

PubMed

Liu, Long; Tian, Jiao; Nan, Hao; Tian, Mengmeng; Li, Yuan; Xu, Xiaodong; Huang, Baicheng; Zhou, Enmin; Hiscox, Julian A; Chen, Hongying

2016-06-01

Porcine reproductive and respiratory syndrome virus (PRRSV) nucleocapsid (N) protein is the main component of the viral capsid to encapsulate viral RNA, and it is also a multifunctional protein involved in the regulation of host cell processes. Nonstructural protein 9 (Nsp9) is the RNA-dependent RNA polymerase that plays a critical role in viral RNA transcription and replication. In this study, we demonstrate that PRRSV N protein is bound to Nsp9 by protein-protein interaction and that the contacting surface on Nsp9 is located in the two predicted α-helixes formed by 48 residues at the C-terminal end of the protein. Mutagenesis analyses identified E646, E608, and E611 on Nsp9 and Q85 on the N protein as the pivotal residues participating in the N-Nsp9 interaction. By overexpressing the N protein binding fragment of Nsp9 in infected Marc-145 cells, the synthesis of viral RNAs, as well as the production of infectious progeny viruses, was dramatically inhibited, suggesting that Nsp9-N protein association is involved in the process of viral RNA production. In addition, we show that PRRSV N interacts with cellular RNA helicase DHX9 and redistributes the protein into the cytoplasm. Knockdown of DHX9 increased the ratio of short subgenomic mRNAs (sgmRNAs); in contrast, DHX9 overexpression benefited the synthesis of longer sgmRNAs and the viral genomic RNA (gRNA). These results imply that DHX9 is recruited by the N protein in PRRSV infection to regulate viral RNA synthesis. We postulate that N and DHX9 may act as antiattenuation factors for the continuous elongation of nascent transcript during negative-strand RNA synthesis. It is unclear whether the N protein of PRRSV is involved in regulation of the viral RNA production process. In this report, we demonstrate that the N protein of the arterivirus PRRSV participates in viral RNA replication and transcription through interacting with Nsp9 and its RdRp and recruiting cellular RNA helicase to promote the production of

[Synthesis and regulation of flavor compounds derived from brewing yeast: Esters].

PubMed

Loviso, Claudia L; Libkind, Diego

2018-04-04

During brewing process yeast produce more than 500 chemical compounds that can negatively and positively impact beer at the organoleptic level. In recent years, and particularly thanks to the advancement of molecular biology and genomics, there has been considerable progress in our understanding about the molecular and cellular basis of the synthesis and regulation of many of these flavor compounds. This article focuses on esters, responsible for the floral and fruity beer flavor. Its formation depends on various enzymes and factors such as the concentration of wort nutrients, the amount of dissolved oxygen and carbon dioxide, fermentation temperature and mainly the genetics of the yeast used. We provide information about how the esters originate and how is the impact of different fermentative parameters on the final concentrations of these compounds and the quality of the end product. Copyright © 2018 The Authors. Publicado por Elsevier España, S.L.U. All rights reserved.

Arrestin-dependent angiotensin AT1 receptor signaling regulates Akt and mTor-mediated protein synthesis.

PubMed

Kendall, Ryan T; Lee, Mi-Hye; Pleasant, Dorea L; Robinson, Katherine; Kuppuswamy, Dhandapani; McDermott, Paul J; Luttrell, Louis M

2014-09-19

Control of protein synthesis is critical to both cell growth and proliferation. The mammalian target of rapamycin (mTOR) integrates upstream growth, proliferation, and survival signals, including those transmitted via ERK1/2 and Akt, to regulate the rate of protein translation. The angiotensin AT1 receptor has been shown to activate both ERK1/2 and Akt in arrestin-based signalsomes. Here, we examine the role of arrestin-dependent regulation of ERK1/2 and Akt in the stimulation of mTOR-dependent protein translation by the AT1 receptor using HEK293 and primary vascular smooth muscle cell models. Nascent protein synthesis stimulated by both the canonical AT1 receptor agonist angiotensin II (AngII), and the arrestin pathway-selective agonist [Sar(1)-Ile(4)-Ile(8)]AngII (SII), is blocked by shRNA silencing of βarrestin1/2 or pharmacological inhibition of Akt, ERK1/2, or mTORC1. In HEK293 cells, SII activates a discrete arrestin-bound pool of Akt and promotes Akt-dependent phosphorylation of mTOR and its downstream effector p70/p85 ribosomal S6 kinase (p70/85S6K). In parallel, SII-activated ERK1/2 helps promote mTOR and p70/85S6K phosphorylation, and is required for phosphorylation of the known ERK1/2 substrate p90 ribosomal S6 kinase (p90RSK). Thus, arrestins coordinate AT1 receptor regulation of ERK1/2 and Akt activity and stimulate protein translation via both Akt-mTOR-p70/85S6K and ERK1/2-p90RSK pathways. These results suggest that in vivo, arrestin pathway-selective AT1 receptor agonists may promote cell growth or hypertrophy through arrestin-mediated mechanisms despite their antagonism of G protein signaling. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

The Interpretation of Cholesterol Balance Derived Synthesis Data and Surrogate Noncholesterol Plasma Markers for Cholesterol Synthesis under Lipid Lowering Therapies

PubMed Central

Stellaard, Frans

2017-01-01

The cholesterol balance procedure allows the calculation of cholesterol synthesis based on the assumption that loss of endogenous cholesterol via fecal excretion and bile acid synthesis is compensated by de novo synthesis. Under ezetimibe therapy hepatic cholesterol is diminished which can be compensated by hepatic de novo synthesis and hepatic extraction of plasma cholesterol. The plasma lathosterol concentration corrected for total cholesterol concentration (R_Lath) as a marker of de novo cholesterol synthesis is increased during ezetimibe treatment but unchanged under treatment with ezetimibe and simvastatin. Cholesterol balance derived synthesis data increase during both therapies. We hypothesize the following. (1) The cholesterol balance data must be applied to the hepatobiliary cholesterol pool. (2) The calculated cholesterol synthesis value is the sum of hepatic de novo synthesis and the net plasma—liver cholesterol exchange rate. (3) The reduced rate of biliary cholesterol absorption is the major trigger for the regulation of hepatic cholesterol metabolism under ezetimibe treatment. Supportive experimental and literature data are presented that describe changes of cholesterol fluxes under ezetimibe, statin, and combined treatments in omnivores and vegans, link plasma R_Lath to liver function, and define hepatic de novo synthesis as target for regulation of synthesis. An ezetimibe dependent direct hepatic drug effect cannot be excluded. PMID:28321334

Synthesis and characterization of TEP-EDTA-regulated bioactive hydroxyapatite

NASA Astrophysics Data System (ADS)

Haders, Daniel Joseph, II

Ca2+ concentration enabled the HA crystallization process to be growth dominated, producing films composed of high crystallinity, hexagonal grains on multiple metallic substrates. TEP regulation of HA crystallization enabled the deposition of an adhesive CaTiO3 intermediate layer, and then HA in a continuous, phase sequenced process on Ti6Al4V substrates, the first such process reported in the hydrothermal HA literature. The HA film was found to be deposited by a passivating competitive growth mechanism that enabled the [0001] crystallographic orientation of hexagonal single crystals to be engineered with synthesis time. Bioactivity analysis demonstrated that films were bioactive and bone bonding. Together, these results suggest that these HA films are candidates for use on metallic orthopedic implants, namely Ti6Al4V.

miR-148a and miR-17-5p synergistically regulate milk TAG synthesis via PPARGC1A and PPARA in goat mammary epithelial cells.

PubMed

Chen, Zhi; Luo, Jun; Sun, Shuang; Cao, Duoyao; Shi, Huaiping; Loor, Juan J

2017-03-04

MicroRNA (miRNA) are a class of '18-25' nt RNA molecules which regulate gene expression and play an important role in several biologic processes including fatty acid metabolism. Here we used S-Poly (T) and high-throughput sequencing to evaluate the expression of miRNA and mRNA during early-lactation and in the non-lactating ("dry") period in goat mammary gland tissue. Results indicated that miR-148a, miR-17-5p, PPARGC1A and PPARA are highly expressed in the goat mammary gland in early-lactation and non-lactating periods. Utilizing a Luciferase reporter assay and Western Blot, PPARA, an important regulator of fatty acid oxidation, and PGC1a (PPARGC1A), a major regulator of fat metabolism, were demonstrated to be targets of miR-148a and miR-17-5p in goat mammary epithelial cells (GMECs). It was also revealed that miR-148a expression can regulate PPARA, and miR-17-5p represses PPARGC1A in GMECs. Furthermore, the overexpression of miR-148a and miR-17-5p promoted triacylglycerol (TAG) synthesis while the knockdown of miR-148a and miR-17-5p impaired TAG synthesis in GMEC. These findings underscore the importance of miR-148a and miR-17-5p as key components in the regulation of TAG synthesis. In addition, miR-148a cooperates with miR-17-5p to regulate fatty acid metabolism by repressing PPARGC1A and PPARA in GMECs. Further studies on the functional role of miRNAs in lipid metabolism of ruminant mammary cells seem warranted.

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

Lipid phosphate phosphatase 3 regulates adipocyte sphingolipid synthesis, but not developmental adipogenesis or diet-induced obesity in mice.

PubMed

Federico, Lorenzo; Yang, Liping; Brandon, Jason; Panchatcharam, Manikandan; Ren, Hongmei; Mueller, Paul; Sunkara, Manjula; Escalante-Alcalde, Diana; Morris, Andrew J; Smyth, Susan S

2018-01-01

Dephosphorylation of phosphatidic acid (PA) is the penultimate step in triglyceride synthesis. Adipocytes express soluble intracellular PA-specific phosphatases (Lipins) and broader specificity membrane-associated lipid phosphate phosphatases (LPPs) that can also dephosphorylate PA. Inactivation of lipin1 causes lipodystrophy in mice due to defective developmental adipogenesis. Triglyceride synthesis is diminished but not ablated by inactivation of lipin1 in differentiated adipocytes implicating other PA phosphatases in this process. To investigate the possible role of LPPs in adipocyte lipid metabolism and signaling we made mice with adipocyte-targeted inactivation of LPP3 encoded by the Plpp3(Ppap2b) gene. Adipocyte LPP3 deficiency resulted in blunted ceramide and sphingomyelin accumulation during diet-induced adipose tissue expansion, accumulation of the LPP3 substrate sphingosine 1- phosphate, and reduced expression of serine palmitoyl transferase. However, adiposity was unaffected by LPP3 deficiency on standard, high fat diet or Western diets, although Western diet-fed mice with adipocyte LPP3 deficiency exhibited improved glucose tolerance. Our results demonstrate functional compartmentalization of lipid phosphatase activity in adipocytes and identify an unexpected role for LPP3 in the regulation of diet-dependent sphingolipid synthesis that may impact on insulin signaling.

Reduction in central venous pressure enhances erythropoietin synthesis: role of volume-regulating hormones.

PubMed

Montero, D; Rauber, S; Goetze, J P; Lundby, C

2016-10-01

Erythropoiesis is a tightly controlled biological event, but its regulation under non-hypoxic conditions, however, remains unresolved. We examined whether acute changes in central venous blood pressure (CVP) elicited by whole-body tilting affect erythropoietin (EPO) concentration according to volume-regulating hormones. Plasma EPO, angiotensin II (ANGII), aldosterone, pro-atrial natriuretic peptide (proANP) and copeptin concentrations were measured at supine rest and up to 3 h during 30° head-up (HUT) and head-down tilt (HDT) in ten healthy male volunteers. Plasma albumin concentration was used to correct for changes in plasma volume and CVP was estimated through the internal jugular vein (IJV) aspect ratio with ultrasonography. From supine rest, the IJV aspect ratio was decreased and increased throughout HUT and HDT respectively. Plasma EPO concentration increased during HUT (13%; P = 0.001, P for linear component = 0.017), independent of changes in albumin concentration. Moreover, ANGII and copeptin concentrations increased during HUT, while proANP decreased. The increase in EPO concentration during HUT disappeared when adjusted for changes in copeptin. During HDT, EPO, ANGII and copeptin concentrations remained unaffected while proANP increased. In regression analyses, EPO was positively associated with copeptin (β = 0.55; 95% CI = 0.18, 0.93; P = 0.004) irrespective of changes in other hormones and albumin concentration. Reduction in CVP prompts an increase in plasma EPO concentration independent of hemoconcentration and hence suggests CVP per se as an acute regulator of EPO synthesis. This effect may be explained by changes in volume-regulating hormones. © 2016 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

Extensive In Vitro Hyphal Growth of Vesicular-Arbuscular Mycorrhizal Fungi in the Presence of CO(2) and Flavonols.

PubMed

Bécard, G; Douds, D D; Pfeffer, P E

1992-03-01

Various flavonoids were tested for their ability to stimulate in vitro growth of germinated spores of vesicular-arbuscular mycorrhizal fungi. Experiments were performed in the presence of 2% CO(2), previously demonstrated to be required for growth of Gigaspora margarita (G. Bécard and Y. Piché, Appl. Environ. Microbiol. 55:2320-2325, 1989). Only the flavonols stimulated fungal growth. The flavones, flavanones, and isoflavones tested were generally inhibitory. Quercetin (10 muM) prolonged hyphal growth from germinated spores of G. margarita from 10 to 42 days. An average of more than 500 mm of hyphal growth and 13 auxiliary cells per spore were obtained. Quercetin also stimulated the growth of Glomus etunicatum. The glycosides of quercetin, rutin, and quercitrin were not stimulatory. The axenic growth of G. margarita achieved here under rigorously defined conditions is the most ever reported for a vesicular-arbuscular mycorrhizal fungus.

Flavonol Activation Defines an Unanticipated Ligand-Binding Site in the Kinase-RNase Domain of IRE1

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

Wiseman, R. Luke; Zhang, Yuhong; Lee, Kenneth P.K.

2010-08-18

Signaling in the most conserved branch of the endoplasmic reticulum (ER) unfolded protein response (UPR) is initiated by sequence-specific cleavage of the HAC1/XBP1 mRNA by the ER stress-induced kinase-endonuclease IRE1. We have discovered that the flavonol quercetin activates yeast IRE1's RNase and potentiates activation by ADP, a natural activating ligand that engages the IRE1 nucleotide-binding cleft. Enzyme kinetics and the structure of a cocrystal of IRE1 complexed with ADP and quercetin reveal engagement by quercetin of an unanticipated ligand-binding pocket at the dimer interface of IRE1's kinase extension nuclease (KEN) domain. Analytical ultracentrifugation and crosslinking studies support the preeminence ofmore » enhanced dimer formation in quercetin's mechanism of action. These findings hint at the existence of endogenous cytoplasmic ligands that may function alongside stress signals from the ER lumen to modulate IRE1 activity and at the potential for the development of drugs that modify UPR signaling from this unanticipated site.« less

De-novo NAD+ synthesis regulates SIRT1-FOXO1 apoptotic pathway in response to NQO1 substrates in lung cancer cells

PubMed Central

Cheng, Xuefang; Li, Qingran; Liu, Fang; Ye, Hui; Zhao, Min; Wang, Hong; Wang, Guangji; Hao, Haiping

2016-01-01

Tryptophan metabolism is essential in diverse kinds of tumors via regulating tumor immunology. However, the direct role of tryptophan metabolism and its signaling pathway in cancer cells remain largely elusive. Here, we establish a mechanistic link from L-type amino acid transporter 1 (LAT1) mediated transport of tryptophan and the subsequent de-novo NAD+ synthesis to SIRT1-FOXO1 regulated apoptotic signaling in A549 cells in response to NQO1 activation. In response to NQO1 activation, SIRT1 is repressed leading to the increased cellular accumulation of acetylated FOXO1 that transcriptionally activates apoptotic signaling. Decreased uptake of tryptophan due to the downregulation of LAT1 coordinates with PARP-1 hyperactivation to induce rapid depletion of NAD+ pool. Particularly, the LAT1-NAD+-SIRT1 signaling is activated in tumor tissues of patients with non-small cell lung cancer. Because NQO1 activation is characterized with oxidative challenge induced DNA damage, these results suggest that LAT1 and de-novo NAD+ synthesis in NSCLC cells may play essential roles in sensing excessive oxidative stress. PMID:27566573

De-novo NAD+ synthesis regulates SIRT1-FOXO1 apoptotic pathway in response to NQO1 substrates in lung cancer cells.

PubMed

Liu, Huiying; Xing, Rong; Cheng, Xuefang; Li, Qingran; Liu, Fang; Ye, Hui; Zhao, Min; Wang, Hong; Wang, Guangji; Hao, Haiping

2016-09-20

Tryptophan metabolism is essential in diverse kinds of tumors via regulating tumor immunology. However, the direct role of tryptophan metabolism and its signaling pathway in cancer cells remain largely elusive. Here, we establish a mechanistic link from L-type amino acid transporter 1 (LAT1) mediated transport of tryptophan and the subsequent de-novo NAD+ synthesis to SIRT1-FOXO1 regulated apoptotic signaling in A549 cells in response to NQO1 activation. In response to NQO1 activation, SIRT1 is repressed leading to the increased cellular accumulation of acetylated FOXO1 that transcriptionally activates apoptotic signaling. Decreased uptake of tryptophan due to the downregulation of LAT1 coordinates with PARP-1 hyperactivation to induce rapid depletion of NAD+ pool. Particularly, the LAT1-NAD+-SIRT1 signaling is activated in tumor tissues of patients with non-small cell lung cancer. Because NQO1 activation is characterized with oxidative challenge induced DNA damage, these results suggest that LAT1 and de-novo NAD+ synthesis in NSCLC cells may play essential roles in sensing excessive oxidative stress.

Application of Differential Colorimetry To Evaluate Anthocyanin-Flavonol-Flavanol Ternary Copigmentation Interactions in Model Solutions.

PubMed

Gordillo, Belén; Rodríguez-Pulido, Francisco J; González-Miret, M Lourdes; Quijada-Morín, Natalia; Rivas-Gonzalo, Julián C; García-Estévez, Ignacio; Heredia, Francisco J; Escribano-Bailón, M Teresa

2015-09-09

The combined effect of anthocyanin-flavanol-flavonol ternary interactions on the colorimetric and chemical stability of malvidin-3-glucoside has been studied. Model solutions with fixed malvidin-3-glucoside/(+)-catechin ratio (MC) and variable quercetin-3-β-d-glucoside concentration (MC+Q) and solutions with fixed malvidin-3-glucoside/quercetin-3-β-d-glucoside ratio (MQ) and variable (+)-catechin concentration (MQ+C) were tested at levels closer to those existing in wines. Color variations during storage were evaluated by differential colorimetry. Changes in the anthocyanin concentration were monitored by HPLC-DAD. CIELAB color-difference formulas were demonstrated to be of practical interest to assess the stronger and more stable interaction of quercetin-3-β-d-glucoside with MC binary mixture than (+)-catechin with MQ mixture. The results imply that MC+Q ternary solutions kept their intensity and bluish tonalities for a longer time in comparison to MQ+C solutions. The stability of malvidin-3-glucoside improves when the concentration of quercetin-3-β-d-glucoside increases in MC+Q mixtures, whereas the addition of (+)-catechin in MQ+C mixtures resulted in an opposite effect.

Insights into the Regulation of DMSP Synthesis in the Diatom Thalassiosira pseudonana through APR Activity, Proteomics and Gene Expression Analyses on Cells Acclimating to Changes in Salinity, Light and Nitrogen

PubMed Central

Kettles, Nicola Louise; Kopriva, Stanislav; Malin, Gill

2014-01-01

Despite the importance of dimethylsulphoniopropionate (DMSP) in the global sulphur cycle and climate regulation, the biological pathways underpinning its synthesis in marine phytoplankton remain poorly understood. The intracellular concentration of DMSP increases with increased salinity, increased light intensity and nitrogen starvation in the diatom Thalassiosira pseudonana. We used these conditions to investigate DMSP synthesis at the cellular level via analysis of enzyme activity, gene expression and proteome comparison. The activity of the key sulphur assimilatory enzyme, adenosine 5′-phosphosulphate reductase was not coordinated with increasing intracellular DMSP concentration. Under all three treatments coordination in the expression of sulphur assimilation genes was limited to increases in sulphite reductase transcripts. Similarly, proteomic 2D gel analysis only revealed an increase in phosphoenolpyruvate carboxylase following increases in DMSP concentration. Our findings suggest that increased sulphur assimilation might not be required for increased DMSP synthesis, instead the availability of carbon and nitrogen substrates may be important in the regulation of this pathway. This contrasts with the regulation of sulphur metabolism in higher plants, which generally involves up-regulation of several sulphur assimilatory enzymes. In T. pseudonana changes relating to sulphur metabolism were specific to the individual treatments and, given that little coordination was seen in transcript and protein responses across the three growth conditions, different patterns of regulation might be responsible for the increase in DMSP concentration seen under each treatment. PMID:24733415

Low Medium pH Value Enhances Anthocyanin Accumulation in Malus Crabapple Leaves

PubMed Central

Tian, Ji; Jin, Kaina; Yao, Yuncong

2014-01-01

Anthocyanin is a critical factor involved in coloration of plant tissues, but the mechanism how medium pH values affect anthocyanin accumulation in woody plants is unknown. We analyzed anthocyanin composition and the expression of elements encoding anthocyanin and flavonols biosynthesis underlying different medium pH values by using three different leave color type cultivars. HPLC analysis demonstrated that high medium pH values treatment induced a dramatic decrease in the concentration of cyaniding in crabapple leaves. Conversely, the high medium pH values induced up-regulation of the content of flavones and flavonols, suggesting that low pH treatment-induced anthocyanin accumulation. Quantitative real time PCR experiment showed the expression level of anthocyanidin synthase (McANS) and uridine diphosphate glucose flavonoid 3-O-glucosyltransferase (McUFGT) was up-regulated by low pH values treatment, and high medium pH value treatment up-regulate the transcription level of flavonol synthase (McFLS). Meanwhile, several MYB TFs have been suggested in the regulation of pH responses. These results strongly indicate that the low pH treatment-induced anthocyanin accumulation is mediated by the variation of mRNA transcription of the anthocyanin biosynthetic genes. PMID:24914811

Long-chain acyl-CoA synthetase 6 regulates lipid synthesis and mitochondrial oxidative capacity in human and rat skeletal muscle.

PubMed

Teodoro, Bruno G; Sampaio, Igor H; Bomfim, Lucas H M; Queiroz, André L; Silveira, Leonardo R; Souza, Anderson O; Fernandes, Anna M A P; Eberlin, Marcos N; Huang, Tai-Yu; Zheng, Donghai; Neufer, P Darrell; Cortright, Ronald N; Alberici, Luciane C

2017-02-01

Long-chain acyl-CoA synthetase 6 (ACSL6) mRNA is present in human and rat skeletal muscle, and is modulated by nutritional status: exercise and fasting decrease ACSL6 mRNA, whereas acute lipid ingestion increase its expression. ACSL6 genic inhibition in rat primary myotubes decreased lipid accumulation, as well as activated the higher mitochondrial oxidative capacity programme and fatty acid oxidation through the AMPK/PGC1-α pathway. ACSL6 overexpression in human primary myotubes increased phospholipid species and decreased oxidative metabolism. Long-chain acyl-CoA synthetases (ACSL 1 to 6) are key enzymes regulating the partitioning of acyl-CoA species toward different metabolic fates such as lipid synthesis or β-oxidation. Despite our understanding of ecotopic lipid accumulation in skeletal muscle being associated with metabolic diseases such as obesity and type II diabetes, the role of specific ACSL isoforms in lipid synthesis remains unclear. In the present study, we describe for the first time the presence of ACSL6 mRNA in human skeletal muscle and the role that ACSL6 plays in lipid synthesis in both rodent and human skeletal muscle. ACSL6 mRNA was observed to be up-regulated by acute high-fat meal ingestion in both rodents and humans. In rats, we also demonstrated that fasting and chronic aerobic training negatively modulated the ACSL6 mRNA and other genes of lipid synthesis. Similar results were obtained following ACSL6 knockdown in rat myotubes, which was associated with a decreased accumulation of TAGs and lipid droplets. Under the same knockdown condition, we further demonstrate an increase in fatty acid content, p-AMPK, mitochondrial content, mitochondrial respiratory rates and palmitate oxidation. These results were associated with increased PGC-1α, UCP2 and UCP3 mRNA and decreased reactive oxygen species production. In human myotubes, ACSL6 overexpression reduced palmitate oxidation and PGC-1α mRNA. In conclusion, ACSL6 drives acyl-CoA toward lipid

Vtc5, a Novel Subunit of the Vacuolar Transporter Chaperone Complex, Regulates Polyphosphate Synthesis and Phosphate Homeostasis in Yeast*

PubMed Central

Desfougères, Yann; Gerasimaitė, R̄uta; Jessen, Henning Jacob

2016-01-01

SPX domains control phosphate homeostasis in eukaryotes. Ten genes in yeast encode SPX-containing proteins, among which YDR089W is the only one of unknown function. Here, we show that YDR089W encodes a novel subunit of the vacuole transporter chaperone (VTC) complex that produces inorganic polyphosphate (polyP). The polyP synthesis transfers inorganic phosphate (Pi) from the cytosol into the acidocalcisome- and lysosome-related vacuoles of yeast, where it can be released again. It was therefore proposed for buffer changes in cytosolic Pi concentration (Thomas, M. R., and O'Shea, E. K. (2005) Proc. Natl. Acad. Sci. U.S.A. 102, 9565–9570). Vtc5 physically interacts with the VTC complex and accelerates the accumulation of polyP synthesized by it. Deletion of VTC5 reduces polyP accumulation in vivo and in vitro. Its overexpression hyperactivates polyP production and triggers the phosphate starvation response via the PHO pathway. Because this Vtc5-induced starvation response can be reverted by shutting down polyP synthesis genetically or pharmacologically, we propose that polyP synthesis rather than Vtc5 itself is a regulator of the PHO pathway. Our observations suggest that polyP synthesis not only serves to establish a buffer for transient drops in cytosolic Pi levels but that it can actively decrease or increase the steady state of cytosolic Pi. PMID:27587415

Requirement for the eIF4E binding proteins for the synergistic down-regulation of protein synthesis by hypertonic conditions and mTOR inhibition.

PubMed

Clemens, Michael J; Elia, Androulla; Morley, Simon J

2013-01-01

The protein kinase mammalian target of rapamycin (mTOR) regulates the phosphorylation and activity of several proteins that have the potential to control translation, including p70S6 kinase and the eIF4E binding proteins 4E-BP1 and 4E-BP2. In spite of this, in exponentially growing cells overall protein synthesis is often resistant to mTOR inhibitors. We report here that sensitivity of wild-type mouse embryonic fibroblasts (MEFs) to mTOR inhibitors can be greatly increased when the cells are subjected to the physiological stress imposed by hypertonic conditions. In contrast, protein synthesis in MEFs with a double knockout of 4E-BP1 and 4E-BP2 remains resistant to mTOR inhibitors under these conditions. Phosphorylation of p70S6 kinase and protein kinase B (Akt) is blocked by the mTOR inhibitor Ku0063794 equally well in both wild-type and 4E-BP knockout cells, under both normal and hypertonic conditions. The response of protein synthesis to hypertonic stress itself does not require the 4E-BPs. These data suggest that under certain stress conditions: (i) translation has a greater requirement for mTOR activity and (ii) there is an absolute requirement for the 4E-BPs for regulation by mTOR. Importantly, dephosphorylation of p70S6 kinase and Akt is not sufficient to affect protein synthesis acutely.

Identification of the C-terminal domain of Daxx acts as a potential regulator of intracellular cholesterol synthesis in HepG2 cells

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

Sun, Shaowei; Medical School, Hunan University of Chinese Medicine, Changsha 410208, Hunan; Wen, Juan

Daxx is a highly conserved nuclear transcriptional factor, which has been implicated in many nuclear processes including transcription and cell cycle regulation. Our previous study demonstrated Daxx also plays a role in regulation of intracellular cholesterol content. Daxx contains several domains that are essential for interaction with a growing number of proteins. To delineate the underlying mechanism of hypocholesterolemic activity of Daxx, we constructed a set of plasmids which can be used to overexpress different fragments of Daxx and transfected to HepG2 cells. We found that the C- terminal region Daxx626–740 clearly reduced intracellular cholesterol levels and inhibited the expressionmore » of SREBPs and SCAP. In GST pull-down experiments and Double immunofluorescence assays, Daxx626–740 was demonstrated to bind directly to androgen receptor (AR). Our findings suggest that the interaction of Daxx626-740 and AR abolishes the AR-mediated activation of SCAP/SREBPs pathway, which suppresses the de novo cholesterol synthesis. Thus, C-terminal domain of Daxx acts as a potential regulator of intracellular cholesterol content in HepG2 cells. - Highlights: • Daxx C-terminal domain reduces cholesterol levels. • Daxx C-terminal domain binds directly to AR. • The interaction of Daxx C-terminal domain and AR suppresses cholesterol synthesis.« less

PAH1-encoded Phosphatidate Phosphatase Plays a Role in the Growth Phase- and Inositol-mediated Regulation of Lipid Synthesis in Saccharomyces cerevisiae*

PubMed Central

Pascual, Florencia; Soto-Cardalda, Aníbal; Carman, George M.

2013-01-01

In the yeast Saccharomyces cerevisiae, the synthesis of phospholipids in the exponential phase of growth occurs at the expense of the storage lipid triacylglycerol. As exponential phase cells progress into the stationary phase, the synthesis of triacylglycerol occurs at the expense of phospholipids. Early work indicates a role of the phosphatidate phosphatase (PAP) in this metabolism; the enzyme produces the diacylglycerol needed for the synthesis of triacylglycerol and simultaneously controls the level of phosphatidate for the synthesis of phospholipids. Four genes (APP1, DPP1, LPP1, and PAH1) encode PAP activity in yeast, and it has been unclear which gene is responsible for the synthesis of triacylglycerol throughout growth. An analysis of lipid synthesis and composition, as well as PAP activity in various PAP mutant strains, showed the essential role of PAH1 in triacylglycerol synthesis throughout growth. Pah1p is a phosphorylated enzyme whose in vivo function is dependent on its dephosphorylation by the Nem1p-Spo7p protein phosphatase complex. nem1Δ mutant cells exhibited defects in triacylglycerol synthesis and lipid metabolism that mirrored those imparted by the pah1Δ mutation, substantiating the importance of Pah1p dephosphorylation throughout growth. An analysis of cells bearing PPAH1-lacZ and PPAH1-DPP1 reporter genes showed that PAH1 expression was induced throughout growth and that the induction in the stationary phase was stimulated by inositol supplementation. A mutant analysis indicated that the Ino2p/Ino4p/Opi1p regulatory circuit and transcription factors Gis1p and Rph1p mediated this regulation. PMID:24196957

Impact of Flavonols on Cardiometabolic Biomarkers:  A Meta-Analysis of Randomized Controlled Human  Trials to Explore the Role of Inter-Individual  Variability.

PubMed

Menezes, Regina; Rodriguez-Mateos, Ana; Kaltsatou, Antonia; González-Sarrías, Antonio; Greyling, Arno; Giannaki, Christoforos; Andres-Lacueva, Cristina; Milenkovic, Dragan; Gibney, Eileen R; Dumont, Julie; Schär, Manuel; Garcia-Aloy, Mar; Palma-Duran, Susana Alejandra; Ruskovska, Tatjana; Maksimova, Viktorija; Combet, Emilie; Pinto, Paula

2017-02-09

Several  epidemiological  studies  have  linked  flavonols  with  decreased  risk  of  cardiovascular  disease  (CVD).  However,  some  heterogeneity  in  the  individual  physiological  responses to the consumption of these compounds has been identified. This meta-analysis aimed to  study the effect of flavonol supplementation on biomarkers of CVD risk such as, blood lipids, blood  pressure and plasma glucose, as well as factors affecting their inter-individual variability. Data from  18 human randomized controlled trials were pooled and the effect was estimated using fixed or  random effects meta-analysis model and reported as difference in means (DM). Variability in the  response of blood lipids to supplementation with flavonols was assessed by stratifying various  population subgroups: age, sex, country, and health status. Results showed significant reductions  in total cholesterol (DM = -0.10 mmol/L; 95% CI: -0.20, -0.01), LDL cholesterol (DM = -0.14 mmol/L;  Nutrients 2017, 9, 117  2 of 21  95% CI: -0.21, 0.07), and triacylglycerol (DM = -0.10 mmol/L; 95% CI: -0.18, 0.03), and a significant  increase in HDL cholesterol (DM = 0.05 mmol/L; 95% CI: 0.02, 0.07). A significant reduction was also  observed in fasting plasma glucose (DM = -0.18 mmol/L; 95%CI: -0.29, -0.08), and in blood pressure  (SBP: DM = -4.84 mmHg; 95% CI: -5.64, -4.04; DBP: DM = -3.32 mmHg; 95% CI: -4.09, -2.55).  Subgroup analysis showed a more pronounced effect of flavonol intake in participants from Asian  countries and in participants with diagnosed disease or dyslipidemia, compared to healthy and  normal baseline values. In conclusion, flavonol consumption improved biomarkers of CVD risk,  however, country of origin and health status may influence the effect of flavonol

Emergence of robust growth laws from optimal regulation of ribosome synthesis.

PubMed

Scott, Matthew; Klumpp, Stefan; Mateescu, Eduard M; Hwa, Terence

2014-08-22

Bacteria must constantly adapt their growth to changes in nutrient availability; yet despite large-scale changes in protein expression associated with sensing, adaptation, and processing different environmental nutrients, simple growth laws connect the ribosome abundance and the growth rate. Here, we investigate the origin of these growth laws by analyzing the features of ribosomal regulation that coordinate proteome-wide expression changes with cell growth in a variety of nutrient conditions in the model organism Escherichia coli. We identify supply-driven feedforward activation of ribosomal protein synthesis as the key regulatory motif maximizing amino acid flux, and autonomously guiding a cell to achieve optimal growth in different environments. The growth laws emerge naturally from the robust regulatory strategy underlying growth rate control, irrespective of the details of the molecular implementation. The study highlights the interplay between phenomenological modeling and molecular mechanisms in uncovering fundamental operating constraints, with implications for endogenous and synthetic design of microorganisms. © 2014 The Authors. Published under the terms of the CC BY 4.0 license.

Discovery of potent α-glucosidase inhibitor flavonols: Insights into mechanism of action through inhibition kinetics and docking simulations.

PubMed

Şöhretoğlu, Didem; Sari, Suat; Barut, Burak; Özel, Arzu

2018-05-17

Beside other pharmaceutical benefits, flavonoids are known for their potent α-glucosidase inhibition. In the present study, we investigated α-glucosidase inhibitory effects of structurally related 11 flavonols, among which quercetin-3-O-(3″-O-galloyl)-β-galactopyranoside (8) and quercetin 3-O-(6″-O-galloyl)-β-glucopyranoside (9) showed significant inhibition compared to the positive control, acarbose, with IC 50 values of 0.97 ± 0.02 and 1.35 ± 0.06 µM, respectively. It was found that while sugar substitution to C3-OH of C ring reduced the α-glucosidase inhibitory effect, galloyl substitution to these sugar units increased it. An enzyme kinetics analysis revealed that 7 was competitive, whereas 1, 2, 8, and 9 were uncompetitive inhibitors. In the light of these findings, we performed molecular docking studies to predict their inhibition mechanisms at atomic level. Copyright © 2018 Elsevier Inc. All rights reserved.

Antibacterial, antioxidant, anti-cholinesterase potential and flavonol glycosides of Biscutella raphanifolia (Brassicaceae).

PubMed

Boudouda, Houria Berhail; Zeghib, Assia; Karioti, Anastazia; Bilia, Anna Rita; Öztürk, Mehmet; Aouni, Mahjoub; Kabouche, Ahmed; Kabouche, Zahia

2015-01-01

Different extracts of the aerial parts of Biscutella raphanifolia (Brassicaceae), which has not been the subject of any study, were screened for the phytochemical content, anti-microbial, antioxidant and anti-cholinesterase activities. We used four methods to identify the antioxidant activity namely, ABTS(•+), DPPH• scavenging, CUPRAC and ferrous-ions chelating methods. Since there is a relationship between antioxidants and cholinesterase enzyme inhibitors, we used two methods to determine the in vitro anti-cholinesterase activity by the use of the basic enzymes that occur in causing Alzheimer's disease: acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). The extracts were also tested in vitro antimicrobial activity against various bacteria. The phytochemical study of B. raphanifolia afforded four flavonol glycosides; namely, quercetin-3-O-β-D-g1ucoside, quercetin-3-O-[β-D-glucosyl(1→2)-O-β-D-glucoside], quercetin-3-O-[β-D-glucosyl(1→3)-O-β-D-glucoside] and kaempferol-3-O-[β-D-glucosyl(1→2)-[(6'''p-coumaroyl)- β-D-glucoside], being isolated here for the first time from Biscutella raphanifolia and the genus. The ethyl acetate extract showed the highest activity in ABTS(•+), DPPH• and CUPRAC assays, while the petroleum ether extract demonstrated optimum efficiency metal chelating activity. The dicloromethane and petroleum ether extracts showed a mild inhibition against AChE and BChE. However, the petroleum ether extract showed a good antibacterial activity against the pathovars Enteropathogenic E. coli (EPEC), Enterotoxigenic E. coli (ETEC) and Enterococcus feacalis, whereas the Enterohemorrhagic E. coli (EHEC) strain was more sensitive to dichloromethane and n-butanol extracts.

Heterogeneous ribonucleoprotein R regulates arylalkylamine N-acetyltransferase synthesis via internal ribosomal entry site-mediated translation in a circadian manner.

PubMed

Lee, Hwa-Rim; Kim, Tae-Don; Kim, Hyo-Jin; Jung, Youngseob; Lee, Dohyun; Lee, Kyung-Ha; Kim, Do-Yeon; Woo, Kyung-Chul; Kim, Kyong-Tai

2015-11-01

Rhythmic arylalkylamine N-acetyltransferase (AANAT) synthesis is a prominent circadian-controlled response that occurs in most mammals. AANAT is the core enzyme in melatonin production; because melatonin participates in many physiological processes, the regulation of AANAT is an important research topic. In this study, we focused on the role of heterogeneous ribonucleoprotein R (hnRNP R) in the translation of AANAT. A novel RNA-binding protein hnRNP R widely interacted with the 5' untranslated region (UTR) of AANAT mRNA and contributed to translation through an internal ribosomal entry site (IRES). Fine-tuning of AANAT protein synthesis occurred in response to knockdown and overexpression of hnRNP R. Nocturnal elevation of AANAT protein was dependent on the rhythmic changes of hnRNP R, whose levels are elevated in the pineal gland during nighttime. Increases in hnRNP R additionally improved AANAT production in rat pinealocytes under norepinephrine (NE) treatment. These results suggest that cap-independent translation of AANAT mRNA plays a role in the rhythmic synthesis of melatonin through the recruitment of translational machinery to hnRNP R-bound AANAT mRNA. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Functional dynamics within the human ribosome regulate the rate of active protein synthesis

PubMed Central

Ferguson, Angelica; Wang, Leyi; Altman, Roger B.; Terry, Daniel S.; Juette, Manuel F.; Burnett, Benjamin J.; Alejo, Jose L.; Dass, Randall A.; Parks, Matthew M.; Vincent, Theresa C.; Blanchard, Scott C.

2015-01-01

SUMMARY The regulation of protein synthesis contributes to gene expression in both normal physiology and disease, yet kinetic investigations of the human translation mechanism are currently lacking. Using single-molecule fluorescence imaging methods, we have quantified the nature and timing of structural processes in human ribosomes during single-turnover and processive translation reactions. These measurements reveal that functional complexes exhibit dynamic behaviors and thermodynamic stabilities distinct from those observed for bacterial systems. Structurally defined sub-states of pre- and post-translocation complexes were sensitive to specific inhibitors of the eukaryotic ribosome demonstrating the utility of this platform to probe drug mechanism. The application of three-color single-molecule FRET methods further revealed a long-distance allosteric coupling between distal tRNA binding sites within ribosomes bearing three tRNAs, which contributed to the rate of processive translation. PMID:26593721

Flavonoid Accumulation Patterns of Transparent Testa Mutants of Arabidopsis1

PubMed Central

Peer, Wendy Ann; Brown, Dana E.; Tague, Brian W.; Muday, Gloria K.; Taiz, Lincoln; Murphy, Angus S.

2001-01-01

Flavonoids have been implicated in the regulation of auxin movements in Arabidopsis. To understand when and where flavonoids may be acting to control auxin movement, the flavonoid accumulation pattern was examined in young seedlings and mature tissues of wild-type Arabidopsis. Using a variety of biochemical and visualization techniques, flavonoid accumulation in mature plants was localized in cauline leaves, pollen, stigmata, and floral primordia, and in the stems of young, actively growing inflorescences. In young Landsberg erecta seedlings, aglycone flavonols accumulated developmentally in three regions, the cotyledonary node, the hypocotyl-root transition zone, and the root tip. Aglycone flavonols accumulated at the hypocotyl-root transition zone in a developmental and tissue-specific manner with kaempferol in the epidermis and quercetin in the cortex. Quercetin localized subcellularly in the nuclear region, plasma membrane, and endomembrane system, whereas kaempferol localized in the nuclear region and plasma membrane. The flavonoid accumulation pattern was also examined in transparent testa mutants blocked at different steps in the flavonoid biosynthesis pathway. The transparent testa mutants were shown to have precursor accumulation patterns similar to those of end product flavonoids in wild-type Landsberg erecta, suggesting that synthesis and end product accumulation occur in the same cells. PMID:11402185

Flavonoid accumulation patterns of transparent testa mutants of arabidopsis

NASA Technical Reports Server (NTRS)

Peer, W. A.; Brown, D. E.; Tague, B. W.; Muday, G. K.; Taiz, L.; Murphy, A. S.

2001-01-01

Flavonoids have been implicated in the regulation of auxin movements in Arabidopsis. To understand when and where flavonoids may be acting to control auxin movement, the flavonoid accumulation pattern was examined in young seedlings and mature tissues of wild-type Arabidopsis. Using a variety of biochemical and visualization techniques, flavonoid accumulation in mature plants was localized in cauline leaves, pollen, stigmata, and floral primordia, and in the stems of young, actively growing inflorescences. In young Landsberg erecta seedlings, aglycone flavonols accumulated developmentally in three regions, the cotyledonary node, the hypocotyl-root transition zone, and the root tip. Aglycone flavonols accumulated at the hypocotyl-root transition zone in a developmental and tissue-specific manner with kaempferol in the epidermis and quercetin in the cortex. Quercetin localized subcellularly in the nuclear region, plasma membrane, and endomembrane system, whereas kaempferol localized in the nuclear region and plasma membrane. The flavonoid accumulation pattern was also examined in transparent testa mutants blocked at different steps in the flavonoid biosynthesis pathway. The transparent testa mutants were shown to have precursor accumulation patterns similar to those of end product flavonoids in wild-type Landsberg erecta, suggesting that synthesis and end product accumulation occur in the same cells.

Fitness advantages conferred by the L20-interacting RNA cis-regulator of ribosomal protein synthesis in Bacillus subtilis.

PubMed

Babina, Arianne M; Parker, Darren J; Li, Gene-Wei; Meyer, Michelle M

2018-06-20

In many bacteria, ribosomal proteins autogenously repress their own expression by interacting with RNA structures typically located in the 5'-UTRs of their mRNA transcripts. This regulation is necessary to maintain a balance between ribosomal proteins and rRNA to ensure proper ribosome production. Despite advances in non-coding RNA discovery and validation of RNA-protein regulatory interactions, the selective pressures that govern the formation and maintenance of such RNA cis-regulators in the context of an organism remain largely undetermined. To examine the impact disruptions to this regulation have on bacterial fitness, we introduced point mutations that abolish ribosomal protein binding and regulation into the RNA structure that controls expression of ribosomal proteins L20 and L35 within the Bacillus subtilis genome. Our studies indicate that removing this regulation results in reduced log phase growth, improper rRNA maturation, and the accumulation of a kinetically trapped or mis-assembled ribosomal particle at low temperatures, suggesting defects in ribosome synthesis. Such work emphasizes the important role regulatory RNAs play in the stoichiometric production of ribosomal components for proper ribosome composition and overall organism viability and reinforces the potential of targeting ribosomal protein production and ribosome assembly with novel antimicrobials. Published by Cold Spring Harbor Laboratory Press for the RNA Society.

Oxygen tension limits nitric oxide synthesis by activated macrophages.

PubMed Central

McCormick, C C; Li, W P; Calero, M

2000-01-01

Previous studies have established that constitutive calcium-dependent ('low-output') nitric oxide synthase (NOS) is regulated by oxygen tension. We have investigated the role of oxygen tension in the synthesis of NO by the 'high-output' calcium-independent NOS in activated macrophages. Hypoxia increased macrophage NOS gene expression in the presence of one additional activator, such as lipopolysaccharide or interferon-gamma, but not in the presence of both. Hypoxia markedly reduced the synthesis of NO by activated macrophages (as measured by accumulation of nitrite and citrulline), such that, at 1% oxygen tension, NO accumulation was reduced by 80-90%. The apparent K(m) for oxygen calculated from cells exposed to a range of oxygen tensions was found to be 10.8%, or 137 microM, O(2) This value is considerably higher than the oxygen tension in tissues, and is virtually identical to that reported recently for purified recombinant macrophage NOS. The decrease in NO synthesis did not appear to be due to diminished arginine or cofactor availability, since arginine transport and NO synthesis during recovery in normoxia were normal. Analysis of NO synthesis during hypoxia as a function of extracellular arginine indicated that an altered V(max), but not K(m)(Arg), accounted for the observed decrease in NO synthesis. We conclude that oxygen tension regulates the synthesis of NO in macrophages by a mechanism similar to that described previously for the calcium-dependent low-output NOS. Our data suggest that oxygen tension may be an important physiological regulator of macrophage NO synthesis in vivo. PMID:10970783

Identification of the Main Regulator Responsible for Synthesis of the Typical Yellow Pigment Produced by Trichoderma reesei

PubMed Central

Derntl, Christian; Rassinger, Alice; Srebotnik, Ewald; Mach, Robert L.

2016-01-01

ABSTRACT The industrially used ascomycete Trichoderma reesei secretes a typical yellow pigment during cultivation, while other Trichoderma species do not. A comparative genomic analysis suggested that a putative secondary metabolism cluster, containing two polyketide-synthase encoding genes, is responsible for the yellow pigment synthesis. This cluster is conserved in a set of rather distantly related fungi, including Acremonium chrysogenum and Penicillium chrysogenum. In an attempt to silence the cluster in T. reesei, two genes of the cluster encoding transcription factors were individually deleted. For a complete genetic proof-of-function, the genes were reinserted into the genomes of the respective deletion strains. The deletion of the first transcription factor (termed yellow pigment regulator 1 [Ypr1]) resulted in the full abolishment of the yellow pigment formation and the expression of most genes of this cluster. A comparative high-pressure liquid chromatography (HPLC) analysis of supernatants of the ypr1 deletion and its parent strain suggested the presence of several yellow compounds in T. reesei that are all derived from the same cluster. A subsequent gas chromatography/mass spectrometry analysis strongly indicated the presence of sorbicillin in the major HPLC peak. The presence of the second transcription factor, termed yellow pigment regulator 2 (Ypr2), reduces the yellow pigment formation and the expression of most cluster genes, including the gene encoding the activator Ypr1. IMPORTANCE Trichoderma reesei is used for industry-scale production of carbohydrate-active enzymes. During growth, it secretes a typical yellow pigment. This is not favorable for industrial enzyme production because it makes the downstream process more complicated and thus increases operating costs. In this study, we demonstrate which regulators influence the synthesis of the yellow pigment. Based on these data, we also provide indication as to which genes are under the control

Regulation of Hemoglobin β-Chain Synthesis in Bone Marrow Erythroid Cells by α Chains

PubMed Central

Wolf, Jeffrey L.; Mason, R. George; Honig, George R.

1973-01-01

Synthesis of α and β chains of hemoglobin was studied in vitro in intact reticulocytes and bone marrow cells. The cells were from rabbits having a variant form of hemoglobin in which L-isoleucine is in the α but not in the β chains. This characteristic permitted a selective inhibition of α-chain synthesis to be produced by addition to the incubation medium of L-O-methylthreonine, an inhibitor of protein synthesis that is a specific antagonist of L-isoleucine. In studies with reticulocytes, 25 mM L-O-methylthreonine produced a 60-70% inhibition of α-chain synthesis, but β-chain synthesis was unaffected even after incubation times for 4 hr. Because reticulocytes contain a pool of uncombined α chains which might have obscured the demonstration of an α chain-dependent mechanism for β-chain synthesis, subsequent studies were done with bone marrow cells. The latter had little or no detectable α-chain pool. A substantial inhibition of α-chain synthesis by the bone marrow cells was produced by the isoleucine antagonist but was also accompanied by a significantly decreased rate of β-chain synthesis. These findings suggest that the coordinated synthesis of the complementary α- and β-globin chains of hemoglobin may reflect in part a modifying effect of α-chain synthesis on the synthesis of β chains. PMID:4519634

Dendritic protein synthesis in the normal and diseased brain

PubMed Central

Swanger, Sharon A.; Bassell, Gary J.

2015-01-01

Synaptic activity is a spatially-limited process that requires a precise, yet dynamic, complement of proteins within the synaptic micro-domain. The maintenance and regulation of these synaptic proteins is regulated, in part, by local mRNA translation in dendrites. Protein synthesis within the postsynaptic compartment allows neurons tight spatial and temporal control of synaptic protein expression, which is critical for proper functioning of synapses and neural circuits. In this review, we discuss the identity of proteins synthesized within dendrites, the receptor-mediated mechanisms regulating their synthesis, and the possible roles for these locally synthesized proteins. We also explore how our current understanding of dendritic protein synthesis in the hippocampus can be applied to new brain regions and to understanding the pathological mechanisms underlying varied neurological diseases. PMID:23262237

Detoxification of polycyclic aromatic hydrocarbons (PAHs) in Arabidopsis thaliana involves a putative flavonol synthase.

PubMed

Hernández-Vega, Juan C; Cady, Brian; Kayanja, Gilbert; Mauriello, Anthony; Cervantes, Natalie; Gillespie, Andrea; Lavia, Lisa; Trujillo, Joshua; Alkio, Merianne; Colón-Carmona, Adán

2017-01-05

Polycyclic aromatic hydrocarbons (PAHs) are environmental contaminants with cytotoxic, teratogenic and carcinogenic properties. Bioremediation studies with bacteria have led to the identification of dioxygenases (DOXs) in the first step to degrade these recalcitrant compounds. In this study, we characterized the role of the Arabidopsis thaliana AT5G05600, a putative DOX of the flavonol synthase family, in the transformation of PAHs. Phenotypic analysis of loss-of-function mutant lines showed that these plant lines were less sensitive to the toxic effects of phenanthrene, suggesting possible roles of this gene in PAH degradation in vivo. Interestingly, these mutant lines showed less accumulation of H 2 O 2 after PAH exposure. Transgenic lines over-expressing At5g05600 showed a hypersensitive response and more oxidative stress after phenanthrene treatments. Moreover, fluorescence spectra results of biochemical assays with the recombinant His-tagged protein AT5G05600 detected chemical modifications of phenanthrene. Taken together, these results support the hypothesis that AT5G05600 is involved in the catabolism of PAHs and the accumulation of toxic intermediates during PAH biotransformation in plants. This research represents the first step in the design of transgenic plants with the potential to degrade PAHs, leading to the development of vigorous plant varieties that can reduce the levels of these pollutants in the environment. Copyright © 2016 Elsevier B.V. All rights reserved.

Regiospecific Methylation of a Dietary Flavonoid Scaffold Selectively Enhances IL-1β Production following Toll-like Receptor 2 Stimulation in THP-1 Monocytes*

PubMed Central

Lim, Eng-Kiat; Mitchell, Paul J.; Brown, Najmeeyah; Drummond, Rebecca A.; Brown, Gordon D.; Kaye, Paul M.; Bowles, Dianna J.

2013-01-01

It is now recognized that innate immunity to intestinal microflora plays a significant role in mediating immune health, and modulation of microbial sensing may underpin the impact of plant natural products in the diet or when used as nutraceuticals. In this context, we have examined five classes of plant-derived flavonoids (flavonols, flavones, flavanones, catechins, and cyanidin) for their ability to regulate cytokine release induced by the Toll-like receptor 2 (TLR2) agonist Pam3CSK4. We found that the flavonols selectively co-stimulated IL-1β secretion but had no impact on the secretion of IL-6. Importantly, this costimulation of TLR2-induced cytokine secretion was dependent on regiospecific methylation of the flavonol scaffold with a rank order of quercetin-3,4′-dimethylether > quercetin-3-methylether > casticin. The mechanism underpinning this costimulation did not involve enhanced inflammasome activation. In contrast, the methylated flavonols enhanced IL-1β gene expression through transcriptional regulation, involving mechanisms that operate downstream of the initial NF-κB and STAT1 activation events. These studies demonstrate an exquisite level of control of scaffold bioactivity by regiospecific methylation, with important implications for understanding how natural products affect innate immunity and for their development as novel immunomodulators for clinical use. PMID:23760261

Saccharomyces Cerevisiae Cho2 Mutants Are Deficient in Phospholipid Methylation and Cross-Pathway Regulation of Inositol Synthesis

PubMed Central

Summers, E. F.; Letts, V. A.; McGraw, P.; Henry, S. A.

1988-01-01

Five allelic Saccharomyces cerevisiae mutants deficient in the methylation of phosphatidylethanolamine (PE) have been isolated, using two different screening techniques. Biochemical analysis suggested that these mutants define a locus, designated CHO2, that may encode a methyltransferase. Membranes of cho2 mutant cells grown in defined medium contain approximately 10% phosphatidylcholine (PC) and 40-50% PE as compared to wild-type levels of 40-45% PC and 15-20% PE. In spite of this greatly altered phospholipid composition, cho2 mutant cells are viable in defined medium and are not auxotrophic for choline or other phospholipid precursors such as monomethylethanolamine (MME). However, analysis of yeast strains carrying more than one mutation affecting phospholipid biosynthesis indicated that some level of methylated phospholipid is essential for viability. The cho2 locus was shown by tetrad analysis to be unlinked to other loci affecting phospholipid synthesis. Interestingly, cho2 mutants and other mutant strains that produce reduced levels of methylated phospholipids are unable to properly repress synthesis of the cytoplasmic enzyme inositol-1-phosphate synthase. This enzyme was previously shown to be regulated at the level of mRNA abundance in response to inositol and choline in the growth medium. We cloned the CHO2 gene on a 3.6-kb genomic DNA fragment and created a null allele of cho2 by disrupting the CHO2 gene in vivo. The cho2 disruptant, like all other cho2 mutants, is viable, exhibits altered regulation of inositol biosynthesis and is not auxotrophic for choline or MME. PMID:3066687

THE SYNTHESIS OF THE STARCH GRANULE.

PubMed

Smith, A. M.; Denyer, K.; Martin, C.

1997-06-01

This review describes and discusses the implications of recent discoveries about how starch polymers are synthesized and organized to form a starch granule. Three issues are highlighted. 1. The role and importance of ADPglucose pyrophosphorylase in the generation of ADPglucose as the substrate for polymer synthesis. 2. The contributions of isoforms of starch-branching enzyme, starch synthase, and debranching enzyme to the synthesis and ordered packing of amylopectin molecules. 3. The requirements for and regulation of the synthesis of amylose.

Flavonol Glycosides in Currant Leaves and Variation with Growth Season, Growth Location, and Leaf Position.

PubMed

Yang, Wei; Alanne, Aino-Liisa; Liu, Pengzhan; Kallio, Heikki; Yang, Baoru

2015-10-28

Flavonol glycosides (FG) were analyzed in the leaves of six currant cultivars (Ribes spp.) with HPLC-DAD, HPLC-MS/MS, and NMR. The average amounts of the 12 major, identified FG constituted 86-93% (9.6-14.1 mg/g DW) of the total of 27 FG found. Quercetin and kaempferol were the major aglycones with trace amounts of myricetin. Quercetin-3-O-(2,6-α-dirhamnopyranosyl-β-glucopyranoside), quercetin-3-O-(2-β-xylopyranosyl-6-α-rhamnopyranosyl-β-glucopyranoside), and kaempferol-3-O-(3,6-α-dirhamnopyranosyl-β-glucopyranoside) were identified for the first time in currant leaves and existed in a white currant cultivar 'White Dutch' only. Kaempferol-3-O-β-(6'-malonyl)glucopyranoside was also a new compound existing in abundance in five cultivars but not in the white one. The results show the primary importance of the genetic background of the cultivars. The content of malonylated FG of special importance in cardiovascular health decreased regularly during summer. Time of collection and leaf position were more prominent factors affecting the composition than were the year of harvest or the growth latitude. Randomly collected leaves differed in their FG profiles from those collected from the middle position of new branches.

Low and moderate photosynthetically active radiation affects the flavonol glycosides and hydroxycinnamic acid derivatives in kale (Brassica oleracea var. sabellica) dependent on two low temperatures.

PubMed

Neugart, Susanne; Fiol, Michaela; Schreiner, Monika; Rohn, Sascha; Zrenner, Rita; Kroh, Lothar W; Krumbein, Angelika

2013-11-01

Kale (Brassica oleracea var. sabellica) contains a large number of naturally occurring structurally different non-acylated and acylated flavonol glycosides as well as hydroxycinnamic acid derivatives. The objective of this study was to determine the effect of low and moderate photosynthetic active radiation (PAR) and how these levels interact with low temperature in these phenolic compounds. Juvenile kale plants were treated with PAR levels from 200 to 800 μmol m(-2) s(-1) at 5 and 10 °C under defined conditions in climate chambers. Of the investigated 20 compounds, 11 and 17 compounds were influenced by PAR and temperature, respectively. In addition, an interaction between PAR and temperature was found for eight compounds. The response of the phenolic compounds to PAR was structure-dependent. While quercetin triglycosides increased with higher PAR at 5 and 10 °C, the kaempferol triglycosides exhibited the highest concentrations at 400 μmol m(-2) s(-1). In contrast, kaempferol diglycosides exhibited the highest concentrations at increased PAR levels of 600 and 800 μmol m(-2) s(-1) at 10 °C. However, key genes of flavonol biosynthesis were influenced by temperature but remained unaffected by PAR. Furthermore, there was no interaction between the PAR level and the low temperature in the response of hydroxycinnamic acid derivatives in kale with the exception of caffeoylquinic acid, which decreased with higher PAR levels of 600 and 800 μmol m(-2) s(-1) and at a lower temperature. In conclusion, PAR and its interaction with temperature could be a suitable tool for modifying the profile of phenolic compounds. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Functional Dynamics within the Human Ribosome Regulate the Rate of Active Protein Synthesis.

PubMed

Ferguson, Angelica; Wang, Leyi; Altman, Roger B; Terry, Daniel S; Juette, Manuel F; Burnett, Benjamin J; Alejo, Jose L; Dass, Randall A; Parks, Matthew M; Vincent, C Theresa; Blanchard, Scott C

2015-11-05

The regulation of protein synthesis contributes to gene expression in both normal physiology and disease, yet kinetic investigations of the human translation mechanism are currently lacking. Using single-molecule fluorescence imaging methods, we have quantified the nature and timing of structural processes in human ribosomes during single-turnover and processive translation reactions. These measurements reveal that functional complexes exhibit dynamic behaviors and thermodynamic stabilities distinct from those observed for bacterial systems. Structurally defined sub-states of pre- and post-translocation complexes were sensitive to specific inhibitors of the eukaryotic ribosome, demonstrating the utility of this platform to probe drug mechanism. The application of three-color single-molecule fluorescence resonance energy transfer (smFRET) methods further revealed a long-distance allosteric coupling between distal tRNA binding sites within ribosomes bearing three tRNAs, which contributed to the rate of processive translation. Copyright © 2015 Elsevier Inc. All rights reserved.

Regulation of ribonucleic acid synthesis by polyamines. Reversal by spermine of inhibition by methylglyoxal bis(guanylhydrazone) of ribonucleic acid synthesis and histone acetylation in rabbit heart.

PubMed

Caldarera, C M; Casti, A; Guarnier, C; Moruzzi, G

1975-10-01

The relationship between polyamines and RNA synthesis was studied by considering the action of spermine on histone acetylation in perfused heart. In addition, the effect of methylglyoxal bis(guanylhydrazone), inhibitor of putrescine-activated S-adenosylmethionine decarboxylase activity, on RNA and polyamine specific radioactivity and on acetylation of histone fractions was also investigated in perfused heart. Different concentrations of spermine and/or methylglyoxas bis(guanylhydrazone) were injected into the heart, 15 min after beginning the perfusion. The results demonstrate that spermine stimulates the specific radioactivity of RNA of subcellular fractions. Acetylation of the arginine-rich histone fractions, involved in the regulation of RNA transcription, is enhanced by spermine. The perfusion with methylglyoxal bis(guanylhydrazone) causes a decrease in the specific radioactivity of polyamines and RNA, and in acetylation of histone fractions. However, spermine is able to reverse the methylglyoxal bis(guanylhydrazone) inhibition when injected simultaneously. From these results we may assume a possible role for spermine in the regulation of RNA transcription.

Regulation of ribonucleic acid synthesis by polyamines. Reversal by spermine of inhibition by methylglyoxal bis(guanylhydrazone) of ribonucleic acid synthesis and histone acetylation in rabbit heart.

PubMed Central

Caldarera, C M; Casti, A; Guarnier, C; Moruzzi, G

1975-01-01

The relationship between polyamines and RNA synthesis was studied by considering the action of spermine on histone acetylation in perfused heart. In addition, the effect of methylglyoxal bis(guanylhydrazone), inhibitor of putrescine-activated S-adenosylmethionine decarboxylase activity, on RNA and polyamine specific radioactivity and on acetylation of histone fractions was also investigated in perfused heart. Different concentrations of spermine and/or methylglyoxas bis(guanylhydrazone) were injected into the heart, 15 min after beginning the perfusion. The results demonstrate that spermine stimulates the specific radioactivity of RNA of subcellular fractions. Acetylation of the arginine-rich histone fractions, involved in the regulation of RNA transcription, is enhanced by spermine. The perfusion with methylglyoxal bis(guanylhydrazone) causes a decrease in the specific radioactivity of polyamines and RNA, and in acetylation of histone fractions. However, spermine is able to reverse the methylglyoxal bis(guanylhydrazone) inhibition when injected simultaneously. From these results we may assume a possible role for spermine in the regulation of RNA transcription. PMID:1212228

Simultaneous characterisation and quantitation of flavonol glycosides and aglycones in noni leaves using a validated HPLC-UV/MS method.

PubMed

Deng, Shixin; West, Brett J; Jensen, C Jarakae

2008-11-15

The leaves of Morinda citrifolia L. (noni) have been utilized in a variety of commercial products marketed for their health benefits. This paper reports on a rapid and selective HPLC method for simultaneous characterization and quantitation of four flavonols in an ethanolic extract of noni leaves by using dual detectors of UV (365nm) and ESI-MS (negative mode). The limits of detection and quantitation were between 0.012 and 0.165μg/mL. The intra- and inter-assay precisions, in terms of percent relative standard deviation, are less than 4.38% and 3.50%, respectively. The accuracy, in terms of recovery percentage, ranged from 96.66% to 100.03%. Good linearity (correlation coefficient >0.999) for each calibration curve of standards was achieved in the range investigated. The contents of four flavonoids in the noni leaves varied from 1.16 to 371.6mg/100g dry weight. Copyright © 2008 Elsevier Ltd. All rights reserved.

Microfluidic Synthesis of Hybrid Nanoparticles with Controlled Lipid Layers: Understanding Flexibility-Regulated Cell-Nanoparticle Interaction.

PubMed

Zhang, Lu; Feng, Qiang; Wang, Jiuling; Zhang, Shuai; Ding, Baoquan; Wei, Yujie; Dong, Mingdong; Ryu, Ji-Young; Yoon, Tae-Young; Shi, Xinghua; Sun, Jiashu; Jiang, Xingyu

2015-10-27

The functionalized lipid shell of hybrid nanoparticles plays an important role for improving their biocompatibility and in vivo stability. Yet few efforts have been made to critically examine the shell structure of nanoparticles and its effect on cell-particle interaction. Here we develop a microfluidic chip allowing for the synthesis of structurally well-defined lipid-polymer nanoparticles of the same sizes, but covered with either lipid-monolayer-shell (MPs, monolayer nanoparticles) or lipid-bilayer-shell (BPs, bilayer nanoparticles). Atomic force microscope and atomistic simulations reveal that MPs have a lower flexibility than BPs, resulting in a more efficient cellular uptake and thus anticancer effect than BPs do. This flexibility-regulated cell-particle interaction may have important implications for designing drug nanocarriers.

Jasmonate Regulates Plant Responses to Postsubmergence Reoxygenation through Transcriptional Activation of Antioxidant Synthesis1

PubMed Central

Dai, Yang-Shuo; Xie, Li-Juan; Yu, Lu-Jun; Zhou, Ying; Lai, Yong-Xia; Yang, Yi-Cong; Xu, Le; Chen, Qin-Fang

2017-01-01

Submergence induces hypoxia in plants; exposure to oxygen following submergence, termed reoxygenation, produces a burst of reactive oxygen species. The mechanisms of hypoxia sensing and signaling in plants have been well studied, but how plants respond to reoxygenation remains unclear. Here, we show that reoxygenation in Arabidopsis (Arabidopsis thaliana) involves rapid accumulation of jasmonates (JAs) and increased transcript levels of JA biosynthesis genes. Application of exogenous methyl jasmonate improved tolerance to reoxygenation in wild-type Arabidopsis; also, mutants deficient in JA biosynthesis and signaling were very sensitive to reoxygenation. Moreover, overexpression of the transcription factor gene MYC2 enhanced tolerance to posthypoxic stress, and myc2 knockout mutants showed increased sensitivity to reoxygenation, indicating that MYC2 functions as a key regulator in the JA-mediated reoxygenation response. MYC2 transcriptionally activates members of the VITAMIN C DEFECTIVE (VTC) and GLUTATHIONE SYNTHETASE (GSH) gene families, which encode rate-limiting enzymes in the ascorbate and glutathione synthesis pathways. Overexpression of VTC1 and GSH1 in the myc2-2 mutant suppressed the posthypoxic hypersensitive phenotype. The JA-inducible accumulation of antioxidants may alleviate oxidative damage caused by reoxygenation, improving plant survival after submergence. Taken together, our findings demonstrate that JA signaling interacts with the antioxidant pathway to regulate reoxygenation responses in Arabidopsis. PMID:28082717

Transcriptomics, Targeted Metabolomics and Gene Expression of Blackberry Leaves and Fruits Indicate Flavonoid Metabolic Flux from Leaf to Red Fruit

PubMed Central

Gutierrez, Enrique; García-Villaraco, Ana; Lucas, José A.; Gradillas, Ana; Gutierrez-Mañero, F. Javier; Ramos-Solano, Beatriz

2017-01-01

Blackberries (Rubus spp.) are among the high added value food products relevant for human health due to the increasing evidence of the beneficial effects of polyphenols, which are very abundant in these fruits. Interestingly, these compounds also play a role on plant physiology, being especially relevant their role in plant defense against biotic and abiotic stress. Hence, we hypothesize that since blackberry fruits have high amounts of flavonols and anthocyanins, leaves would also have high amounts of these compounds, and can be studied as a source of active molecules; furthermore, leaf synthesis would support their high contents in fruits. To explore this hypothesis, the present study reports a de novo transcriptome analysis on field grown blackberry leaves and fruits at the same time point, to establish the metabolic relationship of these compounds in both organs. Transcripts were aligned against Fragaria vesca genome, and genes were identified and annotated in different databases; tissue expression pattern showed 20,463 genes common to leaves and fruits, while 6,604 genes were significantly overexpressed only in fruits, while another 6,599 genes were significantly overexpressed in leaves, among which flavonol-anthocyanin transporter genes were present. Bioactives characterization indicated that total phenolics in leaves were three-fold, and flavonols were six-fold than in fruits, while concentration of anthocyanins was higher in fruits; HPLC-MS analysis indicated different composition in leaves and fruits, with cyanidin-3-glucoside as the only common compound identified. Next, RT-qPCR of the core genes in the flavonol anthocyanin pathway and regulatory MYB genes were carried out. Interestingly, genes in the flavonol-anthocyanin pathway and flavonol-transport families were overexpressed in leaves, consistent with the higher bioactive levels. On the other hand, transcription factors were overexpressed in fruits anticipating an active anthocyanin biosynthesis upon

Seedless Synthesis of Monodispersed Gold Nanorods with Remarkably High Yield: Synergistic Effect of Template Modification and Growth Kinetics Regulation.

PubMed

Liu, Kang; Bu, Yanru; Zheng, Yuanhui; Jiang, Xuchuan; Yu, Aibing; Wang, Huanting

2017-03-08

Gold nanorods (AuNRs) are versatile materials due to their broadly tunable optical properties associated with their anisotropic feature. Conventional seed-mediated synthesis is, however, not only limited by the operational complexity and over-sensitivity towards subtle changes of experimental conditions but also suffers from low yield (≈15 %). A facile seedless method is reported to overcome these challenges. Monodispersed AuNRs with high yield (≈100 %) and highly adjustable longitudinal surface plasmon resonance (LSPR) are reproducibly synthesized. The parameters that influence the AuNRs growth were thoroughly investigated in terms of growth kinetics and soft-template regulation, offering a better understanding of the template-based mechanism. The facile synthesis, broad tunability of LSRP, high reproducibility, high yield, and ease of scale-up make this method promising for the future mass production of monodispersed AuNRs for applications in catalysis, sensing, and biomedicine. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Light-induced Variation in Phenolic Compounds in Cabernet Sauvignon Grapes (Vitis vinifera L.) Involves Extensive Transcriptome Reprogramming of Biosynthetic Enzymes, Transcription Factors, and Phytohormonal Regulators

PubMed Central

Sun, Run-Ze; Cheng, Guo; Li, Qiang; He, Yan-Nan; Wang, Yu; Lan, Yi-Bin; Li, Si-Yu; Zhu, Yan-Rong; Song, Wen-Feng; Zhang, Xue; Cui, Xiao-Di; Chen, Wu; Wang, Jun

2017-01-01

Light environments have long been known to influence grape (Vitis vinifera L.) berry development and biosynthesis of phenolic compounds, and ultimately affect wine quality. Here, the accumulation and compositional changes of hydroxycinnamic acids (HCAs) and flavonoids, as well as global gene expression were analyzed in Cabernet Sauvignon grape berries under sunlight exposure treatments at different phenological stages. Sunlight exposure did not consistently affect the accumulation of berry skin flavan-3-ol or anthocyanin among different seasons due to climatic variations, but increased HCA content significantly at véraison and harvest, and enhanced flavonol accumulation dramatically with its timing and severity degree trend. As in sunlight exposed berries, a highly significant correlation was observed between the expression of genes coding phenylalanine ammonia-lyase, 4-coumarate: CoA ligase, flavanone 3-hydroxylase and flavonol synthase family members and corresponding metabolite accumulation in the phenolic biosynthesis pathway, which may positively or negatively be regulated by MYB, bHLH, WRKY, AP2/EREBP, C2C2, NAC, and C2H2 transcription factors (TFs). Furthermore, some candidate genes required for auxin, ethylene and abscisic acid signal transductions were also identified which are probably involved in berry development and flavonoid biosynthesis in response to enhanced sunlight irradiation. Taken together, this study provides a valuable overview of the light-induced phenolic metabolism and transcriptome changes, especially the dynamic responses of TFs and signaling components of phytohormones, and contributes to the further understanding of sunlight-responsive phenolic biosynthesis regulation in grape berries. PMID:28469625

Jasmonate Hormone: Regulating Synthesis of Reduced Carbon Compounds in Plants

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

Browse, John

Our original interest in understanding the role of jasmonate (JA) in regulating the final stages of stamen and pollen development led to our discovery of the JAZ repressors, and the molecular mechanism of JA action is now a second important focus of our research. The specific goals for this grant period are to: 1. Investigate the generation and clearance of the hormone with emphasis on the regulation of the OPR3 enzyme and the hydrolysis of JA-Ile. 2. Use dominant-negative and overexpression constructs to explore the role of the MYC5 transcription factor in initiating and regulating JA responses. 3. Investigate specificmore » JAZ protein interactions that will help us to recognize and understand the extended network of processes, such as sulfur nutrition, that interface with JA signaling. The COI1 F-Box protein is a JA-Ile coreceptor and coi1 mutant plants lack JA responses. We have tested the possibility that sites of JA action can be probed by using tissue-specific promoters to drive expression of a COI1-YFP fusion protein in coi1 mutant plants deficient in stamen and pollen function. When we expressed COI1 behind a filament-specific promoter (from the DAD1 gene), filament elongation was restored but not anther dehiscence or pollen function. Three tapetum specific promoters, all failed to restore any of these three functions but, unexpectedly, a promoter active in the stomium and epidermal cells, restored both pollen function and anther dehiscence. Most importantly, our results demonstrate the power of promoter::COI1-YFP constructs in revealing the primary sites of JA-regulated gene expression that control developmental and other responses in neighboring tissues. We now plan to use this new tool to test current hypotheses about JA action in other organs of the plant. The MYC2, MYC3, and MYC4 proteins are the primary transcription factors initiating defense and root growth responses to JA signaling. However, transgenic plants overexpressing these proteins do not

Comprehensive evaluation of the flavonol anti-oxidants, alpha-glycosyl isoquercitrin and isoquercitrin, for genotoxic potential.

PubMed

Hobbs, Cheryl A; Koyanagi, Mihoko; Swartz, Carol; Davis, Jeffrey; Kasamoto, Sawako; Maronpot, Robert; Recio, Leslie; Hayashi, Shim-Mo

2018-03-01

Quercetin and its glycosides possess potential benefits to human health. Several flavonols are available to consumers as dietary supplements, promoted as anti-oxidants; however, incorporation of natural quercetin glycosides into food and beverage products has been limited by poor miscibility in water. Enzymatic conjugation of multiple glucose moieties to isoquercitrin to produce alpha-glycosyl isoquercitrin (AGIQ) enhances solubility and bioavailability. AGIQ is used in Japan as a food additive and has been granted generally recognized as safe (GRAS) status. However, although substantial genotoxicity data exist for quercetin, there is very little available data for AGIQ and isoquercitrin. To support expanded global marketing of food products containing AGIQ, comprehensive testing of genotoxic potential of AGIQ and isoquercitrin was conducted according to current regulatory test guidelines. Both chemicals tested positive in bacterial reverse mutation assays, and exposure to isoquercitrin resulted in chromosomal aberrations in CHO-WBL cells. All other in vitro mammalian micronucleus and chromosomal aberration assays, micronucleus and comet assays in male and female B6C3F1 mice and Sprague Dawley rats, and Muta™ Mouse mutation assays evaluating multiple potential target tissues, were negative for both chemicals. These results supplement existing toxicity data to further support the safe use of AGIQ in food and beverage products. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Genetically engineered flavonol enriched tomato fruit modulates chondrogenesis to increase bone length in growing animals.

PubMed

Choudhary, Dharmendra; Pandey, Ashutosh; Adhikary, Sulekha; Ahmad, Naseer; Bhatia, Chitra; Bhambhani, Sweta; Trivedi, Prabodh Kumar; Trivedi, Ritu

2016-02-26

Externally visible body and longitudinal bone growth is a result of proliferation of chondrocytes. In growth disorder, there is delay in the age associated increase in height. The present study evaluates the effect of extract from transgenic tomato fruit expressing AtMYB12 transcription factor on bone health including longitudinal growth. Constitutive expression of AtMYB12 in tomato led to a significantly enhanced biosynthesis of flavonoids in general and the flavonol biosynthesis in particular. Pre-pubertal ovary intact BALB/c mice received daily oral administration of vehicle and ethanolic extract of wild type (WT-TOM) and transgenic AtMYB12-tomato (MYB12-TOM) fruits for six weeks. Animal fed with MYB12-TOM showed no inflammation in hepatic tissues and normal sinusoidal Kupffer cell morphology. MYB12-TOM extract significantly increased tibial and femoral growth and subsequently improved the bone length as compared to vehicle and WT-TOM. Histomorphometry exhibited significantly wider distal femoral and proximal tibial growth plate, increased number and size of hypertrophic chondrocytes in MYB12-TOM which corroborated with micro-CT and expression of BMP-2 and COL-10, marker genes for hypertrophic cells. We conclude that metabolic reprogramming of tomato by AtMYB12 has the potential to improve longitudinal bone growth thus helping in achievement of greater peak bone mass during adolescence.

Leptin regulates MMP-2, TIMP-1 and collagen synthesis via p38 MAPK in HL-1 murine cardiomyocytes.

PubMed

Schram, Kristin; De Girolamo, Sabrina; Madani, Siham; Munoz, Diana; Thong, Farah; Sweeney, Gary

2010-12-01

A clear association between obesity and heart failure exists and a significant role for leptin, the product of the obese gene, has been suggested. One aspect of myocardial remodeling which characterizes heart failure is a disruption in the balance of extracellular matrix synthesis and degradation. Here we investigated the effects of leptin on matrix metalloproteinase (MMP) activity, tissue inhibitor of metalloproteinase (TIMP) expression, as well as collagen synthesis in HL-1 cardiac muscle cells. Gelatin zymographic analysis of MMP activity in conditioned media showed that leptin enhanced MMP-2 activity in a dose- and time-dependent manner. Leptin is known to stimulate phosphorylation of p38 MAPK in cardiac cells and utilization of the p38 MAPK inhibitor, SB203580, demonstrated that this kinase also plays a role in regulating several extracellular matrix components, such that inhibition of p38 MAPK signaling prevented the leptin-induced increase in MMP-2 activation. We also observed that leptin enhanced collagen synthesis determined by both proline incorporation and picrosirius red staining of conditioned media. Pro-collagen type-I and pro-collagen type-III expression, measured by real-time PCR and Western blotting were also increased by leptin, effects which were again attenuated by SB203580. In summary, these results demonstrate the potential for leptin to play a role in mediating myocardial ECM remodeling and that the p38 MAPK pathway plays an important role in mediating these effects.

Biomolecular Specificity Regulated Synthesis of Nanocatalysts and Heterointegration of Photosynthesis Nanodevices

DTIC Science & Technology

2016-01-01

TiO2-Au/Pd nanocomposite composed of P25 TiO2 nanoparticles and Au/Pd nanowheels is applied in benzimidazole synthesis as plasmon-enhanced...activity and selectivity. In this work, TiO2-Au/Pd nanocomposite composed of P25 TiO2 nanoparticles and Au/Pd nanowheels is applied in benzimidazole ...Pd Nanocomposite Photocatalyst for Tandem Synthesis of Benzimidazole 2.1 Approaches: Wet-chemical synthetic routes are explored to create

Relative roles of synthesis and degradation in regulating metallothionein accretion

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

Laurin, D.E.

1989-01-01

Decay kinetics of {sup 35}S-cysteine (cys) in metallothionein (MT) were used to simultaneously measure rates of MT synthesis and degradation in a HD11 chicken-macrophage cell-line. A reverse-phase (RP) high-performance liquid-chromatography procedure was used to purify 2 MT-isoforms from cytosols with approximately 94% purity. The medium that the macrophages were incubated in was validated to ensure that it contained enough unlabeled cys to adequately chase {sup 35}S-cys released by the degradation of labeled protein. The addition of Zn{sup 2+} and unlabeled cys to the medium did not change the fractional-rates of MT synthesis (FRS) and degradation (FRD). These measurements were alsomore » validated by showing that the measured fractional-rate of MT accretion closely approximated the difference between FRS and FRD. When macrophages were incubated in medium supplemented with 50 or 25 {mu}M Zn{sup 2+} the absolute-rate of MT synthesis (ARS) and FRD increased and decreases, respectively. When macrophages were incubated in medium supplemented with 20 or 10 {mu}M Cd{sup 2+}, the ARS increased but the FDR was not changed.« less

Prolyl hydroxylation regulates protein degradation, synthesis, and splicing in human induced pluripotent stem cell-derived cardiomyocytes

PubMed Central

Stoehr, Andrea; Yang, Yanqin; Patel, Sajni; Evangelista, Alicia M.; Aponte, Angel; Wang, Guanghui; Liu, Poching; Boylston, Jennifer; Kloner, Philip H.; Lin, Yongshun; Gucek, Marjan; Zhu, Jun; Murphy, Elizabeth

2016-01-01

Aims Protein hydroxylases are oxygen- and α-ketoglutarate-dependent enzymes that catalyse hydroxylation of amino acids such as proline, thus linking oxygen and metabolism to enzymatic activity. Prolyl hydroxylation is a dynamic post-translational modification that regulates protein stability and protein–protein interactions; however, the extent of this modification is largely uncharacterized. The goals of this study are to investigate the biological consequences of prolyl hydroxylation and to identify new targets that undergo prolyl hydroxylation in human cardiomyocytes. Methods and results We used human induced pluripotent stem cell-derived cardiomyocytes in combination with pulse-chase amino acid labelling and proteomics to analyse the effects of prolyl hydroxylation on protein degradation and synthesis. We identified 167 proteins that exhibit differences in degradation with inhibition of prolyl hydroxylation by dimethyloxalylglycine (DMOG); 164 were stabilized. Proteins involved in RNA splicing such as serine/arginine-rich splicing factor 2 (SRSF2) and splicing factor and proline- and glutamine-rich (SFPQ) were stabilized with DMOG. DMOG also decreased protein translation of cytoskeletal and sarcomeric proteins such as α-cardiac actin. We searched the mass spectrometry data for proline hydroxylation and identified 134 high confidence peptides mapping to 78 unique proteins. We identified SRSF2, SFPQ, α-cardiac actin, and cardiac titin as prolyl hydroxylated. We identified 29 prolyl hydroxylated proteins that showed a significant difference in either protein degradation or synthesis. Additionally, we performed next-generation RNA sequencing and showed that the observed decrease in protein synthesis was not due to changes in mRNA levels. Because RNA splicing factors were prolyl hydroxylated, we investigated splicing ± inhibition of prolyl hydroxylation and detected 369 alternative splicing events, with a preponderance of exon skipping. Conclusions This study

Dynamics in enzymatic protein complexes offer a novel principle for the regulation of melatonin synthesis in the human pineal gland.

PubMed

Maronde, Erik; Saade, Anastasia; Ackermann, Katrin; Goubran-Botros, Hany; Pagan, Cecile; Bux, Roman; Bourgeron, Thomas; Dehghani, Faramarz; Stehle, Jörg H

2011-08-01

Time of day is communicated to the body through rhythmic cues, including pineal gland melatonin synthesis, which is restricted to nighttime. Whereas in most rodents transcriptional regulation of the arylalkylamine N-acetyltransferase (Aanat) gene is essential for rhythmic melatonin synthesis, investigations into nonrodent mammalian species have shown post-transcriptional regulation to be of central importance, with molecular mechanisms still elusive. Therefore, human pineal tissues, taken from routine autopsies were allocated to four time-of-death groups (night/dawn/day/dusk) and analyzed for daytime-dependent changes in phosphorylated AANAT (p31T-AANAT) and in acetyl-serotonin-methyltransferase (ASMT) expression and activity. Protein content, intracellular localization, and colocalization of p31T-AANAT and ASMT were assessed, using immunoblotting, immunofluorescence, and immunoprecipitation techniques. Fresh sheep pineal gland preparations were used for comparative purposes. The amount of p31T-AANAT and ASMT proteins as well as their intracellular localization showed no diurnal variation in autoptic human and fresh sheep pineal glands. Moreover, in human and sheep pineal extracts, AANAT could not be dephosphorylated, which was at variance to data derived from rat pineal extracts. P31T-AANAT and ASMT were often found to colocalize in cellular rod-like structures that were also partly immunoreactive for the pinealocyte process-specific marker S-antigen (arrestin) in both, human and sheep pinealocytes. Protein-protein interaction studies with p31T-AANAT, ASMT, and S-antigen demonstrated a direct association and formation of robust complexes, involving also 14-3-3. This work provides evidence for a regulation principle for AANAT activity in the human pineal gland, which may not be based on a p31T-AANAT phosphorylation/dephosphorylation switch, as described for other mammalian species. © 2011 John Wiley & Sons A/S.

Dihydromyricetin induces mitochondria-mediated apoptosis in HepG2 cells through down-regulation of the Akt/Bad pathway.

PubMed

Zhang, Zhuangwei; Zhang, Huiqin; Chen, Shiyong; Xu, Yan; Yao, Anjun; Liao, Qi; Han, Liyuan; Zou, Zuquan; Zhang, Xiaohong

2017-02-01

The plant flavonol dihydromyricetin (DHM) was reported to induce apoptosis in human hepatocarcinoma HepG2 cells. This study was undertaken to elucidate the underlying molecular mechanism of action of DHM. In the study, DHM down-regulated Akt expression and its phosphorylation at Ser473, up-regulated the levels of mitochondrial proapoptotic proteins Bax and Bad, and inhibited the phosphorylation of Bad at Ser136 and Ser112. It also inhibited the expression of the antiapoptotic protein Bcl-2 and enhanced the cleavage and activation of caspase-3 as well as the degradation of its downstream target poly(ADP-ribose) polymerase. Our results for the first time suggest that DHM-induced apoptosis in HepG2 cells may come about by the inhibition of the Akt/Bad signaling pathway and stimulation of the mitochondrial apoptotic pathway. Dihydromyricetin may be a promising therapeutic medication for hepatocellular carcinoma. Copyright © 2017 Elsevier Inc. All rights reserved.

Flavonoids, alkali earth and rare earth elements affect germination of pecan pollen

USDA-ARS?s Scientific Manuscript database

The factors regulating pecan [Carya illinoinensis (Wangenh.) K. Koch] pollen grain germination on receptive stigmatic flower surfaces in vivo or in vitro in pollen viability assays are poorly understood. While there are many potential regulating factors, there is evidence for involvement of flavonol...

Sugar-mediated semidian oscillation of gene expression in the cassava storage root regulates starch synthesis

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

Jansson, Christer; Baguma, Yona; Sun, Chuanxin

Starch branching enzyme (SBE) activity in the cassava storage root exhibited a diurnal fluctuation, dictated by a transcriptional oscillation of the corresponding SBE genes. The peak of SBE activity coincided with the onset of sucrose accumulation in the storage, and we conclude that the oscillatory mechanism keeps the starch synthetic apparatus in the storage root sink in tune with the flux of sucrose from the photosynthetic source. When storage roots were uncoupled from the source, SBE expression could be effectively induced by exogenous sucrose. Turanose, a sucrose isomer that cannot be metabolized by plants, mimicked the effect of sucrose, demonstratingmore » that downstream metabolism of sucrose was not necessary for signal transmission. Also glucose and glucose-1-P induced SBE expression. Interestingly, induction by sucrose, turanose and glucose but not glucose-1-P sustained an overt semidian (12-h) oscillation in SBE expression and was sensitive to the hexokinase (HXK) inhibitor glucosamine. These results suggest a pivotal regulatory role for HXK during starch synthesis. Abscisic acid (ABA) was another potent inducer of SBE expression. Induction by ABA was similar to that of glucose-1-P in that it bypassed the semidian oscillator. Both the sugar and ABA signaling cascades were disrupted by okadaic acid, a protein phosphatase inhibitor. Based on these findings, we propose a model for sugar signaling in regulation of starch synthesis in the cassava storage root.« less

The laforin-malin complex negatively regulates glycogen synthesis by modulating cellular glucose uptake via glucose transporters.

PubMed

Singh, Pankaj Kumar; Singh, Sweta; Ganesh, Subramaniam

2012-02-01

Lafora disease (LD), an inherited and fatal neurodegenerative disorder, is characterized by increased cellular glycogen content and the formation of abnormally branched glycogen inclusions, called Lafora bodies, in the affected tissues, including neurons. Therefore, laforin phosphatase and malin ubiquitin E3 ligase, the two proteins that are defective in LD, are thought to regulate glycogen synthesis through an unknown mechanism, the defects in which are likely to underlie some of the symptoms of LD. We show here that laforin's subcellular localization is dependent on the cellular glycogen content and that the stability of laforin is determined by the cellular ATP level, the activity of 5'-AMP-activated protein kinase, and the affinity of malin toward laforin. By using cell and animal models, we further show that the laforin-malin complex regulates cellular glucose uptake by modulating the subcellular localization of glucose transporters; loss of malin or laforin resulted in an increased abundance of glucose transporters in the plasma membrane and therefore excessive glucose uptake. Loss of laforin or malin, however, did not affect glycogen catabolism. Thus, the excessive cellular glucose level appears to be the primary trigger for the abnormally higher levels of cellular glycogen seen in LD.

The cochaperone BAG3 coordinates protein synthesis and autophagy under mechanical strain through spatial regulation of mTORC1.

PubMed

Kathage, Barbara; Gehlert, Sebastian; Ulbricht, Anna; Lüdecke, Laura; Tapia, Victor E; Orfanos, Zacharias; Wenzel, Daniela; Bloch, Wilhelm; Volkmer, Rudolf; Fleischmann, Bernd K; Fürst, Dieter O; Höhfeld, Jörg

2017-01-01

The cochaperone BAG3 is a central protein homeostasis factor in mechanically strained mammalian cells. It mediates the degradation of unfolded and damaged forms of the actin-crosslinker filamin through chaperone-assisted selective autophagy (CASA). In addition, BAG3 stimulates filamin transcription in order to compensate autophagic disposal and to maintain the actin cytoskeleton under strain. Here we demonstrate that BAG3 coordinates protein synthesis and autophagy through spatial regulation of the mammalian target of rapamycin complex 1 (mTORC1). The cochaperone utilizes its WW domain to contact a proline-rich motif in the tuberous sclerosis protein TSC1 that functions as an mTORC1 inhibitor in association with TSC2. Interaction with BAG3 results in a recruitment of TSC complexes to actin stress fibers, where the complexes act on a subpopulation of mTOR-positive vesicles associated with the cytoskeleton. Local inhibition of mTORC1 is essential to initiate autophagy at sites of filamin unfolding and damage. At the same time, BAG3-mediated sequestration of TSC1/TSC2 relieves mTORC1 inhibition in the remaining cytoplasm, which stimulates protein translation. In human muscle, an exercise-induced association of TSC1 with the cytoskeleton coincides with mTORC1 activation in the cytoplasm. The spatial regulation of mTORC1 exerted by BAG3 apparently provides the basis for a simultaneous induction of autophagy and protein synthesis to maintain the proteome under mechanical strain. Copyright © 2016 Elsevier B.V. All rights reserved.

A growth-dependent transcription initiation factor (TIF-IA) interacting with RNA polymerase I regulates mouse ribosomal RNA synthesis.

PubMed

Schnapp, A; Pfleiderer, C; Rosenbauer, H; Grummt, I

1990-09-01

Control of mouse ribosomal RNA synthesis in response to extracellular signals is mediated by TIF-IA, a regulatory factor whose amount or activity correlates with cell proliferation. Factor TIF-IA interacts with RNA polymerase I (pol I), thus converting it into a transcriptionally active holoenzyme, which is able to initiate specifically at the rDNA promoter in the presence of the other auxiliary transcription initiation factors, designated TIF-IB, TIF-IC and UBF. With regard to several criteria, the growth-dependent factor TIF-IA behaves like a bacterial sigma factor: (i) it associates physically with pol I, (ii) it is required for initiation of transcription, (iii) it is present in limiting amounts and (iv) under certain salt conditions, it is chromatographically separable from the polymerase. In addition, evidence is presented that dephosphorylation of pol I abolishes in vitro transcription initiation from the ribosomal gene promoter without significantly affecting the polymerizing activity of the enzyme at nonspecific templates. The involvement of both a regulatory factor and post-translational modification of the transcribing enzyme provides an efficient and versatile mechanism of rDNA transcription regulation which enables the cell to adapt ribosome synthesis rapidly to a variety of extracellular signals.

The synthesis, regulation, and functions of sterols in Candida albicans: Well-known but still lots to learn.

PubMed

Lv, Quan-Zhen; Yan, Lan; Jiang, Yuan-Ying

2016-08-17

Sterols are the basal components of the membranes of the fungal pathogen Candida albicans, and these membranes determine the susceptibility of C. albicans cells to a variety of stresses, such as ionic, osmotic and oxidative pressures, and treatment with antifungal drugs. The common antifungal azoles in clinical use are targeted to the biosynthesis of ergosterol. In the past years, the synthesis, storage and metabolism of ergosterol in Saccharomyces cerevisiae has been characterized in some detail; however, these processes has not been as well investigated in the human opportunistic pathogen C. albicans. In this review, we summarize the genes involved in ergosterol synthesis and regulation in C. albicans. As well, genes in S. cerevisiae implicated in ergosterol storage and conversions with other lipids are noted, as these provide us clues and directions for the study of the homologous genes in C. albicans. In this report we have particularly focused on the essential roles of ergosterol in the dynamic process of cell biology and its fundamental status in the biological membrane system that includes lipid rafts, lipid droplets, vacuoles and mitochondria. We believe that a thorough understanding of this classic and essential pathway will give us new ideas about drug resistance and morphological switching in C. albicans.

The synthesis, regulation, and functions of sterols in Candida albicans: Well-known but still lots to learn

PubMed Central

Lv, Quan-zhen; Yan, Lan; Jiang, Yuan-ying

2016-01-01

ABSTRACT Sterols are the basal components of the membranes of the fungal pathogen Candida albicans, and these membranes determine the susceptibility of C. albicans cells to a variety of stresses, such as ionic, osmotic and oxidative pressures, and treatment with antifungal drugs. The common antifungal azoles in clinical use are targeted to the biosynthesis of ergosterol. In the past years, the synthesis, storage and metabolism of ergosterol in Saccharomyces cerevisiae has been characterized in some detail; however, these processes has not been as well investigated in the human opportunistic pathogen C. albicans. In this review, we summarize the genes involved in ergosterol synthesis and regulation in C. albicans. As well, genes in S. cerevisiae implicated in ergosterol storage and conversions with other lipids are noted, as these provide us clues and directions for the study of the homologous genes in C. albicans. In this report we have particularly focused on the essential roles of ergosterol in the dynamic process of cell biology and its fundamental status in the biological membrane system that includes lipid rafts, lipid droplets, vacuoles and mitochondria. We believe that a thorough understanding of this classic and essential pathway will give us new ideas about drug resistance and morphological switching in C. albicans. PMID:27221657

Down-Regulation by Resveratrol of Basic Fibroblast Growth Factor-Stimulated Osteoprotegerin Synthesis through Suppression of Akt in Osteoblasts

PubMed Central

Kuroyanagi, Gen; Otsuka, Takanobu; Yamamoto, Naohiro; Matsushima-Nishiwaki, Rie; Nakakami, Akira; Mizutani, Jun; Kozawa, Osamu; Tokuda, Haruhiko

2014-01-01

It is firmly established that resveratrol, a natural food compound abundantly found in grape skins and red wine, has beneficial properties for human health. In the present study, we investigated the effect of basic fibroblast growth factor (FGF-2) on osteoprotegerin (OPG) synthesis in osteoblast-like MC3T3-E1 cells and whether resveratrol affects the OPG synthesis. FGF-2 stimulated both the OPG release and the expression of OPG mRNA. Resveratrol significantly suppressed the FGF-2-stimulated OPG release and the mRNA levels of OPG. SRT1720, an activator of SIRT1, reduced the FGF-2-induced OPG release and the OPG mRNA expression. PD98059, an inhibitor of upstream kinase activating p44/p42 mitogen-activated protein (MAP) kinase, had little effect on the FGF-2-stimulated OPG release. On the other hand, SB203580, an inhibitor of p38 MAP kinase, SP600125, an inhibitor of stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK), and Akt inhibitor suppressed the OPG release induced by FGF-2. Resveratrol failed to affect the FGF-2-induced phosphorylation of p44/p42 MAP kinase, p38 MAP kinase or SAPK/JNK. The phosphorylation of Akt induced by FGF-2 was significantly suppressed by resveratrol or SRT1720. These findings strongly suggest that resveratrol down-regulates FGF-2-stimulated OPG synthesis through the suppression of the Akt pathway in osteoblasts and that the inhibitory effect of resveratrol is mediated at least in part by SIRT1 activation. PMID:25290095

Regulation of porphyrin synthesis and photodynamic therapy in heavy metal intoxication.

PubMed

Grinblat, Borislava; Pour, Nir; Malik, Zvi

2006-01-01

Protoporphyrin IX (PpIX) synthesis by malignant cells is successfully exploited for photodynamic therapy (PDT) following administration of 5-aminolevulinic acid (ALA) and light irradiation. The influence of two environmental heavy metal poisons, lead and gallium, on PpIX-synthesis and ALA-PDT was studied in two neu-ronal cell lines, SH-SY5Y neuroblastoma and PC12 pheochromocytoma. The heavy metal intoxication affected two of the heme-synthesis enzymes, ALA-dehydratase (ALAD) and porphobilinogen deaminase (PBGD). The present results show that lead poisoning significantly decreased the PBGD cellular level and inhibited its enzymatic activity, whereas the effects of gallium were less prominent. Although, the protein levels were reduced, the mRNA levels of PBGD remained unchanged during metal intoxication. These findings show additional inhibitory activity of lead on top of its classical effect on ALAD. Proteasome activity was enhanced during lead treatment, as measured by the AMC fluorigenic proteasome assay. The reduction in PBGD levels was not a consequence of PBGD mRNA reduced synthesis, which remained unchanged as shown by RT-PCR analysis. As a result of the lead poisoning, marked alterations in the cell cycle were observed, including a decreased G1 phase and an increased number of S phase cells. The efficacy of ALA-PDT was reduced in correlation with decreased activities of the enzymes during lead intoxication. We may conclude that lead poisoning adversely affects the outcome of ALA photodynamic therapy of cancer.

Transcriptional Mechanisms Underlying Hemoglobin Synthesis

PubMed Central

Katsumura, Koichi R.; DeVilbiss, Andrew W.; Pope, Nathaniel J.; Johnson, Kirby D.; Bresnick, Emery H.

2013-01-01

The physiological switch in expression of the embryonic, fetal, and adult β-like globin genes has garnered enormous attention from investigators interested in transcriptional mechanisms and the molecular basis of hemoglobinopathies. These efforts have led to the discovery of cell type-specific transcription factors, unprecedented mechanisms of transcriptional coregulator function, genome biology principles, unique contributions of nuclear organization to transcription and cell function, and promising therapeutic targets. Given the vast literature accrued on this topic, this article will focus on the master regulator of erythroid cell development and function GATA-1, its associated proteins, and its frontline role in controlling hemoglobin synthesis. GATA-1 is a crucial regulator of genes encoding hemoglobin subunits and heme biosynthetic enzymes. GATA-1-dependent mechanisms constitute an essential regulatory core that nucleates additional mechanisms to achieve the physiological control of hemoglobin synthesis. PMID:23838521

Genome-wide Expression Analysis and Metabolite Profiling Elucidate Transcriptional Regulation of Flavonoid Biosynthesis and Modulation under Abiotic Stresses in Banana

PubMed Central

Pandey, Ashutosh; Alok, Anshu; Lakhwani, Deepika; Singh, Jagdeep; Asif, Mehar H.; Trivedi, Prabodh K.

2016-01-01

Flavonoid biosynthesis is largely regulated at the transcriptional level due to the modulated expression of genes related to the phenylpropanoid pathway in plants. Although accumulation of different flavonoids has been reported in banana, a staple fruit crop, no detailed information is available on regulation of the biosynthesis in this important plant. We carried out genome-wide analysis of banana (Musa acuminata, AAA genome) and identified 28 genes belonging to 9 gene families associated with flavonoid biosynthesis. Expression analysis suggested spatial and temporal regulation of the identified genes in different tissues of banana. Analysis revealed enhanced expression of genes related to flavonol and proanthocyanidin (PA) biosynthesis in peel and pulp at the early developmental stages of fruit. Genes involved in anthocyanin biosynthesis were highly expressed during banana fruit ripening. In general, higher accumulation of metabolites was observed in the peel as compared to pulp tissue. A correlation between expression of genes and metabolite content was observed at the early stage of fruit development. Furthermore, this study also suggests regulation of flavonoid biosynthesis, at transcriptional level, under light and dark exposures as well as methyl jasmonate (MJ) treatment in banana. PMID:27539368

Genome-wide Expression Analysis and Metabolite Profiling Elucidate Transcriptional Regulation of Flavonoid Biosynthesis and Modulation under Abiotic Stresses in Banana.

PubMed

Pandey, Ashutosh; Alok, Anshu; Lakhwani, Deepika; Singh, Jagdeep; Asif, Mehar H; Trivedi, Prabodh K

2016-08-19

Flavonoid biosynthesis is largely regulated at the transcriptional level due to the modulated expression of genes related to the phenylpropanoid pathway in plants. Although accumulation of different flavonoids has been reported in banana, a staple fruit crop, no detailed information is available on regulation of the biosynthesis in this important plant. We carried out genome-wide analysis of banana (Musa acuminata, AAA genome) and identified 28 genes belonging to 9 gene families associated with flavonoid biosynthesis. Expression analysis suggested spatial and temporal regulation of the identified genes in different tissues of banana. Analysis revealed enhanced expression of genes related to flavonol and proanthocyanidin (PA) biosynthesis in peel and pulp at the early developmental stages of fruit. Genes involved in anthocyanin biosynthesis were highly expressed during banana fruit ripening. In general, higher accumulation of metabolites was observed in the peel as compared to pulp tissue. A correlation between expression of genes and metabolite content was observed at the early stage of fruit development. Furthermore, this study also suggests regulation of flavonoid biosynthesis, at transcriptional level, under light and dark exposures as well as methyl jasmonate (MJ) treatment in banana.

Alkaline stress and iron deficiency regulate iron uptake and riboflavin synthesis gene expression differently in root and leaf tissue: implications for iron deficiency chlorosis

PubMed Central

Hsieh, En-Jung; Waters, Brian M.

2016-01-01

Iron (Fe) is an essential mineral that has low solubility in alkaline soils, where its deficiency results in chlorosis. Whether low Fe supply and alkaline pH stress are equivalent is unclear, as they have not been treated as separate variables in molecular physiological studies. Additionally, molecular responses to these stresses have not been studied in leaf and root tissues simultaneously. We tested how plants with the Strategy I Fe uptake system respond to Fe deficiency at mildly acidic and alkaline pH by measuring root ferric chelate reductase (FCR) activity and expression of selected Fe uptake genes and riboflavin synthesis genes. Alkaline pH increased cucumber (Cucumis sativus L.) root FCR activity at full Fe supply, but alkaline stress abolished FCR response to low Fe supply. Alkaline pH or low Fe supply resulted in increased expression of Fe uptake genes, but riboflavin synthesis genes responded to Fe deficiency but not alkalinity. Iron deficiency increased expression of some common genes in roots and leaves, but alkaline stress blocked up-regulation of these genes in Fe-deficient leaves. In roots of the melon (Cucumis melo L.) fefe mutant, in which Fe uptake responses are blocked upstream of Fe uptake genes, alkaline stress or Fe deficiency up-regulation of certain Fe uptake and riboflavin synthesis genes was inhibited, indicating a central role for the FeFe protein. These results suggest a model implicating shoot-to-root signaling of Fe status to induce Fe uptake gene expression in roots. PMID:27605716

Phytochrome B Mediates the Regulation of Chlorophyll Biosynthesis through Transcriptional Regulation of ChlH and GUN4 in Rice Seedlings

PubMed Central

Kagawa, Takatoshi; Tanaka, Ayumi; Ueno, Osamu; Shimada, Hiroaki; Takano, Makoto

2015-01-01

Accurate regulation of chlorophyll synthesis is crucial for chloroplast formation during the greening process in angiosperms. In this study, we examined the role of phytochrome B (phyB) in the regulation of chlorophyll synthesis in rice seedlings (Oryza sativa L.) through the characterization of a pale-green phenotype observed in the phyB mutant grown under continuous red light (Rc) irradiation. Our results show that the Rc-induced chlorophyll accumulation can be divided into two components—a phyB-dependent and a phyB-independent component, and that the pale-green phenotype is caused by the absence of the phyB-dependent component. To elucidate the role of the missing component we established an Rc-induced greening experiment, the results of which revealed that several genes encoding proteins on the chlorophyll branch were repressed in the phyB mutant. Notable among them were ChlH and GUN4 genes, which encode subunit H and an activating factor of magnesium chelatase (Mg-chelatase), respectively, that were largely repressed in the mutant. Moreover, the kinetic profiles of chlorophyll precursors suggested that Mg-chelatase activity simultaneously decreased with the reduction in the transcript levels of ChlH and GUN4. These results suggest that phyB mediates the regulation of chlorophyll synthesis through transcriptional regulation of these two genes, whose products exert their action at the branching point of the chlorophyll biosynthesis pathway. Reduction of 5-aminolevulinic acid (5-ALA) synthesis could be detected in the mutant, but the kinetic profiles of chlorophyll precursors indicated that it was an event posterior to the reduction of the Mg-chelatase activity. It means that the repression of 5-ALA synthesis should not be a triggering event for the appearance of the pale-green phenotype. Instead, the repression of 5-ALA synthesis might be important for the subsequent stabilization of the pale-green phenotype for preventing excessive accumulation of hazardous

Molecular cellular mechanisms of peptide regulation of melatonin synthesis in pinealocyte culture.

PubMed

Khavinson, V Kh; Linkova, N S; Kvetnoy, I M; Kvetnaia, T V; Polyakova, V O; Korf, H-W

2012-06-01

The effects of epithalone and vilone peptides on the synthesis of melatonin and factors involved in this process, arylalkylamine-N-acetyltransferase (AANAT) enzyme and pCREB transcription protein, were studied in rat pinealocyte culture. Epithalone stimulated AANAT and pCREB synthesis and increased melatonin level in culture medium. Simultaneous addition of norepinephrine and peptides into the culture potentiated the expression of AANAT and pCREB.

Urinary bladder organ hypertrophy is partially regulated by Akt1-mediated protein synthesis pathway.

PubMed

Qiao, Li-Ya; Xia, Chunmei; Shen, Shanwei; Lee, Seong Ho; Ratz, Paul H; Fraser, Matthew O; Miner, Amy; Speich, John E; Lysiak, Jeffrey J; Steers, William D

2018-05-15

The present study aims to investigate the role of Akt in the regulation of urinary bladder organ hypertrophy caused by partial bladder outlet obstruction (pBOO). Male rats were surgically induced for pBOO. Real-time PCR and western blot were used to examine the levels of mRNA and protein. A phosphoinositide 3-kinase (PI3K) inhibitor LY294002 was used to inhibit the activity of endogenous Akt. The urinary bladder developed hypertrophy at 2 weeks of pBOO. The protein but not mRNA levels of type I collagen and α-smooth muscle actin (αSMA) were increased in pBOO bladder when compared to sham control. The phosphorylation (activation) levels of Akt1 (p-Ser 473 ), mammalian target of rapamycin (mTOR), p70S6 kinase (p70S6K), and 4E-BP1 were also increased in pBOO bladder. LY294002 treatment reduced the phosphorylation levels of Akt1 and 4E-BP1, and the protein levels of type I collagen and αSMA in pBOO bladder. The mRNA and protein levels of proliferating cell nuclear antigen (PCNA) were increased in pBOO bladder, and PCNA up-regulation occurred in urothelial not muscular layer. LY294002 treatment had no effect on the mRNA and protein levels of PCNA in pBOO bladder. LY294002 treatment partially reduced the bladder weight caused by pBOO. pBOO-induced urinary bladder hypertrophy is attributable to fibrosis, smooth muscle cellular hypertrophy, and urothelium cell hyper-proliferation. Akt1-mediated protein synthesis in pBOO bladder contributes to type I collagen and αSMA but not PCNA up-regulation. Target of Akt1 is necessary but not sufficient in treatment of urinary bladder hypertrophy following pBOO. Copyright © 2018 Elsevier Inc. All rights reserved.

MiR-30a-3p Negatively Regulates BAFF Synthesis in Systemic Sclerosis and Rheumatoid Arthritis Fibroblasts

PubMed Central

Philippe, Lucas; Gong, Ya-Zhuo; Bahram, Seiamak; Cetin, Semih; Pfeffer, Sébastien; Gottenberg, Jacques-Eric; Wachsmann, Dominique; Georgel, Philippe; Sibilia, Jean

2014-01-01

We evaluated micro (mi) RNA-mediated regulation of BAFF expression in fibroblasts using two concomitant models: (i) synovial fibroblasts (FLS) isolated from healthy controls (N) or Rheumatoid Arthritis (RA) patients; (ii) human dermal fibroblasts (HDF) isolated from healthy controls (N) or Systemic Sclerosis (SSc) patients. Using RT-qPCR and ELISA, we first showed that SScHDF synthesized and released BAFF in response to Poly(I:C) or IFN-γ treatment, as previously observed in RAFLS, whereas NHDF released BAFF preferentially in response to IFN-γ. Next, we demonstrated that miR-30a-3p expression was down regulated in RAFLS and SScHDF stimulated with Poly(I:C) or IFN-γ. Moreover, we demonstrated that transfecting miR-30a-3p mimic in Poly(I:C)- and IFN-γ-activated RAFLS and SScHDF showed a strong decrease on BAFF synthesis and release and thus B cells survival in our model. Interestingly, FLS and HDF isolated from healthy subjects express higher levels of miR-30a-3p and lower levels of BAFF than RAFLS and SScHDF. Transfection of miR-30a-3p antisense in Poly(I:C)- and IFN-γ-activated NFLS and NHDF upregulated BAFF secretion, confirming that this microRNA is a basal repressors of BAFF expression in cells from healthy donors. Our data suggest a critical role of miR-30a-3p in the regulation of BAFF expression, which could have a major impact in the regulation of the autoimmune responses occurring in RA and SSc. PMID:25360821

Transcriptional and post-transcriptional regulation of the ionizing radiation response by ATM and p53

PubMed Central

Venkata Narayanan, Ishwarya; Paulsen, Michelle T.; Bedi, Karan; Berg, Nathan; Ljungman, Emily A.; Francia, Sofia; Veloso, Artur; Magnuson, Brian; di Fagagna, Fabrizio d’Adda; Wilson, Thomas E.; Ljungman, Mats

2017-01-01

In response to ionizing radiation (IR), cells activate a DNA damage response (DDR) pathway to re-program gene expression. Previous studies using total cellular RNA analyses have shown that the stress kinase ATM and the transcription factor p53 are integral components required for induction of IR-induced gene expression. These studies did not distinguish between changes in RNA synthesis and RNA turnover and did not address the role of enhancer elements in DDR-mediated transcriptional regulation. To determine the contribution of synthesis and degradation of RNA and monitor the activity of enhancer elements following exposure to IR, we used the recently developed Bru-seq, BruChase-seq and BruUV-seq techniques. Our results show that ATM and p53 regulate both RNA synthesis and stability as well as enhancer element activity following exposure to IR. Importantly, many genes in the p53-signaling pathway were coordinately up-regulated by both increased synthesis and RNA stability while down-regulated genes were suppressed either by reduced synthesis or stability. Our study is the first of its kind that independently assessed the effects of ionizing radiation on transcription and post-transcriptional regulation in normal human cells. PMID:28256581

Regulation of expression, activity and localization of fungal chitin synthases

PubMed Central

Rogg, Luise E.; Fortwendel, Jarrod R.; Juvvadi, Praveen R.; Steinbach, William J.

2013-01-01

The fungal cell wall represents an attractive target for pharmacologic inhibition, as many of the components are fungal-specific. Though targeted inhibition of β-glucan synthesis is effective treatment for certain fungal infections, the ability of the cell wall to dynamically compensate via the cell wall integrity pathway may limit overall efficacy. To date, chitin synthesis inhibitors have not been successfully deployed in the clinical setting. Fungal chitin synthesis is a complex and highly regulated process. Regulation of chitin synthesis occurs on multiple levels, thus targeting of these regulatory pathways may represent an exciting alternative approach. A variety of signaling pathways have been implicated in chitin synthase regulation, at both transcriptional and post-transcriptional levels. Recent research suggests that localization of chitin synthases likely represents a major regulatory mechanism. However, much of the regulatory machinery is not necessarily shared among different chitin synthases. Thus, an in depth understanding of the precise roles of each protein in cell wall maintenance and repair will be essential to identifying the most likely therapeutic targets. PMID:21526913

Proteomic analysis of tea plants (Camellia sinensis) with purple young shoots during leaf development

PubMed Central

Zhou, Qiongqiong; Chen, Zhidan; Lee, Jinwook; Li, Xinghui; Sun, Weijiang

2017-01-01

Tea products made from purple leaves are highly preferred by consumers due to the health benefits. This study developed a proteome reference map related to color changes during leaf growth in tea (Camellia sinensis) plant with purple young shoots using two-dimensional electrophoresis (2-DE). Forty-six differentially expressed proteins were detected in the gel and successfully identified by using MALDI-TOF/TOF-MS. The pronounced changes in the proteomic profile between tender purple leaves (TPL) and mature green leaves (MGL) included: 1) the lower activity of proteins associated with CO2 assimilation, energy metabolism and photo flux efficiency and higher content of anthocyanins in TPL than those in MGL may protect tender leaves against photo-damage; 2) the higher abundance of chalcone synthase (CHS), chalcone isomerase (CHI) and flavonol synthase (FLS) likely contributes to the synthesis of anthocyanins, catechins and flavonols in TPL tissues; 3) higher abundance of stress response proteins, such as glutathione S-transferases (GST) and phospholipid hydroperoxide glutathione peroxidase (PHGPx), could enhance the tolerance of TPL tissues to adverse condition in; and 4) the increased abundance of proteins related to protein synthesis, nucleic acids and cell wall proteins should be beneficial for the proliferation and expansion of leaf cell in TPL tissues. qPCR analysis showed that the expression of differentially abundant proteins was regulated at the transcriptional level. Therefore, the results indicated that higher abundance of CHI and CHS may account for the production of the purple-shoot phenotype in Wuyiqizhong 18 and thereby, enhancing the anthocyanin biosynthesis. The higher abundance of glutamine synthetase (GS) proteins related to the theanine biosynthesis may improve the flavor of tea products from TPL materials. Thus, this work should help to understand the molecular mechanisms underlying the changes in leaf color alteration. PMID:28520776

The HD-GYP Domain Protein RpfG of Xanthomonas oryzae pv. oryzicola Regulates Synthesis of Extracellular Polysaccharides that Contribute to Biofilm Formation and Virulence on Rice

PubMed Central

Zhang, Yuanbao; Wei, Chao; Jiang, Wendi; Wang, Lei; Li, Churui; Wang, Yunyue; Dow, John Maxwell; Sun, Wenxian

2013-01-01

Bacterial leaf streak caused by Xanthomonas oryzae pv. oryzicola (Xoc) is one of the most important diseases in rice. However, little is known about the pathogenicity mechanisms of Xoc. Here we have investigated the function of three HD-GYP domain regulatory proteins in biofilm formation, the synthesis of virulence factors and virulence of Xoc. Deletion of rpfG resulted in altered production of extracellular polysaccharides (EPS), abolished virulence on rice and enhanced biofilm formation, but had little effect on the secretion of proteases and motility. In contrast, mutational analysis showed that the other two HD-GYP domain proteins had no effect on virulence factor synthesis and tested phenotypes. Mutation of rpfG led to up-regulation of the type III secretion system and altered expression of three putative glycosyltransferase genes gumD, pgaC and xagB, which are part of operons directing the synthesis of different extracellular polysaccharides. The pgaABCD and xagABCD operons were greatly up-regulated in the Xoc ΔrpfG mutant, whereas the expression of the gum genes was unaltered or slightly enhanced. The elevated biofilm formation of the Xoc ΔrpfG mutant was dramatically reduced upon deletion of gumD, xagA and xagB, but not when pgaA and pgaC were deleted. Interestingly, only the ΔgumD mutant, among these single gene mutants, exhibits multiple phenotype alterations including reduced biofilm and EPS production and attenuated virulence on rice. These data indicate that RpfG is a global regulator that controls biofilm formation, EPS production and bacterial virulence in Xoc and that both gumD- and xagB-dependent EPS contribute to biofilm formation under different conditions. PMID:23544067

Flavonoids such as luteolin, fisetin and apigenin are inhibitors of interleukin-4 and interleukin-13 production by activated human basophils.

PubMed

Hirano, Toru; Higa, Shinji; Arimitsu, Junsuke; Naka, Tetsuji; Shima, Yoshihito; Ohshima, Shiro; Fujimoto, Minoru; Yamadori, Tomoki; Kawase, Ichiro; Tanaka, Toshio

2004-06-01

We have previously shown that fisetin, a flavonol, inhibits IL-4 and IL-13 synthesis by allergen- or anti-IgE-antibody-stimulated basophils. This time, we investigated the inhibition of IL-4 and IL-13 production by basophils by other flavonoids and attempted to determine the fundamental structure of flavonoids related to inhibition. We additionally investigated whether flavonoids suppress leukotriene C4 synthesis by basophils and IL-4 synthesis by T cells in response to anti-CD3 antibody. Highly purified peripheral basophils were stimulated for 12 h with anti-IgE antibody alone or anti-IgE antibody plus IL-3 in the presence of various concentrations of 18 different kinds of flavones and flavonols. IL-4 and IL-13 concentrations in the supernatants were then measured. Leukotriene C4 synthesis was also measured after basophils were stimulated for 1 h in the presence of flavonoids. Regarding the inhibitory activity of flavonoids on IL-4 synthesis by T cells, peripheral blood mononuclear cells were cultured with flavonoids in anti-CD3-antibody-bound plates for 2 days. Luteolin, fisetin and apigenin were found to be the strongest inhibitors of both IL-4 and IL-13 production by basophils but did not affect leukotriene C4 synthesis. At higher concentrations, these flavonoids suppressed IL-4 production by T cells. Based on a hierarchy of inhibitory activity, the basic structure for IL-4 inhibition by basophils was determined. Due to the inhibitory activity of flavonoids on IL-4 and IL-13 synthesis, it can be expected that the intake of flavonoids, depending on the quantity and quality, may ameliorate allergic symptoms or prevent the onset of allergic diseases. Copyright 2004 S. Karger AG, Basel

Synthesis, characterization, and ligand exchange reactivity of a series of first row divalent metal 3-hydroxyflavonolate complexes.

PubMed

Grubel, Katarzyna; Rudzka, Katarzyna; Arif, Atta M; Klotz, Katie L; Halfen, Jason A; Berreau, Lisa M

2010-01-04

A series of divalent metal flavonolate complexes of the general formula [(6-Ph(2)TPA)M(3-Hfl)]X (1-5-X; X = OTf(-) or ClO(4)(-); 6-Ph(2)TPA = N,N-bis((6-phenyl-2-pyridyl)methyl)-N-((2-pyridyl)methyl)amine; M = Mn(II), Co(II), Ni(II), Cu(II), Zn(II); 3-Hfl = 3-hydroxyflavonolate) were prepared and characterized by X-ray crystallography, elemental analysis, FTIR, UV-vis, (1)H NMR or EPR, and cyclic voltammetry. All of the complexes have a bidentate coordinated flavonolate ligand. The difference in M-O distances (Delta(M-O)) involving this ligand varies through the series, with the asymmetry of flavonolate coordination increasing in the order Mn(II) approximately Ni(II) < Cu(II) < Zn(II) < Co(II). The hypsochromic shift of the absorption band I (pi-->pi*) of the coordinated flavonolate ligand in 1-5-OTf (relative to that in free anion) increases in the order Ni(II) < Mn(II) < Cu(II) < Zn(II), Co(II). Previously reported 3-Hfl complexes of divalent metals fit well with this ordering. (1)H NMR studies indicate that the 3-Hfl complexes of Co(II), Ni(II), and Zn(II) exhibit a pseudo-octahedral geometry in solution. EPR studies suggest that the Mn(II) complex 1-OTf may form binuclear structures in solution. The mononuclear Cu(II) complex 4-OTf has a distorted square pyramidal geometry. The oxidation potential of the flavonolate ligand depends on the metal ion present and/or the solution structure of the complex, with the Mn(II) complex 1-OTf exhibiting the lowest potential, followed by the pseudo-octahedral Ni(II) and Zn(II) 3-Hfl complexes, and the distorted square pyramidal Cu(II) complex 4-OTf. The Mn(II) complex [(6-Ph(2)TPA)Mn(3-Hfl)]OTf (1-OTf) is unique in the series in undergoing ligand exchange reactions in the presence of M(ClO(4))(2).6H(2)O (M = Co, Ni, Zn) in CD(3)CN to produce [(6-Ph(2)TPA)M(CD(3)CN)(n)](X)(2), [Mn(3-Hfl)(2).0.5H(2)O], and MnX(2) (X = OTf(-) or ClO(4)(-)). Under similar conditions, the 3-Hfl complexes of Co(II), Ni(II), and Cu(II) undergo

Improving monoterpene geraniol production through geranyl diphosphate synthesis regulation in Saccharomyces cerevisiae.

PubMed

Zhao, Jianzhi; Bao, Xiaoming; Li, Chen; Shen, Yu; Hou, Jin

2016-05-01

Monoterpenes have wide applications in the food, cosmetics, and medicine industries and have recently received increased attention as advanced biofuels. However, compared with sesquiterpenes, monoterpene production is still lagging in Saccharomyces cerevisiae. In this study, geraniol, a valuable acyclic monoterpene alcohol, was synthesized in S. cerevisiae. We evaluated three geraniol synthases in S. cerevisiae, and the geraniol synthase Valeriana officinalis (tVoGES), which lacked a plastid-targeting peptide, yielded the highest geraniol production. To improve geraniol production, synthesis of the precursor geranyl diphosphate (GPP) was regulated by comparing three specific GPP synthase genes derived from different plants and the endogenous farnesyl diphosphate synthase gene variants ERG20 (G) (ERG20 (K197G) ) and ERG20 (WW) (ERG20 (F96W-N127W) ), and controlling endogenous ERG20 expression, coupled with increasing the expression of the mevalonate pathway by co-overexpressing IDI1, tHMG1, and UPC2-1. The results showed that overexpressing ERG20 (WW) and strengthening the mevalonate pathway significantly improved geraniol production, while expressing heterologous GPP synthase genes or down-regulating endogenous ERG20 expression did not show positive effect. In addition, we constructed an Erg20p(F96W-N127W)-tVoGES fusion protein, and geraniol production reached 66.2 mg/L after optimizing the amino acid linker and the order of the proteins. The best strain yielded 293 mg/L geraniol in a fed-batch cultivation, a sevenfold improvement over the highest titer previously reported in an engineered S. cerevisiae strain. Finally, we showed that the toxicity of geraniol limited its production. The platform developed here can be readily used to synthesize other monoterpenes.

PdhR, the pyruvate dehydrogenase repressor, does not regulate lipoic acid synthesis.

PubMed

Feng, Youjun; Cronan, John E

2014-01-01

Lipoic acid is a covalently-bound enzyme cofactor required for central metabolism all three domains of life. In the last 20 years the pathway of lipoic acid synthesis and metabolism has been established in Escherichia coli. Expression of the genes of the lipoic acid biosynthesis pathway was believed to be constitutive. However, in 2010 Kaleta and coworkers (BMC Syst. Biol. 4:116) predicted a binding site for the pyruvate dehydrogenase operon repressor, PdhR (referred to lipA site 1) upstream of lipA, the gene encoding lipoic acid synthase and concluded that PdhR regulates lipA transcription. We report in vivo and in vitro evidence that lipA is not controlled by PdhR and that the putative regulatory site deduced by the prior workers is nonfunctional and physiologically irrelevant. E. coli PdhR was purified to homogeneity and used for electrophoretic mobility shift assays. The lipA site 1 of Kaleta and coworkers failed to bind PdhR. The binding detected by these workers is due to another site (lipA site 3) located far upstream of the lipA promoter. Relative to the canonical PdhR binding site lipA site 3 is a half-palindrome and as expected had only weak PdhR binding ability. Manipulation of lipA site 3 to construct a palindrome gave significantly enhanced PdhR binding affinity. The native lipA promoter and the version carrying the artificial lipA3 palindrome were transcriptionally fused to a LacZ reporter gene to directly assay lipA expression. Deletion of pdhR gave no significant change in lipA promoter-driven β-galactosidase activity with either the native or constructed palindrome upstream sequences, indicating that PdhR plays no physiological role in regulation of lipA expression. Copyright © 2014 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Iron Induction of Ferritin Synthesis in Soybean Cell Suspensions

PubMed Central

Proudhon, Dominique; Briat, Jean-François; Lescure, Anne-Marie

1989-01-01

In animal cells specialized for iron storage, iron-induced accumulation of ferritin is known to result from a shift of stored mRNA from the ribonucleoprotein fraction to polysomes. Previous reports with bean leaves suggested that in plants iron induction of ferritin synthesis would result from a regulation at the transcriptional level (F van der Mark, F Bienfait, H van der Ende [1983] Biochem Biophys Res Commun 115:463-469). Soybean (Glycine max, cv Mandarin) cell suspension cultures have been used here to support these findings. Ferritin induction is obtained by addition of Fe-citrate to the culture medium. A good correlation is found between cellular iron content and the amount of ferritin accumulation. This protein accumulation corresponds to an increase of in vitro translatable ferritin mRNA. Addition of 4 micrograms actinomycin D per milliliter to the cultures inhibits completely in vivo RNA synthesis, whereas protein synthesis was poorly affected, at least for 24 hours. During the same time, this concentration of actinomycin D strongly inhibits the iron-induced synthesis of ferritin. These results show that in soybean cell cultures, the mechanism of regulation of ferritin synthesis in response to iron does not result from recruitment of preexisting mRNA. They confirm that in plant systems, ferritin synthesis results from increased transcription of the corresponding genes. Images Figure 2 Figure 3 Figure 5 PMID:16666812

Iron induction of ferritin synthesis in soybean cell suspensions.

PubMed

Proudhon, D; Briat, J F; Lescure, A M

1989-06-01

In animal cells specialized for iron storage, iron-induced accumulation of ferritin is known to result from a shift of stored mRNA from the ribonucleoprotein fraction to polysomes. Previous reports with bean leaves suggested that in plants iron induction of ferritin synthesis would result from a regulation at the transcriptional level (F van der Mark, F Bienfait, H van der Ende [1983] Biochem Biophys Res Commun 115:463-469). Soybean (Glycine max, cv Mandarin) cell suspension cultures have been used here to support these findings. Ferritin induction is obtained by addition of Fe-citrate to the culture medium. A good correlation is found between cellular iron content and the amount of ferritin accumulation. This protein accumulation corresponds to an increase of in vitro translatable ferritin mRNA. Addition of 4 micrograms actinomycin D per milliliter to the cultures inhibits completely in vivo RNA synthesis, whereas protein synthesis was poorly affected, at least for 24 hours. During the same time, this concentration of actinomycin D strongly inhibits the iron-induced synthesis of ferritin. These results show that in soybean cell cultures, the mechanism of regulation of ferritin synthesis in response to iron does not result from recruitment of preexisting mRNA. They confirm that in plant systems, ferritin synthesis results from increased transcription of the corresponding genes.

Inhibition of Photophosphorylation by Kaempferol 1

PubMed Central

Arntzen, Charles J.; Falkenthal, Scott V.; Bobick, Sandra

1974-01-01

Kaempferol, a naturally occurring flavonol, inhibited coupled electron transport and both cyclic and noncyclic photophosphorylation in isolated pea (Pisum sativum) chloroplasts. Over a concentration range which gave marked inhibition of ATP synthesis, there was no effect on basal or uncoupled electron flow or light-induced proton accumulation by isolated thylakoids. It is suggested that kaempferol acts as an energy transfer inhibitor. PMID:16658695

The nucleolus as a stress sensor: JNK2 inactivates the transcription factor TIF-IA and down-regulates rRNA synthesis.

PubMed

Mayer, Christine; Bierhoff, Holger; Grummt, Ingrid

2005-04-15

Cells respond to a variety of extracellular and intracellular forms of stress by down-regulating rRNA synthesis. We have investigated the mechanism underlying stress-dependent inhibition of RNA polymerase I (Pol I) transcription and show that the Pol I-specific transcription factor TIF-IA is inactivated upon stress. Inactivation is due to phosphorylation of TIF-IA by c-Jun N-terminal kinase (JNK) at a single threonine residue (Thr 200). Phosphorylation at Thr 200 impairs the interaction of TIF-IA with Pol I and the TBP-containing factor TIF-IB/SL1, thereby abrogating initiation complex formation. Moreover, TIF-IA is translocated from the nucleolus into the nucleoplasm. Substitution of Thr 200 by valine as well as knock-out of Jnk2 prevent inactivation and translocation of TIF-IA, leading to stress-resistance of Pol I transcription. Our data identify TIF-IA as a downstream target of the JNK pathway and suggest a critical role of JNK2 to protect rRNA synthesis against the harmful consequences of cellular stress.

ESCRT-II controls retinal axon growth by regulating DCC receptor levels and local protein synthesis

PubMed Central

Konopacki, Filip A.; Dwivedy, Asha; Bellon, Anaïs; Blower, Michael D.

2016-01-01

Endocytosis and local protein synthesis (LPS) act coordinately to mediate the chemotropic responses of axons, but the link between these two processes is poorly understood. The endosomal sorting complex required for transport (ESCRT) is a key regulator of cargo sorting in the endocytic pathway, and here we have investigated the role of ESCRT-II, a critical ESCRT component, in Xenopus retinal ganglion cell (RGC) axons. We show that ESCRT-II is present in RGC axonal growth cones (GCs) where it co-localizes with endocytic vesicle GTPases and, unexpectedly, with the Netrin-1 receptor, deleted in colorectal cancer (DCC). ESCRT-II knockdown (KD) decreases endocytosis and, strikingly, reduces DCC in GCs and leads to axon growth and guidance defects. ESCRT-II-depleted axons fail to turn in response to a Netrin-1 gradient in vitro and many axons fail to exit the eye in vivo. These defects, similar to Netrin-1/DCC loss-of-function phenotypes, can be rescued in whole (in vitro) or in part (in vivo) by expressing DCC. In addition, ESCRT-II KD impairs LPS in GCs and live imaging reveals that ESCRT-II transports mRNAs in axons. Collectively, our results show that the ESCRT-II-mediated endocytic pathway regulates both DCC and LPS in the axonal compartment and suggest that ESCRT-II aids gradient sensing in GCs by coupling endocytosis to LPS. PMID:27248654

Redox regulation of carbon storage and partitioning in response to light and sugars.

PubMed

Geigenberger, Peter; Kolbe, Anna; Tiessen, Axel

2005-06-01

Redox signals generated by the photosynthetic electron transport chain are known to be involved in regulating the Calvin cycle, ATP synthesis, and NADPH export from chloroplasts in response to light. The signal cascade involves transfer of electrons from photosystem I via the ferredoxin-thioredoxin system to target enzymes that are activated by reduction of regulatory disulphide bonds. The purpose of this review is to discuss recent findings showing that this concept can be extended to the regulation of carbon storage and partitioning in plants. Starch is the major carbon store in plants, and ADP-glucose pyrophosphorylase (AGPase) is the key regulatory enzyme of starch synthesis in the plastid. It has been shown that AGPase from potato tubers is subject to post-translational redox modification, and here experimental data will be provided showing that the isozyme from pea leaf chloroplasts is activated by reduced thioredoxin f or m in a similar way. Recent reports will be summarized providing in planta evidence that this mechanism regulates storage starch synthesis in response to light and sugars. Post-translational redox activation of AGPase in response to sugars is part of a signalling mechanism linking the rate of starch synthesis to the availability of carbon in diverse plant tissues. Some of the components of the signalling pathway reporting changes in the cytosolic sugar status to the plastid have been postulated, but detailed work is in progress to confirm the exact mode of action. Recent evidence will be discussed showing that key enzymes of de novo fatty acid synthesis (acetyl-CoA carboxylase) and ammonium assimilation (glutamine synthetase and glutamine:oxoglutarate amino transferase) are regulated by reversible disulphide-bond formation similar to AGPase. Redox regulation is proposed to be the preferred strategy of plastidial enzymes to regulate various metabolic processes such as carbon fixation, starch metabolism, lipid synthesis, and amino acid

Aldo-keto reductase family 1 B10 affects fatty acid synthesis by regulating the stability of acetyl-CoA carboxylase-alpha in breast cancer cells.

PubMed

Ma, Jun; Yan, Ruilan; Zu, Xuyu; Cheng, Ji-Ming; Rao, Krishna; Liao, Duan-Fang; Cao, Deliang

2008-02-08

Recent studies have demonstrated that aldo-keto reductase family 1 B10 (AKR1B10), a novel protein overexpressed in human hepatocellular carcinoma and non-small cell lung carcinoma, may facilitate cancer cell growth by detoxifying intracellular reactive carbonyls. This study presents a novel function of AKR1B10 in tumorigenic mammary epithelial cells (RAO-3), regulating fatty acid synthesis. In RAO-3 cells, Sephacryl-S 300 gel filtration and DEAE-Sepharose ion exchange chromatography demonstrated that AKR1B10 exists in two distinct forms, monomers (approximately 40 kDa) bound to DEAE-Sepharose column and protein complexes (approximately 300 kDa) remaining in flow-through. Co-immunoprecipitation with AKR1B10 antibody and protein mass spectrometry analysis identified that AKR1B10 associates with acetyl-CoA carboxylase-alpha (ACCA), a rate-limiting enzyme of de novo fatty acid synthesis. This association between AKR1B10 and ACCA proteins was further confirmed by co-immunoprecipitation with ACCA antibody and pulldown assays with recombinant AKR1B10 protein. Intracellular fluorescent studies showed that AKR1B10 and ACCA proteins co-localize in the cytoplasm of RAO-3 cells. More interestingly, small interfering RNA-mediated AKR1B10 knock down increased ACCA degradation through ubiquitination-proteasome pathway and resulted in >50% decrease of fatty acid synthesis in RAO-3 cells. These data suggest that AKR1B10 is a novel regulator of the biosynthesis of fatty acid, an essential component of the cell membrane, in breast cancer cells.

Genome-wide mRNA processing in methanogenic archaea reveals post-transcriptional regulation of ribosomal protein synthesis

PubMed Central

Qi, Lei; Yue, Lei; Feng, Deqin; Qi, Fengxia

2017-01-01

Abstract Unlike stable RNAs that require processing for maturation, prokaryotic cellular mRNAs generally follow an ‘all-or-none’ pattern. Herein, we used a 5΄ monophosphate transcript sequencing (5΄P-seq) that specifically captured the 5΄-end of processed transcripts and mapped the genome-wide RNA processing sites (PSSs) in a methanogenic archaeon. Following statistical analysis and stringent filtration, we identified 1429 PSSs, among which 23.5% and 5.4% were located in 5΄ untranslated region (uPSS) and intergenic region (iPSS), respectively. A predominant uridine downstream PSSs served as a processing signature. Remarkably, 5΄P-seq detected overrepresented uPSS and iPSS in the polycistronic operons encoding ribosomal proteins, and the majority upstream and proximal ribosome binding sites, suggesting a regulatory role of processing on translation initiation. The processed transcripts showed increased stability and translation efficiency. Particularly, processing within the tricistronic transcript of rplA-rplJ-rplL enhanced the translation of rplL, which can provide a driving force for the 1:4 stoichiometry of L10 to L12 in the ribosome. Growth-associated mRNA processing intensities were also correlated with the cellular ribosomal protein levels, thereby suggesting that mRNA processing is involved in tuning growth-dependent ribosome synthesis. In conclusion, our findings suggest that mRNA processing-mediated post-transcriptional regulation is a potential mechanism of ribosomal protein synthesis and stoichiometry. PMID:28520982

Neural-genetic synthesis for state-space controllers based on linear quadratic regulator design for eigenstructure assignment.

PubMed

da Fonseca Neto, João Viana; Abreu, Ivanildo Silva; da Silva, Fábio Nogueira

2010-04-01

Toward the synthesis of state-space controllers, a neural-genetic model based on the linear quadratic regulator design for the eigenstructure assignment of multivariable dynamic systems is presented. The neural-genetic model represents a fusion of a genetic algorithm and a recurrent neural network (RNN) to perform the selection of the weighting matrices and the algebraic Riccati equation solution, respectively. A fourth-order electric circuit model is used to evaluate the convergence of the computational intelligence paradigms and the control design method performance. The genetic search convergence evaluation is performed in terms of the fitness function statistics and the RNN convergence, which is evaluated by landscapes of the energy and norm, as a function of the parameter deviations. The control problem solution is evaluated in the time and frequency domains by the impulse response, singular values, and modal analysis.

Prolyl hydroxylation regulates protein degradation, synthesis, and splicing in human induced pluripotent stem cell-derived cardiomyocytes.

PubMed

Stoehr, Andrea; Yang, Yanqin; Patel, Sajni; Evangelista, Alicia M; Aponte, Angel; Wang, Guanghui; Liu, Poching; Boylston, Jennifer; Kloner, Philip H; Lin, Yongshun; Gucek, Marjan; Zhu, Jun; Murphy, Elizabeth

2016-06-01

Protein hydroxylases are oxygen- and α-ketoglutarate-dependent enzymes that catalyse hydroxylation of amino acids such as proline, thus linking oxygen and metabolism to enzymatic activity. Prolyl hydroxylation is a dynamic post-translational modification that regulates protein stability and protein-protein interactions; however, the extent of this modification is largely uncharacterized. The goals of this study are to investigate the biological consequences of prolyl hydroxylation and to identify new targets that undergo prolyl hydroxylation in human cardiomyocytes. We used human induced pluripotent stem cell-derived cardiomyocytes in combination with pulse-chase amino acid labelling and proteomics to analyse the effects of prolyl hydroxylation on protein degradation and synthesis. We identified 167 proteins that exhibit differences in degradation with inhibition of prolyl hydroxylation by dimethyloxalylglycine (DMOG); 164 were stabilized. Proteins involved in RNA splicing such as serine/arginine-rich splicing factor 2 (SRSF2) and splicing factor and proline- and glutamine-rich (SFPQ) were stabilized with DMOG. DMOG also decreased protein translation of cytoskeletal and sarcomeric proteins such as α-cardiac actin. We searched the mass spectrometry data for proline hydroxylation and identified 134 high confidence peptides mapping to 78 unique proteins. We identified SRSF2, SFPQ, α-cardiac actin, and cardiac titin as prolyl hydroxylated. We identified 29 prolyl hydroxylated proteins that showed a significant difference in either protein degradation or synthesis. Additionally, we performed next-generation RNA sequencing and showed that the observed decrease in protein synthesis was not due to changes in mRNA levels. Because RNA splicing factors were prolyl hydroxylated, we investigated splicing ± inhibition of prolyl hydroxylation and detected 369 alternative splicing events, with a preponderance of exon skipping. This study provides the first extensive

Alkaline stress and iron deficiency regulate iron uptake and riboflavin synthesis gene expression differently in root and leaf tissue: implications for iron deficiency chlorosis.

PubMed

Hsieh, En-Jung; Waters, Brian M

2016-10-01

Iron (Fe) is an essential mineral that has low solubility in alkaline soils, where its deficiency results in chlorosis. Whether low Fe supply and alkaline pH stress are equivalent is unclear, as they have not been treated as separate variables in molecular physiological studies. Additionally, molecular responses to these stresses have not been studied in leaf and root tissues simultaneously. We tested how plants with the Strategy I Fe uptake system respond to Fe deficiency at mildly acidic and alkaline pH by measuring root ferric chelate reductase (FCR) activity and expression of selected Fe uptake genes and riboflavin synthesis genes. Alkaline pH increased cucumber (Cucumis sativus L.) root FCR activity at full Fe supply, but alkaline stress abolished FCR response to low Fe supply. Alkaline pH or low Fe supply resulted in increased expression of Fe uptake genes, but riboflavin synthesis genes responded to Fe deficiency but not alkalinity. Iron deficiency increased expression of some common genes in roots and leaves, but alkaline stress blocked up-regulation of these genes in Fe-deficient leaves. In roots of the melon (Cucumis melo L.) fefe mutant, in which Fe uptake responses are blocked upstream of Fe uptake genes, alkaline stress or Fe deficiency up-regulation of certain Fe uptake and riboflavin synthesis genes was inhibited, indicating a central role for the FeFe protein. These results suggest a model implicating shoot-to-root signaling of Fe status to induce Fe uptake gene expression in roots. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Guanosine 3'-diphosphate 5'-diphosphate is not required for growth rate-dependent control of rRNA synthesis in Escherichia coli.

PubMed Central

Gaal, T; Gourse, R L

1990-01-01

rRNA synthesis in Escherichia coli is subject to at least two regulation systems, growth rate-dependent control and stringent control. The inverse correlation between rRNA synthesis rates and guanosine 3'-diphosphate 5'-diphosphate (ppGpp) levels under various physiological conditions has led to the supposition that ppGpp is the mediator of both control mechanisms by inhibiting transcription from rrn P1 promoters. Recently, relA- spoT- strains have been constructed in which both ppGpp synthesis pathways most likely have been removed (M. Cashel, personal communication). We have confirmed that such strains produce no detectable ppGpp and therefore offer a direct means for testing the involvement of ppGpp in the regulation of rRNA synthesis in vivo. Stringent control was determined by measurement of rRNA synthesis after amino acid starvation, while growth rate control was determined by measurement of rRNA synthesis under different nutritional conditions. As expected, the relA- spoT- strain is relaxed for stringent control. However, growth rate-dependent regulation is unimpaired. These results indicate that growth rate regulation can occur in the absence of ppGpp and imply that ppGpp is not the mediator, or at least is not the sole mediator, of growth rate-dependent control. Therefore, growth rate-dependent control and stringent control may utilize different mechanisms for regulating stable RNA synthesis. PMID:2196571

Urotensin II contributes to collagen synthesis and up-regulates Egr-1 expression in cultured pulmonary arterial smooth muscle cells through the ERK1/2 pathway

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

Li, Wei; Cai, Zhifeng; Liu, Mengmeng

Aim: The objective of this study was to investigate the effects of urotensin II (UII) treatment on the proliferation and collagen synthesis of cultured rat pulmonary arterial smooth muscle cells (PASMCs) and to explore whether these effects are mediated by mitogen-activated protein kinase (MAPK) signaling pathways and early growth response 1 (Egr-1). Methods: The proliferation of cultured PASMCs stimulated with different doses of UII was detected by BrdU incorporation. The mRNA expression levels of procollagen I (procol I), procollagen III (procol III), extracellular regulated protein kinase 1/2 (ERK1/2), stress-stimulated protein kinase (Sapk), p38 MAPK (p38), and Egr-1 mRNA in culturedmore » PASMCs after treatment with UII, the UII-specific antagonist urantide, and the ERK1/2 inhibitor PD98059 were detected by real-time polymerase chain reaction (PCR), and the protein expression levels of procol I, procol III, phosphorylated (p)-ERK1/2, p-Sapk, p-p38, and Egr-1 were detected by Western blotting. Results: Treatment with UII increased the proliferation of cultured PASMCs in a dose-dependent manner (P < 0.05). However, treatment with urantide and PD98059 inhibited the promoting effect of UII on PASMC proliferation (P < 0.05). Real-time PCR analysis showed that UII up-regulated the expression of procol I, procol III, ERK1/2, Sapk, and Egr-1 mRNA (P < 0.05), but not p38 mRNA. However, the up-regulating effect of UII was inhibited by PD98059 and urantide. Western blotting analysis showed that UII increased the synthesis of collagen I, collagen III, p-ERK1/2, p-Sapk, and Egr-1, and these effects also were inhibited by PD98059 and urantide (P < 0.05). Conclusions: Egr-1 participates in the UII-mediated proliferation and collagen synthesis of cultured rat PASMCs via activation of the ERK1/2 signaling pathway.« less

Mitochondrial protein acetylation mediates nutrient sensing of mitochondrial protein synthesis and mitonuclear protein balance.

PubMed

Di Domenico, Antonella; Hofer, Annette; Tundo, Federica; Wenz, Tina

2014-11-01

Changes in nutrient supply require global metabolic reprogramming to optimize the utilization of the nutrients. Mitochondria as a central component of the cellular metabolism play a key role in this adaptive process. Since mitochondria harbor their own genome, which encodes essential enzymes, mitochondrial protein synthesis is a determinant of metabolic adaptation. While regulation of cytoplasmic protein synthesis in response to metabolic challenges has been studied in great detail, mechanisms which adapt mitochondrial translation in response to metabolic challenges remain elusive. Our results suggest that the mitochondrial acetylation status controlled by Sirt3 and its proposed opponent GCN5L1 is an important regulator of the metabolic adaptation of mitochondrial translation. Moreover, both proteins modulate regulators of cytoplasmic protein synthesis as well as the mitonuclear protein balance making Sirt3 and GCN5L1 key players in synchronizing mitochondrial and cytoplasmic translation. Our results thereby highlight regulation of mitochondrial translation as a novel component in the cellular nutrient sensing scheme and identify mitochondrial acetylation as a new regulatory principle for the metabolic competence of mitochondrial protein synthesis. © 2014 International Union of Biochemistry and Molecular Biology.

Glucocorticoids Inhibit Basal and Hormone-Induced Serotonin Synthesis in Pancreatic Beta Cells

PubMed Central

Hasni Ebou, Moina; Singh-Estivalet, Amrit; Launay, Jean-Marie; Callebert, Jacques; Tronche, François; Ferré, Pascal; Gautier, Jean-François; Guillemain, Ghislaine; Bréant, Bernadette

2016-01-01

Diabetes is a major complication of chronic Glucocorticoids (GCs) treatment. GCs induce insulin resistance and also inhibit insulin secretion from pancreatic beta cells. Yet, a full understanding of this negative regulation remains to be deciphered. In the present study, we investigated whether GCs could inhibit serotonin synthesis in beta cell since this neurotransmitter has been shown to be involved in the regulation of insulin secretion. To this aim, serotonin synthesis was evaluated in vitro after treatment with GCs of either islets from CD1 mice or MIN6 cells, a beta-cell line. We also explored the effect of GCs on the stimulation of serotonin synthesis by several hormones such as prolactin and GLP 1. We finally studied this regulation in islet in two in vivo models: mice treated with GCs and with liraglutide, a GLP1 analog, and mice deleted for the glucocorticoid receptor in the pancreas. We showed in isolated islets and MIN6 cells that GCs decreased expression and activity of the two key enzymes of serotonin synthesis, Tryptophan Hydroxylase 1 (Tph1) and 2 (Tph2), leading to reduced serotonin contents. GCs also blocked the induction of serotonin synthesis by prolactin or by a previously unknown serotonin activator, the GLP-1 analog exendin-4. In vivo, activation of the Glucagon-like-Peptide-1 receptor with liraglutide during 4 weeks increased islet serotonin contents and GCs treatment prevented this increase. Finally, islets from mice deleted for the GR in the pancreas displayed an increased expression of Tph1 and Tph2 and a strong increased serotonin content per islet. In conclusion, our results demonstrate an original inhibition of serotonin synthesis by GCs, both in basal condition and after stimulation by prolactin or activators of the GLP-1 receptor. This regulation may contribute to the deleterious effects of GCs on beta cells. PMID:26901633

Regulation of complement C3 and C4 synthesis in human peritoneal mesothelial cells by peritoneal dialysis fluid

PubMed Central

TANG, S; LEUNG, J C K; CHAN, L Y Y; TSANG, A W L; CHEN, C X R; ZHOU, W; LAI, K N; SACKS, S H

2004-01-01

Although complement is activated in the peritoneal cavity during chronic peritoneal dialysis (PD), little is known about its role in peritoneal defence and injury related to long-term PD. We examined the impact of glucose and commercial peritoneal dialysis solutions on complement expression in HPMCs obtained by primary culture from omental tissues of consented patients undergoing elective abdominal surgery. Constitutive expression of C3 and C4 mRNA in HPMCs was up-regulated upon exposure to 75 mm glucose in a time-dependent manner. C3 and C4 protein was secreted in both apical and basolateral directions. Glucose doses beyond 100 mm markedly down-regulated C3 and C4 expression, and stimulated LDH release dose-dependently. Such cytotoxic effects were attenuated using equivalent doses of mannitol instead of glucose. Treatment with conventional lactate-buffered dialysis solution gave rise to down-regulation of C3 and C4 expression, and heightened LDH release in HPMCs. These effects correlated with the glucose strength of the solution, persisted despite replacement with a bicarbonate-buffered solution, aggravated by glycated albumin, and were partially abrogated by supplementation with 10% fetal bovine serum in the culture system. Our findings suggest that the artificial conditions imposed by PD lead to alterations in local complement synthesis that have implications for the role of the peritoneal mesothelium in both inflammation and defence. PMID:15030518

Flavonol-rich dark cocoa significantly decreases plasma endothelin-1 and improves cognition in urban children.

PubMed

Calderón-Garcidueñas, Lilian; Mora-Tiscareño, Antonieta; Franco-Lira, Maricela; Cross, Janet V; Engle, Randall; Aragón-Flores, Mariana; Gómez-Garza, Gilberto; Jewells, Valerie; Medina-Cortina, Humberto; Solorio, Edelmira; Chao, Chih-Kai; Zhu, Hongtu; Mukherjee, Partha S; Ferreira-Azevedo, Lara; Torres-Jardón, Ricardo; D'Angiulli, Amedeo

2013-01-01

Air pollution exposures are linked to systemic inflammation, cardiovascular and respiratory morbidity and mortality, neuroinflammation and neuropathology in young urbanites. In particular, most Mexico City Metropolitan Area (MCMA) children exhibit subtle cognitive deficits, and neuropathology studies show 40% of them exhibiting frontal tau hyperphosphorylation and 51% amyloid-β diffuse plaques (compared to 0% in low pollution control children). We assessed whether a short cocoa intervention can be effective in decreasing plasma endothelin 1 (ET-1) and/or inflammatory mediators in MCMA children. Thirty gram of dark cocoa with 680 mg of total flavonols were given daily for 10.11 ± 3.4 days (range 9-24 days) to 18 children (10.55 years, SD = 1.45; 11F/7M). Key metabolite ratios in frontal white matter and in hippocampus pre and during cocoa intervention were quantified by magnetic resonance spectroscopy. ET-1 significantly decreased after cocoa treatment (p = 0.0002). Fifteen children (83%) showed a marginally significant individual improvement in one or both of the applied simple short memory tasks. Endothelial dysfunction is a key feature of exposure to particulate matter (PM) and decreased endothelin-1 bioavailability is likely useful for brain function in the context of air pollution. Our findings suggest that cocoa interventions may be critical for early implementation of neuroprotection of highly exposed urban children. Multi-domain nutraceutical interventions could limit the risk for endothelial dysfunction, cerebral hypoperfusion, neuroinflammation, cognitive deficits, structural volumetric detrimental brain effects, and the early development of the neuropathological hallmarks of Alzheimer's and Parkinson's diseases.

Flavonol-rich dark cocoa significantly decreases plasma endothelin-1 and improves cognition in urban children

PubMed Central

Calderón-Garcidueñas, Lilian; Mora-Tiscareño, Antonieta; Franco-Lira, Maricela; Cross, Janet V.; Engle, Randall; Aragón-Flores, Mariana; Gómez-Garza, Gilberto; Jewells, Valerie; Weili, Lin; Medina-Cortina, Humberto; Solorio, Edelmira; Chao, Chih-kai; Zhu, Hongtu; Mukherjee, Partha S.; Ferreira-Azevedo, Lara; Torres-Jardón, Ricardo; D'Angiulli, Amedeo

2013-01-01

Air pollution exposures are linked to systemic inflammation, cardiovascular and respiratory morbidity and mortality, neuroinflammation and neuropathology in young urbanites. In particular, most Mexico City Metropolitan Area (MCMA) children exhibit subtle cognitive deficits, and neuropathology studies show 40% of them exhibiting frontal tau hyperphosphorylation and 51% amyloid-β diffuse plaques (compared to 0% in low pollution control children). We assessed whether a short cocoa intervention can be effective in decreasing plasma endothelin 1 (ET-1) and/or inflammatory mediators in MCMA children. Thirty gram of dark cocoa with 680 mg of total flavonols were given daily for 10.11 ± 3.4 days (range 9–24 days) to 18 children (10.55 years, SD = 1.45; 11F/7M). Key metabolite ratios in frontal white matter and in hippocampus pre and during cocoa intervention were quantified by magnetic resonance spectroscopy. ET-1 significantly decreased after cocoa treatment (p = 0.0002). Fifteen children (83%) showed a marginally significant individual improvement in one or both of the applied simple short memory tasks. Endothelial dysfunction is a key feature of exposure to particulate matter (PM) and decreased endothelin-1 bioavailability is likely useful for brain function in the context of air pollution. Our findings suggest that cocoa interventions may be critical for early implementation of neuroprotection of highly exposed urban children. Multi-domain nutraceutical interventions could limit the risk for endothelial dysfunction, cerebral hypoperfusion, neuroinflammation, cognitive deficits, structural volumetric detrimental brain effects, and the early development of the neuropathological hallmarks of Alzheimer's and Parkinson's diseases. PMID:23986703

Biosynthesis and engineering of kaempferol in Saccharomyces cerevisiae.

PubMed

Duan, Lijin; Ding, Wentao; Liu, Xiaonan; Cheng, Xiaozhi; Cai, Jing; Hua, Erbing; Jiang, Huifeng

2017-09-26

Kaempferol is a flavonol with broad bioactivity of anti-oxidant, anti-cancer, anti-diabetic, anti-microbial, cardio-protective and anti-asthma. Microbial synthesis of kaempferol is a promising strategy because of the low content in primary plant source. In this study, the biosynthesis pathway of kaempferol was constructed in the budding yeast Saccharomyces cerevisiae to produce kaempferol de novo, and several biological measures were taken for high production. Firstly, a high efficient flavonol synthases (FLS) from Populus deltoides was introduced into the biosynthetic pathway of kaempferol. Secondly, a S. cerevisiae recombinant was constructed for de novo synthesis of kaempferol, which generated about 6.97 mg/L kaempferol from glucose. To further promote kaempferol production, the acetyl-CoA biosynthetic pathway was overexpressed and p-coumarate was supplied as substrate, which improved kaempferol titer by about 23 and 120%, respectively. Finally, a fed-batch process was developed for better kaempferol fermentation performance, and the production reached 66.29 mg/L in 40 h. The titer of kaempferol in our engineered yeast is 2.5 times of the highest reported titer. Our study provides a possible strategy to produce kaempferol using microbial cell factory.

IL-15 regulates memory CD8+ T cell O-glycan synthesis and affects trafficking

PubMed Central

Nolz, Jeffrey C.; Harty, John T.

2014-01-01

Memory and naive CD8+ T cells exhibit distinct trafficking patterns. Specifically, memory but not naive CD8+ T cells are recruited to inflamed tissues in an antigen-independent manner. However, the molecular mechanisms that regulate memory CD8+ T cell trafficking are largely unknown. Here, using murine models of infection and T cell transfer, we found that memory but not naive CD8+ T cells dynamically regulate expression of core 2 O-glycans, which interact with P- and E-selectins to modulate trafficking to inflamed tissues. Following infection, antigen-specific effector CD8+ T cells strongly expressed core 2 O-glycans, but this glycosylation pattern was lost by most memory CD8+ T cells. After unrelated infection or inflammatory challenge, memory CD8+ T cells synthesized core 2 O-glycans independently of antigen restimulation. The presence of core 2 O-glycans subsequently directed these cells to inflamed tissue. Memory and naive CD8+ T cells exhibited the opposite pattern of epigenetic modifications at the Gcnt1 locus, which encodes the enzyme that initiates core 2 O-glycan synthesis. The open chromatin configuration in memory CD8+ T cells permitted de novo generation of core 2 O-glycans in a TCR-independent, but IL-15–dependent, manner. Thus, IL-15 stimulation promotes antigen-experienced memory CD8+ T cells to generate core 2 O-glycans, which subsequently localize them to inflamed tissues. These findings suggest that CD8+ memory T cell trafficking potentially can be manipulated to improve host defense and immunotherapy. PMID:24509081

Ribonucleotide reductase activity is regulated by proliferating cell nuclear antigen (PCNA)

PubMed Central

Salguero, Israel; Guarino, Estrella; Shepherd, Marianne; Deegan, Tom; Havens, Courtney G.; MacNeill, Stuart A.; Walter, Johannes C.; Kearsey, Stephen E.

2014-01-01

Summary Synthesis of dNTPs is required for both DNA replication and DNA repair and is catalyzed by ribonucleotide reductases (RNR), which convert ribonucleotides to their deoxy forms [1, 2]. Maintaining the correct levels of dNTPs for DNA synthesis is important for minimising the mutation rate [3-7], and this is achieved by tight regulation of ribonucleotide reductase [2, 8, 9]. In fission yeast, ribonucleotide reductase is regulated in part by a small protein inhibitor, Spd1, which is degraded in S phase and after DNA damage to allow up-regulation of dNTP supply [10-12]. Spd1 degradation is mediated by the activity of the CRL4Cdt2 ubiquitin ligase complex [5, 13, 14]. This has been reported to be dependent on modulation of Cdt2 levels which are cell cycle regulated, peaking in S phase, and which also increase after DNA damage in a checkpoint-dependent manner [7, 13]. We show here that Cdt2 levels fluctuations are not sufficient to regulate Spd1 proteolysis and that the key step in this event is the interaction of Spd1 with the polymerase processivity factor PCNA, complexed onto DNA. This mechanism thus provides a direct link between DNA synthesis and ribonucleotide reductase regulation. PMID:22464192

Neural Protein Synthesis during Aging: Effects on Plasticity and Memory

PubMed Central

Schimanski, Lesley A.; Barnes, Carol A.

2010-01-01

During aging, many experience a decline in cognitive function that includes memory loss. The encoding of long-term memories depends on new protein synthesis, and this is also reduced during aging. Thus, it is possible that changes in the regulation of protein synthesis contribute to the memory impairments observed in older animals. Several lines of evidence support this hypothesis. For instance, protein synthesis is required for a longer period following learning to establish long-term memory in aged rodents. Also, under some conditions, synaptic activity or pharmacological activation can induce de novo protein synthesis and lasting changes in synaptic transmission in aged, but not young, rodents; the opposite results can be observed in other conditions. These changes in plasticity likely play a role in manifesting the altered place field properties observed in awake and behaving aged rats. The collective evidence suggests a link between memory loss and the regulation of protein synthesis in senescence. In fact, pharmaceuticals that target the signaling pathways required for induction of protein synthesis have improved memory, synaptic plasticity, and place cell properties in aged animals. We suggest that a better understanding of the mechanisms that lead to different protein expression patterns in the neural circuits that change as a function of age will enable the development of more effective therapeutic treatments for memory loss. PMID:20802800

Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running

PubMed Central

Kato, Hiroyuki; Suzuki, Hiromi; Inoue, Yoshiko; Suzuki, Katsuya; Kobayashi, Hisamine

2016-01-01

Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR) at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d3-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control) was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise. PMID:27367725

Changes in N-acetylglutamate are involved in regulating urea synthesis in rats given a low gluten diet supplemented with L-lysine, L-methinone and L-threonine.

PubMed

Tujioka, Kazuyo; Tuchiya, Tamami; Shi, Xianglan; Ohsumi, Miho; Hayase, Kazutoshi; Yokogoshi, Hidehiko

2009-01-01

We have shown that urinary urea excretion decreased in rats fed a low gluten diet supplemented with dietary limiting amino acids. The purpose of present study was to determine whether the addition of dietary limiting amino acids to a low gluten diet affected the synthesis and degradation of N-acetylglutamate and regulated urea synthesis. Experiments were done on two groups of rats, given diets containing 10% gluten or 10% gluten+0.5% L-lysine, 0.2% L-threonine and 0.2% L-methionine for 10 d. The urinary excretion of urea, and the liver concentration of N-acetylglutamate, and the liver activity of N-acetylglutamate synthetase decreased with the addition of dietary L-lysine, L-threonine and L-methionine. N-Acetylglutamate concentration in the liver was closely correlated with the N-acetylglutamate synthetase activity in the liver and excretion of urea. The greater degradation of N-acetylglutamate was observed in the group fed the 10% gluten+L-lysine, L-threonine and L-methionine. The hepatic concentration of glutamate and plasma concentration of arginine were not related to the N-acetylglutamate concentration in the liver. These results suggest that the addition of limiting amino acids to the low gluten diet controls the synthesis and degradation of N-acetylglutamate in the liver and lowers urea synthesis.

Host Pah1p phosphatidate phosphatase limits viral replication by regulating phospholipid synthesis

PubMed Central

Zhang, Zhenlu; He, Guijuan; Catanzaro, Nicholas; Wu, Zujian; Xie, Lianhui

2018-01-01

Replication of positive-strand RNA viruses [(+)RNA viruses] takes place in membrane-bound viral replication complexes (VRCs). Formation of VRCs requires virus-mediated manipulation of cellular lipid synthesis. Here, we report significantly enhanced brome mosaic virus (BMV) replication and much improved cell growth in yeast cells lacking PAH1 (pah1Δ), the sole yeast ortholog of human LIPIN genes. PAH1 encodes Pah1p (phosphatidic acid phosphohydrolase), which converts phosphatidate (PA) to diacylglycerol that is subsequently used for the synthesis of the storage lipid triacylglycerol. Inactivation of Pah1p leads to altered lipid composition, including high levels of PA, total phospholipids, ergosterol ester, and free fatty acids, as well as expansion of the nuclear membrane. In pah1Δ cells, BMV replication protein 1a and double-stranded RNA localized to the extended nuclear membrane, there was a significant increase in the number of VRCs formed, and BMV genomic replication increased by 2-fold compared to wild-type cells. In another yeast mutant that lacks both PAH1 and DGK1 (encodes diacylglycerol kinase converting diacylglycerol to PA), which has a normal nuclear membrane but maintains similar lipid compositional changes as in pah1Δ cells, BMV replicated as efficiently as in pah1Δ cells, suggesting that the altered lipid composition was responsible for the enhanced BMV replication. We further showed that increased levels of total phospholipids play an important role because the enhanced BMV replication required active synthesis of phosphatidylcholine, the major membrane phospholipid. Moreover, overexpression of a phosphatidylcholine synthesis gene (CHO2) promoted BMV replication. Conversely, overexpression of PAH1 or plant PAH1 orthologs inhibited BMV replication in yeast or Nicotiana benthamiana plants. Competing with its host for limited resources, BMV inhibited host growth, which was markedly alleviated in pah1Δ cells. Our work suggests that Pah1p promotes

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

Regulation of Intestinal Mucosal Growth by Amino Acids

PubMed Central

Ray, Ramesh M.; Johnson, Leonard R.

2013-01-01

Amino acids, especially glutamine (GLN) have been known for many years to stimulate the growth of small intestinal mucosa. Polyamines are also required for optimal mucosal growth, and the inhibition of ornithine decarboxylase (ODC), the first rate-limiting enzyme in polyamine synthesis, blocks growth. Certain amino acids, primarily asparagine (ASN) and GLN stimulate ODC activity in a solution of physiological salts. More importantly, their presence is also required before growth factors and hormones such as EGF and insulin are able to increase ODC activity. ODC activity is inhibited by antizyme-1 (AZ) whose synthesis is stimulated by polyamines, thus, providing a negative feedback regulation of the enzyme. In the absence of amino acids mammalian target of rapamycin complex 1 (mTORC1) is inhibited, whereas, mTORC2 is stimulated leading to the inhibition of global protein synthesis but increasing the synthesis of AZ via a cap-independent mechanism. These data, therefore, explain why ASN or GLN is essential for the activation of ODC. Interestingly, in a number of papers, AZ has been shown to inhibit cell proliferation, stimulate apoptosis or increase autophagy. Each of these activities results in decreased cellular growth. AZ binds to and accelerates the degradation of ODC and other proteins shown to regulate proliferation and cell death, such as Aurora-A, Cyclin D1 and Smad1. The correlation between the stimulation of ODC activity and the absence of AZ as influenced by amino acids is high. Not only do amino acids such as ASN and GLN stimulate ODC while inhibiting AZ synthesis, but also amino acids such as lysine, valine and ornithine, which inhibit ODC activity, increase the synthesis of AZ. The question remaining to be answered is whether AZ inhibits growth directly or whether it acts by decreasing the availability of polyamines to the dividing cells. In either case, evidence strongly suggests that the regulation of AZ synthesis is the mechanism through which amino

Regulation of intestinal mucosal growth by amino acids.

PubMed

Ray, Ramesh M; Johnson, Leonard R

2014-03-01

Amino acids, especially glutamine (GLN) have been known for many years to stimulate the growth of small intestinal mucosa. Polyamines are also required for optimal mucosal growth, and the inhibition of ornithine decarboxylase (ODC), the first rate-limiting enzyme in polyamine synthesis, blocks growth. Certain amino acids, primarily asparagine (ASN) and GLN stimulate ODC activity in a solution of physiological salts. More importantly, their presence is also required before growth factors and hormones such as epidermal growth factor and insulin are able to increase ODC activity. ODC activity is inhibited by antizyme-1 (AZ) whose synthesis is stimulated by polyamines, thus, providing a negative feedback regulation of the enzyme. In the absence of amino acids mammalian target of rapamycin complex 1 (mTORC1) is inhibited, whereas, mTORC2 is stimulated leading to the inhibition of global protein synthesis but increasing the synthesis of AZ via a cap-independent mechanism. These data, therefore, explain why ASN or GLN is essential for the activation of ODC. Interestingly, in a number of papers, AZ has been shown to inhibit cell proliferation, stimulate apoptosis, or increase autophagy. Each of these activities results in decreased cellular growth. AZ binds to and accelerates the degradation of ODC and other proteins shown to regulate proliferation and cell death, such as Aurora-A, Cyclin D1, and Smad1. The correlation between the stimulation of ODC activity and the absence of AZ as influenced by amino acids is high. Not only do amino acids such as ASN and GLN stimulate ODC while inhibiting AZ synthesis, but also amino acids such as lysine, valine, and ornithine, which inhibit ODC activity, increase the synthesis of AZ. The question remaining to be answered is whether AZ inhibits growth directly or whether it acts by decreasing the availability of polyamines to the dividing cells. In either case, evidence strongly suggests that the regulation of AZ synthesis is the

Coordinate Regulation of Yeast Sterol Regulatory Element-binding Protein (SREBP) and Mga2 Transcription Factors.

PubMed

Burr, Risa; Stewart, Emerson V; Espenshade, Peter J

2017-03-31

The Mga2 and Sre1 transcription factors regulate oxygen-responsive lipid homeostasis in the fission yeast Schizosaccharomyces pombe in a manner analogous to the mammalian sterol regulatory element-binding protein (SREBP)-1 and SREBP-2 transcription factors. Mga2 and SREBP-1 regulate triacylglycerol and glycerophospholipid synthesis, whereas Sre1 and SREBP-2 regulate sterol synthesis. In mammals, a shared activation mechanism allows for coordinate regulation of SREBP-1 and SREBP-2. In contrast, distinct pathways activate fission yeast Mga2 and Sre1. Therefore, it is unclear whether and how these two related pathways are coordinated to maintain lipid balance in fission yeast. Previously, we showed that Sre1 cleavage is defective in the absence of mga2 Here, we report that this defect is due to deficient unsaturated fatty acid synthesis, resulting in aberrant membrane transport. This defect is recapitulated by treatment with the fatty acid synthase inhibitor cerulenin and is rescued by addition of exogenous unsaturated fatty acids. Furthermore, sterol synthesis inhibition blocks Mga2 pathway activation. Together, these data demonstrate that Sre1 and Mga2 are each regulated by the lipid product of the other transcription factor pathway, providing a source of coordination for these two branches of lipid synthesis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

A Csr-type regulatory system, including small non-coding RNAs, regulates the global virulence regulator RovA of Yersinia pseudotuberculosis through RovM.

PubMed

Heroven, Ann Kathrin; Böhme, Katja; Rohde, Manfred; Dersch, Petra

2008-06-01

The MarR-type regulator RovA controls expression of virulence genes of Yersinia pseudotuberculosis in response to environmental signals. Using a genetic strategy to discover components that influence rovA expression, we identified new regulatory factors with homology to components of the carbon storage regulator system (Csr). We showed that overexpression of a CsrB- or a CsrC-type RNA activates rovA, whereas a CsrA-like protein represses RovA synthesis. We further demonstrate that influence of the Csr system on rovA is indirect and occurs through control of the LysR regulator RovM, which inhibits rovA transcription. The CsrA protein had also a major influence on the motility of Yersinia, which was independent of RovM. The CsrB and CsrC RNAs are differentially expressed in Yersinia. CsrC is highly induced in complex but not in minimal media, indicating that medium-dependent rovM expression is mediated through CsrC. CsrB synthesis is generally very low. However, overexpression of the response regulator UvrY was found to activate CsrB production, which in turn represses CsrC synthesis independent of the growth medium. In summary, the post-transcriptional Csr-type components were shown to be key regulators in the co-ordinated environmental control of physiological processes and virulence factors, which are crucial for the initiation of Yersinia infections.

Ectodomain shedding of TNF receptor 1 induced by protein synthesis inhibitors regulates TNF-{alpha}-mediated activation of NF-{kappa}B and caspase-8

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

Ogura, Hirotsugu; Tsukumo, Yoshinori; Department of Bioengineering, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama 226-8501

2008-04-01

The transcription factor nuclear factor {kappa}B (NF-{kappa}B) plays a major role in the inducible resistance to death receptor-mediated apoptosis. It has been established that the protein synthesis inhibitor cycloheximide (CHX) sensitizes many types of cells to tumor necrosis factor (TNF)-{alpha}-induced apoptosis, mainly due to its ability to block de novo synthesis of cellular FLICE-inhibitory protein (c-FLIP). Nevertheless, we have surprisingly found that CHX, as well as its structural analogue acetoxycycloheximide (Ac-CHX), prevents TNF-{alpha}-mediated activation of NF-{kappa}B and caspase-8 in human lung carcinoma A549 cells. Both CHX and Ac-CHX reduced the expression of cell surface TNF receptor 1 (TNF-R1) in amore » dose-dependent manner, while Ac-CHX was approximately 100-fold more effective than CHX. Consistent with this observation, Ac-CHX induced the proteolytic cleavage of TNF-R1 and its release into the culture medium. CHX and Ac-CHX profoundly decreased constitutive and inducible expression of c-FLIP, whereas these compounds potentiated TNF-{alpha}-induced caspase-8 activation only when metalloprotease inhibitors were present. Thus, our results indicate that ectodomain shedding of TNF-R1 induced by protein synthesis inhibitors regulates TNF-{alpha}-mediated activation of NF-{kappa}B and caspase-8.« less

A melanosomal two-pore sodium channel regulates pigmentation

PubMed Central

Bellono, Nicholas W.; Escobar, Iliana E.; Oancea, Elena

2016-01-01

Intracellular organelles mediate complex cellular functions that often require ion transport across their membranes. Melanosomes are organelles responsible for the synthesis of the major mammalian pigment melanin. Defects in melanin synthesis result in pigmentation defects, visual deficits, and increased susceptibility to skin and eye cancers. Although genes encoding putative melanosomal ion transporters have been identified as key regulators of melanin synthesis, melanosome ion transport and its contribution to pigmentation remain poorly understood. Here we identify two-pore channel 2 (TPC2) as the first reported melanosomal cation conductance by directly patch-clamping skin and eye melanosomes. TPC2 has been implicated in human pigmentation and melanoma, but the molecular mechanism mediating this function was entirely unknown. We demonstrate that the vesicular signaling lipid phosphatidylinositol bisphosphate PI(3,5)P2 modulates TPC2 activity to control melanosomal membrane potential, pH, and regulate pigmentation. PMID:27231233

Genome-wide mRNA processing in methanogenic archaea reveals post-transcriptional regulation of ribosomal protein synthesis.

PubMed

Qi, Lei; Yue, Lei; Feng, Deqin; Qi, Fengxia; Li, Jie; Dong, Xiuzhu

2017-07-07

Unlike stable RNAs that require processing for maturation, prokaryotic cellular mRNAs generally follow an 'all-or-none' pattern. Herein, we used a 5΄ monophosphate transcript sequencing (5΄P-seq) that specifically captured the 5΄-end of processed transcripts and mapped the genome-wide RNA processing sites (PSSs) in a methanogenic archaeon. Following statistical analysis and stringent filtration, we identified 1429 PSSs, among which 23.5% and 5.4% were located in 5΄ untranslated region (uPSS) and intergenic region (iPSS), respectively. A predominant uridine downstream PSSs served as a processing signature. Remarkably, 5΄P-seq detected overrepresented uPSS and iPSS in the polycistronic operons encoding ribosomal proteins, and the majority upstream and proximal ribosome binding sites, suggesting a regulatory role of processing on translation initiation. The processed transcripts showed increased stability and translation efficiency. Particularly, processing within the tricistronic transcript of rplA-rplJ-rplL enhanced the translation of rplL, which can provide a driving force for the 1:4 stoichiometry of L10 to L12 in the ribosome. Growth-associated mRNA processing intensities were also correlated with the cellular ribosomal protein levels, thereby suggesting that mRNA processing is involved in tuning growth-dependent ribosome synthesis. In conclusion, our findings suggest that mRNA processing-mediated post-transcriptional regulation is a potential mechanism of ribosomal protein synthesis and stoichiometry. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Fatty acid regulation of hepatic lipid metabolism

PubMed Central

Jump, Donald B.

2012-01-01

Purpose of review To discuss transcriptional mechanisms regulating hepatic lipid metabolism. Recent findings Humans who are obese or have diabetes (NIDDM) or metabolic syndrome (MetS) have low blood and tissue levels of C20–22 polyunsaturated fatty acids (PUFAs). Although the impact of low C20–22 PUFAs on disease progression in humans is not fully understood, studies with mice have provided clues suggesting that impaired PUFA metabolism may contribute to the severity of risk factors associated with NIDDM and MetS. High fat diets promote hyperglycemia, insulin resistance and fatty liver in C57BL/6J mice, an effect that correlates with suppressed expression of enzymes involved in PUFA synthesis and decreased hepatic C20–22 PUFA content. A/J mice, in contrast, are resistant to diet-induced obesity and diabetes; these mice have elevated expression of hepatic enzymes involved in PUFA synthesis and C20–22 PUFA content. Moreover, loss-of-function and gain-of-function studies have identified fatty acid elongase (Elovl5), a key enzyme involved in PUFA synthesis, as a regulator of hepatic lipid and carbohydrate metabolism. Elovl5 activity regulates hepatic C20–22 PUFA content, signaling pathways (Akt and PP2A) and transcription factors (SREBP-1, PPARα, FoxO1 and PGC1α) that control fatty acid synthesis and gluconeogenesis. Summary These studies may help define novel strategies to control fatty liver and hyperglycemia associated with NIDDM and MetS. PMID:21178610

Fisetin, a flavonol, inhibits TH2-type cytokine production by activated human basophils.

PubMed

Higa, Shinji; Hirano, Toru; Kotani, Mayumi; Matsumoto, Motonobu; Fujita, Akihito; Suemura, Masaki; Kawase, Ichiro; Tanaka, Toshio

2003-06-01

Activation of mast cells and basophils through allergen stimulation releases chemical mediators and synthesizes cytokines. Among these cytokines, IL-4, IL-13, and IL-5 have major roles in allergic inflammation. We sought to determine the potency of flavonoids (astragalin, fisetin, kaempferol, myricetin, quercetin, and rutin) for the inhibition of cytokine expression and synthesis by human basophils. The inhibitory effect of flavonoids on cytokine expression by stimulated KU812 cells, a human basophilic cell line, and freshly purified peripheral blood basophils was measured by means of semiquantitative RT-PCR and ELISA assays. The effects of flavonoids on transcriptional activation of the nuclear factor of activated T cells were assessed by means of electrophoretic mobility shift assays. Fisetin suppressed the induction of IL-4, IL-13, and IL-5 mRNA expression by A23187-stimulated KU812 cells and basophils in response to cross-linkage of the IgE receptor. Fisetin reduced IL-4, IL-13, and IL-5 synthesis (inhibitory concentration of 50% [IC(50)] = 19.4, 17.7, and 17.4 micromol/L, respectively) but not IL-6 and IL-8 production by KU812 cells. In addition, fisetin inhibited IL-4 and IL-13 synthesis by anti-IgE antibody-stimulated human basophils (IC(50) = 5.1 and 6.2 micromol/L, respectively) and IL-4 synthesis by allergen-stimulated basophils from allergic patients (IC(50) = 4.8 micromol/L). Among the flavonoids examined, kaempferol and quercetin showed substantial inhibitory activities in cytokine expression but less so than those of fisetin. Fisetin inhibited nuclear localization of nuclear factor of activated T cells c2 by A23187-stimulated KU812 cells. These results provide evidence of a novel activity of the flavonoid fisetin that suppresses the expression of T(H)2-type cytokines (IL-4, IL-13, and IL-5) by basophils.

Induction of phytic acid synthesis by abscisic acid in suspension-cultured cells of rice.

PubMed

Matsuno, Koya; Fujimura, Tatsuhito

2014-03-01

A pathway of phytic acid (PA) synthesis in plants has been revealed via investigations of low phytic acid mutants. However, the regulation of this pathway is not well understood because it is difficult to control the environments of cells in the seeds, where PA is mainly synthesized. We modified a rice suspension culture system in order to study the regulation of PA synthesis. Rice cells cultured with abscisic acid (ABA) accumulate PA at higher levels than cells cultured without ABA, and PA accumulation levels increase with ABA concentration. On the other hand, higher concentrations of sucrose or inorganic phosphorus do not affect PA accumulation. Mutations in the genes RINO1, OsMIK, OsIPK1 and OsLPA1 have each been reported to confer low phytic acid phenotypes in seeds. Each of these genes is upregulated in cells cultured with ABA. OsITPK4 and OsITPK6 are upregulated in cells cultured with ABA and in developing seeds. These results suggest that the regulation of PA synthesis is similar between developing seeds and cells in this suspension culture system. This system will be a powerful tool for elucidating the regulation of PA synthesis. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Abscisic Acid Synthesis and Response

PubMed Central

Finkelstein, Ruth

2013-01-01

Abscisic acid (ABA) is one of the “classical” plant hormones, i.e. discovered at least 50 years ago, that regulates many aspects of plant growth and development. This chapter reviews our current understanding of ABA synthesis, metabolism, transport, and signal transduction, emphasizing knowledge gained from studies of Arabidopsis. A combination of genetic, molecular and biochemical studies has identified nearly all of the enzymes involved in ABA metabolism, almost 200 loci regulating ABA response, and thousands of genes regulated by ABA in various contexts. Some of these regulators are implicated in cross-talk with other developmental, environmental or hormonal signals. Specific details of the ABA signaling mechanisms vary among tissues or developmental stages; these are discussed in the context of ABA effects on seed maturation, germination, seedling growth, vegetative stress responses, stomatal regulation, pathogen response, flowering, and senescence. PMID:24273463

All-trans retinoic acid regulates hepatic bile acid homeostasis

PubMed Central

Yang, Fan; He, Yuqi; Liu, Hui-Xin; Tsuei, Jessica; Jiang, Xiaoyue; Yang, Li; Wang, Zheng-Tao; Wan, Yu-Jui Yvonne

2014-01-01

Retinoic acid (RA) and bile acids share common roles in regulating lipid homeostasis and insulin sensitivity. In addition, the receptor for RA (retinoid x receptor) is a permissive partner of the receptor for bile acids, farnesoid x receptor (FXR/NR1H4). Thus, RA can activate the FXR-mediated pathway as well. The current study was designed to understand the effect of all-trans RA on bile acid homeostasis. Mice were fed an all-trans RA-supplemented diet and the expression of 46 genes that participate in regulating bile acid homeostasis was studied. The data showed that all-trans RA has a profound effect in regulating genes involved in synthesis and transport of bile acids. All-trans RA treatment reduced the gene expression levels of Cyp7a1, Cyp8b1, and Akr1d1, which are involved in bile acid synthesis. All-trans RA also decreased the hepatic mRNA levels of Lrh-1 (Nr5a2) and Hnf4α (Nr2a1), which positively regulate the gene expression of Cyp7a1 and Cyp8b1. Moreover, all-trans RA induced the gene expression levels of negative regulators of bile acid synthesis including hepatic Fgfr4, Fxr, and Shp (Nr0b2) as well as ileal Fgf15. All-trans RA also decreased the expression of Abcb11 and Slc51b, which have a role in bile acid transport. Consistently, all-trans RA reduced hepatic bile acid levels and the ratio of CA/CDCA, as demonstrated by liquid chromatography-mass spectrometry. The data suggest that all-trans RA-induced SHP may contribute to the inhibition of CYP7A1 and CYP8B1, which in turn reduces bile acid synthesis and affects lipid absorption in the gastrointestinal tract. PMID:25175738

ABA and GA3 regulate the synthesis of primary and secondary metabolites related to alleviation from biotic and abiotic stresses in grapevine.

PubMed

Murcia, Germán; Fontana, Ariel; Pontin, Mariela; Baraldi, Rita; Bertazza, Gianpaolo; Piccoli, Patricia N

2017-03-01

Plants are able to synthesize a large number of organic compounds. Among them, primary metabolites are known to participate in plant growth and development, whereas secondary metabolites are mostly involved in defense and other facultative processes. In grapevine, one of the major fruit crops in the world, secondary metabolites, mainly polyphenols, are of great interest for the wine industry. Even though there is an extensive literature on the content and profile of those compounds in berries, scarce or no information is available regarding polyphenols in other organs. In addition, little is known about the effect of plant growth regulators (PGRs), ABA and GA 3 (extensively used in table grapes) on the synthesis of primary and secondary metabolites in wine grapes. In table grapes, cultural practices include the use of GA 3 sprays shortly before veraison, to increase berry and bunch size, and sugar content in fruits. Meanwhile, ABA applications to the berries on pre-veraison improve the skin coloring and sugar accumulation, anticipating the onset of veraison. Accordingly, the aim of this study was to assess and characterize primary and secondary metabolites in leaves, berries and roots of grapevine plants cv. Malbec at veraison, and changes in compositions after ABA and GA 3 aerial sprayings. Metabolic profiling was conducted using GC-MS, GC-FID and HPLC-MWD. A large set of metabolites was identified: sugars, alditols, organic acids, amino acids, polyphenols (flavonoids and non-flavonoids) and terpenes (mono-, sesqui-, di- and triterpenes). The obtained results showed that ABA applications elicited synthesis of mono- and sesquiterpenes in all assessed tissues, as well as L-proline, acidic amino acids and anthocyanins in leaves. Additionally, applications with GA 3 elicited synthesis of L-proline in berries, and mono- and sesquiterpenes in all the tissues. However, treatment with GA 3 seemed to block polyphenol synthesis, mainly in berries. In conclusion, ABA and GA

Adenosine triphosphate inhibits melatonin synthesis in the rat pineal gland.

PubMed

Souza-Teodoro, Luis Henrique; Dargenio-Garcia, Letícia; Petrilli-Lapa, Camila Lopes; Souza, Ewerton da Silva; Fernandes, Pedro A C M; Markus, Regina P; Ferreira, Zulma S

2016-03-01

Adenosine triphosphate (ATP) is released onto the pinealocyte, along with noradrenaline, from sympathetic neurons and triggers P2Y1 receptors that enhance β-adrenergic-induced N-acetylserotonin (NAS) synthesis. Nevertheless, the biotransformation of NAS into melatonin, which occurs due to the subsequent methylation by acetylserotonin O-methyltransferase (ASMT; EC 2.1.1.4), has not yet been evaluated in the presence of purinergic stimulation. We therefore evaluated the effects of purinergic signaling on melatonin synthesis induced by β-adrenergic stimulation. ATP increased NAS levels, but, surprisingly, inhibited melatonin synthesis in an inverse, concentration-dependent manner. Our results demonstrate that enhanced NAS levels, which depend on phospholipase C (PLC) activity (but not the induction of gene transcription), are a post-translational effect. By contrast, melatonin reduction is related to an ASMT inhibition of expression at both the gene transcription and protein levels. These results were independent of nuclear factor-kappa B (NF-kB) translocation. Neither the P2Y1 receptor activation nor the PLC-mediated pathway was involved in the decrease in melatonin, indicating that ATP regulates pineal metabolism through different mechanisms. Taken together, our data demonstrate that purinergic signaling differentially modulates NAS and melatonin synthesis and point to a regulatory role for ATP as a cotransmitter in the control of ASMT, the rate-limiting enzyme in melatonin synthesis. The endogenous production of melatonin regulates defense responses; therefore, understanding the mechanisms involving ASMT regulation might provide novel insights into the development and progression of neurological disorders since melatonin presents anti-inflammatory, neuroprotective, and neurogenic effects. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

PROTEASOME INHIBITOR TREATMENT REDUCED FATTY ACID, TRIACYLGLYCEROL AND CHOLESTEROL SYNTHESIS

PubMed Central

Oliva, Joan; French, Samuel W.; Li, Jun; Bardag-Gorce, Fawzia

2014-01-01

In the present study, the beneficial effects of proteasome inhibitor treatment in reducing ethanol-induced steatosis were investigated. A microarray analysis was performed on the liver of rats injected with PS-341 (Bortezomib, Velcade®), and the results showed that proteasome inhibitor treatment significantly reduced the mRNA expression of SREBP-1c, and the downstream lipogenic enzymes, such as fatty acid synthase (FAS) and acetyl-CoA carboxylase (ACC), which catalyzes the carboxylation of acetyl-CoA to malonyl-CoA, the rate-limiting step in fatty acid synthesis. ELOVL6, which is responsible for fatty acids long chain elongation, was also significantly down regulated by proteasome inhibitor treatment. Moreover, PS-341 administration significantly reduced the expression of acyl-glycerol-3-phosphate acyltransferase (AGPAT), and diacylglycerol acyltransferase (DGAT), enzyme involved in triacylglycerol (TAG) synthesis. Finally, PS-341 was found to down regulate the enzymes 3-hydroxy-3-methylglutaryl-CoenzymeA synthase (HMG-CoA synthase) that is responsible for cholesterol synthesis. Proteasome inhibitor was also found to play a role in intestinal lipid adsorption because apolipoproteins A (apoA-I, apoAII, apoA-IV and ApoCIII) were down regulated by proteasome inhibitor treatment, especially ApoA-II that is known to be a marker of alcohol consumption. Proteasome inhibitor treatment also decreased apobec-1 complementation factor (ACF) leading to lower level of editing and production of ApoB protein. Moreover apolipoprotein C-III, a major component of chylomicrons was significantly down regulated. However, lipoprotein lipase (Lpl) and High density lipoprotein binding protein (Hdlbp) mRNA levels were increased by proteasome inhibitor treatment. These results suggested that proteasome inhibitor treatment could be used to reduce the alcohol-enhanced lipogenesis and alcohol-induced liver steatosis. A morphologic analysis, performed on the liver of rats fed ethanol for one

Dietary Lipid Levels Influence Lipid Deposition in the Liver of Large Yellow Croaker (Larimichthys crocea) by Regulating Lipoprotein Receptors, Fatty Acid Uptake and Triacylglycerol Synthesis and Catabolism at the Transcriptional Level.

PubMed

Yan, Jing; Liao, Kai; Wang, Tianjiao; Mai, Kangsen; Xu, Wei; Ai, Qinghui

2015-01-01

Ectopic lipid accumulation has been observed in fish fed a high-lipid diet. However, no information is available on the mechanism by which dietary lipid levels comprehensively regulate lipid transport, uptake, synthesis and catabolism in fish. Therefore, the present study aimed to gain further insight into how dietary lipids affect lipid deposition in the liver of large yellow croaker(Larimichthys crocea). Fish (150.00±4.95 g) were fed a diet with a low (6%), moderate (12%, the control diet) or high (18%) crude lipid content for 10 weeks. Growth performance, plasma biochemical indexes, lipid contents and gene expression related to lipid deposition, including lipoprotein assembly and clearance, fatty acid uptake and triacylglycerol synthesis and catabolism, were assessed. Growth performance was not significantly affected. However, the hepato-somatic and viscera-somatic indexes as well as plasma triacylglycerol, non-esterified fatty acids and LDL-cholesterol levels were significantly increased in fish fed the high-lipid diet. In the livers of fish fed the high-lipid diet, the expression of genes related to lipoprotein clearance (LDLR) and fatty acid uptake (FABP11) was significantly up-regulated, whereas the expression of genes involved in lipoprotein assembly (apoB100), triacylglycerol synthesis and catabolism (DGAT2, CPT I) was significantly down-regulated compared with fish fed the control diet, and hepatic lipid deposition increased. In fish fed the low-lipid diet, the expression of genes associated with lipoprotein assembly and clearance (apoB100, LDLR, LRP-1), fatty acid uptake (CD36, FATP1, FABP3) and triacylglycerol synthesis (FAS) was significantly increased, whereas the expression of triacylglycerol catabolism related genes (ATGL, CPT I) was reduced compared with fish fed the control diet. However, hepatic lipid content in fish fed the low-lipid diet decreased mainly due to low dietary lipid intake. In summary, findings of this study provide molecular

Dietary Lipid Levels Influence Lipid Deposition in the Liver of Large Yellow Croaker (Larimichthys crocea) by Regulating Lipoprotein Receptors, Fatty Acid Uptake and Triacylglycerol Synthesis and Catabolism at the Transcriptional Level

PubMed Central

Yan, Jing; Liao, Kai; Wang, Tianjiao; Mai, Kangsen; Xu, Wei; Ai, Qinghui

2015-01-01

Ectopic lipid accumulation has been observed in fish fed a high-lipid diet. However, no information is available on the mechanism by which dietary lipid levels comprehensively regulate lipid transport, uptake, synthesis and catabolism in fish. Therefore, the present study aimed to gain further insight into how dietary lipids affect lipid deposition in the liver of large yellow croaker(Larimichthys crocea). Fish (150.00±4.95 g) were fed a diet with a low (6%), moderate (12%, the control diet) or high (18%) crude lipid content for 10 weeks. Growth performance, plasma biochemical indexes, lipid contents and gene expression related to lipid deposition, including lipoprotein assembly and clearance, fatty acid uptake and triacylglycerol synthesis and catabolism, were assessed. Growth performance was not significantly affected. However, the hepato-somatic and viscera-somatic indexes as well as plasma triacylglycerol, non-esterified fatty acids and LDL-cholesterol levels were significantly increased in fish fed the high-lipid diet. In the livers of fish fed the high-lipid diet, the expression of genes related to lipoprotein clearance (LDLR) and fatty acid uptake (FABP11) was significantly up-regulated, whereas the expression of genes involved in lipoprotein assembly (apoB100), triacylglycerol synthesis and catabolism (DGAT2, CPT I) was significantly down-regulated compared with fish fed the control diet, and hepatic lipid deposition increased. In fish fed the low-lipid diet, the expression of genes associated with lipoprotein assembly and clearance (apoB100, LDLR, LRP-1), fatty acid uptake (CD36, FATP1, FABP3) and triacylglycerol synthesis (FAS) was significantly increased, whereas the expression of triacylglycerol catabolism related genes (ATGL, CPT I) was reduced compared with fish fed the control diet. However, hepatic lipid content in fish fed the low-lipid diet decreased mainly due to low dietary lipid intake. In summary, findings of this study provide molecular

Regulation of Botulinum Neurotoxin Synthesis and Toxin Complex Formation by Arginine and Glucose in Clostridium botulinum ATCC 3502.

PubMed

Fredrick, Chase M; Lin, Guangyun; Johnson, Eric A

2017-07-01

Botulinum neurotoxin (BoNT), produced by neurotoxigenic clostridia, is the most potent biological toxin known and the causative agent of the paralytic disease botulism. The nutritional, environmental, and genetic regulation of BoNT synthesis, activation, stability, and toxin complex (TC) formation is not well studied. Previous studies indicated that growth and BoNT formation were affected by arginine and glucose in Clostridium botulinum types A and B. In the present study, C. botulinum ATCC 3502 was grown in toxin production medium (TPM) with different levels of arginine and glucose and of three products of arginine metabolism, citrulline, proline, and ornithine. Cultures were analyzed for growth (optical density at 600 nm [OD 600 ]), spore formation, and BoNT and TC formation by Western blotting and immunoprecipitation and for BoNT activity by mouse bioassay. A high level of arginine (20 g/liter) repressed BoNT production approximately 1,000-fold, enhanced growth, slowed lysis, and reduced endospore production by greater than 1,000-fold. Similar effects on toxin production were seen with equivalent levels of citrulline but not ornithine or proline. In TPM lacking glucose, levels of formation of BoNT/A1 and TC were significantly decreased, and extracellular BoNT and TC proteins were partially inactivated after the first day of culture. An understanding of the regulation of C. botulinum growth and BoNT and TC formation should be valuable in defining requirements for BoNT formation in foods and clinical samples, improving the quality of BoNT for pharmaceutical preparations, and elucidating the biological functions of BoNTs for the bacterium. IMPORTANCE Botulinum neurotoxin (BoNT) is a major food safety and bioterrorism concern and is also an important pharmaceutical, and yet the regulation of its synthesis, activation, and stability in culture media, foods, and clinical samples is not well understood. This paper provides insights into the effects of critical

FK506-binding protein 10 (FKBP10) regulates lung fibroblast migration via collagen VI synthesis.

PubMed

Knüppel, Larissa; Heinzelmann, Katharina; Lindner, Michael; Hatz, Rudolf; Behr, Jürgen; Eickelberg, Oliver; Staab-Weijnitz, Claudia A

2018-04-19

In idiopathic pulmonary fibrosis (IPF), fibroblasts gain a more migratory phenotype and excessively secrete extracellular matrix (ECM), ultimately leading to alveolar scarring and progressive dyspnea. Here, we analyzed the effects of deficiency of FK506-binding protein 10 (FKBP10), a potential IPF drug target, on primary human lung fibroblast (phLF) adhesion and migration. Using siRNA, FKBP10 expression was inhibited in phLF in absence or presence of 2ng/ml transforming growth factor-β1 (TGF-β1) and 0.1mM 2-phosphoascorbate. Effects on cell adhesion and migration were monitored by an immunofluorescence (IF)-based attachment assay, a conventional scratch assay, and single cell tracking by time-lapse microscopy. Effects on expression of key players in adhesion dynamics and migration were analyzed by qPCR and Western Blot. Colocalization was evaluated by IF microscopy and by proximity ligation assays. FKBP10 knockdown significantly attenuated adhesion and migration of phLF. Expression of collagen VI was decreased, while expression of key components of the focal adhesion complex was mostly upregulated. The effects on migration were 2-phosphoascorbate-dependent, suggesting collagen synthesis as the underlying mechanism. FKBP10 colocalized with collagen VI and coating culture dishes with collagen VI, and to a lesser extent with collagen I, abolished the effect of FKBP10 deficiency on migration. These findings show, to our knowledge for the first time, that FKBP10 interacts with collagen VI and that deficiency of FKBP10 reduces phLF migration mainly by downregulation of collagen VI synthesis. The results strengthen FKBP10 as an important intracellular regulator of ECM remodeling and support the concept of FKBP10 as drug target in IPF.

[Regulation of acetylcholine synthesis in presynaptic endings of cholinergic neurons of the central nervous system].

PubMed

Tuchek, S; Dolezhal, V; Richny, Ia

1984-01-01

Data on the acetylcholine (ACh) synthesis in nerve cells with special attention to its control are summarized in the paper. At rest or during moderate synaptic activity, the concentration of ACh in the compartment of its synthesis probably corresponds to the equilibrium between the substrates and products in the reaction catalysed by choline acetyltransferase. The release of ACh is followed by a transfer of ACh from the compartment of its synthesis to the compartment of release, and, automatically, by the synthesis of new ACh until a new equilibrium is reached in the compartment of synthesis. In addition, synaptic activity and the release of ACh support the synthesis of new ACh in the following ways: choline carriers are disinhibited by lowering the concentration of ACh in the nerve endings, and the transport of choline from the extracellular fluid to the cell interior according to its electro-chemical gradient is thus facilitated; the concentration of choline in the extracellular fluid is increased in the vicinity of the nerve endings as a consequence of the hydrolysis of the released ACh; postactivation hyperpolarization of the nerve endings brings about an increase of the choline transport and concentration in the nerve endings; presumably, the stimulation of muscarinic receptors brings about a further increase in the choline concentration in the vicinity of the nerve endings by the phosphatidylcholine hydrolysis intensification in postsynaptic cells; the decrease in the concentration of acetyl-CoA (as a consequence of the resynthesis of ACh) increases the activity of pyruvate dehydrogenase and the production of acetyl-CoA; conceivably, the increase in the concentration of Ca2+ ions in the nerve endings assists direct passage of acetyl-CoA from the mitochondria to the cytosol of the nerve endings, where the synthesis of ACh occurs.(ABSTRACT TRUNCATED AT 250 WORDS)

Tomato SlAN11 regulates flavonoid biosynthesis and seed dormancy by interaction with bHLH proteins but not with MYB proteins.

PubMed

Gao, Yongfeng; Liu, Jikai; Chen, Yongfu; Tang, Hai; Wang, Yang; He, Yongmei; Ou, Yongbin; Sun, Xiaochun; Wang, Songhu; Yao, Yinan

2018-01-01

The flavonoid compounds are important secondary metabolites with versatile human nutritive benefits and fulfill a multitude of functions during plant growth and development. The abundance of different flavonoid compounds are finely tuned with species-specific pattern by a ternary MBW complex, which consists of a MYB, a bHLH, and a WD40 protein, but the essential role of SlAN11, which is a WD40 protein, is not fully understood in tomato until now. In this study, a tomato WD40 protein named as SlAN11 was characterized as an effective transcription regulator to promote plant anthocyanin and seed proanthocyanidin (PA) contents, with late flavonoid biosynthetic genes activated in 35S::SlAN11 transgenic lines, while the dihydroflavonol flow to the accumulation of flavonols or their glycosylated derivatives was reduced by repressing the expression of SlFLS in this SlAN11 -overexpressed lines. The above changes were reversed in 35S::SlAN11-RNAi transgenic lines except remained levels of flavonol compounds and SlFLS expression. Interestingly, our data revealed that SlAN11 gene could affect seed dormancy by regulating the expressions of abscisic acid (ABA) signaling-related genes SlABI3 and SlABI5 , and the sensitivity to ABA treatment in seed germination is conversely changed by SlAN11 -overexpressed or -downregulated lines. Yeast two-hybrid assays demonstrated that SlAN11 interacted with bHLH but not with MYB proteins in the ternary MBW complex, whereas bHLH interacted with MYB in tomato. Our results indicated that low level of anthocyanins in tomato fruits, with low expression of bHLH ( SlTT8 ) and MYB ( SlANT1 and SlAN2 ) genes, remain unchanged upon modification of SlAN11 gene alone in the transgenic lines. These results suggest that the tomato WD40 protein SlAN11, coordinating with bHLH and MYB proteins, plays a crucial role in the fine adjustment of the flavonoid biosynthesis and seed dormancy in tomato.

Potentiation of luteolin cytotoxicity by flavonols fisetin and quercetin in human chronic lymphocytic leukemia cell lines.

PubMed

Sak, Katrin; Kasemaa, Kristi; Everaus, Hele

2016-09-14

Despite numerous studies chronic lymphocytic leukemia (CLL) still remains an incurable disease. Therefore, all new compounds and novel strategies which are able to eradicate CLL cells should be considered as valuable clues for a potential future remedy against this malignancy. In the present study, the cytotoxic profiles of natural flavonoids were described in two human CLL cell lines, HG-3 and EHEB, indicating the flavone luteolin as the most potent flavonoid with half-maximal inhibitory constants (IC50) of 37 μM and 26 μM, respectively. Luteolin significantly increased the apoptotic cell population in both cell lines by increasing the activities of caspases-3 and -9 and triggering the intrinsic apoptotic pathway. Two flavonols, fisetin and quercetin, were somewhat less efficient in suppressing cellular viability, whereas baicalein, chrysin, (+)-catechin and hesperetin exerted only a small or no response at doses as high as 100 μM. Both fisetin and quercetin were able to augment the cytotoxic activity of luteolin in both cell lines by reducing the IC50 values up to four fold. As a result of this, luteolin displayed cytotoxicity activity already at low micromolar concentrations that could potentially be physiologically achievable through oral ingestion. No other tested flavonoids were capable of sensitizing CLL cells to luteolin pointing to a specific binding of fisetin and quercetin to the cellular targets which interfere with the signaling pathways induced by luteolin. Although further molecular studies to unravel this potentiating mechanism are certainly needed, this phenomenon could contribute to future remedies for prevention and treatment of chronic lymphocytic leukemia.

Hormonal and gravitropic specificity in the regulation of growth and cell wall synthesis in pulvini and internodes from shoots of Avena sativa L. (oat).

PubMed Central

Montague, M J

1995-01-01

Segments can be cut from the peduncular-1 internode of oat (Avena sativa L.) shoots so as to contain the graviresponsive leaf-sheath pulvinus and gibberellin-sensitive internodal tissue. Incorporation of [14C]glucose was used to monitor cell wall synthesis in these two tissues as affected by gravistimulus, indoleacetic acid (IAA), gibberellic acid (GA3), and fusicoccin (FC). Pulvinar cell wall synthesis was promoted by IAA and FC (both within about 1 h), as well as by gravistimulus (starting between 3 and 6 h), whereas GA3 had no effect on nongravistimulated pulvini. In contrast, GA3 and FC promoted internodal cell wall synthesis (initiated between 1 and 2 h), whereas IAA and gravistimulus caused a decrease in internodal uptake. FC preferentially promoted incorporation into the matrix component of the wall in both tissues. Gravistimulus failed to increase responsiveness of pulvinar tissue to IAA, whereas GA3 partially overcame gravistimulus-promoted incorporation into pulvinar cell wall, probably because of preferential movement of label into the rapidly elongating internode. The results demonstrate that these eight stimulus/tissue combinations can be examined easily in an isolated 10-mm stem segment, providing new opportunities for the comparative study of tissue- and stimulus-specific events in gene regulation and signal transduction in agronomically important cereals. PMID:11536686

Inhibition of Estradiol Synthesis Impairs Fear Extinction in Male Rats

ERIC Educational Resources Information Center

Graham, Bronwyn M.; Milad, Mohammed R.

2014-01-01

Emerging research has demonstrated that the sex hormone estradiol regulates fear extinction in female rodents and women. Estradiol may also regulate fear extinction in males, given its role in synaptic plasticity in both sexes. Here we report that inhibition of estradiol synthesis during extinction training, via the aromatase inhibitor fadrozole,…

Role of Fatty-acid Synthesis in Dendritic Cell Generation and Function

PubMed Central

Rehman, Adeel; Hemmert, Keith C.; Ochi, Atsuo; Jamal, Mohsin; Henning, Justin R.; Barilla, Rocky; Quesada, Juan P.; Zambirinis, Constantinos P.; Tang, Kerry; Ego-Osuala, Melvin; Rao, Raghavendra S.; Greco, Stephanie; Deutsch, Michael; Narayan, Suchithra; Pachter, H. Leon; Graffeo, Christopher S.; Acehan, Devrim; Miller, George

2013-01-01

Dendritic cells (DC) are professional antigen presenting cells that regulate innate and adaptive immunity. The role of fatty-acid synthesis in DC development and function is uncertain. We found that blockade of fatty-acid synthesis markedly decreases dendropoiesis in the liver and in primary and secondary lymphoid organs in mice. Human DC development from PBMC precursors was also diminished by blockade of fatty-acid synthesis. This was associated with higher rates of apoptosis in precursor cells and increased expression of Cleaved Caspase 3 and BCL-xL, and down-regulation of Cyclin B1. Further, blockade of fatty-acid synthesis decreased DC expression of MHCII, ICAM-1, B7-1, B7-2 but increased their production of selected pro-inflammatory cytokines including IL-12 and MCP-1. Accordingly, inhibition of fatty-acid synthesis enhanced DC capacityto activate allogeneic as well as antigen-restricted CD4+ and CD8+ T cells and induce CTL responses. Further, blockade of fatty-acid synthesis increased DC expression of Notch ligands and enhanced their ability to activate NK cell immune-phenotype and IFN-γ production. Since endoplasmic reticular (ER)-stress can augment the immunogenic function of APC, we postulated that this may account for the higher DC immunogenicity. We found that inhibition of fatty-acid synthesis resulted in elevated expression of numerous markers of ER stress in humans and mice and was associated with increased MAP kinase and Akt signaling. Further, lowering ER-stress by 4-phenylbutyrate mitigated the enhanced immune-stimulation associated with fatty-acid synthesis blockade. Our findings elucidate the role of fatty-acid synthesis in DC development and function and have implications to the design of DC vaccines for immunotherapy. PMID:23536633

Dopamine Synthesis and D3 Receptor Activation in Pancreatic β-Cells Regulates Insulin Secretion and Intracellular [Ca2+] Oscillations

PubMed Central

Ustione, Alessandro

2012-01-01

Pancreatic islets are critical for glucose homeostasis via the regulated secretion of insulin and other hormones. We propose a novel mechanism that regulates insulin secretion from β-cells within mouse pancreatic islets: a dopaminergic negative feedback acting on insulin secretion. We show that islets are a site of dopamine synthesis and accumulation outside the central nervous system. We show that both dopamine and its precursor l-dopa inhibit glucose-stimulated insulin secretion, and this inhibition correlates with a reduction in frequency of the intracellular [Ca2+] oscillations. We further show that the effects of dopamine are abolished by a specific antagonist of the dopamine receptor D3. Because the dopamine transporter and dopamine receptors are expressed in the islets, we propose that cosecretion of dopamine with insulin activates receptors on the β-cell surface. D3 receptor activation results in changes in intracellular [Ca2+] dynamics, which, in turn, lead to lowered insulin secretion. Because blocking dopaminergic negative feedback increases insulin secretion, expanding the knowledge of this pathway in β-cells might offer a potential new target for the treatment of type 2 diabetes. PMID:22918877

MCK1 is a novel regulator of myo-inositol phosphate synthase (MIPS) that is required for inhibition of inositol synthesis by the mood stabilizer valproate

PubMed Central

Yu, Wenxi; Daniel, Joshua; Mehta, Dhara; Maddipati, Krishna Rao

2017-01-01

Myo-inositol, the precursor of all inositol compounds, is essential for the viability of eukaryotes. Identifying the factors that regulate inositol homeostasis is of obvious importance to understanding cell function and the pathologies underlying neurological and metabolic resulting from perturbation of inositol metabolism. The current study identifies Mck1, a GSK3 homolog, as a novel positive regulator of inositol de novo synthesis in yeast. Mck1 was required for normal activity of myo-inositol phosphate synthase (MIPS), which catalyzes the rate-limiting step of inositol synthesis. mck1Δ cells exhibited a 50% decrease in MIPS activity and a decreased rate of incorporation of [13C6]glucose into [13C6]-inositol-3-phosphate and [13C6]-inositol compared to WT cells. mck1Δ cells also exhibited decreased growth in the presence of the inositol depleting drug valproate (VPA), which was rescued by supplementation of inositol. However, in contrast to wild type cells, which exhibited more than a 40% decrease in MIPS activity in the presence of VPA, the drug did not significantly decrease MIPS activity in mck1Δ cells. These findings indicate that VPA-induced MIPS inhibition is Mck1-dependent, and suggest a model that unifies two current hypotheses of the mechanism of action of VPA—inositol depletion and GSK3 inhibition. PMID:28817575

MCK1 is a novel regulator of myo-inositol phosphate synthase (MIPS) that is required for inhibition of inositol synthesis by the mood stabilizer valproate.

PubMed

Yu, Wenxi; Daniel, Joshua; Mehta, Dhara; Maddipati, Krishna Rao; Greenberg, Miriam L

2017-01-01

Myo-inositol, the precursor of all inositol compounds, is essential for the viability of eukaryotes. Identifying the factors that regulate inositol homeostasis is of obvious importance to understanding cell function and the pathologies underlying neurological and metabolic resulting from perturbation of inositol metabolism. The current study identifies Mck1, a GSK3 homolog, as a novel positive regulator of inositol de novo synthesis in yeast. Mck1 was required for normal activity of myo-inositol phosphate synthase (MIPS), which catalyzes the rate-limiting step of inositol synthesis. mck1Δ cells exhibited a 50% decrease in MIPS activity and a decreased rate of incorporation of [13C6]glucose into [13C6]-inositol-3-phosphate and [13C6]-inositol compared to WT cells. mck1Δ cells also exhibited decreased growth in the presence of the inositol depleting drug valproate (VPA), which was rescued by supplementation of inositol. However, in contrast to wild type cells, which exhibited more than a 40% decrease in MIPS activity in the presence of VPA, the drug did not significantly decrease MIPS activity in mck1Δ cells. These findings indicate that VPA-induced MIPS inhibition is Mck1-dependent, and suggest a model that unifies two current hypotheses of the mechanism of action of VPA-inositol depletion and GSK3 inhibition.

Arabidopsis thaliana G2-LIKE FLAVONOID REGULATOR and BRASSINOSTEROID ENHANCED EXPRESSION1 are low-temperature regulators of flavonoid accumulation.

PubMed

Petridis, Antonios; Döll, Stefanie; Nichelmann, Lars; Bilger, Wolfgang; Mock, Hans-Peter

2016-08-01

Flavonoid synthesis is predominantly regulated at the transcriptional level through the MYB-basic helix-loop-helix (bHLH)-WD40 (MBW) (MYB: transcription factor of the myeloblastosis protein family, WD40: tanscription factor with a short structural motif of 40 amino acids which terminates in an aspartic acid-tryptophan dipeptide) complex, and responds to both environmental and developmental stimuli. Although the developmental regulation of flavonoid accumulation in Arabidopsis thaliana has been examined in great detail, the response of the flavonoid synthesis pathway to abiotic stress (particularly low temperature) remains unclear. A screen of a Dissociation element (Ds) transposon-induced mutation collection identified two lines which exhibited an altered profile of phenylpropanoid accumulation following exposure to low-temperature stress. One of the mutated genes (BRASSINOSTEROID ENHANCED EXPRESSION1 (BEE1)) encoded a brassinosteroid enhanced expression transcription factor, while the other (G2-LIKE FLAVONOID REGULATOR (GFR)) encoded a G2-like flavonoid regulator. Phenylpropanoid-targeted analysis was performed using high-performance LC-MS, and gene expression analysis using quantitative reverse transcription-PCR. In both mutants, the accumulation of quercetins and scopolin was reduced under low-temperature growing conditions, whereas that of anthocyanin was increased. BEE1 and GFR were both shown to negatively regulate anthocyanin accumulation by inhibiting anthocyanin synthesis genes via the suppression of the bHLH (TRANSPARENT TESTA8 (TT8) and GLABROUS3 (GL3)) and/or the MYB (PRODUCTION OF ANTHOCYANIN PIGMENTS2 (PAP2)) components of the MBW complex. Our results provide new insight into the regulatory control of phenylpropanoid metabolism at low temperatures, and reveal that BEE1 and GFR act as important components of the signal transduction chain. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

The soybean R2R3 MYB transcription factor GmMYB100 negatively regulates plant flavonoid biosynthesis.

PubMed

Yan, Junhui; Wang, Biao; Zhong, Yunpeng; Yao, Luming; Cheng, Linjing; Wu, Tianlong

2015-09-01

Soybean flavonoids, a group of important signaling molecules in plant-environment interaction, ubiquitously exist in soybean and are tightly regulated by many genes. Here we reported that GmMYB100, a gene encoding a R2R3 MYB transcription factor, is involved in soybean flavonoid biosynthesis. GmMYB100 is mainly expressed in flowers, leaves and immature embryo, and its level is decreased after pod ripening. Subcellular localization assay indicates that GmMYB100 is a nuclear protein. GmMYB100 has transactivation ability revealed by a yeast functional assay; whereas bioinformatic analysis suggests that GmMYB100 has a negative function in flavonoid biosynthesis. GmMYB100-overexpression represses the transcript levels of flavonoid-related genes in transgenic soybean hairy roots and Arabidopsis, and inhibits isoflavonoid (soybean) and flavonol (Arabidopsis) production in transgenic plants. Furthermore, the transcript levels of six flavonoid-related genes and flavonoid (isoflavonoid and flavone aglycones) accumulation are elevated in the GmMYB100-RNAi transgenic hairy roots. We also demonstrate that GmMYB100 protein depresses the promoter activities of soybean chalcone synthase and chalcone isomerase. These findings indicate that GmMYB100 is a negative regulator in soybean flavonoid biosynthesis pathway.

A single-base deletion in soybean flavonol synthase gene is associated with magenta flower color.

PubMed

Takahashi, Ryoji; Githiri, Stephen M; Hatayama, Kouta; Dubouzet, Emilyn G; Shimada, Norimoto; Aoki, Toshio; Ayabe, Shin-ichi; Iwashina, Tsukasa; Toda, Kyoko; Matsumura, Hisakazu

2007-01-01

The Wm locus of soybean [Glycine max (L.) Merr.] controls flower color. Dominant Wm and recessive wm allele of the locus produce purple and magenta flower, respectively. A putative full-length cDNA of flavonol synthase (FLS), gmfls1 was isolated by 5' RACE and end-to-end PCR from a cultivar Harosoy with purple flower (WmWm). Sequence analysis revealed that gmfls1 consisted of 1,208 nucleotides encoding 334 amino acids. It had 59-72% homology with FLS proteins of other plant species. Conserved dioxygenase domains A and B were found in the deduced polypeptide. Sequence comparison between Harosoy and Harosoy-wm (magenta flower mutant of Harosoy; wmwm) revealed that they differed by a single G deletion in the coding region of Harosoy-wm. The deletion changed the subsequent reading frame resulting in a truncated polypeptide consisting of 37 amino acids that lacked the dioxygenase domains A and B. Extracts of E. coli cells expressing gmfls1 of Harosoy catalyzed the formation of quercetin from dihydroquercetin, whereas cell extracts expressing gmfls1 of Harosoy-wm had no FLS activity. Genomic Southern analysis suggested the existence of three to four copies of the FLS gene in the soybean genome. CAPS analysis was performed to detect the single-base deletion. Harosoy and Clark (WmWm) exhibited longer fragments, while Harosoy-wm had shorter fragments due to the single-base deletion. The CAPS marker co-segregated with genotypes at Wm locus in a F(2) population segregating for the locus. Linkage mapping using SSR markers revealed that the Wm and gmfls1 were mapped at similar position in the molecular linkage group F. The above results strongly suggest that gmfls1 represents the Wm gene and that the single-base deletion may be responsible for magenta flower color.

Evidence for glycosylation as a regulator of the pigmentary system: key roles of sialyl(α2-6)gal/GalNAc-terminated glycans in melanin synthesis and transfer.

PubMed

Diwakar, Ganesh; Klump, Vincent; Lazova, Rossitza; Pawelek, John

2015-08-01

The major regulators of melanogenesis are glycoproteins, however no role for glycosylation in the pathway has yet been described. We stained skin biopsies and melanocyte-keratinocyte co-cultures with a panel of 20 lectins as oligosaccharide markers. Notably, the Elderberry Bark Lectin (EBL/SNA) stained melanocytes in both systems. EBL binds the sequence Neu5Ac(α(2-6)Gal/GalNAc)- at the termini of some oligosaccharide antennae. We used inhibitors of synthesis and/or binding of this sequence to assess effects on pigmentation. Cell culture, lectin histochemistry, siRNA transfection, and assays for dopa oxidase and melanin were carried out by standard techniques. 6'-sialyllactose, a short homolog of the sequence in question, anti-sialyltransferase 6 (ST6) siRNA, and cytidine, a sialyltransferase (ST) inhibitor, each inhibited EBL binding, melanogenesis and melanosome transfer. Unexpectedly, 3'-sialyllactose and siRNA for ST3, chosen as a negative controls, also inhibited these processes. Though strong inhibitors of melanization, none of the agents affected tyrosinase/dopa oxidase activity, indicating previously unrecognized post-tyrosinase regulation of melanization. We report for the first time that Neu5Ac (α(2-6)Gal/GalNAc)- and possibly Neu5Ac(α(2-3)Gal/GalNAc)-terminated oligosaccharides play multiple roles in melanin synthesis and transfer.

Engineering cell wall synthesis mechanism for enhanced PHB accumulation in E. coli.

PubMed

Zhang, Xing-Chen; Guo, Yingying; Liu, Xu; Chen, Xin-Guang; Wu, Qiong; Chen, Guo-Qiang

2018-01-01

The rigidity of bacterial cell walls synthesized by a complicated pathway limit the cell shapes as coccus, bar or ellipse or even fibers. A less rigid bacterium could be beneficial for intracellular accumulation of poly-3-hydroxybutyrate (PHB) as granular inclusion bodies. To understand how cell rigidity affects PHB accumulation, E. coli cell wall synthesis pathway was reinforced and weakened, respectively. Cell rigidity was achieved by thickening the cell walls via insertion of a constitutive gltA (encoding citrate synthase) promoter in front of a series of cell wall synthesis genes on the chromosome of several E. coli derivatives, resulting in 1.32-1.60 folds increase of Young's modulus in mechanical strength for longer E. coli cells over-expressing fission ring FtsZ protein inhibiting gene sulA. Cell rigidity was weakened by down regulating expressions of ten genes in the cell wall synthesis pathway using CRISPRi, leading to elastic cells with more spaces for PHB accumulation. The regulation on cell wall synthesis changes the cell rigidity: E. coli with thickened cell walls accumulated only 25% PHB while cell wall weakened E. coli produced 93% PHB. Manipulation on cell wall synthesis mechanism adds another possibility to morphology engineering of microorganisms. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Is Bacterial Fatty Acid Synthesis a Valid Target for Antibacterial Drug Discovery?

PubMed Central

Parsons, Joshua B.; Rock, Charles O.

2011-01-01

The emergence of resistance against most current drugs emphasizes the need to develop new approaches to control bacterial pathogens, particularly Staphylococcus aureus. Bacterial fatty acid synthesis is one such target that is being actively pursued by several research groups to develop anti-Staphylococcal agents. Recently, the wisdom of this approach has been challenged based on the ability of a Gram-positive bacterium to incorporate extracellular fatty acids and thus circumvent the inhibition of de novo fatty acid synthesis. The generality of this conclusion has been challenged, and there is enough diversity in the enzymes and regulation of fatty acid synthesis in bacteria to conclude that there isn’t a single organism that can be considered typical and representative of bacteria as a whole. We are left without a clear resolution to this ongoing debate and await new basic research to define the pathways for fatty acid uptake and that determine the biochemical and genetic mechanisms for the regulation of fatty acid synthesis in Gram-positive bacteria. These crucial experiments will determine whether diversity in the control of this important pathway accounts for the apparently different responses of Gram-positive bacteria to the inhibition of de novo fatty acid synthesis in presence of extracellular fatty acid supplements. PMID:21862391

Nitric Oxide Regulates Skeletal Muscle Fatigue, Fiber Type, Microtubule Organization, and Mitochondrial ATP Synthesis Efficiency Through cGMP-Dependent Mechanisms.

PubMed

Moon, Younghye; Balke, Jordan E; Madorma, Derik; Siegel, Michael P; Knowels, Gary; Brouckaert, Peter; Buys, Emmanuel S; Marcinek, David J; Percival, Justin M

2017-06-10

Skeletal muscle nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathways are impaired in Duchenne and Becker muscular dystrophy partly because of reduced nNOSμ and soluble guanylate cyclase (GC) activity. However, GC function and the consequences of reduced GC activity in skeletal muscle are unknown. In this study, we explore the functions of GC and NO-cGMP signaling in skeletal muscle. GC1, but not GC2, expression was higher in oxidative than glycolytic muscles. GC1 was found in a complex with nNOSμ and targeted to nNOS compartments at the Golgi complex and neuromuscular junction. Baseline GC activity and GC agonist responsiveness was reduced in the absence of nNOS. Structural analyses revealed aberrant microtubule directionality in GC1 -/- muscle. Functional analyses of GC1 -/- muscles revealed reduced fatigue resistance and postexercise force recovery that were not due to shifts in type IIA-IIX fiber balance. Force deficits in GC1 -/- muscles were also not driven by defects in resting mitochondrial adenosine triphosphate (ATP) synthesis. However, increasing muscle cGMP with sildenafil decreased ATP synthesis efficiency and capacity, without impacting mitochondrial content or ultrastructure. GC may represent a new target for alleviating muscle fatigue and that NO-cGMP signaling may play important roles in muscle structure, contractility, and bioenergetics. These findings suggest that GC activity is nNOS dependent and that muscle-specific control of GC expression and differential GC targeting may facilitate NO-cGMP signaling diversity. They suggest that nNOS regulates muscle fiber type, microtubule organization, fatigability, and postexercise force recovery partly through GC1 and suggest that NO-cGMP pathways may modulate mitochondrial ATP synthesis efficiency. Antioxid. Redox Signal. 26, 966-985.

Fragile X Mental Retardation Protein Is Required to Maintain Visual Conditioning-Induced Behavioral Plasticity by Limiting Local Protein Synthesis

PubMed Central

Liu, Han-Hsuan

2016-01-01

Fragile X mental retardation protein (FMRP) is thought to regulate neuronal plasticity by limiting dendritic protein synthesis, but direct demonstration of a requirement for FMRP control of local protein synthesis during behavioral plasticity is lacking. Here we tested whether FMRP knockdown in Xenopus optic tectum affects local protein synthesis in vivo and whether FMRP knockdown affects protein synthesis-dependent visual avoidance behavioral plasticity. We tagged newly synthesized proteins by incorporation of the noncanonical amino acid azidohomoalanine and visualized them with fluorescent noncanonical amino acid tagging (FUNCAT). Visual conditioning and FMRP knockdown produce similar increases in FUNCAT in tectal neuropil. Induction of visual conditioning-dependent behavioral plasticity occurs normally in FMRP knockdown animals, but plasticity degrades over 24 h. These results indicate that FMRP affects visual conditioning-induced local protein synthesis and is required to maintain the visual conditioning-induced behavioral plasticity. SIGNIFICANCE STATEMENT Fragile X syndrome (FXS) is the most common form of inherited intellectual disability. Exaggerated dendritic protein synthesis resulting from loss of fragile X mental retardation protein (FMRP) is thought to underlie cognitive deficits in FXS, but no direct evidence has demonstrated that FMRP-regulated dendritic protein synthesis affects behavioral plasticity in intact animals. Xenopus tadpoles exhibit a visual avoidance behavior that improves with visual conditioning in a protein synthesis-dependent manner. We showed that FMRP knockdown and visual conditioning dramatically increase protein synthesis in neuronal processes. Furthermore, induction of visual conditioning-dependent behavioral plasticity occurs normally after FMRP knockdown, but performance rapidly deteriorated in the absence of FMRP. These studies show that FMRP negatively regulates local protein synthesis and is required to maintain visual

The cholesterol, fatty acid and triglyceride synthesis pathways regulated by site 1 protease (S1P) are required for efficient replication of severe fever with thrombocytopenia syndrome virus.

PubMed

Urata, Shuzo; Uno, Yukiko; Kurosaki, Yohei; Yasuda, Jiro

2018-06-12

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by the SFTS virus (SFTSV), which has a high mortality rate. Currently, no licensed vaccines or therapeutic agents have been approved for use against SFTSV infection. Here, we report that the cholesterol, fatty acid, and triglyceride synthesis pathways regulated by S1P is involved in SFTSV replication, using CHO-K1 cell line (SRD-12B) that is deficient in site 1 protease (S1P) enzymatic activity, PF-429242, a small compound targeting S1P enzymatic activity, and Fenofibrate and Lovastatin, which inhibit triglyceride and cholesterol synthesis, respectively. These results enhance our understanding of the SFTSV replication mechanism and may contribute to the development of novel therapies for SFTSV infection. Copyright © 2018. Published by Elsevier Inc.

Anticancer activity of flavonol and flavan-3-ol rich extracts from Croton celtidifolius latex.

PubMed

Biscaro, Fernanda; Parisotto, Eduardo Benedetti; Zanette, Vanilde Citadini; Günther, Tania Mara Fischer; Ferreira, Eduardo Antonio; Gris, Eliana Fortes; Correia, João Francisco Gomes; Pich, Claus Tröger; Mattivi, Fulvio; Filho, Danilo Wilhelm; Pedrosa, Rozangela Curi

2013-06-01

Croton celtidifolius Baill (Euphorbiaceae) is a tree found in the Atlantic Forest in Southern Brazil, where it is commonly known as "Sangue-de-Dragão". Its red latex is used traditionally for treating ulcers, diabetes and cancer. To evaluate antitumor activities of Croton celtififolius latex in vitro and in vivo. Phytochemical analyses were conducted using HPLC-DAD-MS. Cytotoxic, nuclease and pro-apoptotic properties were determined using the tetrazolium salt assay (MTT), plasmid DNA damage assay and ethidium bromide (EB)/acridine orange methods, respectively, and antitumor activity was determined in the Ehrlich ascites carcinoma (EAC) mouse model. Phytochemical studies indicated a high phenol content of flavonols (45.67 ± 0.24 and 18.01 ± 0.23 mg/mL of myricetin and quercetin, respectively) and flavan-3-ols (114.12 ± 1.84 and 1527.41 ± 16.42 mg/L of epicatechin and epigallocatechin, respectively) in latex. These compounds reduced MCF-7 and EAC cell viability in the MTT assay (IC50 = 169.0 ± 1.8 and 187.0 ± 2.2 μg/mL, respectively). Latex compounds caused significant DNA fragmentation and increased the number of apoptotic cells (negative control (NC), 12%; latex, 41%) as indicated by differential staining in the EB/acridine orange assay. The in vivo latex treatment at 3.12 mg/kg/day reduced the body weight by 7.57 ± 2.04 g and increased median survival time to 17.5 days when compared to the NC group (13.0 days). In addition, the highest latex concentration inhibited tumor growth by 56%. These results agree with ethno-pharmacological reports showing cytotoxicity and antitumor activity of C. celtidifolius latex. The mechanism of antitumor action may be related to direct DNA fragmentation that reduces survival and induces apoptosis.

NMR Structural Profiling of Transcriptional Intermediates Reveals Riboswitch Regulation by Metastable RNA Conformations.

PubMed

Helmling, Christina; Wacker, Anna; Wolfinger, Michael T; Hofacker, Ivo L; Hengesbach, Martin; Fürtig, Boris; Schwalbe, Harald

2017-02-22

Gene repression induced by the formation of transcriptional terminators represents a prime example for the coupling of RNA synthesis, folding, and regulation. In this context, mapping the changes in available conformational space of transcription intermediates during RNA synthesis is important to understand riboswitch function. A majority of riboswitches, an important class of small metabolite-sensing regulatory RNAs, act as transcriptional regulators, but the dependence of ligand binding and the subsequent allosteric conformational switch on mRNA transcript length has not yet been investigated. We show a strict fine-tuning of binding and sequence-dependent alterations of conformational space by structural analysis of all relevant transcription intermediates at single-nucleotide resolution for the I-A type 2'dG-sensing riboswitch from Mesoplasma florum by NMR spectroscopy. Our results provide a general framework to dissect the coupling of synthesis and folding essential for riboswitch function, revealing the importance of metastable states for RNA-based gene regulation.

Regulation of Replication Fork Advance and Stability by Nucleosome Assembly

PubMed Central

Prado, Felix; Maya, Douglas

2017-01-01

The advance of replication forks to duplicate chromosomes in dividing cells requires the disassembly of nucleosomes ahead of the fork and the rapid assembly of parental and de novo histones at the newly synthesized strands behind the fork. Replication-coupled chromatin assembly provides a unique opportunity to regulate fork advance and stability. Through post-translational histone modifications and tightly regulated physical and genetic interactions between chromatin assembly factors and replisome components, chromatin assembly: (1) controls the rate of DNA synthesis and adjusts it to histone availability; (2) provides a mechanism to protect the integrity of the advancing fork; and (3) regulates the mechanisms of DNA damage tolerance in response to replication-blocking lesions. Uncoupling DNA synthesis from nucleosome assembly has deleterious effects on genome integrity and cell cycle progression and is linked to genetic diseases, cancer, and aging. PMID:28125036

Differential regulation of catecholamine synthesis and transport in rat adrenal medulla by fluoxetine treatment.

PubMed

Spasojevic, Natasa; Jovanovic, Predrag; Dronjak, Sladjana

2015-03-01

We have recently shown that chronic fluoxetine treatment acted significantly increasing plasma norepinephrine and epinephrine concentrations both in control and chronically stressed adult male rats. However, possible effects of fluoxetine on catecholamine synthesis and re-uptake in adrenal medulla have been largely unknown. In the present study the effects of chronic fluoxetine treatment on tyrosine hydroxylase, a rate-limiting enzyme in catecholamine synthesis, as well as a norepinephrine transporter and vesicular monoamine transporter 2 gene expressions in adrenal medulla of animals exposed to chronic unpredictable mild stress (CUMS) for 4 weeks, were investigated. Gene expression analyses were performed using a real-time quantitative reverse transcription-PCR. Chronically stressed animals had increased tyrosine hydroxylase mRNA levels and decreased expression of both transporters. Fluoxetine increased tyrosine hydroxylase and decreased norepinephrine transporter gene expression in both unstressed and CUMS rats. These findings suggest that chronic fluoxetine treatment increased plasma catecholamine levels by affecting opposing changes in catecholamine synthesis and uptake.

Prenatal programming of postnatal obesity: fetal nutrition and the regulation of leptin synthesis and secretion before birth.

PubMed

McMillen, I C; Muhlhausler, B S; Duffield, J A; Yuen, B S J

2004-08-01

Exposure to either an increased or decreased level of intrauterine nutrition can result in an increase in adiposity and in circulating leptin concentrations in later life. In animals such as the sheep and pig in which fat is deposited before birth, leptin is synthesised in fetal adipose tissue and is present in the fetal circulation throughout late gestation. In the sheep a moderate increase or decrease in the level of maternal nutrition does not alter fetal plasma leptin concentrations, but there is evidence that chronic fetal hyperglycaemia and hyperinsulinaemia increase fetal fat mass and leptin synthesis within fetal fat depots. Importantly, there is a positive relationship between the relative mass of the 'unilocular' component of fetal perirenal and interscapular adipose tissue and circulating fetal leptin concentrations in the sheep. Thus, as in the neonate and adult, circulating leptin concentrations may be a signal of fat mass in fetal life. There is also evidence that leptin can act to regulate the lipid storage, leptin synthetic capacity and potential thermogenic functions of fat before birth. Thus, leptin may act as a signal of energy supply and have a 'lipostatic' role before birth. Future studies are clearly required to determine whether the intrauterine and early postnatal nutrient environment programme the endocrine feedback loop between adipose tissue and the central and peripheral neuroendocrine systems that regulate energy balance, resulting in an enhanced risk of obesity in adult life.

Vitamins and aging: pathways to NAD+ synthesis.

PubMed

Denu, John M

2007-05-04

Recent genetic evidence reveals additional salvage pathways for NAD(+) synthesis. In this issue, Belenky et al. (2007) report that nicotinamide riboside, a new NAD(+) precursor, regulates Sir2 deacetylase activity and life span in yeast. The ability of nicotinamide riboside to enhance life span does not depend on calorie restriction.

Role of p70S6K1-mediated phosphorylation of eIF4B and PDCD4 proteins in the regulation of protein synthesis.

PubMed

Dennis, Michael D; Jefferson, Leonard S; Kimball, Scot R

2012-12-14

Modulation of mRNA binding to the 40 S ribosomal subunit during translation initiation controls not only global rates of protein synthesis but also regulates the pattern of protein expression by allowing for selective inclusion, or exclusion, of mRNAs encoding particular proteins from polysomes. The mRNA binding step is modulated by signaling through a protein kinase known as the mechanistic target of rapamycin complex 1 (mTORC1). mTORC1 directly phosphorylates the translational repressors eIF4E binding proteins (4E-BP) 1 and 2, releasing them from the mRNA cap binding protein eIF4E, thereby promoting assembly of the eIF4E·eIF4G complex. mTORC1 also phosphorylates the 70-kDa ribosomal protein S6 kinase 1 (p70S6K1), which subsequently phosphorylates eIF4B, and programmed cell death 4 (PDCD4), which sequesters eIF4A from the eIF4E·eIF4G complex, resulting in repressed translation of mRNAs with highly structured 5'-untranslated regions. In the present study, we compared the role of the 4E-BPs in the regulation of global rates of protein synthesis to that of eIF4B and PDCD4. We found that maintenance of eIF4E interaction with eIF4G was not by itself sufficient to sustain global rates of protein synthesis in the absence of mTORC1 signaling to p70S6K1; phosphorylation of both eIF4B and PDCD4 was additionally required. We also found that the interaction of eIF4E with eIF4G was maintained in the liver of fasted rats as well as in serum-deprived mouse embryo fibroblasts lacking both 4E-BP1 and 4E-BP2, suggesting that the interaction of eIF4G with eIF4E is controlled primarily through the 4E-BPs.

Phosphoproteomic analysis of the posterior silk gland of Bombyx mori provides novel insight into phosphorylation regulating the silk production.

PubMed

Song, Jia; Che, Jiaqian; You, Zhengying; Ye, Lupeng; Li, Jisheng; Zhang, Yuyu; Qian, Qiujie; Zhong, Boxiong

2016-10-04

To understand phosphorylation event regulating silk synthesis in the posterior silk gland of Bombyx mori, phosphoproteome was profiled in a pair of near-isogenic lines, a normally cocooning strain (IC) and a nakedly pupated strain (IN) that the silk production is much lower than IC. In the posterior silk gland of the IC and IN, 714 and 658 phosphosites resided on 554 and 507 phosphopeptides from 431 and 383 phosphoproteins, were identified, respectively. Of all the phosphosites, the single phosphosite was the dominate phosphorylation form, comprising>60% of all the phosphosites in two phenotypic of silk production. All these phosphosites were classified as acidophilic and proline-directed kinase classes, and three motifs were uniquely identified in the IC. The motif S-P-P might be important for regulating phosphorylation network of silk protein synthesis. The dynamically phosphorylated proteins participated in ribosome, protein transport and energy metabolism suggest that phosphorylation may play key roles in regulating silk protein synthesis and secretion. Furthermore, fibroin heavy chain, an important component of silk protein, was specifically phosphorylated in the IC strain, suggesting its role to ensure the normal formation of silk structure and silk secretion. The data gain new understanding of the regulatory processes of silk protein synthesis and offer as starting point for further research on the silk production at phosphoproteome level. Despite the knowledge on regulation of silk protein synthesis in the posterior silk gland has gained at the gene or protein levels, how phosphorylation event influences the silk yield is largely unknown. To this end, we constructed a pair of silkworm near-isogenic lines that showed different cocooning phenotypes, and the phosphoproteome of the posterior silk gland of two isolines was compared. Here, we reported the first phosphoproteome data on the silkworm and found several key pathways related protein synthesis are

Fragile X Mental Retardation Protein Is Required to Maintain Visual Conditioning-Induced Behavioral Plasticity by Limiting Local Protein Synthesis.

PubMed

Liu, Han-Hsuan; Cline, Hollis T

2016-07-06

Fragile X mental retardation protein (FMRP) is thought to regulate neuronal plasticity by limiting dendritic protein synthesis, but direct demonstration of a requirement for FMRP control of local protein synthesis during behavioral plasticity is lacking. Here we tested whether FMRP knockdown in Xenopus optic tectum affects local protein synthesis in vivo and whether FMRP knockdown affects protein synthesis-dependent visual avoidance behavioral plasticity. We tagged newly synthesized proteins by incorporation of the noncanonical amino acid azidohomoalanine and visualized them with fluorescent noncanonical amino acid tagging (FUNCAT). Visual conditioning and FMRP knockdown produce similar increases in FUNCAT in tectal neuropil. Induction of visual conditioning-dependent behavioral plasticity occurs normally in FMRP knockdown animals, but plasticity degrades over 24 h. These results indicate that FMRP affects visual conditioning-induced local protein synthesis and is required to maintain the visual conditioning-induced behavioral plasticity. Fragile X syndrome (FXS) is the most common form of inherited intellectual disability. Exaggerated dendritic protein synthesis resulting from loss of fragile X mental retardation protein (FMRP) is thought to underlie cognitive deficits in FXS, but no direct evidence has demonstrated that FMRP-regulated dendritic protein synthesis affects behavioral plasticity in intact animals. Xenopus tadpoles exhibit a visual avoidance behavior that improves with visual conditioning in a protein synthesis-dependent manner. We showed that FMRP knockdown and visual conditioning dramatically increase protein synthesis in neuronal processes. Furthermore, induction of visual conditioning-dependent behavioral plasticity occurs normally after FMRP knockdown, but performance rapidly deteriorated in the absence of FMRP. These studies show that FMRP negatively regulates local protein synthesis and is required to maintain visual conditioning

The Gcn4 transcription factor reduces protein synthesis capacity and extends yeast lifespan.

PubMed

Mittal, Nitish; Guimaraes, Joao C; Gross, Thomas; Schmidt, Alexander; Vina-Vilaseca, Arnau; Nedialkova, Danny D; Aeschimann, Florian; Leidel, Sebastian A; Spang, Anne; Zavolan, Mihaela

2017-09-06

In Saccharomyces cerevisiae, deletion of large ribosomal subunit protein-encoding genes increases the replicative lifespan in a Gcn4-dependent manner. However, how Gcn4, a key transcriptional activator of amino acid biosynthesis genes, increases lifespan, is unknown. Here we show that Gcn4 acts as a repressor of protein synthesis. By analyzing the messenger RNA and protein abundance, ribosome occupancy and protein synthesis rate in various yeast strains, we demonstrate that Gcn4 is sufficient to reduce protein synthesis and increase yeast lifespan. Chromatin immunoprecipitation reveals Gcn4 binding not only at genes that are activated, but also at genes, some encoding ribosomal proteins, that are repressed upon Gcn4 overexpression. The promoters of repressed genes contain Rap1 binding motifs. Our data suggest that Gcn4 is a central regulator of protein synthesis under multiple perturbations, including ribosomal protein gene deletions, calorie restriction, and rapamycin treatment, and provide an explanation for its role in longevity and stress response.The transcription factor Gcn4 is known to regulate yeast amino acid synthesis. Here, the authors show that Gcn4 also acts as a repressor of protein biosynthesis in a range of conditions that enhance yeast lifespan, such as ribosomal protein knockout, calorie restriction or mTOR inhibition.

WRINKLED1 Rescues Feedback Inhibition of Fatty Acid Synthesis in Hydroxylase-Expressing Seeds1[OPEN

PubMed Central

Browse, John

2016-01-01

Previous attempts at engineering Arabidopsis (Arabidopsis thaliana) to produce seed oils containing hydroxy fatty acids (HFA) have resulted in low yields of HFA compared with the native castor (Ricinus communis) plant and caused undesirable effects, including reduced total oil content. Recent studies have led to an understanding of problems involved in the accumulation of HFA in oils of transgenic plants, which include metabolic bottlenecks and a decrease in the rate of fatty acid synthesis. Focusing on engineering the triacylglycerol assembly mechanisms led to modest increases in the HFA content of seed oil, but much room for improvement still remains. We hypothesized that engineering fatty acid synthesis in the plastids to increase flux would facilitate enhanced total incorporation of fatty acids, including HFA, into seed oil. The transcription factor WRINKLED1 (WRI1) positively regulates the expression of genes involved in fatty acid synthesis and controls seed oil levels. We overexpressed Arabidopsis WRI1 in seeds of a transgenic line expressing the castor fatty acid hydroxylase. The proportion of HFA in the oil, the total HFA per seed, and the total oil content of seeds increased to an average of 20.9%, 1.26 µg, and 32.2%, respectively, across five independent lines, compared with 17.6%, 0.83 µg, and 27.9%, respectively, for isogenic segregants. WRI1 and WRI1-regulated genes involved in fatty acid synthesis were up-regulated, providing for a corresponding increase in the rate of fatty acid synthesis. PMID:27208047

Interaction of flavonols with human serum albumin: a biophysical study showing structure-activity relationship and enhancement when coated on silver nanoparticles.

PubMed

Das, Pratyusa; Chaudhari, Sunil Kumar; Das, Asmita; Kundu, Somashree; Saha, Chabita

2018-04-24

Binding affinities of flavonols namely quercetin, myricetin, and kaempferol to human serum albumin (HSA) were determined fluorimetrically and the order was observed to be myricetin > quercetin > kaempferol demonstrating structure-activity relationship. Quercetin-coated silver nanoparticles (AgNPs) show higher binding affinity to HSA compared to free quercetin with binding constants 6.04 × 10 7  M -1 and 4.2 × 10 6  M -1 , respectively. Using site-specific markers it is concluded that free quercetin and that coated on AgNPs bind at different sites. Significant structural changes in circular dichroism (CD) spectra of HSA were recorded with quercetin-coated AgNPs compared to free quercetin. These results were further substantiated by time-resolved fluorescence spectroscopy where fluorescence life time of the tryptophan residue in HSA-quercetin-coated AgNPs complex decreased to 3.63 ns from 4.22 ns in HSA-quercetin complex. Isothermal calorimetric studies reveal two binding modes for quercetin-coated AgNPs and also higher binding constants compared to free quercetin. These higher binding affinities are attributed to altered properties of quercetin when coated on AgNPs enabling it to reach the binding sites other than site II where free quercetin mainly binds.

OsbZIP58, a basic leucine zipper transcription factor, regulates starch biosynthesis in rice endosperm.

PubMed

Wang, Jie-Chen; Xu, Heng; Zhu, Ying; Liu, Qiao-Quan; Cai, Xiu-Ling

2013-08-01

Starch composition and the amount in endosperm, both of which contribute dramatically to seed yield, cooking quality, and taste in cereals, are determined by a series of complex biochemical reactions. However, the mechanism regulating starch biosynthesis in cereal seeds is not well understood. This study showed that OsbZIP58, a bZIP transcription factor, is a key transcriptional regulator controlling starch synthesis in rice endosperm. OsbZIP58 was expressed mainly in endosperm during active starch synthesis. osbzip58 null mutants displayed abnormal seed morphology with altered starch accumulation in the white belly region and decreased amounts of total starch and amylose. Moreover, osbzip58 had a higher proportion of short chains and a lower proportion of intermediate chains of amylopectin. Furthermore, OsbZIP58 was shown to bind directly to the promoters of six starch-synthesizing genes, OsAGPL3, Wx, OsSSIIa, SBE1, OsBEIIb, and ISA2, and to regulate their expression. These findings indicate that OsbZIP58 functions as a key regulator of starch synthesis in rice seeds and provide new insights into seed quality control.

Regulation of DNA replication during development

PubMed Central

Nordman, Jared; Orr-Weaver, Terry L.

2012-01-01

As development unfolds, DNA replication is not only coordinated with cell proliferation, but is regulated uniquely in specific cell types and organs. This differential regulation of DNA synthesis requires crosstalk between DNA replication and differentiation. This dynamic aspect of DNA replication is highlighted by the finding that the distribution of replication origins varies between differentiated cell types and changes with differentiation. Moreover, differential DNA replication in some cell types can lead to increases or decreases in gene copy number along chromosomes. This review highlights the recent advances and technologies that have provided us with new insights into the developmental regulation of DNA replication. PMID:22223677

Staphylococcus aureus HemX Modulates Glutamyl-tRNA Reductase Abundance To Regulate Heme Biosynthesis

PubMed Central

Choby, Jacob E.; Grunenwald, Caroline M.; Celis, Arianna I.; Gerdes, Svetlana Y.; DuBois, Jennifer L.

2018-01-01

ABSTRACT Staphylococcus aureus is responsible for a significant amount of devastating disease. Its ability to colonize the host and cause infection is supported by a variety of proteins that are dependent on the cofactor heme. Heme is a porphyrin used broadly across kingdoms and is synthesized de novo from common cellular precursors and iron. While heme is critical to bacterial physiology, it is also toxic in high concentrations, requiring that organisms encode regulatory processes to control heme homeostasis. In this work, we describe a posttranscriptional regulatory strategy in S. aureus heme biosynthesis. The first committed enzyme in the S. aureus heme biosynthetic pathway, glutamyl-tRNA reductase (GtrR), is regulated by heme abundance and the integral membrane protein HemX. GtrR abundance increases dramatically in response to heme deficiency, suggesting a mechanism by which S. aureus responds to the need to increase heme synthesis. Additionally, HemX is required to maintain low levels of GtrR in heme-proficient cells, and inactivation of hemX leads to increased heme synthesis. Excess heme synthesis in a ΔhemX mutant activates the staphylococcal heme stress response, suggesting that regulation of heme synthesis is critical to reduce self-imposed heme toxicity. Analysis of diverse organisms indicates that HemX is widely conserved among heme-synthesizing bacteria, suggesting that HemX is a common factor involved in the regulation of GtrR abundance. Together, this work demonstrates that S. aureus regulates heme synthesis by modulating GtrR abundance in response to heme deficiency and through the activity of the broadly conserved HemX. PMID:29437922

Development of a model describing regulation of casein synthesis by the mammalian target of rapamycin (mTOR) signaling pathway in response to insulin, amino acids, and acetate.

PubMed

Castro, J J; Arriola Apelo, S I; Appuhamy, J A D R N; Hanigan, M D

2016-08-01

To improve dietary protein use efficiency in lactating cows, mammary protein synthesis responses to AA, energy substrates, and hormones must be better understood. These entities exert their effects through stimulation of mRNA translation via control of initiation and elongation rates at the cellular level. A central protein kinase of this phenomenon is the mammalian target of rapamycin (mTOR), which transfers the nutritional and hormonal stimuli onto a series of proteins downstream through a cascade of phosphorylation reactions that ultimately affect protein synthesis. The objective of this work was to further develop an existing mechanistic model of mTOR phosphorylation responses to insulin and total essential AA to include the effects of specific essential AA and acetate mediated by signaling proteins including protein kinase B (Akt), adenosine monophosphate activated protein kinase (AMPK), and mTOR and to add a representation of milk protein synthesis. Data from 6 experiments in MAC-T cells and mammary tissue slices previously conducted in our laboratory were assembled and used to parameterize the dynamic system of differential equations representing Akt, AMPK, and mTOR in their phosphorylated and dephosphorylated states and the resulting regulation of milk protein synthesis. The model predicted phosphorylated Akt, mTOR, AMPK, and casein synthesis rates with root mean square prediction errors of 16.8, 28.4, 33.0, and 54.9%, respectively. All other dependent variables were free of mean and slope bias, indicating an adequate representation of the data. Whereas mTOR was not very sensitive to changes in insulin or acetate levels, it was highly sensitive to leucine and isoleucine, and this signal appeared to be effectively transduced to casein synthesis. Although prior work had observed a relationship with additional essential AA, and data supporting those conclusions were present in the data set, we were unable to derive significant relationships with any essential

GosB Inhibits Triacylglycerol Synthesis and Promotes Cell Survival in Mouse Mammary Epithelial Cells.

PubMed

Xu, Gaoxiao; Duan, Saixing; Hou, Jianye; Wei, Zhongxin; Zhao, Guangwei

2017-01-01

It has been demonstrated that the activator protein related transcription factor Finkel-Biskis-Jinkins murine osteosarcoma B (GosB) is involved in preadipocyte differentiation and triacylglycerol synthesis. However, the role of GosB in regulating the synthesis of milk fatty acid in mouse mammary glands remains unclear. This research uncovered potentially new roles of GosB in suppressing milk fatty acid synthesis. Results revealed that GosB had the highest expression in lung tissue and showed a higher expression level during nonlactation than during lactation. GosB inhibited the expression of fatty acid synthase (FASN) , stearoyl-CoA desaturase (SCD) , fatty acid binding protein 4 (FABP4) , diacylglycerol acyltransferase 1 (DGAT1) , perilipin 2 (PLIN2) , perilipin 3 (PLIN3) , and C/EBPα in mouse mammary gland epithelial cells (MEC). In addition, GosB reduced cellular triglyceride content and the accumulation of lipid droplets; in particular, GosB enhanced saturated fatty acid concentration (C16:0 and C18:0). The PPAR γ agonist, rosiglitazone (ROSI), promoted apoptosis and inhibited cell proliferation. GosB increased the expression of Bcl-2 and protected MEC from ROSI-induced apoptosis. Furthermore, MECs were protected from apoptosis through the GosB regulation of intracellular calcium concentrations. These findings suggest that GosB may regulate mammary epithelial cells milk fat synthesis and apoptosis via PPAR γ in mouse mammary glands.

Polyisoprenoid epoxides stimulate the biosynthesis of coenzyme Q and inhibit cholesterol synthesis.

PubMed

Bentinger, Magnus; Tekle, Michael; Brismar, Kerstin; Chojnacki, Tadeusz; Swiezewska, Ewa; Dallner, Gustav

2008-05-23

In our search for compounds that up-regulate the biosynthesis of coenzyme Q (CoQ), we discovered that irradiation of CoQ with ultraviolet light results in the formation of a number of compounds that influence the synthesis of mevalonate pathway lipids by HepG2 cells. Among the compounds that potently stimulated CoQ synthesis while inhibiting cholesterol synthesis, derivatives of CoQ containing 1-4 epoxide moieties in their polyisoprenoid side chains were identified. Subsequently, chemical epoxidation of all-trans-polyprenols of different lengths revealed that the shorter farnesol and geranylgeraniol derivatives were without effect, whereas the longer derivatives of solanesol enhanced CoQ and markedly reduced cholesterol biosynthesis. In contrast, none of the modified trans-trans-poly-cis-polyprenols exerted noticeable effects. Tocotrienol epoxides were especially potent in our system; those with one epoxide moiety in the side-chain generally up-regulated CoQ biosynthesis by 200-300%, whereas those with two such moieties also decreased cholesterol synthesis by 50-90%. Prolonged treatment of HepG2 cells with tocotrienol epoxides for 26 days elevated their content of CoQ by 30%. In addition, the levels of mRNA encoding enzymes involved in CoQ biosynthesis were also elevated by the tocotrienol epoxides. The site of inhibition of cholesterol synthesis was shown to be oxidosqualene cyclase. In conclusion, epoxide derivatives of certain all-trans-polyisoprenoids cause pronounced stimulation of CoQ synthesis and, in some cases, simultaneous reduction of cholesterol biosynthesis by HepG2 cells.

Flavonol and imidazole derivatives block HPV16 E6 activities and reactivate apoptotic pathways in HPV+ cells

PubMed Central

Yuan, C-H; Filippova, M; Krstenansky, J L; Duerksen-Hughes, P J

2016-01-01

High-risk human papillomaviruses (HR-HPVs) cause nearly all cases of cervical cancer, as well as approximately 30% of head and neck cancers. HPV 16 E6, one of two major viral oncogenes, protects cells from apoptosis by binding to and accelerating the degradation of several proteins important in apoptotic signaling, including caspase 8 and p53. We proposed that blocking the interactions between HPV E6 and its partners using small molecules had the potential to re-sensitize HPV+ cells to apoptosis. To test this idea, we screened libraries of small molecules for candidates that could block E6/caspase 8 binding and identified several candidates from different chemical classes. We tested hits for dose-dependency and specificity in vitro and for toxicity in a cell-based assay and then used this information to select the two best candidates for further testing: myricetin, a flavonol, and spinacine, an imidazole amino-acid derivative of histidine. Both compounds clearly inhibited the ability of E6 to bind in vitro to both caspase 8 and E6AP, the protein that mediates p53 degradation. In addition, both compounds were able to increase the level of caspase 8 and p53 in SiHa cervical cancer cells, resulting in an increase of caspase 3/7 activity. Finally, both myricetin and spinacine sensitized HPV+ cervical and oral cancer cells, but not HPV− cervical and oral cancer cells, to apoptosis induced by the cancer-specific ligand TRAIL, as well as the chemotherapeutic agents doxorubicin and cisplatin. New therapies based on this work may improve treatment for HPV+ cancer patients. PMID:26794656

CTC1-STN1 coordinates G- and C-strand synthesis to regulate telomere length.

PubMed

Gu, Peili; Jia, Shuting; Takasugi, Taylor; Smith, Eric; Nandakumar, Jayakrishnan; Hendrickson, Eric; Chang, Sandy

2018-05-17

Coats plus (CP) is a rare autosomal recessive disorder caused by mutations in CTC1, a component of the CST (CTC1, STN1, and TEN1) complex important for telomere length maintenance. The molecular basis of how CP mutations impact upon telomere length remains unclear. The CP CTC1 L1142H mutation has been previously shown to disrupt telomere maintenance. In this study, we used CRISPR/Cas9 to engineer this mutation into both alleles of HCT116 and RPE cells to demonstrate that CTC1:STN1 interaction is required to repress telomerase activity. CTC1 L1142H interacts poorly with STN1, leading to telomerase-mediated telomere elongation. Impaired interaction between CTC1 L1142H :STN1 and DNA Pol-α results in increased telomerase recruitment to telomeres and further telomere elongation, revealing that C:S binding to DNA Pol-α is required to fully repress telomerase activity. CP CTC1 mutants that fail to interact with DNA Pol-α resulted in loss of C-strand maintenance and catastrophic telomere shortening. Our findings place the CST complex as an important regulator of both G-strand extensions by telomerase and C-strand synthesis by DNA Pol-α. © 2018 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

MicroRNA29a regulates IL-33-mediated tissue remodelling in tendon disease

PubMed Central

Millar, Neal L.; Gilchrist, Derek S.; Akbar, Moeed; Reilly, James H.; Kerr, Shauna C.; Campbell, Abigail L.; Murrell, George A. C.; Liew, Foo Y.; Kurowska-Stolarska, Mariola; McInnes, Iain B.

2015-01-01

MicroRNA (miRNA) has the potential for cross-regulation and functional integration of discrete biological processes during complex physiological events. Utilizing the common human condition tendinopathy as a model system to explore the cross-regulation of immediate inflammation and matrix synthesis by miRNA we observed that elevated IL-33 expression is a characteristic of early tendinopathy. Using in vitro tenocyte cultures and in vivo models of tendon damage, we demonstrate that such IL-33 expression plays a pivotal role in the transition from type 1 to type 3 collagen (Col3) synthesis and thus early tendon remodelling. Both IL-33 effector function, via its decoy receptor sST2, and Col3 synthesis are regulated by miRNA29a. Downregulation of miRNA29a in human tenocytes is sufficient to induce an increase in Col3 expression. These data provide a molecular mechanism of miRNA-mediated integration of the early pathophysiologic events that facilitate tissue remodelling in human tendon after injury. PMID:25857925

Thermo-Regulation of Genes Mediating Motility and Plant Interactions in Pseudomonas syringae

PubMed Central

Hockett, Kevin L.; Burch, Adrien Y.; Lindow, Steven E.

2013-01-01

Pseudomonas syringae is an important phyllosphere colonist that utilizes flagellum-mediated motility both as a means to explore leaf surfaces, as well as to invade into leaf interiors, where it survives as a pathogen. We found that multiple forms of flagellum-mediated motility are thermo-suppressed, including swarming and swimming motility. Suppression of swarming motility occurs between 28° and 30°C, which coincides with the optimal growth temperature of P. syringae. Both fliC (encoding flagellin) and syfA (encoding a non-ribosomal peptide synthetase involved in syringafactin biosynthesis) were suppressed with increasing temperature. RNA-seq revealed 1440 genes of the P. syringae genome are temperature sensitive in expression. Genes involved in polysaccharide synthesis and regulation, phage and IS elements, type VI secretion, chemosensing and chemotaxis, translation, flagellar synthesis and motility, and phytotoxin synthesis and transport were generally repressed at 30°C, while genes involved in transcriptional regulation, quaternary ammonium compound metabolism and transport, chaperone/heat shock proteins, and hypothetical genes were generally induced at 30°C. Deletion of flgM, a key regulator in the transition from class III to class IV gene expression, led to elevated and constitutive expression of fliC regardless of temperature, but did not affect thermo-regulation of syfA. This work highlights the importance of temperature in the biology of P. syringae, as many genes encoding traits important for plant-microbe interactions were thermo-regulated. PMID:23527276

Gadd34 Requirement for Normal Hemoglobin Synthesis

PubMed Central

Patterson, Andrew D.; Hollander, M. Christine; Miller, Georgina F.; Fornace, Albert J.

2006-01-01

The protein encoded by growth arrest and DNA damage-inducible transcript 34 (Gadd34) is associated with translation initiation regulation following certain stress responses. Through interaction with the protein phosphatase 1 catalytic subunit (PP1c), Gadd34 recruits PP1c for the removal of an inhibitory phosphate group on the α subunit of elongation initiation factor 2, thereby reversing the shutoff of protein synthesis initiated by stress-inducible kinases. In the absence of stress, the physiologic consequences of Gadd34 function are not known. Initial analysis of Gadd34-null mice revealed several significant findings, including hypersplenism, decreased erythrocyte volume, increased numbers of circulating erythrocytes, and decreased hemoglobin content, resembling some thalassemia syndromes. Biochemical analysis of the hemoglobin-producing reticulocyte (an erythrocyte precursor) revealed that the decreased hemoglobin content in the Gadd34-null erythrocyte is due to the reduced initiation of the globin translation machinery. We propose that an equilibrium state exists between Gadd34/PP1c and the opposing heme-regulated inhibitor kinase during hemoglobin synthesis in the reticulocyte. PMID:16478986

Down-regulation of UDP-glucose dehydrogenase affects glycosaminoglycans synthesis and motility in HCT-8 colorectal carcinoma cells

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

Wang, Tsung-Pao; Pan, Yun-Ru; Fu, Chien-Yu

2010-10-15

UDP-glucose dehydrogenase (UGDH) catalyzes oxidation of UDP-glucose to yield UDP-glucuronic acid, a precursor of hyaluronic acid (HA) and other glycosaminoglycans (GAGs) in extracellular matrix. Although association of extracellular matrix with cell proliferation and migration has been well documented, the importance of UGDH in these behaviors is not clear. Using UGDH-specific small interference RNA to treat HCT-8 colorectal carcinoma cells, a decrease in both mRNA and protein levels of UGDH, as well as the cellular UDP-glucuronic acid and GAG production was observed. Treatment of HCT-8 cells with either UGDH-specific siRNA or HA synthesis inhibitor 4-methylumbelliferone effectively delayed cell aggregation into multicellularmore » spheroids and impaired cell motility in both three-dimensional collagen gel and transwell migration assays. The reduction in cell aggregation and migration rates could be restored by addition of exogenous HA. These results indicate that UGDH can regulate cell motility through the production of GAG. The enzyme may be a potential target for therapeutic intervention of colorectal cancers.« less

Sterol regulatory element-binding protein-1 participates in the regulation of fatty acid synthase expression in colorectal neoplasia.

PubMed

Li, J N; Mahmoud, M A; Han, W F; Ripple, M; Pizer, E S

2000-11-25

Endogenous fatty acid synthesis has been observed in certain rapidly proliferating normal and neoplastic tissues. Sterol regulatory element-binding proteins (SREBPs) are transcription factors that regulate the expression of lipogenic genes including fatty acid synthase (FAS), the major biosynthetic enzyme for fatty acid synthesis. We have previously shown that SREBP-1, FAS, and Ki-67, a proliferation marker, colocalized in the crypts of the fetal gastrointestinal tract epithelium. This study sought to determine whether SREBP-1 participates in the regulation of proliferation-associated fatty acid synthesis in colorectal neoplasia. An immunohistochemical analysis of SREBP-1, FAS, and Ki-67 expression in 25 primary human colorectal carcinoma specimens showed colocalization in 22 of these. To elucidate a functional linkage between SREBP-1 activation and proliferation-associated FA synthesis, SREBP-1 and FAS content were assayed during the adaptive response of cultured HCT116 colon carcinoma cells to pharmacological inhibition of FA synthesis. Cerulenin and TOFA each inhibited the endogenous synthesis of fatty acids in a dose-dependent manner and each induced increases in both precursor and mature forms of SREBP-1. Subsequently, both the transcriptional activity of the FAS promoter in a luciferase reporter gene construct and the FAS expression increased. These results demonstrate that tumor cells recognize and respond to a deficiency in endogenous fatty acid synthesis by upregulating both SREBP-1 and FAS expression and support the model that SREBP-1 participates in the transcriptional regulation of lipogenic genes in colorectal neoplasia. Copyright 2000 Academic Press.

Lipoic acid metabolism and mitochondrial redox regulation.

PubMed

Solmonson, Ashley D; DeBerardinis, Ralph J

2017-11-30

Lipoic acid is an essential cofactor for mitochondrial metabolism and is synthesized de novo using intermediates from mitochondrial fatty acid synthesis type II, S-adenosylmethionine and iron-sulfur clusters. This cofactor is required for catalysis by multiple mitochondrial 2-ketoacid dehydrogenase complexes, including pyruvate dehydrogenase, alpha-ketoglutarate dehydrogenase, and branched-chain ketoacid dehydrogenase. Lipoic acid also plays a critical role in stabilizing and regulating these multi-enzyme complexes.  Many of these dehydrogenases are regulated by reactive oxygen species, mediated through the disulfide bond of the prosthetic lipoyl moiety.  Collectively, its functions explain why lipoic acid is required for cell growth, mitochondrial activity and coordination of fuel metabolism. Lipoic acid is an essential cofactor for mitochondrial metabolism and is synthesized de novo using intermediates from mitochondrial fatty acid synthesis type II, S-adenosylmethionine and iron-sulfur clusters. This cofactor is required for catalysis by multiple mitochondrial 2-ketoacid dehydrogenase complexes, including pyruvate dehydrogenase, alpha-ketoglutarate dehydrogenase, and branched-chain ketoacid dehydrogenase. Lipoic acid also plays a critical role in stabilizing and regulating these multi-enzyme complexes.  Many of these dehydrogenases are regulated by reactive oxygen species, mediated through the disulfide bond of the prosthetic lipoyl moiety.  Collectively, its functions explain why lipoic acid is required for cell growth, mitochondrial activity and coordination of fuel metabolism. Copyright © 2017, The American Society for Biochemistry and Molecular Biology.

Facile synthesis of diverse graphene nanomeshes based on simultaneous regulation of pore size and surface structure.

PubMed

Zhang, Jia; Song, Huaibing; Zeng, Dawen; Wang, Hao; Qin, Ziyu; Xu, Keng; Pang, Aimin; Xie, Changsheng

2016-08-26

Recently, graphene nanomesh (GNM) has attracted great attentions due to its unique porous structure, abundant active sites, finite band gap and possesses potential applications in the fields of electronics, gas sensor/storage, catalysis, etc. Therefore, diverse GNMs with different physical and chemical properties are required urgently to meet different applications. Herein we demonstrate a facile synthetic method based on the famous Fenton reaction to prepare GNM, by using economically fabricated graphene oxide (GO) as a starting material. By precisely controlling the reaction time, simultaneous regulation of pore size from 2.9 to 11.1 nm and surface structure can be realized. Ultimately, diverse GNMs with tunable band gap and work function can be obtained. Specially, the band gap decreases from 4.5-2.3 eV for GO, which is an insulator, to 3.9-1.24 eV for GNM-5 h, which approaches to a semiconductor. The dual nature of electrophilic addition and oxidizability of HO(•) is responsible for this controllable synthesis. This efficient, low-cost, inherently scalable synthetic method is suitable for provide diverse and optional GNMs, and may be generalized to a universal technique.

Facile synthesis of diverse graphene nanomeshes based on simultaneous regulation of pore size and surface structure

NASA Astrophysics Data System (ADS)

Zhang, Jia; Song, Huaibing; Zeng, Dawen; Wang, Hao; Qin, Ziyu; Xu, Keng; Pang, Aimin; Xie, Changsheng

2016-08-01

Recently, graphene nanomesh (GNM) has attracted great attentions due to its unique porous structure, abundant active sites, finite band gap and possesses potential applications in the fields of electronics, gas sensor/storage, catalysis, etc. Therefore, diverse GNMs with different physical and chemical properties are required urgently to meet different applications. Herein we demonstrate a facile synthetic method based on the famous Fenton reaction to prepare GNM, by using economically fabricated graphene oxide (GO) as a starting material. By precisely controlling the reaction time, simultaneous regulation of pore size from 2.9 to 11.1 nm and surface structure can be realized. Ultimately, diverse GNMs with tunable band gap and work function can be obtained. Specially, the band gap decreases from 4.5-2.3 eV for GO, which is an insulator, to 3.9-1.24 eV for GNM-5 h, which approaches to a semiconductor. The dual nature of electrophilic addition and oxidizability of HO• is responsible for this controllable synthesis. This efficient, low-cost, inherently scalable synthetic method is suitable for provide diverse and optional GNMs, and may be generalized to a universal technique.

Regulation of carotenoid and ABA accumulation during the development and germination of Nicotiana plumbaginifolia seeds.

PubMed

Frey, Anne; Boutin, Jean-Pierre; Sotta, Bruno; Mercier, Raphaël; Marion-Poll, Annie

2006-08-01

Abscisic acid (ABA) is derived from epoxycarotenoid cleavage and regulates seed development and maturation. A detailed carotenoid analysis was undertaken to study the contribution of epoxycarotenoid synthesis to the regulation of ABA accumulation in Nicotiana plumbaginifolia developing seeds. Maximal accumulation of xanthophylls occurred at mid-development in wild type seeds, when total ABA levels also peaked. In contrast, in ABA-deficient mutants xanthophyll synthesis was delayed, in agreement with the retardation in seed maturation. Seed dormancy was restored in mutants impaired in the conversion of zeaxanthin into violaxanthin by zeaxanthin epoxidase (ZEP), by the introduction of the Arabidopsis AtZEP gene under the control of promoters inducing expression during later stages of seed development compared to wild type NpZEP, and in dry and imbibed seeds. Alterations in the timing and level of ZEP expression did not highly affect the temporal regulation of ABA accumulation in transgenic seeds, despite notable perturbations in xanthophyll accumulation. Therefore, major regulatory control of ABA accumulation might occur downstream of epoxycarotenoid synthesis.

Synthesis of aromatic cytokinins for plant biotechnology.

PubMed

Plíhalová, Lucie; Vylíčilová, Hana; Doležal, Karel; Zahajská, Lenka; Zatloukal, Marek; Strnad, Miroslav

2016-09-25

Cytokinins represent an important group of plant growth regulators that can modulate several biotechnological processes owing to their ability to influence almost all stages of plant development and growth. In addition, the use of purine based cytokinins with aromatic substituent in C6 position of the purine moiety in tissue culture techniques is currently experiencing a surge in interest, made possible by the ongoing systematic synthesis and study of these compounds. This review article outlines progress in the synthesis of aromatic cytokinins, the in vitro and in vivo effects of these substances and insights gleaned from their synthesis. As the purine moiety in these compounds can be substituted at several positions, we examine each of the substitution possibilities in relation to the derivatives prepared so far. The discussion highlights the gradual simplification of their preparation in relation to their application in practice and summarizes the relevant organic chemistry literature and published patents. Copyright © 2015 Elsevier B.V. All rights reserved.

Sterols regulate 3β-hydroxysterol Δ24-reductase (DHCR24) via dual sterol regulatory elements: cooperative induction of key enzymes in lipid synthesis by Sterol Regulatory Element Binding Proteins.

PubMed

Zerenturk, Eser J; Sharpe, Laura J; Brown, Andrew J

2012-10-01

3β-Hydroxysterol Δ24-reductase (DHCR24) catalyzes a final step in cholesterol synthesis, and has been ascribed diverse functions, such as being anti-apoptotic and anti-inflammatory. How this enzyme is regulated transcriptionally by sterols is currently unclear. Some studies have suggested that its expression is regulated by Sterol Regulatory Element Binding Proteins (SREBPs) while another suggests it is through the Liver X Receptor (LXR). However, these transcription factors have opposing effects on cellular sterol levels, so it is likely that one predominates. Here we establish that sterol regulation of DHCR24 occurs predominantly through SREBP-2, and identify the particular region of the DHCR24 promoter to which SREBP-2 binds. We demonstrate that sterol regulation is mediated by two sterol regulatory elements (SREs) in the promoter of the gene, assisted by two nearby NF-Y binding sites. Moreover, we present evidence that the dual SREs work cooperatively to regulate DHCR24 expression by comparison to two known SREBP target genes, the LDL receptor with one SRE, and farnesyl-diphosphate farnesyltransferase 1, with two SREs. Copyright © 2012 Elsevier B.V. All rights reserved.

Synthesis of 8-Aryl-O-methylcyanidins and Their Usage for Dye-Sensitized Solar Cell Devices

PubMed Central

Kimura, Yuki; Oyama, Kin-ichi; Murata, Yasujiro; Wakamiya, Atsushi; Yoshida, Kumi

2017-01-01

Anthocyanins as natural pigments are colorful and environmentally compatible dyes for dye-sensitized solar cells (DSSCs). To increase the efficiency, we designed and synthesized unnatural O-methylflavonols and O-methylcyanidins that possess an aryl group at the 8-position. We synthesized per-O-methylquercetin from quercetin, then using selective demethylation prepared various O-methylquercetins. Using the Suzuki-Miyaura coupling reaction, 8-arylation of per-O-methylquercetin was achieved. Using a LiAlH4 reduction or Clemmensen reduction, these flavonols were transformed to the corresponding cyanidin derivatives in satisfactory yields. Using these dyes, we fabricated DSSCs, and their efficiency was investigated. The efficiency of tetra-O-methylflavonol was 0.31%. However, the introduction of the 8-aryl residue increased the efficiency to 1.04%. In comparison to these flavonols, O-methylcyanidins exhibited a lower efficiency of 0.05% to 0.52%. The introduction of the 8-aryl group into the cyanidin derivatives did not result in a remarkable increase in the efficiency. These phenomena may be due to the poor fit of the HOMO-LUMO level of the dyes to the TiO2 conduction band. PMID:28212330

Synthesis of 8-Aryl-O-methylcyanidins and Their Usage for Dye-Sensitized Solar Cell Devices.

PubMed

Kimura, Yuki; Oyama, Kin-Ichi; Murata, Yasujiro; Wakamiya, Atsushi; Yoshida, Kumi

2017-02-16

Anthocyanins as natural pigments are colorful and environmentally compatible dyes for dye-sensitized solar cells (DSSCs). To increase the efficiency, we designed and synthesized unnatural O -methylflavonols and O -methylcyanidins that possess an aryl group at the 8-position. We synthesized per - O -methylquercetin from quercetin, then using selective demethylation prepared various O -methylquercetins. Using the Suzuki-Miyaura coupling reaction, 8-arylation of per - O -methylquercetin was achieved. Using a LiAlH₄ reduction or Clemmensen reduction, these flavonols were transformed to the corresponding cyanidin derivatives in satisfactory yields. Using these dyes, we fabricated DSSCs, and their efficiency was investigated. The efficiency of tetra - O -methylflavonol was 0.31%. However, the introduction of the 8-aryl residue increased the efficiency to 1.04%. In comparison to these flavonols, O -methylcyanidins exhibited a lower efficiency of 0.05% to 0.52%. The introduction of the 8-aryl group into the cyanidin derivatives did not result in a remarkable increase in the efficiency. These phenomena may be due to the poor fit of the HOMO-LUMO level of the dyes to the TiO₂ conduction band.

Developmental Regulation across the Life Span: Toward a New Synthesis

ERIC Educational Resources Information Center

Haase, Claudia M.; Heckhausen, Jutta; Wrosch, Carsten

2013-01-01

How can individuals regulate their own development to live happy, healthy, and productive lives? Major theories of developmental regulation across the life span have been proposed (e.g., dual-process model of assimilation and accommodation; motivational theory of life-span development; model of selection, optimization, and compensation), but they…

DNA synthesis in the pituitary gland of the rat: effect of sulpiride and clomiphene.

PubMed

Burdman, J A; Szijan, I; Jahn, G A; Machiavelli, G; Kalbermann, L E

1979-09-15

Sulpiride administration to rats releases prolactin and increases DNA replication in the anterior pituitary gland. Clomiphene prevents the stimulation of DNA synthesis produced by sulpiride, but does not affect prolactin release from the gland. These findings suggest that the intracellular prolactin content of the anterior pituitary gland plays a role in the regulation of DNA synthesis through a mechanism mediated by oestrogens.

The relationship between protein synthesis and heat shock proteins levels in rabbit reticulocyte lysates.

PubMed

Matts, R L; Hurst, R

1992-09-05

Besides heme deficiency, protein synthesis in rabbit reticulocyte lysates becomes inhibited upon exposure to a variety of agents that mimic conditions which induce the heat shock response in cells. This inhibition has been demonstrated to be due primarily to the activation of the heme-regulated eIF-2 alpha kinase (HRI) which causes an arrest in the initiation of translation. In this report, the sensitivity of protein synthesis in hemin-supplemented lysates to inhibition by Hg2+, GSSG, methylene blue, and heat shock was examined in six different reticulocyte lysate preparations. The extent to which translation was inhibited in response to Hg2+, GSSG, methylene blue, and heat shock correlated inversely with the relative levels of the 70-kDa heat shock proteins (hsp 70) and a 56-kDa protein (p56) present in the lysates determined by Western blotting. The ability of hemin to restore protein synthesis upon addition to heme-deficient lysates was also examined. While the restoration of protein synthesis correlated roughly with the levels of hsp 90 present, the results also suggest that the heme regulation of HRI probably involves the interaction of HRI with several factors present in the lysate besides hsp 90. A comparison of two lysate preparations, which had a 2-fold difference in their protein synthesis rates, indicated that the slower translational rate of the one lysate could be accounted for by its low level of constitutive eIF-2 alpha phosphorylation, with its accompanying decrease in the eIF-2B activity and lower level of polyribosome loading. The present study supports the notion that the previously demonstrated interaction of HRI with hsp 90, hsp 70, and p56 in reticulocyte lysates may play a direct role in regulating HRI activation or activity. We hypothesize that the competition of denatured protein and HRI for the binding of hsp 70 may be a molecular signal that triggers the activation of HRI in reticulocyte lysates in response to stress. Possible functions

Current Advances on the Structure, Bioactivity, Synthesis, and Metabolic Regulation of Novel Ubiquinone Derivatives in the Edible and Medicinal Mushroom Antrodia cinnamomea.

PubMed

Zhang, Bo-Bo; Hu, Peng-Fei; Huang, Jing; Hu, Yong-Dan; Chen, Lei; Xu, Gan-Rong

2017-12-06

In recent years, Antrodia cinnamomea has attracted great attention around the world as an extremely precious edible and medicinal mushroom. Ubiquinone derivatives, which are characteristic metabolites of A. cinnamomea, have shown great bioactivities. Some of them have been regarded as promising therapeutic agents and approved into clinical trial by the U.S. Food and Drug Administration. Although some excellent reviews have been published covering different aspects of A. cinnamomea, this review brings, for the first time, complete information about the structure, bioactivity, chemical synthesis, biosynthesis, and metabolic regulation of ubiquinone derivatives in A. cinnamomea. It not only advances our knowledge on the bioactive metabolites, especially the ubiquinone derivatives, in A. cinnamomea but also provides valuable information for the investigation on other edible and medicinal mushrooms.

Endogenous secreted phospholipase A2 group X regulates cysteinyl leukotrienes synthesis by human eosinophils.

PubMed

Hallstrand, Teal S; Lai, Ying; Hooper, Kathryn A; Oslund, Rob C; Altemeier, William A; Matute-Bello, Gustavo; Gelb, Michael H

2016-01-01

Phospholipase A2s mediate the rate-limiting step in the formation of eicosanoids such as cysteinyl leukotrienes (CysLTs). Group IVA cytosolic PLA2α (cPLA2α) is thought to be the dominant PLA2 in eosinophils; however, eosinophils also have secreted PLA2 (sPLA2) activity that has not been fully defined. To examine the expression of sPLA2 group X (sPLA2-X) in eosinophils, the participation of sPLA2-X in the formation of CysLTs, and the mechanism by which sPLA2-X initiates the synthesis of CysLTs in eosinophils. Peripheral blood eosinophils were obtained from volunteers with asthma and/or allergy. A rabbit polyclonal anti-sPLA2-X antibody identified sPLA2-X by Western blot. We used confocal microscopy to colocalize the sPLA2-X to intracellular structures. An inhibitor of sPLA2-X (ROC-0929) that does not inhibit other mammalian sPLA2s, as well as inhibitors of the mitogen-activated kinase cascade (MAPK) and cPLA2α, was used to examine the mechanism of N-formyl-methionyl-leucyl-phenylalanine (fMLP)-mediated formation of CysLT. Eosinophils express the mammalian sPLA2-X gene (PLA2G10). The sPLA2-X protein is located in the endoplasmic reticulum, golgi, and granules of eosinophils and moves to the granules and lipid bodies during fMLP-mediated activation. Selective sPLA2-X inhibition attenuated the fMLP-mediated release of arachidonic acid and CysLT formation by eosinophils. Inhibitors of p38, extracellular-signal-regulated kinases 1/2 (p44/42 MAPK), c-Jun N-terminal kinase, and cPLA2α also attenuated the fMLP-mediated formation of CysLT. The sPLA2-X inhibitor reduced the phosphorylation of p38 and extracellular-signal-regulated kinases 1/2 (p44/42 MAPK) as well as cPLA2α during cellular activation, indicating that sPLA2-X is involved in activating the MAPK cascade leading to the formation of CysLT via cPLA2α. We further demonstrate that sPLA2-X is activated before secretion from the cell during activation. Short-term priming with IL-13 and TNF/IL-1β increased the

Optimization of the Technological Synthesis of Refractory Compounds

NASA Astrophysics Data System (ADS)

Gaidar, S. M.; Karelina, M. Yu.; Prikhod'ko, V. M.; Volkov, A. A.

2017-12-01

The results of experimental studies, which are related to the regulation of the fractional composition of refractory compounds by roll milling in using controlled roll opening and unbalanced peripheral speeds of rollers, are reported. The content of prepared fine, middle, and coarse fractions is within 50-80%; in this case, the milling time of synthesis products is less than the time of ball milling by an order of magnitude. The application of roll milling for refining the products of self-propagating high-temperature synthesis can be most efficient in using together with heat-generating reactor to solve the main problem of self-propagating synthesis (SHS), which is a problem for recent several decades (the problem is the creation of intense automated production of refractory compounds in using continuous manufacturing cycle within a energotechnological system with the recovery of a great quantity of heat released during SHS).

Poly(A) tail length regulates PABPC1 expression to tune translation in the heart.

PubMed

Chorghade, Sandip; Seimetz, Joseph; Emmons, Russell; Yang, Jing; Bresson, Stefan M; Lisio, Michael De; Parise, Gianni; Conrad, Nicholas K; Kalsotra, Auinash

2017-06-27

The rate of protein synthesis in the adult heart is one of the lowest in mammalian tissues, but it increases substantially in response to stress and hypertrophic stimuli through largely obscure mechanisms. Here, we demonstrate that regulated expression of cytosolic poly(A)-binding protein 1 (PABPC1) modulates protein synthetic capacity of the mammalian heart. We uncover a poly(A) tail-based regulatory mechanism that dynamically controls PABPC1 protein synthesis in cardiomyocytes and thereby titrates cellular translation in response to developmental and hypertrophic cues. Our findings identify PABPC1 as a direct regulator of cardiac hypertrophy and define a new paradigm of gene regulation in the heart, where controlled changes in poly(A) tail length influence mRNA translation.

Flavonoid biosynthesis-related genes in grape skin are differentially regulated by temperature and light conditions.

PubMed

Azuma, Akifumi; Yakushiji, Hiroshi; Koshita, Yoshiko; Kobayashi, Shozo

2012-10-01

Temperature and light are important environmental factors that affect flavonoid biosynthesis in grape berry skin. However, the interrelationships between temperature and light effects on flavonoid biosynthesis have not been fully elucidated at the molecular level. Here, we investigated the effects of temperature and light conditions on the biosynthesis of flavonoids (anthocyanins and flavonols) and the expression levels of related genes in an in vitro environmental experiment using detached grape berries. Sufficient anthocyanin accumulation in the grape skin was observed under a low temperature (15 °C) plus light treatment, whereas high temperature (35 °C) or dark treatment severely suppressed anthocyanin accumulation. This indicates that the accumulation of anthocyanins is dependent on both low temperature and light. qRT-PCR analysis showed that the responses of three MYB-related genes (VlMYBA1-3, VlMYBA1-2, and VlMYBA2) to temperature and light differed greatly even though the products of all three genes had the ability to regulate anthocyanin biosynthesis pathway genes. Furthermore, the expression levels of other MYB-related genes and many flavonoid biosynthesis pathway genes were regulated independently by temperature and light. We also found that temperature and light conditions affected the anthocyanin composition in the skin through the regulation of flavonoid biosynthesis pathway genes. Our results suggest that low temperature and light have a synergistic effect on the expression of genes in the flavonoid biosynthesis pathway. These findings provide new information about the relationships between environmental factors and flavonoid accumulation in grape berry skin.

Selective memory generalization by spatial patterning of protein synthesis

PubMed Central

O’Donnell, Cian; Sejnowski, Terrence J.

2014-01-01

Summary Protein synthesis is crucial for both persistent synaptic plasticity and long-term memory. De novo protein expression can be restricted to specific neurons within a population, and to specific dendrites within a single neuron. Despite its ubiquity, the functional benefits of spatial protein regulation for learning are unknown. We used computational modeling to study this problem. We found that spatially patterned protein synthesis can enable selective consolidation of some memories but forgetting of others, even for simultaneous events that are represented by the same neural population. Key factors regulating selectivity include the functional clustering of synapses on dendrites, and the sparsity and overlap of neural activity patterns at the circuit level. Based on these findings we proposed a novel two-step model for selective memory generalization during REM and slow-wave sleep. The pattern-matching framework we propose may be broadly applicable to spatial protein signaling throughout cortex and hippocampus. PMID:24742462

Selective memory generalization by spatial patterning of protein synthesis.

PubMed

O'Donnell, Cian; Sejnowski, Terrence J

2014-04-16

Protein synthesis is crucial for both persistent synaptic plasticity and long-term memory. De novo protein expression can be restricted to specific neurons within a population, and to specific dendrites within a single neuron. Despite its ubiquity, the functional benefits of spatial protein regulation for learning are unknown. We used computational modeling to study this problem. We found that spatially patterned protein synthesis can enable selective consolidation of some memories but forgetting of others, even for simultaneous events that are represented by the same neural population. Key factors regulating selectivity include the functional clustering of synapses on dendrites, and the sparsity and overlap of neural activity patterns at the circuit level. Based on these findings, we proposed a two-step model for selective memory generalization during REM and slow-wave sleep. The pattern-matching framework we propose may be broadly applicable to spatial protein signaling throughout cortex and hippocampus. Copyright © 2014 Elsevier Inc. All rights reserved.

Antioxidant capacity changes and phenolic profile of Echinacea purpurea, nettle (Urtica dioica L.), and dandelion (Taraxacum officinale) after application of polyamine and phenolic biosynthesis regulators.

PubMed

Hudec, Jozef; Burdová, Mária; Kobida, L'ubomír; Komora, Ladislav; Macho, Vendelín; Kogan, Grigorij; Turianica, Ivan; Kochanová, Radka; Lozek, Otto; Habán, Miroslav; Chlebo, Peter

2007-07-11

The changes of the antioxidant (AOA) and antiradical activities (ARA) and the total contents of phenolics, anthocyanins, flavonols, and hydroxybenzoic acid in roots and different aerial sections of Echinacea purpurea, nettle, and dandelion, after treatment with ornithine decarboxylase inhibitor, a polyamine inhibitor (O-phosphoethanolamine, KF), and a phenol biosynthesis stimulator (carboxymethyl chitin glucan, CCHG) were analyzed spectrophotometrically; hydroxycinnamic acids content was analyzed by RP-HPLC with UV detection. Both regulators increased the AOA measured as inhibition of peroxidation (IP) in all herb sections, with the exception of Echinacea stems after treatment with KF. In root tissues IP was dramatically elevated mainly after CCHG application: 8.5-fold in Echinacea, 4.14-fold in nettle, and 2.08-fold in dandelion. ARA decrease of Echinacea leaves treated with regulators was in direct relation only with cichoric acid and caftaric acid contents. Both regulators uphold the formation of cinnamic acid conjugates, the most expressive being that of cichoric acid after treatment with CCHG in Echinacea roots from 2.71 to 20.92 mg g(-1). There was a strong relationship between increase of the total phenolics in all sections of Echinacea, as well as in the studied sections of dandelion, and the anthocyanin content.

Evidence that intracellular magnesium is present in cells at a regulatory concentration for protein synthesis.

PubMed Central

Terasaki, M; Rubin, H

1985-01-01

When extracellular magnesium is reduced by a factor of 50 (from 1.0 to 0.02 mM), the total intracellular magnesium of a spontaneously transformed clone of 3T3 cells decreases by 30-50%. Protein synthesis rates in these cells were measured as the intracellular magnesium decreased. Protein synthesis rates and magnesium content were found to decrease in parallel with each other. At 3 hr, a decrease to 84% of control values of magnesium content was accompanied by a decrease to 85% of control values of leucine incorporation rates. A larger inhibition had occurred by 12 hr, when the magnesium had decreased to 67% and leucine incorporation rates had decreased to 57%. When magnesium was restored to magnesium-deprived cells, both magnesium content and leucine incorporation increased about 2-fold by 1 hr. In the experiments reported here, initial small changes in magnesium content are associated with changes in protein synthesis rates. This strongly suggests that magnesium is present at a regulatory rather than excess concentration for protein synthesis. The results are consistent with a role for intracellular magnesium in the regulation of protein synthesis and support the hypothesis that magnesium has a central role in the regulation of metabolism and growth. PMID:2997785

Variable effects of dexamethasone on protein synthesis in clonal rat osteosarcoma cells

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

Hodge, B.O.; Kream, B.E.

1988-05-01

We examined the effects of dexamethasone on protein synthesis in clonal rat osteoblastic osteosarcoma (ROS) cell lines by measuring the incorporation of (/sup 3/H)proline into collagenase-digestible and noncollagen protein in the cell layer and medium of the cultures. In ROS 17/2 and subclone C12 of ROS 17/2.8, dexamethasone decreased collagen synthesis with no change in DNA content of the cultures. In ROS 17/2.8 and its subclone G2, dexamethasone stimulated collagen and noncollagen protein synthesis, with a concomitant decrease in the DNA content of the cells. These data indicate that ROS cell lines are phenotypically heterogeneous and suggest that in normalmore » bone there may be distinct subpopulations of osteoblasts with varying phenotypic traits with respect to the regulation of protein synthesis.« less

Loss of Cystic Fibrosis Transmembrane Conductance Regulator Function Enhances Activation of p38 and ERK MAPKs, Increasing Interleukin-6 Synthesis in Airway Epithelial Cells Exposed to Pseudomonas aeruginosa*