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Sample records for del inositol fosfoglicano

  1. Phosphate, inositol and polyphosphates.

    PubMed

    Livermore, Thomas M; Azevedo, Cristina; Kolozsvari, Bernadett; Wilson, Miranda S C; Saiardi, Adolfo

    2016-02-15

    Eukaryotic cells have ubiquitously utilized the myo-inositol backbone to generate a diverse array of signalling molecules. This is achieved by arranging phosphate groups around the six-carbon inositol ring. There is virtually no biological process that does not take advantage of the uniquely variable architecture of phosphorylated inositol. In inositol biology, phosphates are able to form three distinct covalent bonds: phosphoester, phosphodiester and phosphoanhydride bonds, with each providing different properties. The phosphoester bond links phosphate groups to the inositol ring, the variable arrangement of which forms the basis of the signalling capacity of the inositol phosphates. Phosphate groups can also form the structural bridge between myo-inositol and diacylglycerol through the phosphodiester bond. The resulting lipid-bound inositol phosphates, or phosphoinositides, further expand the signalling potential of this family of molecules. Finally, inositol is also notable for its ability to host more phosphates than it has carbons. These unusual organic molecules are commonly referred to as the inositol pyrophosphates (PP-IPs), due to the presence of high-energy phosphoanhydride bonds (pyro- or diphospho-). PP-IPs themselves constitute a varied family of molecules with one or more pyrophosphate moiety/ies located around the inositol. Considering the relationship between phosphate and inositol, it is no surprise that members of the inositol phosphate family also regulate cellular phosphate homoeostasis. Notably, the PP-IPs play a fundamental role in controlling the metabolism of the ancient polymeric form of phosphate, inorganic polyphosphate (polyP). Here we explore the intimate links between phosphate, inositol phosphates and polyP, speculating on the evolution of these relationships. PMID:26862212

  2. Inositol pyrophosphate pyrotechnics.

    PubMed

    Bhandari, Rashna; Chakraborty, Anutosh; Snyder, Solomon H

    2007-05-01

    Physiologic roles of highly phosphorylated inositol phosphates, including those containing pyrophosphate groups, have been the focus of much recent interest. In the April 6, 2007 issue of Science, two papers (Lee et al., 2007; Mulugu et al., 2007) demonstrate the occurrence of a novel inositol pyrophosphate molecule in yeast and elucidate its role in phosphate homeostasis. PMID:17488633

  3. Two inositol hexakisphosphate kinases drive inositol pyrophosphate synthesis in plants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Inositol pyrophosphates are novel cellular signaling molecules with newly discovered roles in energy sensing and metabolic control. Studies in eukaryotes have revealed that these compounds turn over rapidly, and thus only small amounts accumulate. Inositol pyrophosphates have not been the subject of...

  4. Preparation of quality inositol pyrophosphates.

    PubMed

    Loss, Omar; Azevedo, Cristina; Szijgyarto, Zsolt; Bosch, Daniel; Saiardi, Adolfo

    2011-01-01

    Myo-inositol is present in nature either unmodified or in more complex phosphorylated derivates. Of the latest, the two most abundant in eukaryotic cells are inositol pentakisphosphate (IP(5;)) and inositol hexakisphosphate (phytic acid or IP(6;)). IP(5;) and IP(6;) are the precursors of inositol pyrophosphate molecules that contain one or more pyrophosphate bonds(1). Phosphorylation of IP(6;) generates diphoshoinositolpentakisphosphate (IP(7;) or PP-IP(5;)) and bisdiphoshoinositoltetrakisphosphate (IP(8;) or (PP)(2;)-IP(4;)). Inositol pyrophosphates have been isolated from all eukaryotic organisms so far studied. In addition, the two distinct classes of enzymes responsible for inositol pyrophosphate synthesis are highly conserved throughout evolution(2-4). The IP(6;) kinases (IP(6;)Ks) posses an enormous catalytic flexibility, converting IP(5;) and IP(6;) to PP-IP(4;) and IP(7;) respectively and subsequently, by using these products as substrates, promote the generation of more complex molecules(5,6). Recently, a second class of pyrophosphate generating enzymes was identified in the form of the yeast protein VIP(1;) (also referred as PP-IP(5;)K), which is able to convert IP(6;) to IP(7;) and IP(8;)(7,8). Inositol pyrophosphates regulate many disparate cellular processes such as insulin secretion(9), telomere length(10,11), chemotaxis(12), vesicular trafficking(13), phosphate homeostasis(14) and HIV-1 gag release(15). Two mechanisms of actions have been proposed for this class of molecules. They can affect cellular function by allosterically interacting with specific proteins like AKT(16). Alternatively, the pyrophosphate group can donate a phosphate to pre-phosphorylated proteins(17). The enormous potential of this research field is hampered by the absence of a commercial source of inositol pyrophosphates, which is preventing many scientists from studying these molecules and this new post-translational modification. The methods currently available to isolate

  5. Myo-inositol transport in Saccharomyces cerevisiae.

    PubMed

    Nikawa, J; Nagumo, T; Yamashita, S

    1982-05-01

    myo-Inositol uptake in Saccharomyces cerevisiae was dependent on temperature, time, and substrate concentration. The transport obeyed saturation kinetics with an apparent Km for myo-inositol of 0.1 mM, myo-Inositol analogs, such as scyllo-inositol, 2-inosose, mannitol, and 1,2-cyclohexanediol, had no effect on myo-inositol uptake, myo-Inositol uptake required metabolic energy. Removal of D-glucose resulted in a loss of activity, and azide and cyanide ions were inhibitory. In the presence of D-glucose, myo-inositol was accumulated in the cells against a concentration gradient. A myo-inositol transport mutant was isolated from UV-mutagenized S. cerevisiae cells using the replica-printing technique. The defect in myo-inositol uptake was due to a single nuclear gene mutation. The activities of L-serine and D-glucose transport were not affected by the mutation. Thus it was shown that S. cerevisiae grown under the present culture conditions possessed a single and specific myo-inositol transport system. myo-Inositol transport activity was reduced by the addition of myo-inositol to the culture medium. The activity was reversibly restored by the removal of myo-inositol from the medium. This restoration of activity was completely abolished by cycloheximide. PMID:7040334

  6. 21 CFR 582.5370 - Inositol.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Inositol. 582.5370 Section 582.5370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements 1 § 582.5370 Inositol. (a) Product. Inositol....

  7. Measurement of phospholipase C by monitoring inositol phosphates using [³H]inositol labeling protocols in permeabilized cells.

    PubMed

    Skippen, Alison; Swigart, Philip; Cockcroft, Shamshad

    2013-01-01

    Data on the production of inositol phosphates is a useful complement to measurements of intracellular Ca(2+). The basic principle is labeling of the inositol lipids by growing the appropriate cell line in culture in the presence of [3H]inositol for 2-3 days to reach labeling equilibrium. Lithium ions at 10 mM inhibits the degradation of inositol phosphates to free inositol and is used to trap the inositol in the inositol polyphosphate forms. Inositol phosphates can be separated with ease from free inositol by using anion exchange chromatography. A method capable of easily processing approximately 40-60 samples in a single day is presented. PMID:23007585

  8. How versatile are inositol phosphate kinases?

    PubMed Central

    Shears, Stephen B

    2004-01-01

    This review assesses the extent and the significance of catalytic versatility shown by several inositol phosphate kinases: the inositol phosphate multikinase, the reversible Ins(1,3,4) P (3)/Ins(3,4,5,6) P (4) kinase, and the kinases that synthesize diphosphoinositol polyphosphates. Particular emphasis is placed upon data that are relevant to the situation in vivo. It will be shown that catalytic promiscuity towards different inositol phosphates is not typically an evolutionary compromise, but instead is sometimes exploited to facilitate tight regulation of physiological processes. This multifunctionality can add to the complexity with which inositol signalling pathways interact. This review also assesses some proposed additional functions for the catalytic domains, including transcriptional regulation, protein kinase activity and control by molecular 'switching', all in the context of growing interest in 'moonlighting' (gene-sharing) proteins. PMID:14567754

  9. Does ovary need D-chiro-inositol?

    PubMed Central

    2012-01-01

    Backgroud Polycystic Ovary Syndrome (PCOS) is a multifactorial pathology that affects 10% of the women in reproductive age being the main cause of infertility due to menstrual dysfunction. Since 1980, it is known that PCOS is associated with insulin resistance (IR). The recognition of this association has prompted extensive investigation on the relationship between insulin and gonadal function, and has turned insulin sensitizer agent as the main therapeutic choice. In particular two different polyalcohol myo-inositol and D-chiro-inositol have been shown to improve insulin resistance, hyperandrogenism and to induce ovulation in PCOS women. In particular, while data on myo-inositol and restored ovulation were consistent, data on D-chiro-inositol were not . Recently, a comparative study, proposed a D-chiro-inositol paradox in the ovary of PCOS patients hypothesizing that only myo-inositol has a specific ovarian action. In the present study we aim to further study the role played by D-chiro-inositol at ovarian level. Methods A total of 54 women, aged <40 years and diagnosed with PCOS were enrolled in this study. Patients with insulin resistance and/or hyperglycaemia were excluded from the study. Patients were randomly divided into 5 groups (n=10-12): a placebo group, and 4 groups (A-D) that received 300-600-1200-2400 mg of DCI daily respectively. All treatments were carried out for 8 weeks before follicle stimulating hormone (rFSH) administration. Results Total r-FSH units increased significantly in the two groups that received the higher doses of DCI. The number of immature oocytes was significantly increased in the three groups that received the higher doses of DCI. Concurrently, the number of MII oocytes was significantly lower in the D group compared to placebo group. Noteworthy, the number of grade I embryos was significantly reduced by DCI supplementation. Conclusions Indeed, increasing DCI dosage progressively worsens oocyte quality and ovarian response. PMID

  10. Inositol-Requiring Mutants of SACCHAROMYCES CEREVISIAE

    PubMed Central

    Culbertson, Michael R.; Henry, Susan A.

    1975-01-01

    Fifty-two inositol-requiring mutants of Saccharomyces cerevisiae were isolated following mutagenesis with ethyl methanesulfonate. Complementation and tetrad analysis revealed ten major complementation classes, representing ten independently segregating loci (designated ino1 through ino10) which recombined freely with their respective centromeres. Members of any given complementation class segregated as alleles of a single locus. Thirteen complementation subclasses were identified among thirty-six mutants which behaved as alleles of the ino1 locus. The complementation map for these mutants was circular.—Dramatic cell viability losses indicative of unbalanced growth were observed in liquid cultures of representative mutants under conditions of inositol starvation. Investigation of the timing, kinetics, and extent of cell death revealed that losses in cell viability in the range of 2-4 log orders could be prevented by the addition of inositol to the medium or by disruption of protein synthesis with cycloheximide. Mutants defective in nine of the ten loci identified in this study displayed these unusual characteristics. The results suggest an important physiological role for inositol that may be related to its cellular localization and function in membrane phospholipids. The possibility is discussed that inositol deficiency initiates the process of unbalanced growth leading to cell death through the loss of normal assembly, function, or integrity of biomembranes.—Part of this work has been reported in preliminary form (Culbertson and Henry 1974). PMID:1093935

  11. Myo-inositol content of common foods: development of a high-myo-inositol diet.

    PubMed

    Clements, R S; Darnell, B

    1980-09-01

    Since virtually no information is available concerning the myo-inositol content of dietary constituents, we have measured the amount of this material present in 487 foods by gas-liquid chromatography. We observed that the greatest amounts of myo-inositol were present in fruits, beans, grains, and nuts. Fresh vegetables and fruits were found to contain more myo-inositol than did frozen, canned, or salt-free products. The data provided in this report were used to develop diets that contained varying, but known amounts of myo-inositol. The myo-inositol intake that could be provided by such diets ranged from 225 to 1500 mg/day per 1800 kcal and within this range the agreement between the calculated and measured amounts of this material was excellent (r = 0.98). Since abnormalities in the metabolism of myo-inositol have been speculated to play a role in the pathogenesis of the polyneuropathies associated with diabetes mellitus and chronic renal failure, it is possible that the natural history of these neuropahties can be altered by modifying the amount of dietary myo-inositol that is ingested by patients with these diseases. PMID:7416064

  12. Both myo-inositol to chiro-inositol epimerase activities and chiro-inositol to myo-inositol ratios are decreased in tissues of GK type 2 diabetic rats compared to Wistar controls.

    PubMed

    Sun, Tie-hua; Heimark, Douglas B; Nguygen, Thang; Nadler, Jerry L; Larner, Joseph

    2002-05-10

    Previous data from our and other labs demonstrated a decreased chiro-inositol content in urine and tissues of human subjects and animals with type 2 diabetes. In urine this decrease in chiro-inositol was accompanied by an increase in myo-inositol content. Decreased urine levels of chiro-inositol in monkeys were next correlated with the severity of underlying insulin resistance determined by five separate assays. To investigate the decreased chiro-inositol and the accompanying increased myo-inositol excretions in urine in humans and monkeys, we postulated a defect in the epimerization of myo-inositol to chiro-inositol. [(3)H]Myo-inositol was then shown to be converted to [(3)H]chiro-inositol in rats in vivo and in fibroblasts in vitro in a process stimulated by insulin. We next demonstrated that the conversion of [(3)H]myo-inositol to [(3)H]chiro-inositol in vivo was markedly decreased in GK type 2 diabetic rats compared to Wistar controls in liver, muscle, and fat, insulin sensitive tissues. Decreases of 20-25% conversion to baseline levels of under 5% conversion were observed. In the present work, we initially compared the total contents of myo-inositol and chiro-inositol in GK type 2 diabetic rat kidney, liver, and muscle compared to Wistar controls. We demonstrated a consistent decreased total chiro-inositol to myo-inositol ratio in kidney, liver, and muscle compared to controls. We next established the presence of a myo-inositol to chiro-inositol epimerase activity in rat liver cytosol. Enzyme activity was shown to be time and enzyme concentration dependent with a broad pH optimum. It required NADH and NADPH for full activity, which is compatible with its action via an oxido-reductive mechanism. Lastly, we demonstrated that the epimerase enzyme bioactivity was significantly decreased in muscle, liver, and fat cytosolic extracts of GK type 2 diabetic rats versus Wistar controls. Decreased myo-inositol to chiro-inositol epimerase activity may therefore play a

  13. myo-Inositol uptake is essential for bulk inositol phospholipid but not glycosylphosphatidylinositol synthesis in Trypanosoma brucei.

    PubMed

    Gonzalez-Salgado, Amaia; Steinmann, Michael E; Greganova, Eva; Rauch, Monika; Mäser, Pascal; Sigel, Erwin; Bütikofer, Peter

    2012-04-13

    myo-Inositol is an essential precursor for the production of inositol phosphates and inositol phospholipids in all eukaryotes. Intracellular myo-inositol is generated by de novo synthesis from glucose 6-phosphate or is provided from the environment via myo-inositol symporters. We show that in Trypanosoma brucei, the causative pathogen of human African sleeping sickness and nagana in domestic animals, myo-inositol is taken up via a specific proton-coupled electrogenic symport and that this transport is essential for parasite survival in culture. Down-regulation of the myo-inositol transporter using RNA interference inhibited uptake of myo-inositol and blocked the synthesis of the myo-inositol-containing phospholipids, phosphatidylinositol and inositol phosphorylceramide; in contrast, it had no effect on glycosylphosphatidylinositol production. This together with the unexpected localization of the myo-inositol transporter in both the plasma membrane and the Golgi demonstrate that metabolism of endogenous and exogenous myo-inositol in T. brucei is strictly segregated. PMID:22351763

  14. 21 CFR 582.5370 - Inositol.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Inositol. 582.5370 Section 582.5370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  15. 21 CFR 582.5370 - Inositol.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Inositol. 582.5370 Section 582.5370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  16. 21 CFR 184.1370 - Inositol.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... ingredient meets the specifications of the Food Chemicals Codex, 3d Ed. (1981), p. 150, which is incorporated... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Inositol. 184.1370 Section 184.1370 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR...

  17. Myo-Inositol content determined by myo-inositol biosynthesis and oxidation in blueberry fruit.

    PubMed

    Song, Fangyuan; Su, Hongyan; Yang, Nan; Zhu, Luying; Cheng, Jieshan; Wang, Lei; Cheng, Xianhao

    2016-11-01

    Myo-inositol metabolism in plant edible organs has become the focus of many recent studies because of its benefits to human health and unique functions in plant development. In this study, myo-inositol contents were analyzed during the development of two blueberry cultivars, cv 'Berkeley' and cv 'Bluecrop'. Furthermore, two VcMIPS 1/2 (Vaccinium corymbosum MIPS) genes, one VcIMP (Vaccinium corymbosum IMP) gene and one VcMIOX (Vaccinium corymbosum MIOX) gene were isolated for the first time from blueberry. The expression patterns of VcMIPS2, VcIMP and VcMIOX genes showed a relationship with the change profiles of myo-inositol content during fruit ripening. The results were further confirmed by the analyses of the enzyme activity. Results indicated that both myo-inositol biosynthesis and oxidation played important roles in determining of myo-inositol levels during the development of blueberry. To our knowledge, this report is the first to discuss myo-inositol levels in fruits in terms of biosynthesis and catabolism. PMID:27211661

  18. Turnover of inositol pentakisphosphates, inositol hexakisphosphate and diphosphoinositol polyphosphates in primary cultured hepatocytes.

    PubMed Central

    Glennon, M C; Shears, S B

    1993-01-01

    We have used a non-transformed cell model, the primary cultured hepatocyte, to explore the turnover of inositol hexakisphosphate, multiple isomers of inositol pentakisphosphate and two novel diphosphoinositol polyphosphates. All of these compounds gradually accumulated radioactivity throughout a 70 h period of labelling with [3H]inositol. However, a rapid metabolic rate was revealed upon inhibition of diphosphoinositol polyphosphate biphosphatase(s) with 1 mM fluoride for 40 min: this treatment elevated levels of [3H]diphosphoinositol polyphosphates up to 10-fold, indicating that their cellular pools were normally turning over at least 10 times every 40 min. This was accompanied by a turnover of about 10% of the pool of inositol hexakisphosphate. Control experiments established that 200 nM vasopressin brought about a typical activation of phospholipase C in hepatocytes after 62 h of primary culture. This agonist treatment did not affect steady-state levels of [3H]inositol pentakisphosphates, [3H]inositol hexakisphosphate or [3H]diphosphoinositol polyphosphates. However, prolonged treatment of hepatocytes with 2 microM thapsigargin reduced steady-state levels of [3H]diphosphoinositol polyphosphates by 50-70%. This effect of thapsigargin was also observed in the presence of fluoride, indicating that thapsigargin inhibited the rate of synthesis of diphosphoinositol polyphosphates. PMID:8343137

  19. Inositol pyrophosphates inhibit synaptotagmin-dependent exocytosis.

    PubMed

    Lee, Tae-Sun; Lee, Joo-Young; Kyung, Jae Won; Yang, Yoosoo; Park, Seung Ju; Lee, Seulgi; Pavlovic, Igor; Kong, Byoungjae; Jho, Yong Seok; Jessen, Henning J; Kweon, Dae-Hyuk; Shin, Yeon-Kyun; Kim, Sung Hyun; Yoon, Tae-Young; Kim, Seyun

    2016-07-19

    Inositol pyrophosphates such as 5-diphosphoinositol pentakisphosphate (5-IP7) are highly energetic inositol metabolites containing phosphoanhydride bonds. Although inositol pyrophosphates are known to regulate various biological events, including growth, survival, and metabolism, the molecular sites of 5-IP7 action in vesicle trafficking have remained largely elusive. We report here that elevated 5-IP7 levels, caused by overexpression of inositol hexakisphosphate (IP6) kinase 1 (IP6K1), suppressed depolarization-induced neurotransmitter release from PC12 cells. Conversely, IP6K1 depletion decreased intracellular 5-IP7 concentrations, leading to increased neurotransmitter release. Consistently, knockdown of IP6K1 in cultured hippocampal neurons augmented action potential-driven synaptic vesicle exocytosis at synapses. Using a FRET-based in vitro vesicle fusion assay, we found that 5-IP7, but not 1-IP7, exhibited significantly higher inhibitory activity toward synaptic vesicle exocytosis than IP6 Synaptotagmin 1 (Syt1), a Ca(2+) sensor essential for synaptic membrane fusion, was identified as a molecular target of 5-IP7 Notably, 5-IP7 showed a 45-fold higher binding affinity for Syt1 compared with IP6 In addition, 5-IP7-dependent inhibition of synaptic vesicle fusion was abolished by increasing Ca(2+) levels. Thus, 5-IP7 appears to act through Syt1 binding to interfere with the fusogenic activity of Ca(2+) These findings reveal a role of 5-IP7 as a potent inhibitor of Syt1 in controlling the synaptic exocytotic pathway and expand our understanding of the signaling mechanisms of inositol pyrophosphates. PMID:27364007

  20. Disruption of inositol biosynthesis through targeted mutagenesis in Dictyostelium discoideum: generation and characterization of inositol-auxotrophic mutants.

    PubMed

    Fischbach, Andreas; Adelt, Stephan; Müller, Alexander; Vogel, Günter

    2006-08-01

    myo-Inositol and its downstream metabolites participate in diverse physiological processes. Nevertheless, considering their variety, it is likely that additional roles are yet to be uncovered. Biosynthesis of myo-inositol takes place via an evolutionarily conserved metabolic pathway and is strictly dependent on inositol-3-phosphate synthase (EC 5.5.1.4). Genetic manipulation of this enzyme will disrupt the cellular inositol supply. Two methods, based on gene deletion and antisense strategy, were used to generate mutants of the cellular slime mould Dictyostelium discoideum. These mutants are inositol-auxotrophic and show phenotypic changes under inositol starvation. One remarkable attribute is their inability to live by phagocytosis of bacteria, which is the exclusive nutrient source in their natural environment. Cultivated on fluid medium, the mutants lose their viability when deprived of inositol for longer than 24 h. Here, we report a study of the alterations in the first 24 h in cellular inositol, inositol phosphate and phosphoinositide concentrations, whereby a rapidly accumulating phosphorylated compound was detected. After its identification as 2,3-BPG (2,3-bisphosphoglycerate), evidence could be found that the internal disturbances of inositol homoeostasis trigger the accumulation. In a first attempt to characterize this as a physiologically relevant response, the efficient in vitro inhibition of a D. discoideum inositol-polyphosphate 5-phosphatase (EC 3.1.3.56) by 2,3-BPG is presented. PMID:16599905

  1. Disruption of inositol biosynthesis through targeted mutagenesis in Dictyostelium discoideum: generation and characterization of inositol-auxotrophic mutants

    PubMed Central

    Fischbach, Andreas; Adelt, Stephan; Müller, Alexander; Vogel, Günter

    2006-01-01

    myo-Inositol and its downstream metabolites participate in diverse physiological processes. Nevertheless, considering their variety, it is likely that additional roles are yet to be uncovered. Biosynthesis of myo-inositol takes place via an evolutionarily conserved metabolic pathway and is strictly dependent on inositol-3-phosphate synthase (EC 5.5.1.4). Genetic manipulation of this enzyme will disrupt the cellular inositol supply. Two methods, based on gene deletion and antisense strategy, were used to generate mutants of the cellular slime mould Dictyostelium discoideum. These mutants are inositol-auxotrophic and show phenotypic changes under inositol starvation. One remarkable attribute is their inability to live by phagocytosis of bacteria, which is the exclusive nutrient source in their natural environment. Cultivated on fluid medium, the mutants lose their viability when deprived of inositol for longer than 24 h. Here, we report a study of the alterations in the first 24 h in cellular inositol, inositol phosphate and phosphoinositide concentrations, whereby a rapidly accumulating phosphorylated compound was detected. After its identification as 2,3-BPG (2,3-bisphosphoglycerate), evidence could be found that the internal disturbances of inositol homoeostasis trigger the accumulation. In a first attempt to characterize this as a physiologically relevant response, the efficient in vitro inhibition of a D. discoideum inositol-polyphosphate 5-phosphatase (EC 3.1.3.56) by 2,3-BPG is presented. PMID:16599905

  2. Partial purification and characterization of indol-3-ylacetylglucose:myo-inositol indol-3-ylacetyltransferase (indoleacetic acid-inositol synthase)

    NASA Technical Reports Server (NTRS)

    Kesy, J. M.; Bandurski, R. S.

    1990-01-01

    A procedure is described for the purification of the enzyme indol-3-ylacetylglucose:myo-inositol indol-3-ylacetyltransferase (IAA-myo-inositol synthase). This enzyme catalyzes the transfer of indol-3-ylacetate from 1-0-indol-3-ylacetyl-beta-d-glucose to myo-inositol to form indol-3-ylacetyl-myo-inositol and glucose. A hexokinase or glucose oxidase based assay system is described. The enzyme has been purified approximately 16,000-fold, has an isoelectric point of pH 6.1 and yields three catalytically inactive bands upon acrylamide gel electrophoresis of the native protein. The enzyme shows maximum transferase activity with myo-inositol but shows some transferase activity with scyllo-inositol and myo-inosose-2. No transfer of IAA occurs with myo-inositol-d-galactopyranose, cyclohexanol, mannitol, or glycerol as acyl acceptor. The affinity of the enzyme for 1-0-indol-3-ylacetyl-beta-d-glucose is, Km = 30 micromolar, and for myo-inositol is, Km = 4 millimolar. The enzyme does not catalyze the exchange incorporation of glucose into IAA-glucose indicating the reaction mechanism involves binding of IAA glucose to the enzyme with subsequent hydrolytic cleavage of the acyl moiety by the hydroxyl of myo-inositol to form IAA myo-inositol ester.

  3. Extraction and analysis of soluble inositol polyphosphates from yeast.

    PubMed

    Azevedo, Cristina; Saiardi, Adolfo

    2006-01-01

    Soluble inositol polyphosphates are implicated in the regulation of many important cellular functions. This protocol to extract and separate inositol polyphosphates from Saccharomyces cerevisiae is divided into three steps: labeling of yeast, extraction of soluble inositol polyphosphates and chromatographic separation. Yeast cells are incubated with tritiated inositol, which is taken up and metabolized into different phosphorylated forms. Soluble inositol polyphosphates are then acid-extracted and fractionated by high-performance liquid chromatography. The radioactivity of each fraction is determined by scintillation counting. This highly sensitive and reproducible method allows the accurate detection of subtle changes in the inositol polyphosphate profile and takes less than 48 h. It can easily be applied to other systems and we have included two adaptations of the protocol, one optimized for mammalian cells and the other for Arabidopsis thaliana. PMID:17406485

  4. Characterization of the inositol monophosphatase gene family in Arabidopsis

    PubMed Central

    Nourbakhsh, Aida; Collakova, Eva; Gillaspy, Glenda E.

    2015-01-01

    Synthesis of myo-inositol is crucial in multicellular eukaryotes for production of phosphatidylinositol and inositol phosphate signaling molecules. The myo-inositol monophosphatase (IMP) enzyme is required for the synthesis of myo-inositol, breakdown of inositol (1,4,5)-trisphosphate, a second messenger involved in Ca2+ signaling, and synthesis of L-galactose, a precursor of ascorbic acid. Two myo-inositol monophosphatase -like (IMPL) genes in Arabidopsis encode chloroplast proteins with homology to the prokaryotic IMPs and one of these, IMPL2, can complement a bacterial histidinol 1-phosphate phosphatase mutant defective in histidine synthesis, indicating an important role for IMPL2 in amino acid synthesis. To delineate how this small gene family functions in inositol synthesis and metabolism, we sought to compare recombinant enzyme activities, expression patterns, and impact of genetic loss-of-function mutations for each. Our data show that purified IMPL2 protein is an active histidinol-phosphate phosphatase enzyme in contrast to the IMPL1 enzyme, which has the ability to hydrolyze D-galactose 1-phosphate, and D-myo-inositol 1-phosphate, a breakdown product of D-inositol (1,4,5) trisphosphate. Expression studies indicated that all three genes are expressed in multiple tissues, however, IMPL1 expression is restricted to above-ground tissues only. Identification and characterization of impl1 and impl2 mutants revealed no viable mutants for IMPL1, while two different impl2 mutants were identified and shown to be severely compromised in growth, which can be rescued by histidine. Analyses of metabolite levels in impl2 and complemented mutants reveals impl2 mutant growth is impacted by alterations in the histidine biosynthesis pathway, but does not impact myo-inositol synthesis. Together, these data indicate that IMPL2 functions in the histidine biosynthetic pathway, while IMP and IMPL1 catalyze the hydrolysis of inositol- and galactose-phosphates in the plant cell

  5. Characterization of inositol phosphates in carrot (Daucus carota L. ) cells

    SciTech Connect

    Rincon, M.; Chen, Q.; Boss, W.F. )

    1989-01-01

    We have shown previously that inositol-1,4,5-trisphosphate (IP{sub 3}) stimulates an efflux of {sup 45}Ca{sup 2+} from fusogenic carrot protoplasts. In light of these results, we suggested that IP{sub 3} might serve as a second messenger for the mobilization of intracellular Ca{sup 2+} in higher plant cells. To determine whether or not IP{sub 3} and other inositol phosphates were present in the carrot cells, the cells were labeled with myo-(2-{sup 3}H)inositol for 18 hours and extracted with ice-cold 10% trichloroacetic acid. The inositol metabolites were separated by anion exchange chromatography and by paper electrophoresis. We found that ({sup 3}H)inositol metabolites coeluted with inositol bisphosphate (IP{sub 2}) and IP{sub 3} when separated by anion exchange chromatography. However, we could not detect IP{sub 2} or IP{sub 3} when the inositol metabolites were analyzed by paper electrophoresis even though the polyphosphoinositides, which are the source of IP{sub 2} and IP{sub 3}, were present in these cells. Thus, ({sup 3}H)inositol metabolites other than IP{sub 2} and IP{sub 3} had coeluted on the anion exchange columns. The data indicate that either IP{sub 3} is rapidly metabolized or that it is not present at a detectable level in the carrot cells.

  6. Formation of inositol 1,4,5-trisphosphate and inositol 1,3,4-trisphosphate from inositol 1,3,4,5-tetrakisphosphate and their pathways of degradation in RBL-2H3 cells

    SciTech Connect

    Cunha-Melo, J.R.; Dean, N.M.; Ali, H.; Beaven, M.A.

    1988-10-05

    Previous studies with antigen-stimulated rat basophilic leukemia (RBL-2H3) cells indicated the formation of multiple isomers of each of the various categories of inositol phosphates. The identities of the different isomers have been elucidated by selective labeling of (3H)inositol 1,3,4,5-tetrakisphosphate with (32P)phosphate in the 3'-or 4',5'-positions and by following the metabolism of different radiolabeled inositol phosphates in extracts of RBL-2H3 cells. We report here that inositol 1,3,4,5-tetrakisphosphate, when incubated with the membrane fraction of extracts of RBL-2H3 cells, was converted to inositol 1,4,5-trisphosphate and inositol 1,3,4-trisphosphate. Further dephosphorylation of the inositol polyphosphates proceeded rapidly in whole extracts of cells, although the process was significantly retarded when ATP (2 mM) levels were maintained by an ATP-regenerating system. The degradation of inositol 1,4,5-trisphosphate proceeded with the sequential formation of inositol 1,4-bisphosphate, the inositol 4-monophosphate (with smaller amounts of the 1-monophosphate), and finally inositol. Inositol 1,3,4-trisphosphate, on the other hand, was converted to inositol 1,3-bisphosphate and inositol 3,4-bisphosphate and subsequently to inositol 4-monophosphate and inositol 1-monophosphate (stereoisomeric forms were undetermined). The possible implications of the apparent interconversion between inositol 1,4,5-trisphosphate and inositol 1,3,4,5-tetrakisphosphate in regulating histamine secretion in the RBL-2H3 cells are discussed.

  7. Behavioral evidence for the existence of two pools of cellular inositol.

    PubMed

    Bersudsky, Y; Kaplan, Z; Shapiro, Y; Agam, G; Kofman, O; Belmaker, R H

    1994-12-01

    Lithium reduces brain inositol levels by inhibiting inositol monophosphatase. In a previous study it was found that administration of pilocarpine to Li-treated rats causes limbic seizure behavior which can be reversed by i.c.v. myo-inositol but not chiro-inositol, suggesting that this behavior is related to inositol depletion in the PI cycle. Hyponatremia can lower brain inositol and hypernatremia can raise brain inositol. We now report that induction of low brain inositol by hyponatremia followed by pilocarpine did not cause limbic seizures. Induction of high brain inositol using hypernatremia followed by Li-pilocarpine administration did not reverse limbic seizures. These data support the concept that inositol available for P1 synthesis and inositol for osmotic function are sequestered in different cellular pools. PMID:7894256

  8. Myo-Inositol-Dependent Sodium Uptake in Ice Plant1

    PubMed Central

    Nelson, Donald E.; Koukoumanos, Michelle; Bohnert, Hans J.

    1999-01-01

    In salt-stressed ice plants (Mesembryanthemum crystallinum), sodium accumulates to high concentrations in vacuoles, and polyols (myo-inositol, d-ononitol, and d-pinitol) accumulate in the cytosol. Polyol synthesis is regulated by NaCl and involves induction and repression of gene expression (D.E. Nelson, B. Shen, and H.J. Bohnert [1998] Plant Cell 10: 753–764). In the study reported here we found increased phloem transport of myo-inositol and reciprocal increased transport of sodium and inositol to leaves under stress. To determine the relationship between increased translocation and sodium uptake, we analyzed the effects of exogenous application of myo-inositol: The NaCl-inducible ice plant myo-inositol 1-phosphate synthase is repressed in roots, and sodium uptake from root to shoot increases without stimulating growth. Sodium uptake and transport through the xylem was coupled to a 10-fold increase of myo-inositol and ononitol in the xylem. Seedlings of the ice plant are not salt-tolerant, and yet the addition of exogenous myo-inositol conferred upon them patterns of gene expression and polyol accumulation observed in mature, salt-tolerant plants. Sodium uptake and transport through the xylem was enhanced in the presence of myo-inositol. The results indicate an interdependence of sodium uptake and alterations in the distribution of myo-inositol. We hypothesize that myo-inositol could serve not only as a substrate for the production of compatible solutes but also as a leaf-to-root signal that promotes sodium uptake. PMID:9880357

  9. Content of methylated inositols in familiar edible plants.

    PubMed

    Negishi, Osamu; Mun'im, Abdul; Negishi, Yukiko

    2015-03-18

    Familiar plants contain large amounts of inositols; soybean, white clover, red clover, bush clover, locust tree, wisteria, and kudzu of the legume family contain pinitol (3-O-methyl-chiro-inositol) at approximately 200-600 mg/100 g fresh weight (FW). The contents of pinitol in other plants were 260 mg/100 g FW for sticky mouse-ear, 275 mg/100 g FW for chickweed, and 332 mg/100 g FW for ginkgo. chiro-Inositol of 191 and 156 mg/100 g FW was also found in dandelion and Japanese mallotus, respectively. Ononitol (4-O-methyl-myo-inositol) of 166 mg/100 g FW was found in sticky mouse-ear. Furthermore, young leaves of ginkgo contained sequoyitol (5-O-methyl-myo-inositol) of 287 mg/100 g FW. Hydroxyl radical scavenging activities of the methylated inositols were higher than those of the original inositols. Effective uses of these familiar edible plants are expected to promote good health. PMID:25734537

  10. Phosphoinositide and Inositol Phosphate Analysis in Lymphocyte Activation

    PubMed Central

    Sauer, Karsten; Huang, Yina Hsing; Lin, Hongying; Sandberg, Mark; Mayr, Georg W.

    2015-01-01

    Lymphocyte antigen receptor engagement profoundly changes the cellular content of phosphoinositide lipids and soluble inositol phosphates. Among these, the phosphoinositides phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylinositol 3,4,5-trisphosphate (PIP3) play key signaling roles by acting as pleckstrin homology (PH) domain ligands that recruit signaling proteins to the plasma membrane. Moreover, PIP2 acts as a precursor for the second messenger molecules diacylglycerol and soluble inositol 1,4,5-trisphosphate (IP3), essential mediators of PKC, Ras/Erk, and Ca2+ signaling in lymphocytes. IP3 phosphorylation by IP3 3-kinases generates inositol 1,3,4,5-tetrakisphosphate (IP4), an essential soluble regulator of PH domain binding to PIP3 in developing T cells. Besides PIP2, PIP3, IP3, and IP4, lymphocytes produce multiple other phosphoinositides and soluble inositol phosphates that could have important physiological functions. To aid their analysis, detailed protocols that allow one to simultaneously measure the levels of multiple different phosphoinositide or inositol phosphate isomers in lymphocytes are provided here. They are based on thin layer, conventional and high-performance liquid chromatographic separation methods followed by radiolabeling or non-radioactive metal-dye detection. Finally, less broadly applicable nonchromatographic methods for detection of specific phosphoinositide or inositol phosphate isomers are discussed. Support protocols describe how to obtain pure unstimulated CD4+CD8+ thymocyte populations for analyses of inositol phosphate turnover during positive and negative selection, key steps in T cell development. PMID:19918943

  11. Biosynthesis and possible functions of inositol pyrophosphates in plants

    PubMed Central

    Williams, Sarah P.; Gillaspy, Glenda E.; Perera, Imara Y.

    2015-01-01

    Inositol phosphates (InsPs) are intricately tied to lipid signaling, as at least one portion of the inositol phosphate signaling pool is derived from hydrolysis of the lipid precursor, phosphatidyl inositol (4,5) bisphosphate. The focus of this review is on the inositol pyrophosphates, which are a novel group of InsP signaling molecules containing diphosphate or triphosphate chains (i.e., PPx) attached to the inositol ring. These PPx-InsPs are emerging as critical players in the integration of cellular metabolism and stress signaling in non-plant eukaryotes. Most eukaryotes synthesize the precursor molecule, myo-inositol (1,2,3,4,5,6)-hexakisphosphate (InsP6), which can serve as a signaling molecule or as storage compound of inositol, phosphorus, and minerals (referred to as phytic acid). Even though plants produce huge amounts of precursor InsP6 in seeds, almost no attention has been paid to whether PPx-InsPs exist in plants, and if so, what roles these molecules play. Recent work has delineated that Arabidopsis has two genes capable of PP-InsP5 synthesis, and PPx-InsPs have been detected across the plant kingdom. This review will detail the known roles of PPx-InsPs in yeast and animal systems, and provide a description of recent data on the synthesis and accumulation of these novel molecules in plants, and potential roles in signaling. PMID:25729385

  12. Inositol-deficient food augments a behavioral effect of long-term lithium treatment mediated by inositol monophosphatase inhibition: an animal model with relevance for bipolar disorder.

    PubMed

    Shtein, Liza; Agam, Galila; Belmaker, R H; Bersudsky, Yuly

    2015-04-01

    Lithium treatment in rodents markedly enhances cholinergic agonists such as pilocarpine. This effect can be reversed in a stereospecific manner by administration of inositol, suggesting that the effect of lithium is caused by inositol monophosphatase inhibition and consequent inositol depletion. If so, inositol-deficient food would be expected to enhance lithium effects. Inositol-deficient food was prepared from inositol-free ingredients. Mice with a homozygote knockout of the inositol monophosphatase 1 gene unable to synthesize inositol endogenously and mimicking lithium-treated animals were fed this diet or a control diet. Lithium-treated wild-type animals were also treated with the inositol-deficient diet or control diet. Pilocarpine was administered after 1 week of treatment, and behavior including seizures was assessed using rating scale. Inositol-deficient food-treated animals, both lithium treated and with inositol monophosphatase 1 knockout, had significantly elevated cholinergic behavior rating and significantly increased or earlier seizures compared with the controls. The effect of inositol-deficient food supports the role of inositol depletion in the effects of lithium on pilocarpine-induced behavior. However, the relevance of this behavior to other more mood-related effects of lithium is not clear. PMID:25679134

  13. Purification and biochemical characterization of Mycobacterium tuberculosis SuhB, an inositol monophosphatase involved in inositol biosynthesis.

    PubMed

    Nigou, Jérôme; Dover, Lynn G; Besra, Gurdyal S

    2002-04-01

    Phosphatidylinositol is an essential component of mycobacteria, and phosphatidylinositol-based lipids such as phosphatidylinositolmannosides, lipomannan, and lipoarabinomannan are major immunomodulatory components of the Mycobacterium tuberculosis cell wall. Inositol monophosphatase (EC 3.1.3.25) is a crucial enzyme in the biosynthesis of free myo-inositol from inositol-1-phosphate, a key substrate for the phosphatidylinositol synthase in mycobacteria. Analysis of the M. tuberculosis genome suggested the presence of four M. tuberculosis gene products that exhibit an inositol monophosphatase signature. In the present report, we have focused on SuhB, which possesses the highest degree of homology with human inositol monophosphatase. SuhB gene was cloned into an E. coli expression vector to over-produce a His-tagged protein, which was purified and characterized. SuhB required divalent metal ions for functional inositol monophosphatase activity, with Mg(2+) being the strongest activator. Inositol monophosphatase activity catalyzed by SuhB was inhibited by the monovalent cation lithium (IC(50) = 0.9 mM). As anticipated, inositol-1-phosphate was the preferred substrate (K(m) = 0.177 +/- 0.025 mM; k(cat) = 3.6 +/- 0.2 s(-)(1)); however, SuhB was also able to hydrolyze a variety of polyol phosphates such as glucitol-6-phosphate, glycerol-2-phosphate, and 2'-AMP. To provide further insight into the structure-function relationship of SuhB, different mutant proteins were generated (E83D, D104N, D107N, W234L, and D235N). These mutations almost completely abrogated inositol monophosphatase activity, thus underlining the importance of these residues in inositol-1-phosphate dephosphorylation. We also identified L81 as a key residue involved in sensitivity to lithium. The L81A mutation rendered SuhB inositol monophosphatase activity 10-fold more resistant to inhibition by lithium (IC(50) = 10 mM). These studies provide the first steps in the delineation of the biosynthesis of the

  14. Convenient synthesis of 4,6-di-O-benzyl-myo-inositol and myo-inositol 1,3,5-orthoesters.

    PubMed

    Praveen, T; Shashidhar, M S

    2001-02-15

    Convenient high yielding methods for the preparation of 4,6-di-O-benzyl-myo-inositol, myo-inositol 1,3,5-orthoformate and myo-inositol 1,3,5-orthoacetate, without involving chromatography are described. Myo-inositol was converted to racemic 2,4-di-O-benzoyl-myo-inositol 1,3,5-orthoformate by successive treatment with triethyl orthoformate and benzoyl chloride. The dibenzoate obtained on benzylation with benzyl bromide and silver(I) oxide gave 2-O-benzoyl-4,6-di-O-benzyl-myo-inositol 1,3,5-orthoformate. Deprotection of the benzoate and the orthoformate with isobutylamine and aqueous trifluoroacetic acid, respectively gave 4,6-di-O-benzyl-myo-inositol in an overall yield of 67%. Myo-inositol orthoformate and orthoacetate were prepared and isolated as their tribenzoates. The free orthoesters were regenerated by deprotection of the benzoates by aminolysis with isobutylamine. PMID:11270820

  15. Inositol Depletion Restores Vesicle Transport in Yeast Phospholipid Flippase Mutants

    PubMed Central

    Yamagami, Kanako; Yamamoto, Takaharu; Sakai, Shota; Mioka, Tetsuo; Sano, Takamitsu; Igarashi, Yasuyuki; Tanaka, Kazuma

    2015-01-01

    In eukaryotic cells, type 4 P-type ATPases function as phospholipid flippases, which translocate phospholipids from the exoplasmic leaflet to the cytoplasmic leaflet of the lipid bilayer. Flippases function in the formation of transport vesicles, but the mechanism remains unknown. Here, we isolate an arrestin-related trafficking adaptor, ART5, as a multicopy suppressor of the growth and endocytic recycling defects of flippase mutants in budding yeast. Consistent with a previous report that Art5p downregulates the inositol transporter Itr1p by endocytosis, we found that flippase mutations were also suppressed by the disruption of ITR1, as well as by depletion of inositol from the culture medium. Interestingly, inositol depletion suppressed the defects in all five flippase mutants. Inositol depletion also partially restored the formation of secretory vesicles in a flippase mutant. Inositol depletion caused changes in lipid composition, including a decrease in phosphatidylinositol and an increase in phosphatidylserine. A reduction in phosphatidylinositol levels caused by partially depleting the phosphatidylinositol synthase Pis1p also suppressed a flippase mutation. These results suggest that inositol depletion changes the lipid composition of the endosomal/TGN membranes, which results in vesicle formation from these membranes in the absence of flippases. PMID:25781026

  16. Dictyostelium uses ether-linked inositol phospholipids for intracellular signalling

    PubMed Central

    Clark, Jonathan; Kay, Robert R; Kielkowska, Anna; Niewczas, Izabella; Fets, Louise; Oxley, David; Stephens, Len R; Hawkins, Phillip T

    2014-01-01

    Inositol phospholipids are critical regulators of membrane biology throughout eukaryotes. The general principle by which they perform these roles is conserved across species and involves binding of differentially phosphorylated inositol head groups to specific protein domains. This interaction serves to both recruit and regulate the activity of several different classes of protein which act on membrane surfaces. In mammalian cells, these phosphorylated inositol head groups are predominantly borne by a C38:4 diacylglycerol backbone. We show here that the inositol phospholipids of Dictyostelium are different, being highly enriched in an unusual C34:1e lipid backbone, 1-hexadecyl-2-(11Z-octadecenoyl)-sn-glycero-3-phospho-(1'-myo-inositol), in which the sn-1 position contains an ether-linked C16:0 chain; they are thus plasmanylinositols. These plasmanylinositols respond acutely to stimulation of cells with chemoattractants, and their levels are regulated by PIPKs, PI3Ks and PTEN. In mammals and now in Dictyostelium, the hydrocarbon chains of inositol phospholipids are a highly selected subset of those available to other phospholipids, suggesting that different molecular selectors are at play in these organisms but serve a common, evolutionarily conserved purpose. PMID:25180230

  17. Inositol hexaphosphate and inositol inhibit DMBA-induced rat mammary cancer.

    PubMed

    Vucenik, I; Yang, G Y; Shamsuddin, A M

    1995-05-01

    Because inositol hexaphosphate (InsP6) and inositol (Ins), contained in plants and most mammalian cells, have been demonstrated to have anti-cancer and anti-cell proliferative action in several experimental models of carcinogenesis we have examined the effect of InsP6 +/- Ins on DMBA-induced rat mammary tumor model. Starting two weeks prior to induction with DMBA, the drinking water of female Sprague-Dawley rats was supplemented with either: 15 mM InsP6, 15 mM Ins, or 15 mM InsP6 + 15 mM Ins; a control group received no inositol compounds. Animals (49-day-old) were given a single intragastric dose of DMBA (5 mg/rat) in 1 ml of corn oil administered by oral intubation. After 45 weeks of treatment, the animals in all the three treatment regimens showed a significant reduction (P < 0.05) in tumor incidence. Tumor number, multiplicity and tumor burden were also significantly (P < 0.05) reduced by InsP6 +/- Ins. When all the parameters were taken into consideration, the best results were obtained by the combination treatment of InsP6 + Ins. Four additional groups not receiving DMBA, but drinking tap water, InsP6, Ins, or InsP6 + Ins of the same molarity as experimental groups were observed for the duration of the study to monitor for any toxicity following this long-term treatment; no significant toxicity as evaluated by body weight gain, serum and bone mineral levels was detected. We demonstrate that InsP6 +/- Ins reproducibly inhibits experimental mammary carcinoma, therefore having great potential as a chemopreventive and adjuvant therapeutic agent for this disease as well. PMID:7767964

  18. The Inositol-3-Phosphate Synthase Biosynthetic Enzyme Has Distinct Catalytic and Metabolic Roles

    PubMed Central

    Frej, Anna D.; Clark, Jonathan; Le Roy, Caroline I.; Lilla, Sergio; Thomason, Peter A.; Otto, Grant P.; Churchill, Grant; Insall, Robert H.; Claus, Sandrine P.; Hawkins, Phillip; Stephens, Len

    2016-01-01

    Inositol levels, maintained by the biosynthetic enzyme inositol-3-phosphate synthase (Ino1), are altered in a range of disorders, including bipolar disorder and Alzheimer's disease. To date, most inositol studies have focused on the molecular and cellular effects of inositol depletion without considering Ino1 levels. Here we employ a simple eukaryote, Dictyostelium discoideum, to demonstrate distinct effects of loss of Ino1 and inositol depletion. We show that loss of Ino1 results in an inositol auxotrophy that can be rescued only partially by exogenous inositol. Removal of inositol supplementation from the ino1− mutant resulted in a rapid 56% reduction in inositol levels, triggering the induction of autophagy, reduced cytokinesis, and substrate adhesion. Inositol depletion also caused a dramatic generalized decrease in phosphoinositide levels that was rescued by inositol supplementation. However, loss of Ino1 triggered broad metabolic changes consistent with the induction of a catabolic state that was not rescued by inositol supplementation. These data suggest a metabolic role for Ino1 that is independent of inositol biosynthesis. To characterize this role, an Ino1 binding partner containing SEL1L1 domains (Q54IX5) and having homology to mammalian macromolecular complex adaptor proteins was identified. Our findings therefore identify a new role for Ino1, independent of inositol biosynthesis, with broad effects on cell metabolism. PMID:26951199

  19. The Inositol-3-Phosphate Synthase Biosynthetic Enzyme Has Distinct Catalytic and Metabolic Roles.

    PubMed

    Frej, Anna D; Clark, Jonathan; Le Roy, Caroline I; Lilla, Sergio; Thomason, Peter A; Otto, Grant P; Churchill, Grant; Insall, Robert H; Claus, Sandrine P; Hawkins, Phillip; Stephens, Len; Williams, Robin S B

    2016-05-15

    Inositol levels, maintained by the biosynthetic enzyme inositol-3-phosphate synthase (Ino1), are altered in a range of disorders, including bipolar disorder and Alzheimer's disease. To date, most inositol studies have focused on the molecular and cellular effects of inositol depletion without considering Ino1 levels. Here we employ a simple eukaryote, Dictyostelium discoideum, to demonstrate distinct effects of loss of Ino1 and inositol depletion. We show that loss of Ino1 results in an inositol auxotrophy that can be rescued only partially by exogenous inositol. Removal of inositol supplementation from the ino1(-) mutant resulted in a rapid 56% reduction in inositol levels, triggering the induction of autophagy, reduced cytokinesis, and substrate adhesion. Inositol depletion also caused a dramatic generalized decrease in phosphoinositide levels that was rescued by inositol supplementation. However, loss of Ino1 triggered broad metabolic changes consistent with the induction of a catabolic state that was not rescued by inositol supplementation. These data suggest a metabolic role for Ino1 that is independent of inositol biosynthesis. To characterize this role, an Ino1 binding partner containing SEL1L1 domains (Q54IX5) and having homology to mammalian macromolecular complex adaptor proteins was identified. Our findings therefore identify a new role for Ino1, independent of inositol biosynthesis, with broad effects on cell metabolism. PMID:26951199

  20. Inositol Trisphosphate Receptor Ca2+ Release Channels

    PubMed Central

    FOSKETT, J. KEVIN; WHITE, CARL; CHEUNG, KING-HO; MAK, DON-ON DANIEL

    2010-01-01

    The inositol 1,4,5-trisphosphate (InsP3) receptors (InsP3Rs) are a family of Ca2+ release channels localized predominately in the endoplasmic reticulum of all cell types. They function to release Ca2+ into the cytoplasm in response to InsP3 produced by diverse stimuli, generating complex local and global Ca2+ signals that regulate numerous cell physiological processes ranging from gene transcription to secretion to learning and memory. The InsP3R is a calcium-selective cation channel whose gating is regulated not only by InsP3, but by other ligands as well, in particular cytoplasmic Ca2+. Over the last decade, detailed quantitative studies of InsP3R channel function and its regulation by ligands and interacting proteins have provided new insights into a remarkable richness of channel regulation and of the structural aspects that underlie signal transduction and permeation. Here, we focus on these developments and review and synthesize the literature regarding the structure and single-channel properties of the InsP3R. PMID:17429043

  1. Synthesis of inositol phosphate-based competitive antagonists of inositol 1,4,5-trisphosphate receptors.

    PubMed

    Konieczny, Vera; Stefanakis, John G; Sitsanidis, Efstratios D; Ioannidou, Natalia-Anastasia T; Papadopoulos, Nikolaos V; Fylaktakidou, Konstantina C; Taylor, Colin W; Koumbis, Alexandros E

    2016-02-16

    Inositol 1,4,5-trisphosphate receptors (IP3Rs) are intracellular Ca(2+) channels that are widely expressed in animal cells, where they mediate the release of Ca(2+) from intracellular stores evoked by extracellular stimuli. A diverse array of synthetic agonists of IP3Rs has defined structure-activity relationships, but existing antagonists have severe limitations. We combined analyses of Ca(2+) release with equilibrium competition binding to IP3R to show that (1,3,4,6)IP4 is a full agonist of IP3R1 with lower affinity than (1,4,5)IP3. Systematic manipulation of this meso-compound via a versatile synthetic scheme provided a family of dimeric analogs of 2-O-butyryl-(1,3,4,6)IP4 and (1,3,4,5,6)IP5 that compete with (1,4,5)IP3 for binding to IP3R without evoking Ca(2+) release. These novel analogs are the first inositol phosphate-based competitive antagonists of IP3Rs with affinities comparable to that of the only commonly used competitive antagonist, heparin, the utility of which is limited by off-target effects. PMID:26818818

  2. Reflections on inositol(s) for PCOS therapy: steps toward success.

    PubMed

    Nestler, John E; Unfer, Vittorio

    2015-07-01

    In polycystic ovary syndrome (PCOS) pathogenesis, both the insulin resistance and the related compensatory hyperinsulinemia are involved. Despite their similarities, Myo-inositol (MI) and d-chiro-inositol (DCI) play different roles in PCOS etiology and therapy. Indeed, in tissue such as the liver both molecules are involved in the insulin signaling, i.e. MI promotes glucose uptake and DCI glycogen synthesis. In reproductive tissue such as the ovary, MI regulates glucose uptake and follicle stimulating hormone (FSH) signaling, whereas DCI is devoted to the insulin-mediated androgen production. The new hypothesis on "DCI paradox" in the ovary has provided the key for a better understanding. Unlike other tissues, ovary is not insulin resistant, indeed because the epimerase enzyme, which converts MI to DCI, is insulin dependent, the "DCI paradox" hypothesis suggests that in the ovary of PCOS women, an increased epimerase activity leads to a DCI overproduction and MI depletion. This imbalance could be the cause of the poor oocyte quality and the impairment in the FSH signaling. Owing to this situation, the focal point is the administration of both MI and DCI in a proper ratio for treating PCOS. This topic, with several other "hot" issues, was the driving thread in the discussion between the two scientists. PMID:26177098

  3. Serotonergic agonists stimulate inositol lipid metabolism in rabbit platelets

    SciTech Connect

    Schaechter, M.; Godfrey, P.P.; Minchin, M.C.W.; McClue, S.J.; Young, M.M.

    1985-10-28

    The metabolism of inositol phospholipids in response to serotonergic agonists was investigated in rabbit platelets. In platelets prelabelled with (/sup 3/H)-inositol, in a medium containing 10 mM LiCl which blocks the enzyme inositol-1-phosphatase, 5-hydroxytryptamine (5-HT) caused a dose-dependent accumulation of inositol phosphates (IP). This suggests a phospholipase-C-mediated breakdown of phosphoinositides. Ketanserin, a selective 5-HT/sub 2/ antagonist, was a potent inhibitor of the 5-HT response, with a Ki of 28 nM, indicating that 5-HT is activating receptors of the 5-HT/sub 2/ type in the platelet. Lysergic acid diethylamide (LSD) and quipazine also caused dose-related increases in inositol phosphate levels, though these were considerably less than those produced by 5-HT. These results show that relatively small changes in phosphoinositide metabolism induced by serotonergic agonists can be investigated in the rabbit platelet, and this cell may therefore be a useful model for the study of some 5-HT receptors. 30 references, 4 figures.

  4. New Functions of the Inositol Polyphosphate 5-Phosphatases in Cancer.

    PubMed

    Erneux, Christophe; Ghosh, Somadri; Ramos, Ana Raquel; Edimo, William's Elong

    2016-01-01

    Inositol polyphosphate 5-phosphatases act on inositol phosphates and phosphoinositides as substrates. They are 10 different isoenzymes and several splice variants in the human genome that are involved in a series of human pathologies such as the Lowe syndrome, the Joubert and MORM syndromes, breast cancer, glioblastoma, gastric cancer and several other type of cancers. Inositol 5-phosphatases can be amplified in human cancer cells, whereas the 3- and 4- phosphatase tumor suppressor PTEN and INPP4B, repectively are often repressed or deleted. The inositol 5-phosphatases are critically involved in a complex network of higly regulated phosphoinositides, affecting the lipid content of PI(3, 4, 5)P3, PI(4, 5)P2 and PI(3, 4)P2. This has an impact on the normal behavior of many intracellular target proteins e.g. protein kinase B (PKB/Akt) or actin binding proteins and final biological responses. The production of PI(3, 4P)2 by dephosphorylation of the substrate PI(3, 4, 5)P3 is particularly important as it produces a new signal messenger in the control of cell migration, invasion and endocytosis. New inhibitors/activators of inositol 5- phosphatases have recently been identified for the possible control of their activity in several human pathologies such as inflamation and cancer. PMID:26916021

  5. Inositol and Sugars in Adaptation of Tomato to Salt 1

    PubMed Central

    Sacher, Robert F.; Staples, Richard C.

    1985-01-01

    Tomato (Lycopersicon esculentum Mill. cv New Yorker) plants subjected to 100 millimolar NaCl plus Hoagland nutrients exhibited a pattern of wilting, recovery of turgor, and finally recovery of growth at a reduced level, which required 3 days. During the nongrowing, adaptation phase there were immediate increases in free hexoses and sucrose which declined to near control levels as growth resumed. There was a steady increase in myo-inositol content which reached its maximal level at the time of growth resumption. The myo-inositol level then remained elevated for the remainder of the experiment. Myo-inositol constituted two-thirds of the soluble carbohydrate in leaves and three-fourths of the soluble carbohydrate in roots of salt-adapted plants. Plants which were alternated daily between salt and control solutions accumulated less myo-inositol and exhibited less growth than the continuously salt-treated plants. In L. pennellii and in salt-tolerant and salt-sensitive breeding lines selected from L. esculentum × L. pennellii BC(1) and F(8), myo-inositol content was highest in the most tolerant genotypes, intermediate in the normal cultivar, and lowest in the sensitive genotype after treatment with salt. PMID:16664009

  6. Inositols affect the mating circadian rhythm of Drosophila melanogaster

    PubMed Central

    Sakata, Kazuki; Kawasaki, Haruhisa; Suzuki, Takahiro; Ito, Kumpei; Negishi, Osamu; Tsuno, Takuo; Tsuno, Hiromi; Yamazaki, Youta; Ishida, Norio

    2015-01-01

    Accumulating evidence indicates that the molecular circadian clock underlies the mating behavior of Drosophila melanogaster. However, information about which food components affect circadian mating behavior is scant. The ice plant, Mesembryanthemum crystallinum has recently become a popular functional food. Here, we showed that the close-proximity (CP) rhythm of D. melanogaster courtship behavior was damped under low-nutrient conditions, but significantly enhanced by feeding the flies with powdered ice plant. Among various components of ice plants, we found that myo-inositol increased the amplitude and slightly shortened the period of the CP rhythm. Real-time reporter assays showed that myo-inositol and D-pinitol shortened the period of the circadian reporter gene Per2-luc in NIH 3T3 cells. These data suggest that the ice plant is a useful functional food and that the ability of inositols to shorten rhythms is a general phenomenon in insects as well as mammals. PMID:26097456

  7. Inositol Hexaphosphate and Inositol Inhibit Colorectal Cancer Metastasis to the Liver in BALB/c Mice

    PubMed Central

    Fu, Min; Song, Yang; Wen, Zhaoxia; Lu, Xingyi; Cui, Lianhua

    2016-01-01

    Inositol hexaphosphate (IP6) and inositol (Ins), naturally occurring carbohydrates present in most mammals and plants, inhibit the growth of numerous cancers both in vitro and in vivo. In this study, we first examined the anti-metastatic effects of IP6 and Ins using a liver metastasis model of colorectal cancer (CRC) in BALB/c mice. CT-26 cells were injected into the splenic capsule of 48 BALB/c mice. The mice were then randomly divided into four groups: IP6, Ins, IP6 + Ins and normal saline control (n = 12 per group). IP6 and/or Ins (80 mg/kg each, 0.2 mL/day) were injected into the gastrointestinal tracts of the mice on the second day after surgery. All mice were sacrificed after 20 days, and the tumor inhibition rates were determined. The results demonstrated that the tumor weights of liver metastases and the tumor inhibition rates were reduced in the experimental groups compared to the control group and that treatment with the combination of IP6 and Ins resulted in greater inhibition of tumor growth than treatment with either compound alone. These findings suggest that IP6 and Ins prevent the development and metastatic progression of colorectal cancer to the liver in mice by altering expression of the extracellular matrix proteins collagen IV, fibronectin and laminin; the adhesion factor receptor integrin-β1; the proteolytic enzyme matrix metalloproteinase 9; and the angiogenic factors vascular endothelial growth factor, basic fibroblast growth factor, and transforming growth factor beta in the tumor metastasis microenvironment. In conclusion, IP6 and Ins inhibited the development and metastatic progression of colorectal cancer to the liver in BALB/c mice, and the effect of their combined application was significantly greater than the effect of either compound alone. This evidence supports further testing of the combined application of IP6 and Ins for the prevention of colorectal cancer metastasis to the liver in clinical studies. PMID:27187454

  8. Inositol Hexaphosphate and Inositol Inhibit Colorectal Cancer Metastasis to the Liver in BALB/c Mice.

    PubMed

    Fu, Min; Song, Yang; Wen, Zhaoxia; Lu, Xingyi; Cui, Lianhua

    2016-01-01

    Inositol hexaphosphate (IP6) and inositol (Ins), naturally occurring carbohydrates present in most mammals and plants, inhibit the growth of numerous cancers both in vitro and in vivo. In this study, we first examined the anti-metastatic effects of IP6 and Ins using a liver metastasis model of colorectal cancer (CRC) in BALB/c mice. CT-26 cells were injected into the splenic capsule of 48 BALB/c mice. The mice were then randomly divided into four groups: IP6, Ins, IP6 + Ins and normal saline control (n = 12 per group). IP6 and/or Ins (80 mg/kg each, 0.2 mL/day) were injected into the gastrointestinal tracts of the mice on the second day after surgery. All mice were sacrificed after 20 days, and the tumor inhibition rates were determined. The results demonstrated that the tumor weights of liver metastases and the tumor inhibition rates were reduced in the experimental groups compared to the control group and that treatment with the combination of IP6 and Ins resulted in greater inhibition of tumor growth than treatment with either compound alone. These findings suggest that IP6 and Ins prevent the development and metastatic progression of colorectal cancer to the liver in mice by altering expression of the extracellular matrix proteins collagen IV, fibronectin and laminin; the adhesion factor receptor integrin-β1; the proteolytic enzyme matrix metalloproteinase 9; and the angiogenic factors vascular endothelial growth factor, basic fibroblast growth factor, and transforming growth factor beta in the tumor metastasis microenvironment. In conclusion, IP6 and Ins inhibited the development and metastatic progression of colorectal cancer to the liver in BALB/c mice, and the effect of their combined application was significantly greater than the effect of either compound alone. This evidence supports further testing of the combined application of IP6 and Ins for the prevention of colorectal cancer metastasis to the liver in clinical studies. PMID:27187454

  9. Synthesis of fagopyritols A1 and B1 from D-chiro-inositol.

    PubMed

    Cid, M Belén; Alfonso, Francisco; Martín-Lomas, Manuel

    2004-09-13

    Fagopyritol A1 (3-O-alpha-d-galactopyranosyl-d-chiro-inositol) and fagopyritol B1 (2-O-alpha-d-galactopyranosyl-d-chiro-inositol) have been synthesized by glycosylation of the diequatorial diol 1,4,5,6-tetra-O-benzoyl-d-chiro-inositol, readily obtained from d-chiro-inositol, with 2,3,4,6-tetra-O-benzyl-d-galactopyranosyl trichloroacetimidate. PMID:15337459

  10. Functional properties of Drosophila inositol trisphosphate receptors.

    PubMed Central

    Swatton, J E; Morris, S A; Wissing, F; Taylor, C W

    2001-01-01

    The functional properties of the only inositol trisphosphate (IP(3)) receptor subtype expressed in Drosophila were examined in permeabilized S2 cells. The IP(3) receptors of S2 cells bound (1,4,5)IP(3) with high affinity (K(d)=8.5+/-1.1 nM), mediated positively co-operative Ca(2+) release from a thapsigargin-sensitive Ca(2+) store (EC(50)=75+/-4 nM, Hill coefficient=2.1+/-0.2), and they were recognized by an antiserum to a peptide conserved in all IP(3) receptor subtypes in the same way as mammalian IP(3) receptors. As with mammalian IP(3) receptors, (2,4,5)IP(3) (EC(50)=2.3+/-0.3 microM) and (4,5)IP(2) (EC(50) approx. 10 microM) were approx. 20- and 100-fold less potent than (1,4,5)IP(3). Adenophostin A, which is typically approx. 10-fold more potent than IP(3) at mammalian IP(3) receptors, was 46-fold more potent than IP(3) in S2 cells (EC(50)=1.67+/-0.07 nM). Responses to submaximal concentrations of IP(3) were quantal and IP(3)-evoked Ca(2+) release was biphasically regulated by cytosolic Ca(2+). Using rapid superfusion to examine the kinetics of IP(3)-evoked Ca(2+) release from S2 cells, we established that IP(3) (10 microM) maximally activated Drosophila IP(3) receptors within 400 ms. The activity of the receptors then slowly decayed (t(1/2)=2.03+/-0.07 s) to a stable state which had 47+/-1% of the activity of the maximally active state. We conclude that the single subtype of IP(3) receptor expressed in Drosophila has similar functional properties to mammalian IP(3) receptors and that analyses of IP(3) receptor function in this genetically tractable organism are therefore likely to contribute to understanding the roles of mammalian IP(3) receptors. PMID:11583592

  11. Using inositol as a biocompatible ligand for efficient transgene expression.

    PubMed

    Zhang, Lei; Bellis, Susan L; Fan, Yiwen; Wu, Yunkun

    2015-01-01

    Transgene transfection techniques using cationic polymers such as polyethylenimines (PEIs) and PEI derivatives as gene vectors have shown efficacy, although they also have shortcomings. PEIs have decent DNA-binding capability and good cell internalization performance, but they cannot deliver gene payloads very efficiently to cell nuclei. In this study, three hyperbranched polyglycerol-polyethylenimine (PG6-PEI) polymers conjugated with myo-inositol (INO) molecules were developed. The three resulting PG6-PEI-INO polymers have an increased number of INO ligands per molecule. PG6-PEI-INO 1 had only 14 carboxymethyl INO (CMINO) units per molecule. PG6-PEI-INO 2 had approximately 130 CMINO units per molecule. PG6-PEI-INO 3 had as high as 415 CMINO units approximately. Mixing PG6-PEI-INO polymers with DNA produced compact nanocomposites. We then performed localization studies using fluorescent microscopy. As the number of conjugated inositol ligands increased in PG6-PEI-INO polymers, there was a corresponding increase in accumulation of the polymers within 293T cell nuclei. Transfection performed with spherical 293T cells yielded 82% of EGFP-positive cells when using PG6-PEI-INO 3 as the vehicle. Studies further revealed that extracellular adenosine triphosphate (eATP) can inhibit the transgene efficiency of PG6-PEI-INO polymers, as compared with PEI and PG6-PEI that were not conjugated with inositol. Our work unveiled the possibility of using inositol as an effective ligand for transgene expression. PMID:25926732

  12. Using inositol as a biocompatible ligand for efficient transgene expression

    PubMed Central

    Zhang, Lei; Bellis, Susan L; Fan, Yiwen; Wu, Yunkun

    2015-01-01

    Transgene transfection techniques using cationic polymers such as polyethylenimines (PEIs) and PEI derivatives as gene vectors have shown efficacy, although they also have shortcomings. PEIs have decent DNA-binding capability and good cell internalization performance, but they cannot deliver gene payloads very efficiently to cell nuclei. In this study, three hyperbranched polyglycerol-polyethylenimine (PG6-PEI) polymers conjugated with myo-inositol (INO) molecules were developed. The three resulting PG6-PEI-INO polymers have an increased number of INO ligands per molecule. PG6-PEI-INO 1 had only 14 carboxymethyl INO (CMINO) units per molecule. PG6-PEI-INO 2 had approximately 130 CMINO units per molecule. PG6-PEI-INO 3 had as high as 415 CMINO units approximately. Mixing PG6-PEI-INO polymers with DNA produced compact nanocomposites. We then performed localization studies using fluorescent microscopy. As the number of conjugated inositol ligands increased in PG6-PEI-INO polymers, there was a corresponding increase in accumulation of the polymers within 293T cell nuclei. Transfection performed with spherical 293T cells yielded 82% of EGFP-positive cells when using PG6-PEI-INO 3 as the vehicle. Studies further revealed that extracellular adenosine triphosphate (eATP) can inhibit the transgene efficiency of PG6-PEI-INO polymers, as compared with PEI and PG6-PEI that were not conjugated with inositol. Our work unveiled the possibility of using inositol as an effective ligand for transgene expression. PMID:25926732

  13. Inositol phospholipid hydrolysis in cultured astrocytes and oligodendrocytes

    SciTech Connect

    Ritchie, T.; Cole, R.; Kim, H.S.; de Vellis, J.; Noble, E.P.

    1987-07-06

    Cultures of astrocytes and oligodendrocytes were prelabeled with /sup 3/H-inositol and the accumulation of /sup 3/H-inositol phosphates was determined following stimulation with a number of neuroactive substances. In astrocytes, norepinephrine (NE) produced the greatest stimulation with significant increase also observed with bradykinin. In oligodendrocytes, the greatest stimulation was produced by carbachol with significant increase also produced by bradykinin, histamine and NE. Carbachol was found to be ineffective in producing stimulation in astrocytes. The accumulation of /sup 3/H-inositol phosphates in astrocytes in response to NE was found to be dependent on the presence of Li/sup +/. The NE stimulation in astrocytes was dose-dependent and had an EC/sub 50/ of 1.2 ..mu..M. This stimulation was blocked by the low concentration of the ..cap alpha../sub 1/-adrenergic antagonist prazosin but not by the ..cap alpha../sub 2/-adrenergic antagonist yohimbine. The NE-stimulated accumulation of /sup 3/H-inositol phosphates in astrocytes was inhibited by the cyclic nucleotide phosphodiesterase inhibitor isobutylmethylxanthine as well as by the cAMP analog dibutyryl cAMP. 34 references, 4 figures, 4 tables.

  14. Probing myo-inositol 1-phosphate synthase with multisubstrate adducts

    PubMed Central

    Deranieh, Rania M.; Greenberg, Miriam L.; Le Calvez, Pierre-B.; Mooney, Maura C.; Migaud, Marie E.

    2015-01-01

    The synthesis of a series of carbohydrate-nucleotide hybrids, designed to be multisubstrate adducts mimicking myo-inositol 1-phosphate synthase first oxidative transition state, is reported. Their ability to inhibit the synthase has been assessed and results have been rationalised computationally to estimate their likely binding mode. PMID:23132282

  15. Lithium and valproate decrease inositol mass and increase expression of the yeast INO1 and INO2 genes for inositol biosynthesis.

    PubMed

    Vaden, D L; Ding, D; Peterson, B; Greenberg, M L

    2001-05-01

    Bipolar affective disorder (manic-depressive illness) is a chronic, severe, debilitating illness affecting 1-2% of the population. The Food and Drug Administration-approved drugs lithium and valproate are not completely effective in the treatment of this disorder, and the mechanisms underlying their therapeutic effects have not been established. We are employing genetic and molecular approaches to identify common targets of lithium and valproate in the yeast Saccharomyces cerevisiae. We show that both drugs affect molecular targets in the inositol metabolic pathway. Lithium and valproate cause a decrease in intracellular myo-inositol mass and an increase in expression of both a structural (INO1) and a regulatory (INO2) gene required for inositol biosynthesis. The opi1 mutant, which exhibits constitutive expression of INO1, is more resistant to inhibition of growth by lithium but not by valproate, suggesting that valproate may inhibit the Ino1p-catalyzed synthesis of inositol 1-phosphate. Consistent with this possibility, growth in valproate leads to decreased synthesis of inositol monophosphate. Thus, both lithium and valproate perturb regulation of the inositol biosynthetic pathway, albeit via different mechanisms. This is the first demonstration of increased expression of genes in the inositol biosynthetic pathway by both lithium and valproate. Because inositol is a key regulator of many cellular processes, the effects of lithium and valproate on inositol synthesis have far-reaching implications for predicting genetic determinants of responsiveness and resistance to these agents. PMID:11278273

  16. Perturbation of the Vacuolar ATPase: A NOVEL CONSEQUENCE OF INOSITOL DEPLETION.

    PubMed

    Deranieh, Rania M; Shi, Yihui; Tarsio, Maureen; Chen, Yan; McCaffery, J Michael; Kane, Patricia M; Greenberg, Miriam L

    2015-11-13

    Depletion of inositol has profound effects on cell function and has been implicated in the therapeutic effects of drugs used to treat epilepsy and bipolar disorder. We have previously shown that the anticonvulsant drug valproate (VPA) depletes inositol by inhibiting myo-inositol-3-phosphate synthase, the enzyme that catalyzes the first and rate-limiting step of inositol biosynthesis. To elucidate the cellular consequences of inositol depletion, we screened the yeast deletion collection for VPA-sensitive mutants and identified mutants in vacuolar sorting and the vacuolar ATPase (V-ATPase). Inositol depletion caused by starvation of ino1Δ cells perturbed the vacuolar structure and decreased V-ATPase activity and proton pumping in isolated vacuolar vesicles. VPA compromised the dynamics of phosphatidylinositol 3,5-bisphosphate (PI3,5P2) and greatly reduced V-ATPase proton transport in inositol-deprived wild-type cells. Osmotic stress, known to increase PI3,5P2 levels, did not restore PI3,5P2 homeostasis nor did it induce vacuolar fragmentation in VPA-treated cells, suggesting that perturbation of the V-ATPase is a consequence of altered PI3,5P2 homeostasis under inositol-limiting conditions. This study is the first to demonstrate that inositol depletion caused by starvation of an inositol synthesis mutant or by the inositol-depleting drug VPA leads to perturbation of the V-ATPase. PMID:26324718

  17. Inositol-phosphodihydroceramides in the periodontal pathogen Tannerella forsythia: Structural analysis and incorporation of exogenous myo-inositol

    PubMed Central

    Megson, Zoë Anne; Pittenauer, Ernst; Duda, Katarzyna Anna; Engel, Regina; Ortmayr, Karin; Koellensperger, Gunda; Mach, Lukas; Allmaier, Günter; Holst, Otto; Messner, Paul; Schäffer, Christina

    2015-01-01

    Background Unique phosphodihydroceramides containing phosphoethanolamine and glycerol have been previously described in Porphyromonas gingivalis. Importantly, they were shown to possess pro-inflammatory properties. Other common human bacteria were screened for the presence of these lipids, and they were found, amongst others, in the oral pathogen Tannerella forsythia. To date, no detailed study into the lipids of this organism has been performed. Methods Lipids were extracted, separated and purified by HPTLC, and analyzed using GC-MS, ESI–MS and NMR. Of special interest was how T. forsythia acquires the metabolic precursors for the lipids studied here. This was assayed by radioactive and stable isotope incorporation using carbon-14 and deuterium labeled myo-inositol, added to the growth medium. Results T. forsythia synthesizes two phosphodihydroceramides (Tf GL1, Tf GL2) which are constituted by phospho-myo-inositol linked to either a 17-, 18-, or 19-carbon sphinganine, N-linked to either a branched 17:0(3-OH) or a linear 16:0(3-OH) fatty acid which, in Tf GL2, is, in turn, ester-substituted with a branched 15:0 fatty acid. T. forsythia lacks the enzymatic machinery required for myo-inositol synthesis but was found to internalize inositol from the medium for the synthesis of both Tf GL1 and Tf GL2. Conclusion The study describes two novel glycolipids in T. forsythia which could be essential in this organism. Their synthesis could be reliant on an external source of myo-inositol. General significance The effects of these unique lipids on the immune system and their role in bacterial virulence could be relevant in the search for new drug targets. PMID:26277409

  18. Identification of a functional 2-keto-myo-inositol dehydratase gene of Sinorhizobium fredii USDA191 required for myo-inositol utilization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sinorhizobium fredii USDA191 is a Gram-negative bacterium capable of forming nitrogen-fixing nodules on soybean roots. The USDA191 idhA gene encoding myo-inositol dehydrogenase, an enzyme necessary for myo-inositol utilization, is known to be involved in competitive nodulation and nitrogen fixation....

  19. Metabolism of myo-[2-3H]Inositol and scyllo-[R-3H]Inositol in Ripening Wheat Kernels 1

    PubMed Central

    Sasaki, Ken; Loewus, Frank A.

    1980-01-01

    Injection of myo-[2-3H]inositol or scyllo-[R-3H]inositol into the peduncular cavity of wheat stalks about 2 to 4 weeks postanthesis led to rapid translocation into the spike and accumulation of label in developing kernels, especially the bran fraction. With myo-[2-3H]inositol, about 50 to 60% of the label was incorporated into high molecular weight cell wall substance in the region of the injection. That portion translocated to the kernels was utilized primarily for cell wall polysaccharide formation and phytate biosynthesis. A small amount was recovered as free myo-inositol and galactinol. When scyllo-[R-3H]inositol was supplied, most of the label was translocated into the developing kernels where it accumulated as free scyllo-inositol and O-α-d-galactopyranosyl-scyllo-inositol in approximately equal amount. None of the label from scyllo-[R-3H]inositol was utilized for either phytate biosynthesis or cell wall polysaccharide formation. PMID:16661513

  20. Crystal Structures of Type-II Inositol Polyphosphate 5-Phosphatase INPP5B with Synthetic Inositol Polyphosphate Surrogates Reveal New Mechanistic Insights for the Inositol 5-Phosphatase Family

    PubMed Central

    2016-01-01

    The inositol polyphosphate 5-phosphatase INPP5B hydrolyzes the 5-phosphate group from water- and lipid-soluble signaling messengers. Two synthetic benzene and biphenyl polyphosphates (BzP/BiPhPs), simplified surrogates of inositol phosphates and phospholipid headgroups, were identified by thermodynamic studies as potent INPP5B ligands. The X-ray structure of the complex between INPP5B and biphenyl 3,3′,4,4′,5,5′-hexakisphosphate [BiPh(3,3′,4,4′,5,5′)P6, IC50 5.5 μM] was determined at 2.89 Å resolution. One inhibitor pole locates in the phospholipid headgroup binding site and the second solvent-exposed ring binds to the His-Tag of another INPP5B molecule, while a molecule of inorganic phosphate is also present in the active site. Benzene 1,2,3-trisphosphate [Bz(1,2,3)P3] [one ring of BiPh(3,3′,4,4′,5,5′)P6] inhibits INPP5B ca. 6-fold less potently. Co-crystallization with benzene 1,2,4,5-tetrakisphosphate [Bz(1,2,4,5)P4, IC50 = 6.3 μM] yielded a structure refined at 2.9 Å resolution. Conserved residues among the 5-phosphatase family mediate interactions with Bz(1,2,4,5)P4 and BiPh(3,3′,4,4′,5,5′)P6 similar to those with the polar groups present in positions 1, 4, 5, and 6 on the inositol ring of the substrate. 5-Phosphatase specificity most likely resides in the variable zone located close to the 2- and 3-positions of the inositol ring, offering insights to inhibitor design. We propose that the inorganic phosphate present in the INPP5B–BiPh(3,3′,4,4′,5,5′)P6 complex mimics the postcleavage substrate 5-phosphate released by INPP5B in the catalytic site, allowing elucidation of two new key features in the catalytic mechanism proposed for the family of phosphoinositide 5-phosphatases: first, the involvement of the conserved Arg-451 in the interaction with the 5-phosphate and second, identification of the water molecule that initiates 5-phosphate hydrolysis. Our model also has implications for the proposed “moving metal” mechanism

  1. Crystal Structures of Type-II Inositol Polyphosphate 5-Phosphatase INPP5B with Synthetic Inositol Polyphosphate Surrogates Reveal New Mechanistic Insights for the Inositol 5-Phosphatase Family.

    PubMed

    Mills, Stephen J; Silvander, Camilla; Cozier, Gyles; Trésaugues, Lionel; Nordlund, Pär; Potter, Barry V L

    2016-03-01

    The inositol polyphosphate 5-phosphatase INPP5B hydrolyzes the 5-phosphate group from water- and lipid-soluble signaling messengers. Two synthetic benzene and biphenyl polyphosphates (BzP/BiPhPs), simplified surrogates of inositol phosphates and phospholipid headgroups, were identified by thermodynamic studies as potent INPP5B ligands. The X-ray structure of the complex between INPP5B and biphenyl 3,3',4,4',5,5'-hexakisphosphate [BiPh(3,3',4,4',5,5')P6, IC50 5.5 μM] was determined at 2.89 Å resolution. One inhibitor pole locates in the phospholipid headgroup binding site and the second solvent-exposed ring binds to the His-Tag of another INPP5B molecule, while a molecule of inorganic phosphate is also present in the active site. Benzene 1,2,3-trisphosphate [Bz(1,2,3)P3] [one ring of BiPh(3,3',4,4',5,5')P6] inhibits INPP5B ca. 6-fold less potently. Co-crystallization with benzene 1,2,4,5-tetrakisphosphate [Bz(1,2,4,5)P4, IC50 = 6.3 μM] yielded a structure refined at 2.9 Å resolution. Conserved residues among the 5-phosphatase family mediate interactions with Bz(1,2,4,5)P4 and BiPh(3,3',4,4',5,5')P6 similar to those with the polar groups present in positions 1, 4, 5, and 6 on the inositol ring of the substrate. 5-Phosphatase specificity most likely resides in the variable zone located close to the 2- and 3-positions of the inositol ring, offering insights to inhibitor design. We propose that the inorganic phosphate present in the INPP5B-BiPh(3,3',4,4',5,5')P6 complex mimics the postcleavage substrate 5-phosphate released by INPP5B in the catalytic site, allowing elucidation of two new key features in the catalytic mechanism proposed for the family of phosphoinositide 5-phosphatases: first, the involvement of the conserved Arg-451 in the interaction with the 5-phosphate and second, identification of the water molecule that initiates 5-phosphate hydrolysis. Our model also has implications for the proposed "moving metal" mechanism. PMID:26854536

  2. Inositol lipid phosphatases in membrane trafficking and human disease.

    PubMed

    Billcliff, Peter G; Lowe, Martin

    2014-07-15

    The specific interaction of phosphoinositides with proteins is critical for a plethora of cellular processes, including cytoskeleton remodelling, mitogenic signalling, ion channel regulation and membrane traffic. The spatiotemporal restriction of different phosphoinositide species helps to define compartments within the cell, and this is particularly important for membrane trafficking within both the secretory and endocytic pathways. Phosphoinositide homoeostasis is tightly regulated by a large number of inositol kinases and phosphatases, which respectively phosphorylate and dephosphorylate distinct phosphoinositide species. Many of these enzymes have been implicated in regulating membrane trafficking and, accordingly, their dysregulation has been linked to a number of human diseases. In the present review, we focus on the inositol phosphatases, concentrating on their roles in membrane trafficking and the human diseases with which they have been associated. PMID:24966051

  3. Huntington's disease: Neural dysfunction linked to inositol polyphosphate multikinase.

    PubMed

    Ahmed, Ishrat; Sbodio, Juan I; Harraz, Maged M; Tyagi, Richa; Grima, Jonathan C; Albacarys, Lauren K; Hubbi, Maimon E; Xu, Risheng; Kim, Seyun; Paul, Bindu D; Snyder, Solomon H

    2015-08-01

    Huntington's disease (HD) is a progressive neurodegenerative disease caused by a glutamine repeat expansion in mutant huntingtin (mHtt). Despite the known genetic cause of HD, the pathophysiology of this disease remains to be elucidated. Inositol polyphosphate multikinase (IPMK) is an enzyme that displays soluble inositol phosphate kinase activity, lipid kinase activity, and various noncatalytic interactions. We report a severe loss of IPMK in the striatum of HD patients and in several cellular and animal models of the disease. This depletion reflects mHtt-induced impairment of COUP-TF-interacting protein 2 (Ctip2), a striatal-enriched transcription factor for IPMK, as well as alterations in IPMK protein stability. IPMK overexpression reverses the metabolic activity deficit in a cell model of HD. IPMK depletion appears to mediate neural dysfunction, because intrastriatal delivery of IPMK abates the progression of motor abnormalities and rescues striatal pathology in transgenic murine models of HD. PMID:26195796

  4. Structure of inositol monophosphatase, the putative target of lithium therapy.

    PubMed Central

    Bone, R; Springer, J P; Atack, J R

    1992-01-01

    Inositol monophosphatase (EC 3.1.3.25), the putative molecular site of action of lithium therapy for manic-depressive illness, plays a key role in the phosphatidylinositol signaling pathway by catalyzing the hydrolysis of inositol monophosphates. To provide a structural basis from which to design better therapeutic agents for manic-depressive illness, the structure of human inositol monophosphatase has been determined to 2.1-A resolution by using x-ray crystallography. The enzyme exists as a dimer of identical subunits, each folded into a five-layered sandwich of three pairs of alpha-helices and two beta-sheets. Sulfate and an inhibitory lanthanide cation (Gd3+) are bound at identical sites on each subunit and establish the positions of the active sites. Each site is located in a large hydrophilic cavern that is at the base of the two central helices where several segments of secondary structure intersect. Comparison of the phosphatase aligned sequences of several diverse genes with the phosphatase structure suggests that the products of these genes and the phosphatase form a structural family with a conserved metal binding site. Images PMID:1332026

  5. myo-Inositol 1,3-acetals as early intermediates during the synthesis of cyclitol derivatives.

    PubMed

    Gurale, Bharat P; Sardessai, Richa S; Shashidhar, Mysore S

    2014-11-18

    Synthetic sequences starting from commercially available myo-inositol necessarily involve protection-deprotection strategies of its six hydroxyl groups. Several strategies have been developed/attempted over the last several decades leading to the synthesis of naturally occurring phosphoinositols, their analogs, and cyclitol derivatives. Of late, myo-inositol 1,3-acetals, which can be obtained by the reductive cleavage of myo-inositol orthoesters have emerged as early intermediates for the synthesis of phosphorylated and other inositol derivatives. This mini-review is an attempt to illustrate the economy and convenience of using myo-inositol 1,3-acetals as early intermediates during syntheses from myo-inositol. PMID:25216930

  6. Regulation of cell-specific inositol metabolism and transport in plant salinity tolerance.

    PubMed Central

    Nelson, D E; Rammesmayer, G; Bohnert, H J

    1998-01-01

    myo-Inositol and its derivatives are commonly studied with respect to cell signaling and membrane biogenesis, but they also participate in responses to salinity in animals and plants. In this study, we focused on L-myo-inositol 1-phosphate synthase (INPS), which commits carbon to de novo synthesis, and myo-inositol O-methyltransferase (IMT), which uses myo-inositol for stress-induced accumulation of a methylinositol, D-ononitol. The Imt and Inps promoters are transcriptionally controlled. We determined that the transcription rates, transcript levels, and protein abundance are correlated. During normal growth, INPS is present in all cells, but IMT is repressed. After salinity stress, the amount of INPS was enhanced in leaves but repressed in roots. IMT was induced in all cell types. The absence of myo-inositol synthesis in roots is compensated by inositol/ononitol transport in the phloem. The mobilization of photosynthate through myo-inositol translocation links root metabolism to photosynthesis. Our model integrates the transcriptional control of a specialized metabolic pathway with physiological reactions in different tissues. The tissue-specific differential regulation of INPS, which leads to a gradient of myo-inositol synthesis, supports root growth and sodium uptake. By inducing expression of IMT and increasing myo-inositol synthesis, metabolic end products accumulate, facilitating sodium sequestration and protecting photosynthesis. PMID:9596634

  7. Results from the International Consensus Conference on Myo-inositol and d-chiro-inositol in Obstetrics and Gynecology: the link between metabolic syndrome and PCOS.

    PubMed

    Facchinetti, Fabio; Bizzarri, Mariano; Benvenga, Salvatore; D'Anna, Rosario; Lanzone, Antonio; Soulage, Christophe; Di Renzo, Gian Carlo; Hod, Moshe; Cavalli, Pietro; Chiu, Tony T; Kamenov, Zdravko A; Bevilacqua, Arturo; Carlomagno, Gianfranco; Gerli, Sandro; Oliva, Mario Montanino; Devroey, Paul

    2015-12-01

    In recent years, interest has been focused to the study of the two major inositol stereoisomers: myo-inositol (MI) and d-chiro-inositol (DCI), because of their involvement, as second messengers of insulin, in several insulin-dependent processes, such as metabolic syndrome and polycystic ovary syndrome. Although these molecules have different functions, very often their roles have been confused, while the meaning of several observations still needs to be interpreted under a more rigorous physiological framework. With the aim of clarifying this issue, the 2013 International Consensus Conference on MI and DCI in Obstetrics and Gynecology identified opinion leaders in all fields related to this area of research. They examined seminal experimental papers and randomized clinical trials reporting the role and the use of inositol(s) in clinical practice. The main topics were the relation between inositol(s) and metabolic syndrome, polycystic ovary syndrome (with a focus on both metabolic and reproductive aspects), congenital anomalies, gestational diabetes. Clinical trials demonstrated that inositol(s) supplementation could fruitfully affect different pathophysiological aspects of disorders pertaining Obstetrics and Gynecology. The treatment of PCOS women as well as the prevention of GDM seem those clinical conditions which take more advantages from MI supplementation, when used at a dose of 2g twice/day. The clinical experience with MI is largely superior to the one with DCI. However, the existence of tissue-specific ratios, namely in the ovary, has prompted researchers to recently develop a treatment based on both molecules in the proportion of 40 (MI) to 1 (DCI). PMID:26479434

  8. Characterization of Inositol-containing Phosphosphingolipids from Tobacco Leaves

    PubMed Central

    Kaul, Karan; Lester, Robert L.

    1975-01-01

    A method for a large scale extraction of phosphoglycosphingolipids from the leaves of Nicotiana tabacum L. has been developed. The phosphosphingolipid concentrate consists of a dozen or more polar lipids as judged by thin layer chromatography. Two of these lipids were purified by chromatography on porous silica beads and partially characterized. These lipids are formulated as: N-acetylglucosamidoglucuronidoinositol phosphorylceramide and glucosamidoglucuronidoinositol phosphorylceramide. Although not fully characterized, the other lipids in the concentrate are inositol-containing phosphosphingolipids with a higher carbohydrate content. PMID:16659016

  9. Myo-Inositol Supplementation to Prevent Gestational Diabetes Mellitus.

    PubMed

    Celentano, Claudio; Matarrelli, Barbara; Mattei, Peter A; Pavone, Giulia; Vitacolonna, Ester; Liberati, Marco

    2016-03-01

    Gestational diabetes mellitus (GDM) is a common complication characterized by increased insulin resistance, and by increased risk for adverse pregnancy outcomes affecting both the mother and the fetus. International guidelines describe optimal ways to recognize it, and the recommended treatment of patients affected to reduce adverse outcomes. Improving insulin resistance could reduce incidence of GDM and its complications. Recently, a few trials have been published on the possible prevention of GDM. Inositol has been proposed as a food supplement that might reduce gestational diabetes incidence in high-risk pregnant women. PMID:26898405

  10. Chloride secretagogues stimulate inositol phosphate formation in shark rectal gland tubules cultured in suspension

    SciTech Connect

    Ecay, T.W.; Valentich, J.D. )

    1991-03-01

    Neuroendocrine activation of transepithelial chloride secretion by shark rectal gland cells is associated with increases in cellular cAMP, cGMP, and free calcium concentrations. We report here on the effects of several chloride secretagogues on inositol phosphate formation in cultured rectal gland tubules. Vasoactive intestinal peptide (VIP), atriopeptin (AP), and ionomycin increase the total inositol phosphate levels of cultured tubules, as measured by ion exchange chromatography. Forskolin, a potent chloride secretagogue, has no effect on inositol phosphate formation. The uptake of {sup 3}H-myo-inositol into phospholipids is very slow, preventing the detection of increased levels of inositol trisphosphate. However, significant increases in inositol monophosphate (IP1) and inositol biphosphate (IP2) were measured. The time course of VIP- and AP-stimulated IP1 and IP2 formation is similar to the effects of these agents on the short-circuit current responses of rectal gland monolayer cultures. In addition, aluminum fluoride, an artificial activator of guanine nucleotide-binding proteins, stimulates IP1 and IP2 formation. We conclude that rectal gland cells contain VIP and AP receptors coupled to the activation of phospholipase C. Coupling may be mediated by G-proteins. Receptor-stimulated increases in inositol phospholipid metabolism is one mechanism leading to increased intracellular free calcium concentrations, an important regulatory event in the activation of transepithelial chloride secretion by shark rectal gland epithelial cells.

  11. Inositol induces mesenchymal-epithelial reversion in breast cancer cells through cytoskeleton rearrangement.

    PubMed

    Dinicola, Simona; Fabrizi, Gianmarco; Masiello, Maria Grazia; Proietti, Sara; Palombo, Alessandro; Minini, Mirko; Harrath, Abdel Halim; Alwasel, Saleh H; Ricci, Giulia; Catizone, Angela; Cucina, Alessandra; Bizzarri, Mariano

    2016-07-01

    Inositol displays multi-targeted effects on many biochemical pathways involved in epithelial-mesenchymal transition (EMT). As Akt activation is inhibited by inositol, we investigated if such effect could hamper EMT in MDA-MB-231 breast cancer cells. In cancer cells treated with pharmacological doses of inositol E-cadherin was increased, β-catenin was redistributed behind cell membrane, and metalloproteinase-9 was significantly reduced, while motility and invading capacity were severely inhibited. Those changes were associated with a significant down-regulation of PI3K/Akt activity, leading to a decrease in downstream signaling effectors: NF-kB, COX-2, and SNAI1. Inositol-mediated inhibition of PS1 leads to lowered Notch 1 release, thus contributing in decreasing SNAI1 levels. Overall, these data indicated that inositol inhibits the principal molecular pathway supporting EMT. Similar results were obtained in ZR-75, a highly metastatic breast cancer line. These findings are coupled with significant changes on cytoskeleton. Inositol slowed-down vimentin expression in cells placed behind the wound-healing edge and stabilized cortical F-actin. Moreover, lamellipodia and filopodia, two specific membrane extensions enabling cell migration and invasiveness, were no longer detectable after inositol addiction. Additionally, fascin and cofilin, two mandatory required components for F-actin assembling within cell protrusions, were highly reduced. These data suggest that inositol may induce an EMT reversion in breast cancer cells, suppressing motility and invasiveness through cytoskeleton modifications. PMID:27237097

  12. Retinoic acid treatment of fibroblasts causes a rapid decrease in ( sup 3 H)inositol uptake

    SciTech Connect

    Sinha, R.; Creek, K.E.; Silverman-Jones, C.; de Luca, L.M. )

    1989-04-01

    NIH 3T3 fibroblasts treated with all-trans-retinoic acid (RA) showed a dramatic decrease in the uptake of ({sup 3}H)inositol compared to solvent-treated controls. The onset of RA-induced inhibition of ({sup 3}H)inositol uptake was rapid with a 10-15% decrease occurring after 2-3 h of RA exposure and 60-70% reduction after 16 h of RA treatment. A progressive dose-dependent decrease in inositol uptake was found as the concentration of RA increased from 10{sup {minus}8} to 10{sup {minus}5} M and the effect was fully reversible within 48 h after RA removal. RA inhibition of inositol uptake was also observed in 3T3-Swiss and Balb/3T3 cells but not in two virally transformed 3T3 cell lines. Phlorizin, amiloride, and monensin inhibited inositol uptake by 66, 74, and 58%, respectively, and this inhibition was additive when the cells were treated with RA as well as these inhibitors. A decreased incorporation of ({sup 3}H)inositol into polyphosphoinositides was also observed in RA-treated cells but not to the same extent as for ({sup 3}H)inositol uptake. In conclusion, RA treatment of 3T3 fibroblasts decreases the uptake of ({sup 3}H)inositol by up to 70% within 8 to 10 h at near physiological concentrations in a reversible and specific manner.

  13. A functional myo-inositol catabolism pathway is essential for rhizopine utilization by Sinorhizobium meliloti.

    PubMed

    Galbraith, M P; Feng, S F; Borneman, J; Triplett, E W; de Bruijn, F J; Rossbach, S

    1998-10-01

    Rhizopine (L-3-O-methyl-scyllo-inosamine) is a symbiosis-specific compound found in alfalfa nodules induced by specific Sinorhizobium meliloti strains. It has been postulated that rhizobial strains able to synthesize and catabolize rhizopine gain a competitive advantage in the rhizosphere. The pathway of rhizopine degradation is analysed here. Since rhizopine is an inositol derivative, it was tested whether inositol catabolism is involved in rhizopine utilization. A genetic locus required for the catabolism of inositol as sole carbon source was cloned from S. meliloti. This locus was delimited by transposon Tn5 mutagenesis and its DNA sequence was determined. Based on DNA similarity studies and enzyme assays, this genetic region was shown to encode an S. meliloti myo-inositol dehydrogenase. Strains that harboured a mutation in the myo-inositol dehydrogenase gene (idhA) did not display myo-inositol dehydrogenase activity, were unable to utilize myo-inositol as sole carbon/energy source, and were unable to catabolize rhizopine. Thus, myo-inositol dehydrogenase activity is essential for rhizopine utilization in S. meliloti. PMID:9802033

  14. Abnormalities in myo-inositol metabolism associated with type 2 diabetes in mice fed a high-fat diet: benefits of a dietary myo-inositol supplementation.

    PubMed

    Croze, Marine L; Géloën, Alain; Soulage, Christophe O

    2015-06-28

    We previously reported that a chronic supplementation with myo-inositol (MI) improved insulin sensitivity and reduced fat accretion in mice. We then tested the potency of such dietary intervention in the prevention of insulin resistance in C57BL/6 male mouse fed a high-fat diet (HFD). In addition, some abnormalities in inositol metabolism were reported to be associated with insulin resistance in several animal and human studies. We then investigated the presence of such anomalies (i.e. inosituria and an inositol intra-tissue depletion) in this diet-induced obesity (DIO) mouse model, as well as the potential benefit of a MI supplementation for inositol intra-tissue deficiency correction. HFD (60 % energy from fat) feeding was associated with inosituria and inositol intra-tissue depletion in the liver and kidneys. MI supplementation (0·58 mg/g per d) restored inositol pools in kidneys (partially) and liver (fully). HFD feeding for 4 months induced ectopic lipid redistribution to liver and muscles, fasting hyperglycaemia and hyperinsulinaemia, insulin resistance and obesity that were not prevented by MI supplementation, despite a significant improvement in insulin sensitivity parameter K insulin tolerance test and a reduction in white adipose tissue (WAT) mass ( - 17 %, P< 0·05). MI supplementation significantly reduced fatty acid synthase activity in epididymal WAT, which might explain its beneficial, but modest, effect on WAT accretion in HFD-fed mice. Finally, we found some abnormalities in inositol metabolism in association with a diabetic phenotype (i.e. insulin resistance and fasting hyperglycaemia) in a DIO mouse model. Dietary MI supplementation was efficient in the prevention of inositol intra-tissue depletion, but did not prevent insulin resistance or obesity efficiently in this mouse model. PMID:25990651

  15. Functional expression of a myo-inositol/H+ symporter from Leishmania donovani.

    PubMed Central

    Drew, M E; Langford, C K; Klamo, E M; Russell, D G; Kavanaugh, M P; Landfear, S M

    1995-01-01

    The vast majority of surface molecules in such kinetoplastid protozoa as members of the genus Leishmania contain inositol and are either glycosyl inositol phospholipids or glycoproteins that are tethered to the external surface of the plasma membrane by glycosylphosphatidylinositol anchors. We have shown that the biosynthetic precursor for these abundant glycolipids, myo-inositol, is translocated across the parasite plasma membrane by a specific transporter that is structurally related to mammalian facilitative glucose transporters. This myo-inositol transporter has been expressed and characterized in Xenopus laevis oocytes. Two-electrode voltage clamp experiments demonstrate that this protein is a sodium-independent electrogenic symporter that appears to utilize a proton gradient to concentrate myo-inositol within the cell. Immunolocalization experiments with a transporter-specific polyclonal antibody reveal the presence of this protein in the parasite plasma membrane. PMID:7565702

  16. Generation of phytate-free seeds in Arabidopsis through disruption of inositol polyphosphate kinases

    PubMed Central

    Stevenson-Paulik, Jill; Bastidas, Robert J.; Chiou, Shean-Tai; Frye, Roy A.; York, John D.

    2005-01-01

    Phytate (inositol hexakisphosphate, IP6) is a regulator of intracellular signaling, a highly abundant animal antinutrient, and a phosphate store in plant seeds. Here, we report a requirement for inositol polyphosphate kinases, AtIPK1 and AtIPK2β, for the later steps of phytate synthesis in Arabidopsis thaliana. Coincident disruption of these kinases nearly ablates seed phytate without accumulation of phytate precursors, increases seed-free phosphate by 10-fold, and has normal seed yield. Additionally, we find a requirement for inositol tetrakisphosphate (IP4)/inositol pentakisphosphate (IP5) 2-kinase activity in phosphate sensing and root hair elongation. Our results define a commercially viable strategy for the genetic engineering of phytate-free grain and provide insights into the role of inositol polyphosphate kinases in phosphate signaling biology. PMID:16107538

  17. Effects of tachykinins on inositol phospholipid hydrolysis in slices of hamster urinary bladder.

    PubMed Central

    Bristow, D. R.; Curtis, N. R.; Suman-Chauhan, N.; Watling, K. J.; Williams, B. J.

    1987-01-01

    Tachykinin-stimulated inositol phospholipid hydrolysis was examined in slices of hamster urinary bladder. In the presence of lithium, to inhibit inositol monophosphatase activity, substance P, eledoisin and related tachykinins induced large, dose-dependent increases in [3H]-inositol monophosphate accumulation. The responses to substance P and eledoisin were not antagonized by the cholinoceptor antagonist, atropine. The rank order of potency for various tachykinins was kassinin greater than neurokinin A greater than neurokinin B greater than eledoisin greater than physaelamin greater than substance P greater than substance P methyl ester. The synthetic analogue [p-Glu6, D-Pro9]SP (6-11) was considerably more potent than its L-prolyl stereoisomer at stimulating inositol phospholipid hydrolysis. These results suggest that in the hamster urinary bladder, tachykinin-induced inositol phospholipid breakdown is mediated via tachykinin receptors of the SP-E type, as opposed to the SP-P type. PMID:3028559

  18. 1 L-myo-Inositol 1-Phosphate Synthase from Arabidopsis thaliana.

    PubMed Central

    Johnson, M. D.; Sussex, I. M.

    1995-01-01

    A recombinant phage containing an Arabidopsis thaliana cDNA sequence encoding a protein with 1L-myo-inositol 1-phosphate synthase (EC 5.5.1.4) activity has been isolated and used for transcriptional and translational studies. The identification of the recombinant phage relied on the observations that (a) the clone complements a mutation in the structural gene for 1L-myo-inositol 1-phosphate synthase in the yeast Saccharomyces cerevisiae, (b) the in vitro synthesized polypeptide enzymatically converts glucose 6-phosphate into inositol 1-phosphate, (c) in vitro transcription and translation of this cDNA sequence produces a polypeptide that is recognized by anti-yeast myo-inositol 1-phosphate synthase antiserum, and (d) inositol regulates the expression of the corresponding gene in Arabidopsis. PMID:12228386

  19. Insights into the activation mechanism of class I HDAC complexes by inositol phosphates

    PubMed Central

    Watson, Peter J.; Millard, Christopher J.; Riley, Andrew M.; Robertson, Naomi S.; Wright, Lyndsey C.; Godage, Himali Y.; Cowley, Shaun M.; Jamieson, Andrew G.; Potter, Barry V. L.; Schwabe, John W. R.

    2016-01-01

    Histone deacetylases (HDACs) 1, 2 and 3 form the catalytic subunit of several large transcriptional repression complexes. Unexpectedly, the enzymatic activity of HDACs in these complexes has been shown to be regulated by inositol phosphates, which bind in a pocket sandwiched between the HDAC and co-repressor proteins. However, the actual mechanism of activation remains poorly understood. Here we have elucidated the stereochemical requirements for binding and activation by inositol phosphates, demonstrating that activation requires three adjacent phosphate groups and that other positions on the inositol ring can tolerate bulky substituents. We also demonstrate that there is allosteric communication between the inositol-binding site and the active site. The crystal structure of the HDAC1:MTA1 complex bound to a novel peptide-based inhibitor and to inositol hexaphosphate suggests a molecular basis of substrate recognition, and an entropically driven allosteric mechanism of activation. PMID:27109927

  20. Inositol Hexakisphosphate Kinase 3 Regulates Metabolism and Lifespan in Mice.

    PubMed

    Moritoh, Yusuke; Oka, Masahiro; Yasuhara, Yoshitaka; Hozumi, Hiroyuki; Iwachidow, Kimihiko; Fuse, Hiromitsu; Tozawa, Ryuichi

    2016-01-01

    Inositol hexakisphosphate kinase 3 (IP6K3) generates inositol pyrophosphates, which regulate diverse cellular functions. However, little is known about its own physiological role. Here, we show the roles of IP6K3 in metabolic regulation. We detected high levels of both mouse and human IP6K3 mRNA in myotubes and muscle tissues. In human myotubes, IP6K3 was upregulated by dexamethasone treatment, which is known to inhibit glucose metabolism. Furthermore, Ip6k3 expression was elevated under diabetic, fasting, and disuse conditions in mouse skeletal muscles. Ip6k3(-/-) mice demonstrated lower blood glucose, reduced circulating insulin, deceased fat mass, lower body weight, increased plasma lactate, enhanced glucose tolerance, lower glucose during an insulin tolerance test, and reduced muscle Pdk4 expression under normal diet conditions. Notably, Ip6k3 deletion extended animal lifespan with concomitant reduced phosphorylation of S6 ribosomal protein in the heart. In contrast, Ip6k3(-/-) mice showed unchanged skeletal muscle mass and no resistance to the effects of high fat diet. The current observations suggest novel roles of IP6K3 in cellular regulation, which impact metabolic control and lifespan. PMID:27577108

  1. Structure of HDAC3 bound to corepressor and inositol tetraphosphate

    PubMed Central

    Watson, Peter J.; Fairall, Louise; Santos, Guilherme M.; Schwabe, John W.R.

    2011-01-01

    Summary Histone deacetylase enzymes (HDACs) are emerging cancer drug targets. They regulate gene expression by removing acetyl groups from lysine residues in histone tails resulting in chromatin condensation. The enzymatic activity of most class I HDACs requires recruitment to corepressor complexes. We report the first structure of an HDAC:corepressor complex - HDAC3 with the deacetylase-activation-domain (DAD) from the SMRT corepressor. The structure reveals two remarkable features. First the SMRT-DAD undergoes a large structural rearrangement on forming the complex. Second there is an essential inositol tetraphosphate molecule, Ins(1,4,5,6)P4, acting as an ‘intermolecular glue’ between the two proteins. Assembly of the complex is clearly dependent on the Ins(1,4,5,6)P4, which may act as a regulator – potentially explaining why inositol phosphates and their kinases have been found to act as transcriptional regulators. This mechanism for the activation of HDAC3 appears to be conserved in class I HDACs from yeast to man and opens novel therapeutic opportunities. PMID:22230954

  2. Inositol Hexakisphosphate Kinase 3 Regulates Metabolism and Lifespan in Mice

    PubMed Central

    Moritoh, Yusuke; Oka, Masahiro; Yasuhara, Yoshitaka; Hozumi, Hiroyuki; Iwachidow, Kimihiko; Fuse, Hiromitsu; Tozawa, Ryuichi

    2016-01-01

    Inositol hexakisphosphate kinase 3 (IP6K3) generates inositol pyrophosphates, which regulate diverse cellular functions. However, little is known about its own physiological role. Here, we show the roles of IP6K3 in metabolic regulation. We detected high levels of both mouse and human IP6K3 mRNA in myotubes and muscle tissues. In human myotubes, IP6K3 was upregulated by dexamethasone treatment, which is known to inhibit glucose metabolism. Furthermore, Ip6k3 expression was elevated under diabetic, fasting, and disuse conditions in mouse skeletal muscles. Ip6k3−/− mice demonstrated lower blood glucose, reduced circulating insulin, deceased fat mass, lower body weight, increased plasma lactate, enhanced glucose tolerance, lower glucose during an insulin tolerance test, and reduced muscle Pdk4 expression under normal diet conditions. Notably, Ip6k3 deletion extended animal lifespan with concomitant reduced phosphorylation of S6 ribosomal protein in the heart. In contrast, Ip6k3−/− mice showed unchanged skeletal muscle mass and no resistance to the effects of high fat diet. The current observations suggest novel roles of IP6K3 in cellular regulation, which impact metabolic control and lifespan. PMID:27577108

  3. Trypanosoma brucei Bloodstream Forms Depend upon Uptake of myo-Inositol for Golgi Complex Phosphatidylinositol Synthesis and Normal Cell Growth

    PubMed Central

    González-Salgado, Amaia; Steinmann, Michael; Major, Louise L.; Sigel, Erwin; Reymond, Jean-Louis

    2015-01-01

    myo-Inositol is a building block for all inositol-containing phospholipids in eukaryotes. It can be synthesized de novo from glucose-6-phosphate in the cytosol and endoplasmic reticulum. Alternatively, it can be taken up from the environment via Na+- or H+-linked myo-inositol transporters. While Na+-coupled myo-inositol transporters are found exclusively in the plasma membrane, H+-linked myo-inositol transporters are detected in intracellular organelles. In Trypanosoma brucei, the causative agent of human African sleeping sickness, myo-inositol metabolism is compartmentalized. De novo-synthesized myo-inositol is used for glycosylphosphatidylinositol production in the endoplasmic reticulum, whereas the myo-inositol taken up from the environment is used for bulk phosphatidylinositol synthesis in the Golgi complex. We now provide evidence that the Golgi complex-localized T. brucei H+-linked myo-inositol transporter (TbHMIT) is essential in bloodstream-form T. brucei. Downregulation of TbHMIT expression by RNA interference blocked phosphatidylinositol production and inhibited growth of parasites in culture. Characterization of the transporter in a heterologous expression system demonstrated a remarkable selectivity of TbHMIT for myo-inositol. It tolerates only a single modification on the inositol ring, such as the removal of a hydroxyl group or the inversion of stereochemistry at a single hydroxyl group relative to myo-inositol. PMID:25888554

  4. Trypanosoma brucei Bloodstream Forms Depend upon Uptake of myo-Inositol for Golgi Complex Phosphatidylinositol Synthesis and Normal Cell Growth.

    PubMed

    González-Salgado, Amaia; Steinmann, Michael; Major, Louise L; Sigel, Erwin; Reymond, Jean-Louis; Smith, Terry K; Bütikofer, Peter

    2015-06-01

    myo-Inositol is a building block for all inositol-containing phospholipids in eukaryotes. It can be synthesized de novo from glucose-6-phosphate in the cytosol and endoplasmic reticulum. Alternatively, it can be taken up from the environment via Na(+)- or H(+)-linked myo-inositol transporters. While Na(+)-coupled myo-inositol transporters are found exclusively in the plasma membrane, H(+)-linked myo-inositol transporters are detected in intracellular organelles. In Trypanosoma brucei, the causative agent of human African sleeping sickness, myo-inositol metabolism is compartmentalized. De novo-synthesized myo-inositol is used for glycosylphosphatidylinositol production in the endoplasmic reticulum, whereas the myo-inositol taken up from the environment is used for bulk phosphatidylinositol synthesis in the Golgi complex. We now provide evidence that the Golgi complex-localized T. brucei H(+)-linked myo-inositol transporter (TbHMIT) is essential in bloodstream-form T. brucei. Downregulation of TbHMIT expression by RNA interference blocked phosphatidylinositol production and inhibited growth of parasites in culture. Characterization of the transporter in a heterologous expression system demonstrated a remarkable selectivity of TbHMIT for myo-inositol. It tolerates only a single modification on the inositol ring, such as the removal of a hydroxyl group or the inversion of stereochemistry at a single hydroxyl group relative to myo-inositol. PMID:25888554

  5. Effects of aluminium on the hepatic inositol polyphosphate phosphatase.

    PubMed Central

    Ali, N; Craxton, A; Sumner, M; Shears, S B

    1995-01-01

    There is speculation that some of the toxic effects of Al3+ may originate from it perturbing inositol phosphate/Ca2+ signalling. For example, in permeabilized L1210 mouse lymphoma cells, 10-50 microM Al3+ activated Ins(1,3,4,5)P4-dependent Ca2+ mobilization and Ins(1,3,4,5)P4 3-phosphatase activity [Loomis-Husselbee, Cullen, Irvine and Dawson (1991) Biochem. J. 277, 883-885]. Ins(1,3,4,5)P4 3-phosphatase activity is performed by a multiple inositol polyphosphate phosphatase (MIPP) that also attacks Ins(1,3,4,5,6)P5 and InsP6 [Craxton, Ali and Shears (1995) Biochem. J. 305, 491-498]: 5-50 microM Al3+ increased MIPP activity towards both Ins(1,3,4,5)P4 (by 30%) and Ins(1,3,4,5,6)P5 (by up to 500%), without affecting metabolism of InsP6. Higher concentrations of Al3+ inhibited metabolism of all three substrates, and in the case of InsP6, Al3+ altered the pattern of accumulating products. When 1-50 microM Al3+ was present, InsP6 became a less effective inhibitor of Ins(1,3,4,5)P4 3-phosphatase activity; this effect did not depend on the presence of cellular membranes, contrary to a previous proposal. The latter phenomenon largely explains how, in a cell-free system where Ins(1,3,4,5)P4 3-phosphatase is inhibited by endogenous InsP6, the addition of Al3+ can apparently increase the enzyme activity. However, there was no effect of either 10 or 25 microM Al3+ (in either the presence or absence of apotransferrin) on inositol phosphate profiles in either Jurkat E6-1 lymphoma cells or AR4-2J pancreatoma cells. PMID:7832774

  6. Changes in inositol phosphates in wild carrot cells upon initiation of cell wall digestion

    SciTech Connect

    Rincon, M.; Boss, W.F.

    1987-04-01

    Previous studies have shown that inositol trisphosphate (IP/sub 3/) stimulated /sup 45/Ca/sup +2/ efflux from fusogenic carrot protoplasts and it was suggested that IP/sub 3/ may serve as a second messenger for the mobilization of intracellular Ca/sup +2/ in higher plant cells. To determine whether or not inositol phosphate metabolism changes in response to external stimuli, the cells were labeled with myo-(2-/sup 3/H) inositol for 18 h and exposed to cell wall digestion enzymes, Driselase. The inositol phosphates were extracted with ice cold 10% TCA and separated by anion exchange chromatography. The radioactivity of the fraction that contained IP/sub 3/ increased 2-3.8 fold and that which contained inositol bisphosphate increased 1.9-2.6 fold within 1.5 min of exposure to Driselase. After 6 min, the radioactivity of both fractions increased 6-7.7 fold and an increase in inositol monophosphate was observed. These data indicate that inositol phosphate metabolism is stimulated by Driselase and suggest polyphosphoinositide hydrolysis occurs upon initiation of cell wall digestion.

  7. Regulation of myo-inositol biosynthesis by p53-ISYNA1 pathway.

    PubMed

    Koguchi, Tomoyuki; Tanikawa, Chizu; Mori, Jinichi; Kojima, Yoshiyuki; Matsuda, Koichi

    2016-06-01

    In response to various cellular stresses, p53 exerts its tumor suppressive effects such as apoptosis, cell cycle arrest, and senescence through the induction of its target genes. Recently, p53 was shown to control cellular homeostasis by regulating energy metabolism, glycolysis, antioxidant effect, and autophagy. However, its function in inositol synthesis was not reported. Through a microarray screening, we found that five genes related with myo-inositol metabolism were induced by p53. DNA damage enhanced intracellular myo-inositol content in HCT116 p53+/+ cells, but not in HCT116 p53-/- cells. We also indicated that inositol 3-phosphate synthase (ISYNA1) which encodes an enzyme essential for myo-inositol biosynthesis as a direct target of p53. Activated p53 regulated ISYNA1 expression through p53 response element in the seventh exon. Ectopic ISYNA1 expression increased myo-inositol levels in the cells and suppressed tumor cell growth. Knockdown of ISYNA1 caused resistance to adriamycin treatment, demonstrating the role of ISYNA1 in p53-mediated growth suppression. Furthermore, ISYNA1 expression was significantly associated with p53 mutation in bladder, breast cancer, head and neck squamous cell carcinoma, lung squamous cell carcinoma, and pancreatic adenocarcinoma. Our findings revealed a novel role of p53 in myo-inositol biosynthesis which could be a potential therapeutic target. PMID:27035231

  8. D-chiro-inositol--its functional role in insulin action and its deficit in insulin resistance.

    PubMed

    Larner, Joseph

    2002-01-01

    In this review we discuss the biological significance of D-chiro-inositol, originally discovered as a component of a putative mediator of intracellular insulin action, where as a putative mediator, it accelerates the dephosphorylation of glycogen synthase and pyruvate dehydrogenase, rate limiting enzymes of non-oxidative and oxidative glucose disposal. Early studies demonstrated a linear relationship between its decreased urinary excretion and the degree of insulin resistance present. When tissue contents, including muscle, of type 2 diabetic subjects were assayed, they demonstrated a more general body deficiency. Administration of D-chiro-inositol to diabetic rats, Rhesus monkeys and now to humans accelerated glucose disposal and sensitized insulin action. A defect in vivo in the epimerization of myo-inositol to chiro-inositol in insulin sensitive tissues of the GK type 2 diabetic rat has been elucidated. Thus, administered D-chiro-inositol may act to bypass a defective normal epimerization of myo-inositol to D-chiro-inositol associated with insulin resistance and act to at least partially restore insulin sensitivity and glucose disposal. PMID:11900279

  9. Effects of inositol supplementation in a cohort of mothers at risk of producing an NTD pregnancy.

    PubMed

    Cavalli, Pietro; Tonni, Gabriele; Grosso, Enrico; Poggiani, Carlo

    2011-11-01

    Neural tube defects (NTDs), most commonly spina bifida and anencephaly, can be prevented with periconceptional intake of folic acid in about 70% of cases. Recurrence of NTDs despite supplementation of high dose of folic acid further suggests that a proportion of NTD cases might be resistant to folic acid. Moreover, heterogeneity of NTDs has been suggested in animal studies, indicating that only some sub-type of NTDs should be considered sensitive to folate intake. Inositol isomers (particularly myo- and chiro-inositol) can prevent folate-resistant NTDs in the curly-tail mutant mouse, suggesting that some cases of human NTDs might benefit from inositol supplementation. In humans, lower inositol blood concentration was found in pregnant women carrying NTD fetuses, whereas a periconceptional combination therapy with folic acid associated with inositol has been linked to normal live births, despite high NTD recurrence risk. Fifteen pregnancies from 12 Caucasian women from different parts of Italy with at least one previous NTD-affected pregnancy underwent periconceptional combined myo-inositol and folic acid supplementation. Maternal serum α-feto-protein levels were found in the normal range, and normal results on ultrasound examination were found in all the pregnancies that followed. No collateral effects or intense uterine contractions were demonstrated in this pilot study in any of the pregnancies after inositol supplementation, and seventeen babies were born without any type of NTD. PMID:21956977

  10. An essential role for an inositol polyphosphate multikinase, Ipk2, in mouse embryogenesis and second messenger production

    PubMed Central

    Frederick, Joshua P.; Mattiske, Deidre; Wofford, Jessica A.; Megosh, Louis C.; Drake, Li Yin; Chiou, Shean-Tai; Hogan, Brigid L. M.; York, John D.

    2005-01-01

    Phospholipase C and several inositol polyphosphate kinase (IPK) activities generate a branched ensemble of inositol polyphosphate second messengers that regulate cellular signaling pathways in the nucleus and cytoplasm. Here, we report that mice deficient for Ipk2 (also known as inositol polyphosphate multikinase), an inositol trisphosphate and tetrakisphosphate 6/5/3-kinase active at several places in the inositol metabolic pathways, die around embryonic day 9.5 with multiple morphological defects, including abnormal folding of the neural tube. Metabolic analysis of Ipk2-deficient cells demonstrates that synthesis of the majority of inositol pentakisphosphate, hexakisphosphate and pyrophosphate species are disrupted, although the presence of 10% residual inositol hexakisphosphate indicates the existence of a minor alternative pathway. Agonist induced inositol tris- and bis-phosphate production and calcium release responses are present in homozygous mutant cells, indicating that the observed mouse phenotypes are a result of failure to produce higher inositol polyphosphates. Our data demonstrate that Ipk2 plays a major role in the synthesis of inositol polyphosphate messengers derived from inositol 1,4,5-trisphosphate and uncovers a role for their production in embryogenesis and normal development. PMID:15939867

  11. Inositol for the prevention of neural tube defects: a pilot randomised controlled trial.

    PubMed

    Greene, Nicholas D E; Leung, Kit-Yi; Gay, Victoria; Burren, Katie; Mills, Kevin; Chitty, Lyn S; Copp, Andrew J

    2016-03-01

    Although peri-conceptional folic acid (FA) supplementation can prevent a proportion of neural tube defects (NTD), there is increasing evidence that many NTD are FA non-responsive. The vitamin-like molecule inositol may offer a novel approach to preventing FA-non-responsive NTD. Inositol prevented NTD in a genetic mouse model, and was well tolerated by women in a small study of NTD recurrence. In the present study, we report the Prevention of Neural Tube Defects by Inositol (PONTI) pilot study designed to gain further experience of inositol usage in human pregnancy as a preliminary trial to a future large-scale controlled trial to evaluate efficacy of inositol in NTD prevention. Study subjects were UK women with a previous NTD pregnancy who planned to become pregnant again. Of 117 women who made contact, ninety-nine proved eligible and forty-seven agreed to be randomised (double-blind) to peri-conceptional supplementation with inositol plus FA or placebo plus FA. In total, thirty-three randomised pregnancies produced one NTD recurrence in the placebo plus FA group (n 19) and no recurrences in the inositol plus FA group (n 14). Of fifty-two women who declined randomisation, the peri-conceptional supplementation regimen and outcomes of twenty-two further pregnancies were documented. Two NTD recurred, both in women who took only FA in their next pregnancy. No adverse pregnancy events were associated with inositol supplementation. The findings of the PONTI pilot study encourage a large-scale controlled trial of inositol for NTD prevention, but indicate the need for a careful study design in view of the unwillingness of many high-risk women to be randomised. PMID:26847388

  12. Transport and metabolism of indole-3-acetyl-myo-inositol-galactoside in seedlings of Zea mays

    NASA Technical Reports Server (NTRS)

    Komoszynski, M.; Bandurski, R. S.

    1986-01-01

    Indole-3-acetyl-myo-inositol galactoside labeled with 3H in the indole and 14C in the galactose moieties was applied to kernels of 5 day old germinating seedlings of Zea mays. Indole-3-acetyl-myo-inositol galactoside was not transported into either the shoot or root tissue as the intact molecule but was instead hydrolyzed to yield [3H]indole-3-acetyl-myo-inositol and [3H]indole-3-acetic acid which were then transported to the shoot with little radioactivity going to the root. With certain assumption concerning the equilibration of applied [3H]indole-3-acetyl-myo-inositol-[U-14C]galactose with the endogenous pool, it may be concluded that indole-3-acetyl-myo-inositol galactoside in the endosperm supplies about 2 picomoles per plant per hour of indole-3-acetyl-myo-inositol and 1 picomole per plant per hour of indole-3-acetic acid to the shoot and thus is comparable to indole-3-acetyl-myo-inositol as a source of indole-acetic acid for the shoot. Quantitative estimates of the amount of galactose in the kernels suggest that [3H]indole-3-acetyl-myo-inositol-[14C]galactose is hydrolyzed after the compound leaves the endosperm but before it reaches the shoot. In addition, [3H]indole-3-acetyl-myo-inositol-[14C]galactose supplies appreciable amounts of 14C to the shoot and both 14C and 3H to an uncharacterized insoluble fraction of the endosperm.

  13. Measurement of Inositol Triphosphate Levels from Rat Hippocampal Slices

    PubMed Central

    Tabatadze, Nino; Woolley, Catherine

    2016-01-01

    Inositol triphosphate (IP3) is an important second messenger that participates in signal transduction pathways in diverse cell types including hippocampal neurons. Stimulation of phospholipase C in response to various stimuli (hormones, growth factors, neurotransmitters, neurotrophins, neuromodulators, odorants, light, etc) results in hydrolysis of phosphatidylinositol 4, 5-bisphosphate (PIP2), a phospholipid that is located in the plasma membrane, and leads to the production of IP3 and diacylglycerol. Binding of IP3 to the IP3 receptor (IP3R) induces Ca2+ release from intracellular stores and enables the initiation of intracellular Ca2+-dependent signaling. Here we describe a procedure for the measurement of cellular IP3 levels in tissue homogenates prepared from rat hippocampal slices.

  14. The biological activity of structurally defined inositol glycans

    PubMed Central

    Goel, Meenakshi; Azev, Viatcheslav N; d’Alarcao, Marc

    2009-01-01

    Background The inositol glycans (IGs) are glycolipid-derived carbohydrates produced by insulin-sensitive cells in response to insulin treatment. IGs exhibit an array of insulin-like activities including stimulation of lipogenesis, glucose transport and glycogen synthesis, suggesting that they may be involved in insulin signal transduction. However, because the natural IGs are structurally heterogeneous and difficult to purify to homogeneity, an understanding of the relationship between structure and biological activity has relied principally on synthetic IGs of defined structure. Discussion This article briefly describes what is known about the role of IGs in signal transduction and reviews the specific biological activities of the structurally defined IGs synthesized and tested to date. Conclusion A pharmacophore for IG activity begins to emerge from the reviewed data and the structural elements necessary for activity are summarized. PMID:20390053

  15. Inositol 1,4,5-trisphosphate induced calcium waves

    NASA Astrophysics Data System (ADS)

    Falcke, M.

    Traveling waves of high concentration of Ca2+ are observed in many different cells and have attracted great interest in experimental and theoretical biological research in recent years. They are created by the nonlinear dynamics of the release and uptake of Ca2+ by intracellular Ca2+ stores like the endoplasmatic or sarcoplasmatic reticulum. Their characteristics depend on other cellular organelles and components like mitochondria and Ca2+ buffers too. Here, we present some mathematical models and results of recent research on intracellular Ca2+ waves generated by the inositol 1,4,5-trisphosphate receptor channel including the modeling of Calcium induced Calcium release, buffer dynamics, impact of mitochondria on wave formation and the effect of the spatial discreteness of the channels releasing Ca2+. Modeling of the communication of Ca2+ waves to adjacent cells through gap junctions concludes this report.

  16. Control of eukaryotic phosphate homeostasis by inositol polyphosphate sensor domains.

    PubMed

    Wild, Rebekka; Gerasimaite, Ruta; Jung, Ji-Yul; Truffault, Vincent; Pavlovic, Igor; Schmidt, Andrea; Saiardi, Adolfo; Jessen, Henning Jacob; Poirier, Yves; Hothorn, Michael; Mayer, Andreas

    2016-05-20

    Phosphorus is a macronutrient taken up by cells as inorganic phosphate (P(i)). How cells sense cellular P(i) levels is poorly characterized. Here, we report that SPX domains--which are found in eukaryotic phosphate transporters, signaling proteins, and inorganic polyphosphate polymerases--provide a basic binding surface for inositol polyphosphate signaling molecules (InsPs), the concentrations of which change in response to P(i) availability. Substitutions of critical binding surface residues impair InsP binding in vitro, inorganic polyphosphate synthesis in yeast, and P(i) transport in Arabidopsis In plants, InsPs trigger the association of SPX proteins with transcription factors to regulate P(i) starvation responses. We propose that InsPs communicate cytosolic P(i) levels to SPX domains and enable them to interact with a multitude of proteins to regulate P(i) uptake, transport, and storage in fungi, plants, and animals. PMID:27080106

  17. Alterations in Lipid and Inositol Metabolisms in Two Dopaminergic Disorders

    PubMed Central

    Berger, Hannah S.; Do, Kieu Trinh; Kastenmüller, Gabi; Wahl, Simone; Adamski, Jerzy; Peters, Annette; Krumsiek, Jan; Suhre, Karsten; Haslinger, Bernhard; Ceballos-Baumann, Andres; Gieger, Christian; Winkelmann, Juliane

    2016-01-01

    Background Serum metabolite profiling can be used to identify pathways involved in the pathogenesis of and potential biomarkers for a given disease. Both restless legs syndrome (RLS) and Parkinson`s disease (PD) represent movement disorders for which currently no blood-based biomarkers are available and whose pathogenesis has not been uncovered conclusively. We performed unbiased serum metabolite profiling in search of signature metabolic changes for both diseases. Methods 456 metabolites were quantified in serum samples of 1272 general population controls belonging to the KORA cohort, 82 PD cases and 95 RLS cases by liquid-phase chromatography and gas chromatography separation coupled with tandem mass spectrometry. Genetically determined metabotypes were calculated using genome-wide genotyping data for the 1272 general population controls. Results After stringent quality control, we identified decreased levels of long-chain (polyunsaturated) fatty acids of individuals with PD compared to both RLS (PD vs. RLS: p = 0.0001 to 5.80x10-9) and general population controls (PD vs. KORA: p = 6.09x10-5 to 3.45x10-32). In RLS, inositol metabolites were increased specifically (RLS vs. KORA: p = 1.35x10-6 to 3.96x10-7). The impact of dopaminergic drugs was reflected in changes in the phenylalanine/tyrosine/dopamine metabolism observed in both individuals with RLS and PD. Conclusions A first discovery approach using serum metabolite profiling in two dopamine-related movement disorders compared to a large general population sample identified significant alterations in the polyunsaturated fatty acid metabolism in PD and implicated the inositol metabolism in RLS. These results provide a starting point for further studies investigating new perspectives on factors involved in the pathogenesis of the two diseases as well as possible points of therapeutic intervention. PMID:26808974

  18. Relationship Between Myo-Inositol Supplementary and Gestational Diabetes Mellitus

    PubMed Central

    Zheng, Xiangqin; Liu, Zhaozhen; Zhang, Yulong; Lin, Yuan; Song, Jianrong; Zheng, Lianghui; Lin, Sheng

    2015-01-01

    Abstract To determine whether myo-inositol supplement will increase the action of endogenous insulin, which is mainly measured by markers of insulin resistance such as homeostasis model assessment of insulin resistance. PubMed, Cochrane Library, Embase, and web of science were comprehensively searched using “gestational diabetes mellitus” and “myo-inositol” to identify relevant studies. Both subject headings and free texts were adopted. The methodological quality of the included studies were assessed and pooled analyzed by the methods recommended by the Cochrane collaboration. A total of 5 trials containing 513 participants were included. There was a significant reduction in aspects of gestational diabetes incidence (risk ratio [RR], 0.29; 95% confidence interval (95% CI), 0.19–0.44), birth weight (mean difference [MD], −116.98; 95% CI, −208.87 to −25.09), fasting glucose oral glucose tolerance test (OGTT) (MD, −0.36; 95% CI, −0.51 to −0.21), 1-h glucose OGTT (MD, −0.63; 95% CI, −1.01 to −0.26), 2-h glucose OGTT (MD, −0.45; 95% CI, −0.75 to −0.16), and related complications (odds ratio [OR], 0.28; 95% CI 0.14–0.58). On the basis of current evidence, myo-inositol supplementation reduces the development of gestational diabetes mellitus (GDM), although this conclusion requires further evaluation in large-scale, multicenter, blinded randomized controlled trials. PMID:26496267

  19. Design and Synthesis of an Inositol Phosphate Analog Based on Computational Docking Studies

    PubMed Central

    Peng, Zhenghong; Maxwell, David; Sun, Duoli; Ying, Yunming; Schuber, Paul T.; Bhanu Prasad, Basvoju A.; Gelovani, Juri; Yung, Wai-Kwan Alfred; Bornmann, William G.

    2014-01-01

    A virtual library of 54 inositol analog mimics of In(1,4,5)P3 has been docked, scored, and ranked within the binding site of human inositol 1,4,5-trisphosphate 3-kinase A (IP3-3KA). Chemical synthesis of the best scoring structure that also met distance criteria for 3′-OH to -P in Phosphate has been attempted along with the synthesis of (1S,2R,3S,4S)-3-fluoro-2,4-dihydroxycyclohexanecarboxylic acid as an inositol analog, useful for non-invasive visualization and quantitation of IP3-3KA enzymatic activity PMID:25110363

  20. Improving D-glucaric acid production from myo-inositol in E. coli by increasing MIOX stability and myo-inositol transport.

    PubMed

    Shiue, Eric; Prather, Kristala L J

    2014-03-01

    D-glucaric acid has been explored for a myriad of potential uses, including biopolymer production and cancer treatment. A biosynthetic route to produce D-glucaric acid from glucose has been constructed in Escherichia coli (Moon et al., 2009b), and analysis of the pathway revealed myo-inositol oxygenase (MIOX) to be the least active enzyme. To increase pathway productivity, we explored protein fusion tags for increased MIOX solubility and directed evolution for increased MIOX activity. An N-terminal SUMO fusion to MIOX resulted in a 75% increase in D-glucaric acid production from myo-inositol. While our directed evolution efforts did not yield an improved MIOX variant, our screen isolated a 941 bp DNA fragment whose expression led to increased myo-inositol transport and a 65% increase in D-glucaric acid production from myo-inositol. Overall, we report the production of up to 4.85 g/L of D-glucaric acid from 10.8 g/L myo-inositol in recombinant E. coli. PMID:24333274

  1. Light induces a rapid and transient increase in inositol-trisphosphate in toad rod outer segments

    SciTech Connect

    Brown, J.E.; Blazynski, C.; Cohen, A.I.

    1987-08-14

    The sub-second time course of changes in the content of (/sup 3/H)inositol-1,4,5-trisphosphate was determined in rod outer segments from very rapidly frozen Bufo retinas that had been incubated with (/sup 3/H)inositol. Rod outer segments were cut off frozen specimens with a cryostat microtome and the water soluble extracts were analyzed. The content of (/sup 3/H)inositol-1,4,5-trisphosphate rose after approximately 250 msec of bright illumination, but returned to the unstimulated level after 1 sec, whether the stimulus remained on or not. That is, there was rapid but transient change in the content of (/sup 3/H)inositol-1,4,5-trisphosphate after the onset of stimulation.

  2. Inositol prevents folate-resistant neural tube defects in the mouse.

    PubMed

    Greene, N D; Copp, A J

    1997-01-01

    Clinical trials demonstrate that up to 70% of neural tube defects (NTDs) can be prevented by folic acid supplementation in early pregnancy, whereas the remaining NTDs are resistant to folate. Here, we show that a second vitamin, myo-inositol, is capable of significantly reducing the incidence of spinal NTDs in curly tail mice, a genetic model of folate-resistant NTDs. Inositol increases flux through the inositol/lipid cycle, stimulating protein kinase C activity and upregulating expression of retinoic acid receptor beta, specifically in the caudal portion of the embryonic hindgut. This reduces the delay in closure of the posterior neuropore, the embryonic defect that is known to lead directly to spina bifida in curly tail embryos. Our findings reveal a molecular pathway of NTD prevention and suggest the possible efficacy of combined treatment with folate and inositol in overcoming the majority of human NTDs. PMID:8986742

  3. "Inosaminoacids": novel inositol-amino acid hybrid structures accessed by microbial arene oxidation.

    PubMed

    Pilgrim, Sarah; Kociok-Köhn, Gabriele; Lloyd, Matthew D; Lewis, Simon E

    2011-04-28

    Microbial 1,2-dihydroxylation of sodium benzoate permits the rapid construction of novel inositol-amino acid hybrid structures. Both β- and γ-amino acids are accessible by means of an acylnitroso Diels-Alder cycloaddition. PMID:21409268

  4. Inositol metabolism in WRK-1 cells. Relationship of hormone-sensitive to -insensitive pools of phosphoinositides

    SciTech Connect

    Monaco, M.E.

    1987-09-25

    Previous studies have indicated the existence of two separate pools of phosphoinositides in WRK-1 cells; one is labile and hormone-sensitive with respect to turnover, while the other is stable. Hormonal stimulation results in a rapid increase in /sup 32/Pi incorporation into the sensitive pool, while in the absence of hormone, incorporation of /sup 32/Pi into this pool is slow. Results are quite different when (/sup 3/H)inositol is the precursor utilized. Incorporation of (/sup 3/H)inositol into hormone-sensitive phosphoinositides is not stimulated in the presence of hormone, suggesting entry of this exogenous precursor into the cycle by a route other than the resynthetic phase of the cycle. Furthermore, failure of hormone to induce loss of (/sup 3/H)phosphoinositide in pulse-chase experiments in the absence of lithium suggests reutilization of the (/sup 3/H)inositol moiety generated by phosphodiesteratic cleavage of hormone-sensitive phosphoinositide. Time course studies indicate that the relative rates of incorporation of (/sup 3/H)inositol into sensitive and insensitive phosphoinositide remain constant from 2 to 24 h. Several factors are capable of increasing (/sup 3/H)inositol incorporation into hormone-insensitive phosphoinositide including vasopressin, calcium ionophores, and manganese. On the other hand, vasopressin treatment appears to decrease incorporation of (/sup 3/H)inositol into the hormone-sensitive pool, probably by shifting the equilibrium between phosphoinositides and inositol phosphates, since the decrease in radioactivity observed in the phosphoinositides is equaled by the increase observed in that in the inositol phosphates.

  5. Enantiomers of myo-inositol-1,3,4-trisphosphate and myo-inositol-1,4,6 -trisphosphate: stereospecific recognition by cerebellar and platelet myo-inositol-1,4,5-trisphosphate receptors.

    PubMed

    Murphy, C T; Bullock, A J; Lindley, C J; Mills, S J; Riley, A M; Potter, B V; Westwick, J

    1996-11-01

    The naturally occurring tetrakisphosphate myo-inositol-1,3,4, 6-tetrakisphosphate [Ins(1,3,4,6)P4] was able to release Ca2+ from the intracellular stores of permeabilized rabbit platelets but was 40-fold less potent than D-myo-inositol-1,4,5-trisphosphate [Ins(1,4,5)P3]. The Ca2+ releasing activity of Ins(1,3,4,6)P4 was rationalized by envisaging two alternative receptor binding orientations in which the vicinal D-1,6-bisphosphate of Ins(1,3,4,6)P4 mimics the D-4,5-bisphosphate in the Ins(1,4,5)P3 binding conformation. This rationalization predicted that Ins(1,4,5)P3 regioisomers [i.e, D-myo-inositol -1,4,6-trisphosphate [D-Ins(1,4,6)P3] and D-myo-inositol-1,3,6 -trisphosphate [D-Ins(1,3,6)P3

  6. Regioselective Opening of myo-Inositol Orthoesters: Mechanism and Synthetic Utility

    PubMed Central

    2013-01-01

    Acid hydrolysis of myo-inositol 1,3,5-orthoesters, apart from orthoformates, exclusively affords the corresponding 2-O-acyl myo-inositol products via a 1,2-bridged five-membered ring dioxolanylium ion intermediate observed by NMR spectroscopy. These C-2-substituted inositol derivatives provide valuable precursors for rapid and highly efficient routes to 2-O-acyl inositol 1,3,4,5,6-pentakisphosphates and myo-inositol 1,3,4,5,6-pentakisphosphate with biologically interesting and anticancer properties. Deuterium incorporation into the α-methylene group of such alkyl ester products (2-O-C(O)CD2R), when the analogous alkyl orthoester is treated with deuterated acid, is established utilizing the novel orthoester myo-inositol 1,3,5-orthobutyrate as an example. Such deuterated ester products provide intermediates for deuterium-labeled synthetic analogues. Investigation into this selective formation of 2-O-ester products and the deuterium incorporation is presented with proposed mechanisms from NMR experiments. PMID:23438216

  7. New compatible solutes related to Di-myo-inositol-phosphate in members of the order Thermotogales.

    PubMed Central

    Martins, L O; Carreto, L S; Da Costa, M S; Santos, H

    1996-01-01

    The accumulation of intracellular organic solutes was examined in six species of the order Thermotogales by nuclear magnetic resonance spectroscopy. The newly discovered compounds di-2-O-beta-mannosyl-di-myo-inositol-1,1'(3,3')-phosphate and di-myo-inositol-1,3'-phosphate were identified in Thermotoga maritima and Thermotoga neapolitana. In the latter species, at the optimum temperature and salinity the organic solute pool was composed of di-myo-inositol-1,1'(3,3')-phosphate, beta-glutamate, and alpha-glutamate in addition to di-myo-inositol-1,3'-phosphate and di-2-O-beta-mannosyl-di-myo-inositol-1,1'(3,3')-phosphate. The concentrations of the last two solutes increased dramatically at supraoptimal growth temperatures, whereas beta-glutamate increased mainly in response to a salinity stress. Nevertheless, di-myo-inositol-1,1'(3,3')-phosphate was the major compatible solute at salinities above the optimum for growth. The amino acids alpha-glutamate and proline were identified under optimum growth conditions in Thermosipho africanus, and beta-mannosylglycerate, trehalose, and glycine betaine were detected in Petrotoga miotherma. Organic solutes were not detected, under optimum growth conditions, in Thermotoga thermarum and Fervidobacterium islandicum, which have a low salt requirement or none. PMID:8824608

  8. Does myo-inositol effect on PCOS follicles involve cytoskeleton regulation?

    PubMed

    Bizzarri, Mariano; Cucina, Alessandra; Dinicola, Simona; Harrath, Abdel Halim; Alwasel, Saleh H; Unfer, Vittorio; Bevilacqua, Arturo

    2016-06-01

    Inositol metabolism is severely impaired in follicles obtained from cystic ovaries, leading to deregulated insulin transduction and steroid synthesis. On the contrary, inositol administration to women suffering from polycystic ovary syndrome (PCOS) has been proven to efficiently counteract most of the clinical hallmarks displayed by PCOS patients, including insulin resistance, hyperandrogenism and oligo-amenorrhea. We have recently observed that myo-inositol induces significant changes in cytoskeletal architecture of breast cancer cells, by modulating different biochemical pathways, eventually modulating the epithelial-mesenchymal transition. We hypothesize that inositol and its monophosphate derivatives, besides their effects on insulin transduction, may efficiently revert histological and functional features of cystic ovary by inducing cytoskeleton rearrangements. We propose an experimental model that could address not only whether inositol modulates cytoskeleton dynamics in both normal and cystic ovary cells, but also whether this effect may interfere with ovarian steroidogenesis. A more compelling understanding of the mechanisms of action of inositol (and its derivatives) would greatly improve its therapeutic utilization, by conferring to current treatments a well-grounded scientific rationale. PMID:27142131

  9. Determination of phytic acid and inositol pentakisphosphates in foods by high-performance ion chromatography.

    PubMed

    Chen, Qingchuan

    2004-07-28

    A high-performance anion exchange chromatographic method was adapted for the quantitative determination of phytic acid and inositol pentakisphosphate isomers (excluding enantiomers) in foods. Because of the cost and limited availability of inositol phosphate standards, a phytic acid sodium salt standard was used for the calculation of an average relative response factor for the quantification of inositol pentakisphosphate isomers, and the purity of phytic acid sodium salt standard was also accurately established. The detection limits (S/N = 3) for phytic acid and inositol pentakisphosphates were in the range of 1.5-3.4 microM (0.1-0.2 microg/100 microL). This method has been successfully applied to the determination of phytic acid and inositol pentakisphosphates in a variety of beans and nuts after extraction with 0.5 M HCl and cleanup with solid phase extraction cartridges. The results demonstrated that there was a strong correlation between either the phytic acid content or the total content of phytic acid together with inositol pentakisphosphates and the total dietary fiber content in the group of all raw dry beans and in the group of raw dry black beans but not in the group of raw dry red kidney beans, which was probably due to the insufficient number of the raw dry red kidney bean samples. PMID:15264889

  10. Purification, cloning and characterization of a GPI inositol deacylase from Trypanosoma brucei

    PubMed Central

    Güther, Maria Lucia Sampaio; Leal, Simone; Morrice, Nicholas A.; Cross, George A.M.; Ferguson, Michael A.J.

    2001-01-01

    Inositol acylation is an obligatory step in glycosylphosphatidylinositol (GPI) biosynthesis whereas mature GPI anchors often lack this modification. The GPI anchors of Trypanosoma brucei variant surface glycoproteins (VSGs) undergo rounds of inositol acylation and deacylation during GPI biosynthesis and the deacylation reactions are inhibited by diisopropylfluorophosphate (DFP). Inositol deacylase was affinity labelled with [3H]DFP and purified. Peptide sequencing was used to clone GPIdeAc, which encodes a protein with significant sequence and hydropathy similarity to mammalian acyloxyacyl hydrolase, an enzyme that removes fatty acids from bacterial lipopolysaccharide. Both contain a signal sequence followed by a saposin domain and a GDSL-lipase domain. GPIdeAc–/– trypanosomes were viable in vitro and in animals. Affinity-purified HA-tagged GPIdeAc was shown to have inositol deacylase activity. However, total inositol deacylase activity was only reduced in GPIdeAc–/– trypanosomes and the VSG GPI anchor was indistinguishable from wild type. These results suggest that there is redundancy in T.brucei inositol deacylase activity and that there is another enzyme whose sequence is not recognizably related to GPIdeAc. PMID:11532956

  11. Effects of inositol trisphosphate on calcium mobilization in high-voltage and saponin-permeabilized platelets

    SciTech Connect

    Gear, A.R.L.; Hallam, T.J.

    1986-03-01

    Interest in phosphatidylinositol metabolism has been greatly stimulated by the findings that diglyceride and inositol phosphates may serve as second messengers in modulating cellular function. Formation of 1,4,5-inositol trisphosphate (IP/sub 3/), in particular, has been linked to mobilization of intracellular calcium in a number of cell types. The authors have examined the ability of IP/sub 3/ to mobilize calcium in human platelets permeabilized by either saponin or high-voltage discharge. Saponin at 15 ..mu..g/ml effectively permeabilized platelets to exogenous inositol 1,4,5-trisphosphate which released bound (/sup 45/Ca) within 1 min and with a Ka of 7.4 +/- 4.1 ..mu..M. A small (25%) azide-sensitive pool was also responsive to inositol trisphosphate. The calcium pools were completely discharged by A-23187 and the ATP-dependent uptake was prevented by dinitrophenol. In contrast to the result with saponin, platelets accessed by high-voltage discharge were insensitive to challenge by inositol 1,4,5-trisphosphate. The data suggest that while inositol 1,4,5-trisphosphate can rapidly mobilize platelet calcium, the ability to demonstrate this depends on the method of permeabilization.

  12. Enzyme-assisted total synthesis of the optical antipodes D-myo-inositol 3,4,5-trisphosphate and D-myo-inositol 1,5, 6-trisphosphate: aspects of their structure-activity relationship to biologically active inositol phosphates.

    PubMed

    Adelt, S; Plettenburg, O; Stricker, R; Reiser, G; Altenbach, H J; Vogel, G

    1999-04-01

    Unambiguous total syntheses of both optical antipodes of the enantiomeric pair D-myo-inositol 3,4,5-trisphosphate (Ins(3,4,5)P3) and D-myo-inositol 1,5,6-trisphosphate (Ins(1,5,6)P3) are described. The ring system characteristic of myo-inositol was constructed de novo from p-benzoquinone. X-ray data for the enzymatically resolved (1S,2R,3R,4S)-1,4-diacetoxy-2,3-dibromocyclohex-5-ene enabled the unequivocal assignment of the absolute configuration. Subsequent transformations under stereocontrolled conditions led to enantiopure C2-symmetrical 1,4-(di-O-benzyldiphospho)conduritol B derivatives. Their synthetic potential was exploited to prepare Ins(3,4,5,6)P4 and Ins(1,4,5,6)P4 in three steps. With a recently identified and partially purified InsP5/InsP4 phosphohydrolase from Dictyostelium discoideum, these enantiomers could be converted to the target compounds, Ins(3,4,5)P3 and Ins(1,5,6)P3, on a preparative scale. An HPLC system employed for both purification of the inositol phosphates and analytical runs ensured that the products were isomerically homogeneous. The sensitivity of detection achieved by a complexometric postcolumn derivatization method indicates that the complexation properties of Ins(3,4,5)P3/Ins(1,5,6)P3 resemble those of Ins(1,2,3)P3, a compound with antioxidant potential. The set of inositol phosphates synthesized was used to clarify structural motifs important for molecular recognition by p42(IP4), a high-affinity Ins(1,3,4,5)P4/PtdIns(3,4,5)P3-specific binding protein from pig cerebellum. PMID:10197969

  13. The intracellular distribution of inositol polyphosphates in HL60 promyeloid cells.

    PubMed Central

    Stuart, J A; Anderson, K L; French, P J; Kirk, C J; Michell, R H

    1994-01-01

    1. HL60 promyeloid cells contain high intracellular concentrations of inositol polyphosphates, notably inositol 1,3,4,5,6-pentakisphosphate (InsP5) and inositol hexakisphosphate (InsP6). To determine their intracellular location(s), we studied the release of inositol (poly)phosphates, of ATP, and of cytosolic and granule-enclosed enzymes from cells permeabilized by four different methods. 2. When cells were treated with digitonin, all of the inositol phosphates were released in parallel with the cytosolic constituents. Most of the InsP5 and InsP6 was released before significant permeabilization of azurophil granules. 3. Similar results were obtained from cells preloaded with ethylene glycol and permeabilized by osmotic lysis. 4. Electroporation at approximately 500 V/cm caused rapid release of free inositol. Higher field strengths provoked release of most of the ATP, InsP5 and InsP6, but only slight release of the intracellular enzymes. Multiple discharges released approximately 80-90% of total InsP5 and InsP6. In the absence of bivalent-cation chelators, InsP5 and InsP6 were released less readily than ATP. 5. Treatment of cells with Staphylococcus aureus alpha-toxin caused quantitative release of inositol and ATP, without release of intracellular enzymes. However, inositol phosphates were released much less readily than inositol or ATP. Even after prolonged incubation with a high concentration of alpha-toxin, only approximately 50-70% of InsP2, InsP3 and InsP4 and < or = 20% of InsP5 and InsP6 were released, indicating that the high charge or large hydrated radius of InsP5 and InsP6 might limit their release through small toxin-induced pores. 6. These results indicate that most intracellular inositol metabolites are either in, or in rapid exchange with, the cytosolic compartment of HL60 cells. However, they leave open the possibility that a small proportion of cellular InsP5 and InsP6 (< or = 10-20%) might be in some intracellular bound form. Images Figure 2 PMID

  14. Intracellular calcium channels: inositol-1,4,5-trisphosphate receptors

    PubMed Central

    Fedorenko, Olena A.; Popugaeva, Elena; Enomoto, Masahiro; Stathopulos, Peter B.; Ikura, Mitsuhiko; Bezprozvanny, Ilya

    2014-01-01

    The inositol-1,4,5-trisphosphate receptors (InsP3Rs) are the major intracellular Ca2+-release channels in cells. Activity of InsP3Rs is essential for elementary and global Ca2+ events in the cell. There are three InsP3Rs isoforms that are present in mammalian cells. In this review review we will focus primarily on InsP3R type 1. The InsP3R1 is a predominant isoform in neurons and it is most extensively studied isoform. Combination of biophysical and structural methods revealed key mechanisms of InsP3R function and modulation. Cell biological and biochemical studies lead to identification of a large number of InsP3R-binding proteins. InsP3Rs are involved in the regulation of numerous physiological processes, including learning and memory, proliferation, differentiation, development and cell death. Malfunction of InsP3R1 play a role in a number of neurodegenerative disorders and other disease states. InsP3Rs represent a potentially valuable drug target for treatment of these disorders and for modulating activity of neurons and other cells. Future studies will provide better understanding of physiological functions of InsP3Rs in health and disease. PMID:24300389

  15. Prometabolites of 5-Diphospho-myo-inositol Pentakisphosphate.

    PubMed

    Pavlovic, Igor; Thakor, Divyeshsinh T; Bigler, Laurent; Wilson, Miranda S C; Laha, Debabrata; Schaaf, Gabriel; Saiardi, Adolfo; Jessen, Henning J

    2015-08-10

    Diphospho-myo-inositol phosphates (PP-InsP(y)) are an important class of cellular messengers. Thus far, no method for the transport of PP-InsP(y) into living cells is available. Owing to their high negative charge density, PP-InsP(y) will not cross the cell membrane. A strategy to circumvent this issue involves the generation of precursors in which the negative charges are masked with biolabile groups. A PP-InsP(y) prometabolite would require twelve to thirteen biolabile groups, which need to be cleaved by cellular enzymes to release the parent molecules. Such densely modified prometabolites of phosphate esters and anhydrides have never been reported to date. This study discloses the synthesis of such agents and an analysis of their metabolism in tissue homogenates by gel electrophoresis. The acetoxybenzyl-protected system is capable of releasing 5-PP-InsP5 in mammalian cell/tissue homogenates within a few minutes and can be used to release 5-PP-InsP5 inside cells. These molecules will serve as a platform for the development of fundamental tools required to study PP-InsP(y) physiology. PMID:26014370

  16. Cellular Cations Control Conformational Switching of Inositol Pyrophosphate Analogues.

    PubMed

    Hager, Anastasia; Wu, Mingxuan; Wang, Huanchen; Brown, Nathaniel W; Shears, Stephen B; Veiga, Nicolás; Fiedler, Dorothea

    2016-08-22

    The inositol pyrophosphate messengers (PP-InsPs) are emerging as an important class of cellular regulators. These molecules have been linked to numerous biological processes, including insulin secretion and cancer cell migration, but how they trigger such a wide range of cellular responses has remained unanswered in many cases. Here, we show that the PP-InsPs exhibit complex speciation behaviour and propose that a unique conformational switching mechanism could contribute to their multifunctional effects. We synthesised non-hydrolysable bisphosphonate analogues and crystallised the analogues in complex with mammalian PPIP5K2 kinase. Subsequently, the bisphosphonate analogues were used to investigate the protonation sequence, metal-coordination properties, and conformation in solution. Remarkably, the presence of potassium and magnesium ions enabled the analogues to adopt two different conformations near physiological pH. Understanding how the intrinsic chemical properties of the PP-InsPs can contribute to their complex signalling outputs will be essential to elucidate their regulatory functions. PMID:27460418

  17. Enigmatic ion-exchange behavior of myo-inositol phosphates.

    PubMed

    Shelor, C Phillip; Liao, Hongzhu; Kadjo, Akinde Florence; Dasgupta, Purnendu K

    2015-05-01

    The separation of myo-inositol mono-, di-, tri-, tetra-, pentakis-, and hexakisphosphate (InsP1, InsP2, InsP3, InsP4, InsP5, InsP6) was carried out using hydroxide eluent ion chromatography. Acid hydrolysis of InsP6 (phytate) was used to prepare a distribution of InsPs, ranging from InsP1 to InsP5's and including unhydrolyzed InsP6. Counting all possible positional isomers (many of which have stereoisomers that will not be separable by conventional ion exchange), 40 chromatographically separable peaks are possible; up to 22 were separated and identified by mass spectrometry. InsPs show unusual ion-exchange behavior in two respects: (a) the retention order is not monotonically related with the charge on the ion and (b) at the same hydroxide eluent concentration, retention is greatly dependent on the eluent metal cation. The retention of InsP3-InsP6 was determined to be controlled by steric factors while elution was influenced by eluent cation complexation. These highly phosphorylated InsPs have a much greater affinity for alkali metals (Li(+) > Na(+) > K(+)) than quaternary ammonium ions. This difference in cation affinity was exploited to improve separation through the use of a tetramethylammonium hydroxide-sodium hydroxide gradient. PMID:25865157

  18. Interconversion of inositol (1,4,5)-trisphosphate to inositol (1,3,4,5)-tetrakisphosphate and (1,3,4)-trisphosphate in permeabilized adrenal glomerulosa cells is calcium-sensitive and ATP-dependent

    SciTech Connect

    Rossier, M.F.; Dentand, I.A.; Lew, P.D.; Capponi, A.M.; Vallotton, M.B.

    1986-08-29

    The metabolism of (/sub 3/H)inositol (1,4,5)-trisphosphate was followed in permeabilized bovine adrenal glomerulosa cells. At low Ca++ concentration (pCa = 7.2), more than 90% of (/sub 3/H)inositol (1,4,5)-trisphosphate had disappeared within 2 min, while two other metabolites, (/sub 3/H)inositol (1,3,4)-trisphosphate and (/sub 3/H)inositol (1,3,4,5)-tetrakisphosphate appeared progressively. At higher Ca++ concentrations (pCa = 5.7 and 4.8), the formation of these two metabolites was markedly increased, but completely abolished if the medium was ATP-depleted. The peak levels for the generation of (/sub 3/H)inositol (1,3,4,5)-tetrakisphosphate (1 min) preceded those of (3H)inositol (1,3,4)-trisphosphate and were closely correlated. These results suggest that, in adrenal glomerulosa cells, the isomer inositol (1,3,4)-trisphosphate is generated from inositol (1,4,5)-trisphosphate via a calcium-sensitive and ATP-dependent phosphorylation/dephosphorylation pathway involving the formation of inositol (1,3,4,5)-tetrakisphosphate.

  19. Metabolic evidence for the order of addition of individual phosphate esters in the myo-inositol moiety of inositol hexakisphosphate in the duckweed Spirodela polyrhiza L.

    PubMed Central

    Brearley, C A; Hanke, D E

    1996-01-01

    The aquatic monocotyledonous plant Spirodela polyrhiza was labelled with [33P]Pi for short periods under non-equilibrium conditions. An InsP6 fraction was obtained and dissected by using enantiospecific (enzymic) and non-enantiospecific (chemical) means to determine the relative labelling of individual phosphate substituents on the inositol ring of InsP6. Phosphates in positions D-1, -2, -3, -4, -5 and -6 contained approx. 21%, 32-39%, 9-10%, 14-16%, 19-23% and 16-18% of the label respectively. We conclude from the foregoing, together with identities [described in the preceding paper, Brearley and Hanke (1996) Biochem. J. 314, 215-225] of inositol phosphates found in this plant at a developmental stage associated with massive accumulation of InsP6, that synthesis of InsP6 from myo-inositol proceeds according to the sequence Ins3P-->Ins(3,4)P2-->Ins(3,4,6)P3-->Ins(3,4,5,6)P4-->Ins(1,3,4,5,6 ) P5-->InsP6 in Spirodela polyrhiza. These results represent the first description of the synthetic sequence to InsP6 in the plant kingdom and the only comprehensive description of endogenous inositol phosphates in any plant tissue. The sequence described differs from that reported in the slime mould Dictyostelium discoideum. PMID:8660287

  20. Regiospecific phosphohydrolases from Dictyostelium as tools for the chemoenzymatic synthesis of the enantiomers D-myo-inositol 1,2,4-trisphosphate and D-myo-inositol 2,3,6-trisphosphate: non-physiological, potential analogues of biologically active D-myo-inositol 1,3,4-trisphosphate.

    PubMed

    Adelt, S; Plettenburg, O; Dallmann, G; Ritter, F P; Shears, S B; Altenbach, H J; Vogel, G

    2001-10-22

    A new de novo synthesis of the enantiomeric pair D-myo-inositol 1,2,4-trisphosphate and D-myo-inositol 2,3,6-trisphosphate is described. Starting from enantiopure dibromocyclohexenediol, several C2 symmetrical building blocks were synthesized which gave access to D-myo-inositol 1,2,4,5-tetrakisphosphate and D-myo-inositol 1,2,3,6-tetrakisphosphate. Exploiting the high regiospecificity of two partially purified phosphohydrolases from Dictyostelium, a 5-phosphatase and a phytase, the inositol tetrakisphosphates were converted enzymatically to the target compounds. Their potential to modulate the activity of Ins3,4,5,6P4 1-kinase was investigated and compared with the effects of D-myo-inositol 1,3,4-trisphosphate. PMID:11591506

  1. Early effects of Escherichia coli endotoxin infusion on vasopressin-stimulated breakdown and metabolism of inositol lipids in rat hepatocytes

    SciTech Connect

    Rodriguez de Turco, E.B.; Spitzer, J.A.

    1988-08-30

    The turnover of vasopressin-stimulated 32P-phosphoinositides and 32P-phosphatidic acid and accumulation of (2-3H)-inositol phosphates were examined in hepatocytes from rats infused i.v. with saline and E. coli endotoxin for 3 hrs. Within 60s of VP stimulation the decrease in phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate labeling as well as the increased uptake of 32P into phosphatidic acid were similar in both groups. However, at a later time (300s) the 32P-phosphatidylinositol turnover was greatly decreased concomitantly with a higher labeling of phosphatidic acid. The accumulation of (2-3H)-inositol phosphates in ET-cells was significantly decreased both at 30s and 600s after VP addition. The distribution of (2-3H)-inositol labeling accumulated in the different inositol phosphate fractions over the first 30s of VP stimulation showed a tendency to lower accumulation of inositol trisphosphate, and a significantly lower accumulation of inositol bisphosphate simultaneously with a higher labeling of the inositol tetrakisphosphate fraction. These observations reflect an early effect of ET-infusion on VP-stimulated inositol lipid turnover and on the subsequent metabolism of the released inositol phosphates.

  2. Metabolism of the phospholipid precursor inositol and its relationship to growth and viability in the natural auxotroph Schizosaccharomyces pombe.

    PubMed Central

    Fernandez, S; Homann, M J; Henry, S A; Carman, G M

    1986-01-01

    Phospholipid metabolism in the fission yeast Schizosaccharomyces pombe was examined. Three enzymes of phospholipid biosynthesis, cytidine diphosphate diacylglycerol synthase (CDP-DG), phosphatidylinositol (PI) synthase, and phosphatidylserine (PS) synthase, were characterized in extracts of S. pombe cells. Contrary to an earlier report, we were able to demonstrate that CDP-DG served as a precursor for PI and PS biosynthesis in S. pombe. S. pombe is naturally auxotrophic for the phospholipid precursor inositol. We found that S. pombe was much more resistant to loss of viability during inositol starvation than artificially generated inositol auxotrophs of Saccharomyces cerevisiae. The phospholipid composition of S. pombe cells grown in inositol-rich medium (50 microM) was similar to that of S. cerevisiae cells grown under similar conditions. However, growth of S. pombe at low inositol concentrations (below 30 microM) affected the ratio of the anionic phospholipids PI and PS, while the relative proportions of other glycerophospholipids remained unchanged. During inositol starvation, the rate of PI synthesis decreased rapidly, and there was a concomitant increase in the rate of PS synthesis. Phosphatidic acid and CDP-DG, which are precursors to these phospholipids, also increased when PI synthesis was blocked by lack of exogenous inositol. The major product of turnover of inositol-containing phospholipids in S. pombe was found to be free inositol, which accumulated in the medium and could be reused by the cell. Images PMID:3011744

  3. Complexation studies on inositol-phosphates: IV. Ca(II) complexes of myo-inositol 1,4,5-trisphosphate.

    PubMed

    Schmitt, L; Schlewer, G; Spiess, B

    1992-01-01

    The stability constants of the complexes formed between Ca2+ and the myo-inositol 1,4,5-triphosphate (Ins(1,4,5)P3) were determined by potentiometric titration in two different media and temperature conditions (medium 1: I = 0.1 M But4NBr, 25 degrees C; medium 2: I = 0.2 M KCl, 37 degrees C). Mainly because of the presence of potassium the results obtained in these media show large differences in both the nature and the stability of the complexes. In medium 1, MH2L and M2L species are formed along with the ML and MHL species which also exist in medium 2. In addition, the stability of the latter species decreases by more than one log unit in going from medium 1 to medium 2. In an attempt to assess the biological significance of the metal binding to Ins(1,4,5)P3, the results were compared to the Ca2+-ATP complexes that form in the same media conditions. Taking into account the relative stability of the complexes of both systems, it is likely that the action or metabolism of Ins(1,4,5)P3 may be influenced by coordination of either alkali or alkali-earth cations. PMID:1588343

  4. Inositol Metabolism in Plants. V. Conversion of Myo-inositol to Uronic Acid and Pentose Units of Acidic Polysaccharides in Root-tips of Zea mays 1

    PubMed Central

    Roberts, R. M.; Deshusses, J.; Loewus, F.

    1968-01-01

    The metabolism of myo-inositol-2-14C, d-glucuronate-1-14C, d-glucuronate-6-14C, and l-methionine-methyl-14C to cell wall polysaccharides was investigated in excised root-tips of 3 day old Zea mays seedlings. From myo-inositol, about one-half of incorporated label was recovered in ethanol insoluble residues. Of this label, about 90% was solubilized by treatment, first with a preparation of pectinase-EDTA, then with dilute hydrochloric acid. The only labeled constituents in these hydrolyzates were d-galacturonic acid, d-glucuronic acid, 4-O-methyl-d-glucuronic acid, d-xylose, and l-arabinose, or larger oligosaccharide fragments containing these units. Medium external to excised root-tips grown under sterile conditions in myo-inositol-2-14C contained labeled polysaccharide. When label was supplied in the form of d-glucuronate, the pattern of labeled uronic acid and pentose units in cell wall polysaccharides resembled that obtained from labeled myo-inositol, indicating that both substances were metabolized along a common path during polysaccharide formation, and that methylation occurred at a step subsequent to uronic acid formation. When label was supplied in the form of l-methionine-methyl-14C, 4-O-methyl-d-glucuronic acid was the only labeled monosaccharide component that survived enzymatic or acid hydrolysis. Zea mays endosperm, a known source of phytin, developed maximal phytase activity after the third day of germination. Results obtained here suggest that myo-inositol released by hydrolysis of phytin represents the initial precursor of a normal, possibly predominant pathway for the formation of uronic acids in plants. PMID:16656871

  5. Inositol trisphosphate metabolism in carrot (Daucus carota L. ) cells

    SciTech Connect

    Memon, A.R.; Rincon, M.; Boss, W.F. )

    1989-10-01

    The metabolism of exogenously added D-myo-(1-{sup 3}H)inositol 1,4,5-trisphosphate (IP{sub 3}) has been examined in microsomal membrane and soluble fractions of carrot cells grown in suspension culture. When ({sup 3}H)IP{sub 3} was added to a microsomal membrane fraction, ({sup 3}H)IP{sub 2} was the primary metabolite consisting of approximately 83% of the total recovered ({sup 3}H) by electrophoresis. ({sup 3}H)IP was only 6% of the ({sup 3}H) recovered, and 10% of the ({sup 3}H)IP{sub 3} was not further metabolized. In contrast, when ({sup 3}H)IP{sub 3} was added to the soluble fraction, approximately equal amounts of ({sup 3}H)IP{sub 2} and ({sup 3}H)IP were recovered. Ca{sup 2+} (100 micromolar) tended to enhance IP{sub 3} dephosphorylation but inhibited the IP{sub 2} dephosphorylation in the soluble fraction by about 20%. MoO{sub 4}{sup 2{minus}} (1 millimolar) inhibited the dephosphorylation of IP{sub 3} by the microsomal fraction and the dephosphorylation of IP{sub 2} by the soluble fraction. MoO{sub 4}{sup 2{minus}}, however, did not inhibit the dephosphorylation of IP{sub 3} by the soluble fraction. Li{sup +} (10 and 50 millimolar) had no effect on IP{sub 3} metabolism in either the soluble or membrane fraction; however, Li{sup +} (50 millimolar) inhibited IP{sub 2} dephosphorylation in the soluble fraction about 25%.

  6. Stereo- and regiospecificity of yeast phytases-chemical synthesis and enzymatic conversion of the substrate analogues neo- and L-chiro-inositol hexakisphosphate.

    PubMed

    Adelt, Stephan; Podeschwa, Michael; Dallmann, Guido; Altenbach, Hans-Josef; Vogel, Günter

    2003-02-01

    Phytases are enzymes that catalyze the hydrolysis of phosphate esters in myo-inositol hexakisphosphate (phytic acid). The precise routes of enzymatic dephosphorylation by phytases of the yeast strains Saccharomyces cerevisiae and Pichia rhodanensis have been investigated up to the myo-inositol trisphosphate level, including the absolute configuration of the intermediates. Stereoselective assignment of the myo-inositol pentakisphosphates (D-myo-inositol 1,2,4,5,6-pentakisphosphate and D-myo-inositol 1,2,3,4,5-pentakisphosphate) generated was accomplished by a new method based on enantiospecific enzymatic conversion and HPLC analysis. Via conduritol B or E derivatives the total syntheses of two epimers of myo-inositol hexakisphosphate, neo-inositol hexakisphosphate and L-chiro-inositol hexakisphosphate were performed to examine the specificity of the yeast phytases with these substrate analogues. A comparison of kinetic data and the degradation pathways determined gave the first hints about the molecular recognition of inositol hexakisphosphates by the enzymes. Exploitation of the high stereo- and regiospecificity observed in the dephosphorylation of neo- and L-chiro-inositol hexakisphosphate made it possible to establish enzyme-assisted steps for the synthesis of D-neo-inositol 1,2,5,6-tetrakisphosphate, L-chiro-inositol 1,2,3,5,6-pentakisphosphate and L-chiro-inositol 1,2,3,6-tetrakisphosphate. PMID:12697168

  7. Inositol lipid metabolism in vasopressin stimulated hepatocytes from rats infused with tumor necrosis factor

    SciTech Connect

    Spitzer, J.A.; Rodriguez de Turco, E.B. )

    1989-05-30

    We studied the effect of i.v. infusion of human recombinant tumor necrosis factor alpha (rHuTNF alpha, Cetus, 15 micrograms/100 g bw over 3 h) on vasopressin (VP)-stimulated {sup 32}P-inositol lipid turnover and the release of {sup 3}H-inositol phosphates in isolated rat hepatocytes. The early VP-induced decrease (within 30 s) in {sup 32}P-phosphatidylinositol 4-phosphate and {sup 32}P-phosphatidylinositol 4,5-bisphosphate labeling was significantly reduced (-40%) and at the same time the uptake of {sup 32}P into phosphatidic acid was 50% lower than in saline-infused (matched control) rats. Within 5 min of VP-stimulation, lower {sup 32}P phosphatidylinositol (-40%) and higher {sup 32}P-phosphatidic acid (+30%) labeling were observed in rHuTNF alpha-infused rats. Infusion of rHuTNF alpha also affected the VP-induced release of {sup 3}H-inositol phosphates. The accumulation of {sup 3}H-inositol-labeled water soluble products was decreased by 25% and 17% at 30 s and 10 min, respectively. These data show that rHuTNF alpha mimics early perturbations induced by Escherichia coli endotoxin infusion in VP-stimulated inositol lipid metabolism in rat hepatocytes.

  8. Osmoregulation of Na(+)-inositol cotransporter activity and mRNA levels in brain glial cells.

    PubMed

    Paredes, A; McManus, M; Kwon, H M; Strange, K

    1992-12-01

    During plasma hypertonicity brain volume is regulated acutely by electrolyte uptake and chronically by accumulation of organic solutes such as inositol. Cultured rat C6 glioma cells, an astrocyte-like cell line, show a similar pattern of volume control. Volume regulatory accumulation of inositol requires external inositol, indicating that membrane transport plays a central role in this process. The inositol uptake pathway is Na+ dependent and exhibits Michaelis-Menten kinetics. Chronic hypertonic acclimation results in a twofold increase in the maximum velocity of the transporter without changing the Km. Hypertonic stress also results in a 17-fold increase in transporter mRNA. Elevation of mRNA levels precedes activation of the transporter by 4-6 h, suggesting that increased inositol uptake is mediated by synthesis and membrane insertion of new transport proteins. Reacclimation of hypertonic cells to isotonicity causes a rapid reduction of transporter mRNA levels to control levels within 4 h. In contrast, downregulation of transport activity does not begin until between 10 and 24 h after reexposure to isotonicity. PMID:1476169

  9. Asymmetric distribution of glucose and indole-3-acetyl-myo-inositol in geostimulated Zea mays seedlings

    NASA Technical Reports Server (NTRS)

    Momonoki, Y. S.; Bandurski, R. S. (Principal Investigator)

    1988-01-01

    Indole-3-acetyl-myo-inositol occurs in both the kernel and vegetative shoot of germinating Zea mays seedlings. The effect of a gravitational stimulus on the transport of [3H]-5-indole-3-acetyl-myo-inositol and [U-14C]-D-glucose from the kernel to the seedling shoot was studied. Both labeled glucose and labeled indole-3-acetyl-myo-inositol become asymmetrically distributed in the mesocotyl cortex of the shoot with more radioactivity occurring in the bottom half of a horizontally placed seedling. Asymmetric distribution of [3H]indole-3-acetic acid, derived from the applied [3H]indole-3-acetyl-myo-inositol, occurred more rapidly than distribution of total 3H-radioactivity. These findings demonstrate that the gravitational stimulus can induce an asymmetric distribution of substances being transported from kernel to shoot. They also indicate that, in addition to the transport asymmetry, gravity affects the steady state amount of indole-3-acetic acid derived from indole-3-acetyl-myo-inositol.

  10. Simulations of inositol phosphate metabolism and its interaction with InsP(3)-mediated calcium release.

    PubMed Central

    Mishra, Jyoti; Bhalla, Upinder S

    2002-01-01

    Inositol phosphates function as second messengers for a variety of extracellular signals. Ins(1,4,5)P(3) generated by phospholipase C-mediated hydrolysis of phosphatidylinositol bisphosphate, triggers numerous cellular processes by regulating calcium release from internal stores. The Ins(1,4,5)P(3) signal is coupled to a complex metabolic cascade involving a series of phosphatases and kinases. These enzymes generate a range of inositol phosphate derivatives, many of which have signaling roles of their own. We have integrated published biochemical data to build a mass action model for InsP(3) metabolism. The model includes most inositol phosphates that are currently known to interact with each other. We have used this model to study the effects of a G-protein coupled receptor stimulus that activates phospholipase C on the inositol phosphates. We have also monitored how the metabolic cascade interacts with Ins(1,4,5)P(3)-mediated calcium release. We find temporal dynamics of most inositol phosphates to be strongly influenced by the elaborate networking. We also show that Ins(1,3,4,5)P(4) plays a key role in InsP(3) dynamics and allows for paired pulse facilitation of calcium release. Calcium oscillations produce oscillatory responses in parts of the metabolic network and are in turn temporally modulated by the metabolism of InsP(3). PMID:12202356

  11. Asymmetric distribution of glucose and indole-3-acetyl-myo-inositol in geostimulated Zea mays seedlings.

    PubMed

    Momonoki, Y S

    1988-01-01

    Indole-3-acetyl-myo-inositol occurs in both the kernel and vegetative shoot of germinating Zea mays seedlings. The effect of a gravitational stimulus on the transport of [3H]-5-indole-3-acetyl-myo-inositol and [U-14C]-D-glucose from the kernel to the seedling shoot was studied. Both labeled glucose and labeled indole-3-acetyl-myo-inositol become asymmetrically distributed in the mesocotyl cortex of the shoot with more radioactivity occurring in the bottom half of a horizontally placed seedling. Asymmetric distribution of [3H]indole-3-acetic acid, derived from the applied [3H]indole-3-acetyl-myo-inositol, occurred more rapidly than distribution of total 3H-radioactivity. These findings demonstrate that the gravitational stimulus can induce an asymmetric distribution of substances being transported from kernel to shoot. They also indicate that, in addition to the transport asymmetry, gravity affects the steady state amount of indole-3-acetic acid derived from indole-3-acetyl-myo-inositol. PMID:11537873

  12. Metabolism and Ovarian Function in PCOS Women: A Therapeutic Approach with Inositols.

    PubMed

    Laganà, Antonio Simone; Rossetti, Paola; Buscema, Massimo; La Vignera, Sandro; Condorelli, Rosita Angela; Gullo, Giuseppe; Granese, Roberta; Triolo, Onofrio

    2016-01-01

    Polycystic ovary syndrome (PCOS) is characterized by chronical anovulation and hyperandrogenism which may be present in a different degree of severity. Insulin-resistance and hyperinsulinemia are the main physiopathological basis of this syndrome and the failure of inositol-mediated signaling may concur to them. Myo (MI) and D-chiro-inositol (DCI), the most studied inositol isoforms, are classified as insulin sensitizers. In form of glycans, DCI-phosphoglycan and MI-phosphoglycan control key enzymes were involved in glucose and lipid metabolism. In form of phosphoinositides, they play an important role as second messengers in several cellular biological functions. Considering the key role played by insulin-resistance and androgen excess in PCOS patients, the insulin-sensitizing effects of both MI and DCI were tested in order to ameliorate symptoms and signs of this syndrome, including the possibility to restore patients' fertility. Accumulating evidence suggests that both isoforms of inositol are effective in improving ovarian function and metabolism in patients with PCOS, although MI showed the most marked effect on the metabolic profile, whereas DCI reduced hyperandrogenism better. The purpose of this review is to provide an update on inositol signaling and correlate data on biological functions of these multifaceted molecules, in view of a rational use for the therapy in women with PCOS. PMID:27579037

  13. Metabolism and Ovarian Function in PCOS Women: A Therapeutic Approach with Inositols

    PubMed Central

    Rossetti, Paola; Buscema, Massimo; Condorelli, Rosita Angela; Gullo, Giuseppe; Triolo, Onofrio

    2016-01-01

    Polycystic ovary syndrome (PCOS) is characterized by chronical anovulation and hyperandrogenism which may be present in a different degree of severity. Insulin-resistance and hyperinsulinemia are the main physiopathological basis of this syndrome and the failure of inositol-mediated signaling may concur to them. Myo (MI) and D-chiro-inositol (DCI), the most studied inositol isoforms, are classified as insulin sensitizers. In form of glycans, DCI-phosphoglycan and MI-phosphoglycan control key enzymes were involved in glucose and lipid metabolism. In form of phosphoinositides, they play an important role as second messengers in several cellular biological functions. Considering the key role played by insulin-resistance and androgen excess in PCOS patients, the insulin-sensitizing effects of both MI and DCI were tested in order to ameliorate symptoms and signs of this syndrome, including the possibility to restore patients' fertility. Accumulating evidence suggests that both isoforms of inositol are effective in improving ovarian function and metabolism in patients with PCOS, although MI showed the most marked effect on the metabolic profile, whereas DCI reduced hyperandrogenism better. The purpose of this review is to provide an update on inositol signaling and correlate data on biological functions of these multifaceted molecules, in view of a rational use for the therapy in women with PCOS. PMID:27579037

  14. Vasopressin induced production of inositol trisphosphate and calcium efflux in a smooth muscle cell line

    SciTech Connect

    Doyle, V.M.; Rueegg, U.T.

    1985-08-30

    Phosphatidylinositol metabolism and /sup 45/Ca/sup 2 +/ efflux were examined in a vascular smooth muscle cell line (A7r5). (Arg 8)Vasopressin stimulated the rapid formation (measurable at 1 sec) of inositol phosphates in a concentration-dependent manner. The time course for formation of inositol phosphates was similar to that for /sup 45/Ca/sup 2 +/ efflux from preloaded cells. The efflux of /sup 45/Ca/sup 2 +/ in response to (Arg8)vasopressin could be inhibited by a vasopressin antagonist. This supports the hypothesis that inositol 1,4,5-trisphosphate plays a role in vasopressin stimulated calcium mobilization from an intracellular source in cultured vascular smooth muscle cells.

  15. Inositol stimulates DNA and protein synthesis, and expansion by rabbit blastocysts in vitro.

    PubMed

    Fahy, M M; Kane, M T

    1992-04-01

    The effect of different concentrations (0, 0.6, 3, 15, 75 and 375 microM) of myo-inositol on the development of rabbit morulae to expanded blastocysts was investigated in terms of blastocyst expansion and synthesis of DNA and protein, as measured by incorporation of [3H]thymidine and [14C]amino acids into acid-precipitable material. A concentration of 15 microM inositol caused a 2.8-fold increase in blastocyst expansion (P less than 0.01), a 9.9-fold increase in thymidine incorporation into DNA (P less than 0.01) and a 3.6-fold increase in amino acid incorporation into protein (P less than 0.01). There were no significant differences in the range from 15 to 375 microM inositol. PMID:1522201

  16. Inositol Pentakisphosphate Isomers Bind PH Domains with Varying Specificity and Inhibit Phosphoinositide Interactions

    SciTech Connect

    S Jackson; S Al-Saigh; C Schultz; M Junop

    2011-12-31

    PH domains represent one of the most common domains in the human proteome. These domains are recognized as important mediators of protein-phosphoinositide and protein-protein interactions. Phosphoinositides are lipid components of the membrane that function as signaling molecules by targeting proteins to their sites of action. Phosphoinositide based signaling pathways govern a diverse range of important cellular processes including membrane remodeling, differentiation, proliferation and survival. Myo-Inositol phosphates are soluble signaling molecules that are structurally similar to the head groups of phosphoinositides. These molecules have been proposed to function, at least in part, by regulating PH domain-phosphoinositide interactions. Given the structural similarity of inositol phosphates we were interested in examining the specificity of PH domains towards the family of myo-inositol pentakisphosphate isomers. In work reported here we demonstrate that the C-terminal PH domain of pleckstrin possesses the specificity required to discriminate between different myo-inositol pentakisphosphate isomers. The structural basis for this specificity was determined using high-resolution crystal structures. Moreover, we show that while the PH domain of Grp1 does not possess this high degree of specificity, the PH domain of protein kinase B does. These results demonstrate that some PH domains possess enough specificity to discriminate between myo-inositol pentakisphosphate isomers allowing for these molecules to differentially regulate interactions with phosphoinositides. Furthermore, this work contributes to the growing body of evidence supporting myo-inositol phosphates as regulators of important PH domain-phosphoinositide interactions. Finally, in addition to expanding our knowledge of cellular signaling, these results provide a basis for developing tools to probe biological pathway.

  17. Anti-inflammatory Inositol Derivatives from the Whole Plant of Inula cappa.

    PubMed

    Wu, Jiewei; Tang, Chunping; Yao, Sheng; Zhang, Lei; Ke, Changqiang; Feng, Linyin; Lin, Ge; Ye, Yang

    2015-10-23

    Twelve new inositol derivatives, classified into myoinositol (1-6) and l-inositol (10-15) types, along with five known analogues were isolated from the whole plant of Inula cappa. The structures of the new compounds were established by extensive analysis of mass spectrometric and 1D and 2D NMR spectroscopic data. All the tested compounds showed anti-inflammatory activities against the production of NO in RAW264.7 macrophages stimulated by lipopolysaccharide, with IC50 values ranging from 7 to 23 μM. PMID:26444098

  18. Phospholipid biosynthesis in Candida albicans: Regulation by the precursors inositol and choline

    SciTech Connect

    Klig, L.S.; Friedli, L.; Schmid, E. )

    1990-08-01

    Phospholipid metabolism in the pathogenic fungus Candida albicans was examined. The phospholipid biosynthetic pathways of C. albicans were elucidated and were shown to be similar to those of Saccharomyces cerevisiae. However, marked differences were seen between these two fungi in the regulation of the pathways in response to exogenously provided precursors inositol and choline. In S. cerevisiae, the biosynthesis of phosphatidylcholine via methylation of phosphatidylethanolamine appears to be regulated in response to inositol and choline; provision of choline alone does not repress the activity of this pathway. The same pathway in C. albicans responds to the exogenous provision of choline. Possible explanations for the observed differences in regulation are discussed.

  19. Low myo-inositol indicating astrocytic damage in a case series of neuromyelitis optica.

    PubMed

    Ciccarelli, Olga; Thomas, David L; De Vita, Enrico; Wheeler-Kingshott, Claudia A M; Kachramanoglou, Carolina; Kapoor, Raj; Leary, Siobhan; Matthews, Lucy; Palace, Jacqueline; Chard, Declan; Miller, David H; Toosy, Ahmed T; Thompson, Alan J

    2013-08-01

    Astrocytic necrosis is a prominent pathological feature of neuromyelitis optica (NMO) lesions and is clinically relevant. We report 5 NMO-related cases, all with longitudinally extensive lesions in the upper cervical cord, who underwent cervical cord (1) H-magnetic resonance spectroscopy. Lower myo-inositol/creatine values, suggesting astrocytic damage, were consistently found within the NMO lesions when compared with healthy controls and patients with multiple sclerosis (MS), who showed at least 1 demyelinating lesion at the same cord level. Therefore, the in vivo quantification of myo-inositol may distinguish NMO from MS. This is an important step toward developing imaging markers for clinical trials in NMO. PMID:23553900

  20. Surface complexation modeling of inositol hexaphosphate sorption onto gibbsite.

    PubMed

    Ruyter-Hooley, Maika; Larsson, Anna-Carin; Johnson, Bruce B; Antzutkin, Oleg N; Angove, Michael J

    2015-02-15

    The sorption of Inositol hexaphosphate (IP6) onto gibbsite was investigated using a combination of adsorption experiments, (31)P solid-state MAS NMR spectroscopy, and surface complexation modeling. Adsorption experiments conducted at four temperatures showed that IP6 sorption decreased with increasing pH. At pH 6, IP6 sorption increased with increasing temperature, while at pH 10 sorption decreased as the temperature was raised. (31)P MAS NMR measurements at pH 3, 6, 9 and 11 produced spectra with broad resonance lines that could be de-convoluted with up to five resonances (+5, 0, -6, -13 and -21ppm). The chemical shifts suggest the sorption process involves a combination of both outer- and inner-sphere complexation and surface precipitation. Relative intensities of the observed resonances indicate that outer-sphere complexation is important in the sorption process at higher pH, while inner-sphere complexation and surface precipitation are dominant at lower pH. Using the adsorption and (31)P MAS NMR data, IP6 sorption to gibbsite was modeled with an extended constant capacitance model (ECCM). The adsorption reactions that best described the sorption of IP6 to gibbsite included two inner-sphere surface complexes and one outer-sphere complex: ≡AlOH + IP₆¹²⁻ + 5H⁺ ↔ ≡Al(IP₆H₄)⁷⁻ + H₂O, ≡3AlOH + IP₆¹²⁻ + 6H⁺ ↔ ≡Al₃(IP₆H₃)⁶⁻ + 3H₂O, ≡2AlOH + IP₆¹²⁻ + 4H⁺ ↔ (≡AlOH₂)₂²⁺(IP₆H₂)¹⁰⁻. The inner-sphere complex involving three surface sites may be considered to be equivalent to a surface precipitate. Thermodynamic parameters were obtained from equilibrium constants derived from surface complexation modeling. Enthalpies for the formation of inner-sphere surface complexes were endothermic, while the enthalpy for the outer-sphere complex was exothermic. The entropies for the proposed sorption reactions were large and positive suggesting that changes in solvation of species play a major role in driving

  1. Novel compound heterozygous mutations in inositol polyphosphate phosphatase-like 1 in a family with severe opsismodysplasia.

    PubMed

    Feist, Cori; Holden, Paul; Fitzgerald, Jamie

    2016-10-01

    This study aimed to identify the genetic basis of a severe skeletal lethal dysplasia. The main clinical features of two affected fetuses included short limbs with flared metaphyses, bowed radii, femora and tibiae, irregular ossification of hands and feet, and marked platyspondyly. Affected and nonaffected family members were subjected to whole-exome sequencing, followed by immunoblot analysis on amniocytes isolated from one of the affected individuals. Unique compound heterozygous variants in the inositol polyphosphate phosphatase-like 1 (INPPL1) gene encoding the SHIP2 protein were identified in both affected individuals. One variant was inherited from each unaffected parent. Both allelic variants, c.(2327-1G>C);(1150_1151delGA), are predicted to result in premature stop codons leading to nonsense-mediated mRNA decay of the mutant alleles and no production of SHIP2. The absence of SHIP2 was confirmed by immunoblot analysis of proband amniocytes. This skeletal disorder is caused by the complete absence of the SHIP2 protein. INPPL1 mutations have been reported in opsismodysplasia, an autosomal recessive skeletal dysplasias with significant delayed bone formation. Our finding highlights the critical role that INPPL1/SHIP2 plays in skeletal development. PMID:27233067

  2. Lactic Acid and Thermal Treatments Trigger the Hydrolysis of Myo-Inositol Hexakisphosphate and Modify the Abundance of Lower Myo-Inositol Phosphates in Barley (Hordeum vulgare L.)

    PubMed Central

    Metzler-Zebeli, Barbara U.; Deckardt, Kathrin; Schollenberger, Margit; Rodehutscord, Markus; Zebeli, Qendrim

    2014-01-01

    Barley is an important source of dietary minerals, but it also contains myo-inositol hexakisphosphate (InsP6) that lowers their absorption. This study evaluated the effects of increasing concentrations (0.5, 1, and 5%, vol/vol) of lactic acid (LA), without or with an additional thermal treatment at 55°C (LA-H), on InsP6 hydrolysis, formation of lower phosphorylated myo-inositol phosphates, and changes in chemical composition of barley grain. Increasing LA concentrations and thermal treatment linearly reduced (P<0.001) InsP6-phosphate (InsP6-P) by 0.5 to 1 g compared to the native barley. In particular, treating barley with 5% LA-H was the most efficient treatment to reduce the concentrations of InsP6-P, and stimulate the formation of lower phosphorylated myo-inositol phosphates such as myo-inositol tetraphosphate (InsP4) and myo-inositol pentaphosphates (InsP5). Also, LA and thermal treatment changed the abundance of InsP4 and InsP5 isomers with Ins(1,2,5,6)P4 and Ins(1,2,3,4,5)P5 as the dominating isomers with 5% LA, 1% LA-H and 5% LA-H treatment of barley, resembling to profiles found when microbial 6-phytase is applied. Treating barley with LA at room temperature (22°C) increased the concentration of resistant starch and dietary fiber but lowered those of total starch and crude ash. Interestingly, total phosphorus (P) was only reduced (P<0.05) in barley treated with LA-H but not after processing of barley with LA at room temperature. In conclusion, LA and LA-H treatment may be effective processing techniques to reduce InsP6 in cereals used in animal feeding with the highest degradation of InsP6 at 5% LA-H. Further in vivo studies are warranted to determine the actual intestinal P availability and to assess the impact of changes in nutrient composition of LA treated barley on animal performance. PMID:24967651

  3. Inositol 1,2,3-trisphosphate and inositol 1,2- and/or 2,3-bisphosphate are normal constituents of mammalian cells.

    PubMed Central

    Barker, C J; French, P J; Moore, A J; Nilsson, T; Berggren, P O; Bunce, C M; Kirk, C J; Michell, R H

    1995-01-01

    1. An inositol trisphosphate (InsP3) distinct from Ins(1,4,5)P3 and Ins(1,3,4)P3, which we previously observed in myeloid and lymphoid cells [French, Bunce, Stephens, Lord, McConnell, Brown, Creba and Michell (1991) Proc R. Soc. London B 245, 193-201; Bunce, French, Allen, Mountford, Moore, Greaves, Michell and Brown (1993) Biochem. J. 289, 667-673], is present in WRK1 rat mammary tumour cells and pancreatic endocrine beta-cells. 2. It has been identified as Ins(1,2,3)P3 by a combination of oxidation to ribitol, a structurally diagnostic polyol, and ammoniacal hydrolysis to identified inositol monophosphates. 3. Ins(1,2,3)P3 concentration in HL60 cells changed little during stimulation by ATP or fMetLeuPhe or during neutrophilic or monocytic differentiation, and Ins(1,2,3)P3 was unresponsive to vasopressin in WRK1 cells. 4. Ins(1,2,3)P3 was usually more abundant than Ins(1,4,5)P3, often being present at concentrations between approximately 1 microM and approximately 10 microM. 5. HL60, WRK-1 and lymphoid cells also contain Ins(1,2)P2 or Ins(2,3)P2, or a mixture of these two enantiomers, as a major InsP2 species. 6. Ins(1,2,3)P3 and Ins(1,2)P2/Ins(2,3)P2 are readily detected in cells labelled for long periods, but not in acutely labelled cells. This behaviour resembles that of InsP6, the most abundant cellular inositol polyphosphate that includes the 1,2,3-trisphosphate motif, which also achieves isotopic equilibrium with inositol only slowly. 7. Ins(1,2,3)P3 is the major InsP3 that accumulates during metabolism of InsP6 by WRK-1 cell homogenates. 8. Possible metabolic relationships between Ins(1,2,3)P3, Ins(1,2)P2/Ins(2,3)P2 and other inositol polyphosphates in cells, and a possible role for Ins(1,2,3)P3 in cellular iron handling, are considered. PMID:7887911

  4. Lipoarabinomannans: characterization of the multiacylated forms of the phosphatidyl-myo-inositol anchor by NMR spectroscopy.

    PubMed Central

    Nigou, J; Gilleron, M; Puzo, G

    1999-01-01

    Lipoarabinomannans, which exhibit a large spectrum of immunological activities, emerge as the major antigens of mycobacterial envelopes. The lipoarabinomannan structure is based on a phosphatidyl-myo-inositol anchor whose integrity has been shown to be crucial for lipoarabinomannan biological activity and particularly for presentation to CD4/CD8 double-negative alphabetaT cells by CD1 molecules. In this report, an analytical approach was developed for high-resolution 31P-NMR analysis of native, i.e. multiacylated, lipoarabinomannans. The one-dimensional 31P spectrum of cellular lipoarabinomannans, from Mycobacterium bovis Bacillus Calmette-Guérin, exhibited four 31P resonances typifying four types of lipoarabinomannans. Two-dimensional 1H-31P heteronuclear multiple-quantum-correlation/homonuclear Hartmann-Hahn analysis of the native molecules showed that these four types of lipoarabinomannan differed in the number and localization of fatty acids (from 1 to 4) esterifying the anchor. Besides the three acylation sites previously described, i.e. positions 1 and 2 of glycerol and 6 of the mannosyl unit linked to the C-2 of myo-inositol, we demonstrate the existence of a fourth acylation position at the C-3 of myo-inositol. We report here the first structural study of native multiacylated lipoarabinomannans, establishing the structure of the intact phosphatidyl-myo-inositol anchor. Our findings would help gain more understanding of the molecular basis of lipoarabinomannan discrimination in the binding process to CD1 molecules. PMID:9895288

  5. Blast neurotrauma impairs working memory and disrupts prefrontal myo-inositol levels in rats.

    PubMed

    Sajja, Venkata Siva Sai Sujith; Perrine, Shane A; Ghoddoussi, Farhad; Hall, Christina S; Galloway, Matthew P; VandeVord, Pamela J

    2014-03-01

    Working memory, which is dependent on higher-order executive function in the prefrontal cortex, is often disrupted in patients exposed to blast overpressure. In this study, we evaluated working memory and medial prefrontal neurochemical status in a rat model of blast neurotrauma. Adult male Sprague-Dawley rats were anesthetized with 3% isoflurane and exposed to calibrated blast overpressure (17 psi, 117 kPa) while sham animals received only anesthesia. Early neurochemical effects in the prefrontal cortex included a significant decrease in betaine (trimethylglycine) and an increase in GABA at 24 h, and significant increases in glycerophosphorylcholine, phosphorylethanolamine, as well as glutamate/creatine and lactate/creatine ratios at 48 h. Seven days after blast, only myo-inositol levels were altered showing a 15% increase. Compared to controls, short-term memory in the novel object recognition task was significantly impaired in animals exposed to blast overpressure. Working memory in control animals was negatively correlated with myo-inositol levels (r=-.759, p<0.05), an association that was absent in blast exposed animals. Increased myo-inositol may represent tardive glial scarring in the prefrontal cortex, a notion supported by GFAP changes in this region after blast overexposure as well as clinical reports of increased myo-inositol in disorders of memory. PMID:24534010

  6. MECHANISMS UNDERLYING ALC13 INHIBITION OF AGONIST-STIMULATED INOSITOL PHOSPHATE ACCUMULATION

    EPA Science Inventory

    Possible mechanisms of AlC13-induced inhibition of agonist-stimulated inositol phosphate (IP) accumulation were investigated using rat brain cortex slices, synaptosomes or homogenates. nder conditions in which AlC13 inhibits carbachol (CARB) stimulated IP accumulation (Gp-mediate...

  7. Extraction and analysis of inositols and other carbohydrates from soybean plant tissues

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An outstanding characteristic of soybean plants is their ability to produce large amounts of the carbohydrate pinitol. Pinitol and the closely related inositols are currently undergoing widespread investigation for their biological and nutritional value. These and all the carbohydrates are typical...

  8. Production of glucaric acid from myo-inositol in engineered Pichia pastoris.

    PubMed

    Liu, Ye; Gong, Xu; Wang, Cui; Du, Guocheng; Chen, Jian; Kang, Zhen

    2016-09-01

    A potential myo-inositol oxygenase (ppMIOX) was identified as a functional enzyme and a glucaric acid synthetic pathway was firstly constructed in Pichia pastoris. Coexpression of the native ppMIOX and the urinate dehydrogenase (Udh) from Pseudomonas putida KT2440 led to obvious accumulation of glucaric acid (90.46±0.04mg/L) from myo-inositol whereas no glucaric acid was detected from glucose. In comparison, coexpression of the heterologous mouse MIOX (mMIOX) and Udh resulted in higher titers of glucaric acid from glucose and myo-inositol, 107.19±11.91mg/L and 785.4±1.41mg/L, respectively. By applying a fusion expression strategy with flexible peptides, the mMIOX specific activity and the glucaric acid concentration were significantly increased. Using glucose and myo-inositol as carbon substrates, the production of glucaric acid was substantially enhanced to 6.61±0.30g/L in fed-batch cultures. To the best of our knowledge, this is the highest reported value to date. PMID:27444324

  9. Muscarinic receptor-mediated inositol tetrakisphosphate response in bovine adrenal chromaffin cells

    SciTech Connect

    Sanborn, B.B.; Schneider, A.S. )

    1990-01-01

    Inositol trisphosphate (IP{sub 3}), a product of the phosphoinositide cycle, mobilizes intracellular Ca{sup 2+} in many cell types. New evidence suggests that inositol tetrakisphosphate (IP{sub 4}), an IP{sub 3} derivative, may act as another second messenger to further alter calcium homeostasis. However, the function and mechanism of action of IP{sub 4} are presently unresolved. We now report evidence of muscarinic receptor-mediated accumulation of IP{sub 4} in bovine adrenal chromaffin cells, a classic neurosecretory system in which calcium movements have been well studied. Muscarine stimulated an increase in ({sup 3}H)IP{sub 4} and ({sup 3}H)IP{sub 3} accumulation in chromaffin cells and this effect was completely blocked by atropine. ({sup 3}H)IP{sub 4} accumulation was detectable within 15 sec, increased to a maximum by 30 sec and thereafter declined. 2,3-diphosphoglycerate, an inhibitor of IP{sub 3} and IP{sub 4} hydrolysis, enhanced accumulation of these inositol polyphosphates. The results provide the first evidence of a rapid inositol tetrakisphosphate response in adrenal chromaffin cells, which should facilitate the future resolution of the relationship between IP{sub 4} and calcium homeostasis.

  10. Genetic Control of Lithium Sensitivity and Regulation of Inositol Biosynthetic Genes

    PubMed Central

    King, Jason; Keim, Melanie; Teo, Regina; Weening, Karin E.; Kapur, Mridu; McQuillan, Karina; Ryves, Jonathan; Rogers, Ben; Dalton, Emma; Williams, Robin S. B.; Harwood, Adrian J.

    2010-01-01

    Lithium (Li+) is a common treatment for bipolar mood disorder, a major psychiatric illness with a lifetime prevalence of more than 1%. Risk of bipolar disorder is heavily influenced by genetic predisposition, but is a complex genetic trait and, to date, genetic studies have provided little insight into its molecular origins. An alternative approach is to investigate the genetics of Li+ sensitivity. Using the social amoeba Dictyostelium, we previously identified prolyl oligopeptidase (PO) as a modulator of Li+ sensitivity. In a link to the clinic, PO enzyme activity is altered in bipolar disorder patients. Further studies demonstrated that PO is a negative regulator of inositol(1,4,5)trisphosphate (IP3) synthesis, a Li+ sensitive intracellular signal. However, it was unclear how PO could influence either Li+ sensitivity or risk of bipolar disorder. Here we show that in both Dictyostelium and cultured human cells PO acts via Multiple Inositol Polyphosphate Phosphatase (Mipp1) to control gene expression. This reveals a novel, gene regulatory network that modulates inositol metabolism and Li+ sensitivity. Among its targets is the inositol monophosphatase gene IMPA2, which has also been associated with risk of bipolar disorder in some family studies, and our observations offer a cellular signalling pathway in which PO activity and IMPA2 gene expression converge. PMID:20567601

  11. Barley (Hordeum vulgare L.) inositol monophosphatase: gene structure and enzyme characteristics

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The de novo synthesis of myo-inositol (Ins) is catalyzed by two enzymatic activities; Ins(3)P1 synthase (MIPS; EC. 5.5.1.4) and Ins monophosphatase (IMPase; EC 3.1.3.25). The barley IMP-1 gene and gene products were studied to facilitate research into the regulation of Ins synthesis and supply. In m...

  12. GATA4-mediated cardiac hypertrophy induced by D-myo-inositol 1,4,5-tris-phosphate

    SciTech Connect

    Zhu Zhiming . E-mail: zhuzming@mail.dph-fsi.com; Zhu Shanjun; Liu Daoyan; Yu Zengping; Yang Yongjian; Giet, Markus van der; Tepel, Martin . E-mail: Martin.Tepel@charite.de

    2005-12-16

    We evaluated the effects of D-myo-inositol 1,4,5-tris-phosphate on cardiac hypertrophy. D-myo-inositol 1,4,5-tris-phosphate augmented cardiac hypertrophy as evidenced by its effects on DNA synthesis, protein synthesis, and expression of immediate-early genes c-myc and c-fos, {beta}-myosin heavy chain, and {alpha}-actin. The administration of D-myo-inositol 1,4,5-tris-phosphate increased the expression of nuclear factor of activated T-cells and cardiac-restricted zinc finger transcription factor (GATA4). Real-time quantitative RT-PCR showed that D-myo-inositol 1,4,5-tris-phosphate-induced GATA4 mRNA was significantly enhanced even in the presence of the calcineurin inhibitor, cyclosporine A. The effect of D-myo-inositol 1,4,5-tris-phosphate was blocked after inhibition of inositol-trisphosphate receptors but not after inhibition of c-Raf/mitogen-activated protein kinase kinase (MEK)/mitogen-activated protein kinase (ERK) or p38 mitogen-activated protein kinase pathways. The study shows that D-myo-inositol 1,4,5-tris-phosphate-induced cardiac hypertrophy is mediated by GATA4 but independent from the calcineurin pathway.

  13. Inositol deficiency increases the susceptibility to neural tube defects of genetically predisposed (curly tail) mouse embryos in vitro.

    PubMed

    Cockroft, D L; Brook, F A; Copp, A J

    1992-02-01

    Curly tail (ct/ct) mouse embryos, which have a genetic predisposition for neural tube defects (NTD), were grown in culture from the 2-5 somite stage, before the initiation of neurulation, up to the 22-24 somite stage, when closure of the anterior neural tube is normally complete. The embryos were cultured in whole rat serum or in extensively dialysed serum supplemented with glucose, amino acids, and vitamins, with inositol omitted or added at concentrations of 2, 10, 20, and 50 mg/l. Two strains were used as controls; CBA mice, which are related to curly tails, and an unrelated PO stock. It was found that ct/ct embryos were particularly sensitive to inositol deficiency; both they and the CBA embryos showed a similar high incidence of cranial NTD after culture in inositol deficient medium (12/17 and 11/18, respectively). Furthermore, the lowest dose of inositol had no effect on the frequency of head defects in ct/ct mice, though it halved the incidence in CBA embryos. With higher inositol concentrations, the majority of ct/ct embryos completed head closure normally, and their development was generally similar to that obtained in whole serum. PO embryos showed a lower proportion (5/19) of cranial NTD in the inositol deficient medium than the other two strains, and this was further reduced by even the lowest inositol dose. PMID:1615432

  14. Formation and metabolism of inositol 1,4,5-trisphosphate in human platelets.

    PubMed Central

    Daniel, J L; Dangelmaier, C A; Smith, J B

    1987-01-01

    1. myo-[3H]Inositol 1,4,5-trisphosphate [Ins(1,4,5)P3], when added to lysed platelets, was rapidly converted into [3H]inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P4], which was in turn converted into [3H]inositol 1,3,4-trisphosphate [Ins(1,3,4)P3]. This result demonstrates that platelets have the same metabolic pathways for interconversion of inositol polyphosphates that are found in other cells. 2. Labelling of platelets with [32P]Pi, followed by h.p.l.c., was used to measure thrombin-induced changes in the three inositol polyphosphates. Interfering compounds were removed by a combination of enzymic and non-enzymic techniques. 3. Ins(1,4,5)P3 was formed rapidly, and reached a maximum at about 4 s. It was also rapidly degraded, and was no longer detectable after 30-60 s. 4. Formation of Ins(1,3,4,5)P4 was almost as rapid as that of Ins(1,4,5)P3, and it remained detectable for a longer time. 5. Ins(1,3,4)P3 was formed after an initial lag, and this isomer reached its maximum, which was 10-fold higher than that of Ins(1,4,5)P3, at 30 s. 6. Comparison of the intracellular Ca2+ concentration as measured with fura-2 indicates that agents other than Ins(1,4,5)P3 are responsible for the sustained maintenance of a high concentration of intracellular Ca2+. It is proposed that either Ins(1,3,4)P3 or Ins(1,3,4,5)P4 may also be Ca2+-mobilizing agents. PMID:3499898

  15. PTH (parathyroid hormone) elevates inositol polyphosphates and diacylglycerol in a rat osteoblast-like cell line

    SciTech Connect

    Civitelli, R.; Reid, I.R.; Westbrook, S.; Avioli, L.V.; Hruska, K.A. )

    1988-11-01

    Parathyroid hormone (PTH)-stimulated signal transduction through mechanisms alternate to adenosine 3{prime},5{prime}-cyclic monophosphate (cAMP) production were studied in UMR 106-01 cells, a cell line with an osteoblastic phenotype. PTH produced transient, dose-related increases in cytosolic calcium ((Ca{sup 2+}){sub i}), inositol trisphosphates, and diacylglycerol (DAG). Both inositol 1,4,5-trisphosphate (Ins-1,4,5P{sub 3}) and inositol 1,3,4-trisphosphate (Ins-1,3,4P{sub 3}) production were rapidly stimulated by PTH. Consistent with the production of Ins-1,3,4P{sub 3}, rapid stimulation of late eluting inositol tetrakisphosphate was observed. The effects on the inositol phosphates were induced rapidly, consistent with roles as signals for changes in (Ca{sup 2+}){sub i}. In saponin-permeabilized UMR 106-01 cells, Ins-1,4,5P{sub 3} stimulated {sup 45}Ca release from a nonmitochondrial intracellular pool. Thus the hypothesis that PTH-stimulated Ins-1,4,5P{sub 3} production initiates Ca{sup 2+} release and contributes to transient elevations of (Ca{sup 2+}){sub i} is supported. These data suggest that stimulation of cAMP production during PTH stimulation may negatively affect production of rises in (Ca{sup 2+}){sub i} during PTH stimulation. The inactivation of the inhibitory G protein of adenylate cyclase by pertussis toxin could explain its action similar to cAMP analogues. Cyclci nucleotides diminish the effects of PTH on (Ca{sup 2+}){sub i}, probably interacting on a biochemical step subsequent to or independent of Ins-1,4,5P{sub 3} release.

  16. Sodium- and energy-dependent uptake of myo-inositol by rabbit peripheral nerve. Competitive inhibition by glucose and lack of an insulin effect.

    PubMed Central

    Greene, D A; Lattimer, S A

    1982-01-01

    Experimental diabetes consistently reduces the concentration of free myo-inositol in peripheral nerve, which usually exceeds that of plasma by 90-100-fold. This phenomenon has been explicitly linked to the impairment of nerve conduction in the acutely diabetic streptozocin-treated rat. However, the mechanism by which acute experimental diabetes lowers nerve myo-inositol content and presumably alters nerve myo-inositol content and presumably alters nerve myo-inositol metabolism is unknown. Therefore, the effects of insulin and elevated medium glucose concentration of 2-[3H]myo-inositol uptake were studied in a metabolically-defined in vitro peripheral nerve tissue preparation derived from rabbit sciatic nerve, whose free myo-inositol content is reduced by experimental diabetes. The results demonstrate that myo-inositol uptake occurs by at least two distinct transport systems in the normal endoneurial preparation. A sodium- and energy-dependent saturable transport system is responsible for at least 94% of the measured uptake at medium myo-inositol concentrations approximating that present in plasma. This carrier-mediated transport system has a high affinity for myo-inositol (Kt = 63 microM), and is not influenced acutely by physiological concentrations of insulin; it is, however, inhibited by hyperglycemic concentrations of glucose added to the incubation medium in a primarily competitive fashion. Thus, competitive inhibition of peripheral nerve myo-inositol uptake by glucose may constitute a mechanism by which diabetes produces physiologically significant alterations in peripheral nerve myo-inositol metabolism. PMID:6813354

  17. Mannose, glucosamine and inositol monophosphate inhibit the effects of insulin on lipogenesis. Further evidence for a role for inositol phosphate-oligosaccharides in insulin action.

    PubMed Central

    Machicao, F; Mushack, J; Seffer, E; Ermel, B; Häring, H U

    1990-01-01

    The mechanism of insulin signalling is not yet understood in detail. Recently, a role for inositol phosphate (IP)-oligosaccharides as second messengers transmitting the insulin signal at the post-kinase level was proposed. To evaluate this hypothesis further, we studied whether IP-oligosaccharides isolated from 'haemodialysate' have insulin-like activity. We found that these compounds mimic, in a dose-dependent fashion, the following effects of insulin in adipocytes. (1) Lipogenesis. Incorporation of [3H]glucose into lipids (expressed in nmol/min per 10(6) cells): basal, 0.74 +/- 0.05; insulin (1 mu unit/ml), 4.43 +/- 0.21; IP-oligosaccharide (2 micrograms/ml), 4.07 +/- 0.19. (2) Inhibition of isoprenaline (isoproterenol) (1 microM)-stimulated cyclic AMP levels and lipolysis. Cyclic AMP (pmol/10(5) cells): basal 0.84 +/- 0.05; isoprenaline, 4.03 +/- 0.19; isoprenaline + insulin (200 mu units/ml), 2.06 +/- 0.7; isoprenaline + IP-oligosaccharides (2 micrograms/ml), 2.4 +/- 0.29. Inhibition of lipolysis (mumol of glycerol/mg of protein): isoprenaline (1 microM), 166 +/- 11; isoprenaline+insulin (150 mu units/ml), 53 +/- 3.5; isoprenaline+IP-oligosaccharides (2 micrograms/ml), 58 +/- 5. (3) Stimulation of 3-O-methylglucose transport; basal, 9 +/- 3%; insulin (1 mu unit/ml), 67 +/- 4%, IP-oligosaccharides (2 micrograms/ml), 54 +/- 2%. To identify the active molecules of the IP-oligosaccharide fraction, competition experiments were performed. IP-oligosaccharide effects on lipogenesis were blocked by inositol monophosphate, glucosamine and mannose. In contrast, these compounds did not inhibit IP-oligosaccharide effects on membrane-mediated functions (3-O-methylglucose transport, cyclic AMP levels, lipolysis). We also found that the effect of insulin on lipogenesis was blocked by mannose, glucosamine and inositol monophosphate, whereas the insulin effects on 3-O-methylglucose, cyclic AMP and lipolysis were unaffected. The following conclusions were reached. (1) IP

  18. RETROGRADE AXONAL TRANSPORT OF PHOSPHOINOSITIDES AFTER INTRANEURAL INJECTION OF [3H]MYO-INOSITOL INTO THE RAT SCIATIC NERVE

    EPA Science Inventory

    Although autoradiography has demonstrated local incorporation of [3H]inositol into axonal phospholipids after intraneural injection (Gould, 1976; Gould et at., 1987b), retrograde axonal transport of phosphatidylinositol has only been demonstrated after injection of lipid precurso...

  19. Osmoregulatory inositol transporter SMIT1 modulates electrical activity by adjusting PI(4,5)P2 levels.

    PubMed

    Dai, Gucan; Yu, Haijie; Kruse, Martin; Traynor-Kaplan, Alexis; Hille, Bertil

    2016-06-01

    Myo-inositol is an important cellular osmolyte in autoregulation of cell volume and fluid balance, particularly for mammalian brain and kidney cells. We find it also regulates excitability. Myo-inositol is the precursor of phosphoinositides, key signaling lipids including phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2]. However, whether myo-inositol accumulation during osmoregulation affects signaling and excitability has not been fully explored. We found that overexpression of the Na(+)/myo-inositol cotransporter (SMIT1) and myo-inositol supplementation enlarged intracellular PI(4,5)P2 pools, modulated several PI(4,5)P2-dependent ion channels including KCNQ2/3 channels, and attenuated the action potential firing of superior cervical ganglion neurons. Further experiments using the rapamycin-recruitable phosphatase Sac1 to hydrolyze PI(4)P and the P4M probe to visualize PI(4)P suggested that PI(4)P levels increased after myo-inositol supplementation with SMIT1 expression. Elevated relative levels of PIP and PIP2 were directly confirmed using mass spectrometry. Inositol trisphosphate production and release of calcium from intracellular stores also were augmented after myo-inositol supplementation. Finally, we found that treatment with a hypertonic solution mimicked the effect we observed with SMIT1 overexpression, whereas silencing tonicity-responsive enhancer binding protein prevented these effects. These results show that ion channel function and cellular excitability are under regulation by several "physiological" manipulations that alter the PI(4,5)P2 setpoint. We demonstrate a previously unrecognized linkage between extracellular osmotic changes and the electrical properties of excitable cells. PMID:27217553

  20. Agonist-induced desensitization of histamine H1 receptor-mediated inositol phospholipid hydrolysis in human umbilical vein endothelial cells.

    PubMed Central

    McCreath, G; Hall, I P; Hill, S J

    1994-01-01

    1. The regulation of histamine-induced [3H]-inositol phosphate formation was studied in human cultured umbilical vein endothelial cells (HUVEC). 2. Histamine (EC50 4.8 microM) produced a 12.7 fold increase in [3H]-inositol phosphate formation over basal levels. Prior exposure to 0.1 mM histamine (2 h) produced a 78% reduction in the response to subsequent histamine (0.1 mM) challenge. The IC50 for this histamine-induced desensitization was 0.9 microM. 3. The inositol phosphate response to histamine (0.1 mM) was inhibited by phorbol dibutyrate (IC50 40 nM; maximal reduction 64%). This effect was antagonized by both staurosporine (100 nM) and Ro 31-8220 (10 microM). However, the histamine-induced desensitization of the H1-receptor-mediated inositol phosphate response was insensitive to the protein kinase inhibitors, staurosporine, Ro 31-8220, K252a and KN62. 4. Prior exposure to sodium nitroprusside (100 microM), forskolin (10 microM) or dibutyryl cyclic AMP (1 mM) had no effect upon histamine-induced [3H]-inositol phosphate formation. 5. NaF (20 mM) and thrombin (EC50 0.4 u ml-1) also induced inositol phosphate formation in HUVEC. Histamine pretreatment (0.1 mM, 10-120 min) failed to modify the inositol phosphate response to a subsequent NaF or thrombin challenge. 6. We conclude that the desensitization of histamine H1-receptor-mediated [3H]-inositol phosphate formation occurs at the level of the receptor and involves a mechanism independent of activation of protein kinase A, G, or C, or calcium calmodulin-dependent protein kinase II. PMID:7858873

  1. Probing the promiscuous active site of myo-inositol dehydrogenase using synthetic substrates, homology modeling, and active site modification.

    PubMed

    Daniellou, Richard; Zheng, Hongyan; Langill, David M; Sanders, David A R; Palmer, David R J

    2007-06-26

    The active site of myo-inositol dehydrogenase (IDH, EC 1.1.1.18) from Bacillus subtilis recognizes a variety of mono- and disaccharides, as well as 1l-4-O-substituted inositol derivatives. It catalyzes the NAD+-dependent oxidation of the axial alcohol of these substrates with comparable kinetic constants. We have found that 4-O-p-toluenesulfonyl-myo-inositol does not act as a substrate for IDH, in contrast to structurally similar compounds such as those bearing substituted benzyl substituents in the same position. X-ray crystallographic analysis of 4-O-p-toluenesulfonyl-myo-inositol and 4-O-(2-naphthyl)methyl-myo-inositol, which is a substrate for IDH, shows a distinct difference in the preferred conformation of the aryl substituent. Conformational analysis of known substrates of IDH suggests that this conformational difference may account for the difference in reactivity of 4-O-p-toluenesulfonyl-myo-inositol in the presence of IDH. A sequence alignment of IDH with the homologous glucose-fructose oxidoreductase allowed the construction of an homology model of inositol dehydrogenase, to which NADH and 4-O-benzyl-scyllo-inosose were docked and the active site energy minimized. The active site model is consistent with all experimental results and suggests that a conserved tyrosine-glycine-tyrosine motif forms the hydrophobic pocket adjoining the site of inositol recognition. Y233F and Y235F retain activity, while Y233R and Y235R do not. A histidine-aspartate pair, H176 and D172, are proposed to act as a dyad in which H176 is the active site acid/base. The enzyme is inactivated by diethyl pyrocarbonate, and the mutants H176A and D172N show a marked loss of activity. Kinetic isotope effect experiments with D172N indicate that chemistry is rate-determining for this mutant. PMID:17539607

  2. A Role of Arabidopsis Inositol Polyphosphate Kinase, AtIPK2α, in Pollen Germination and Root Growth1

    PubMed Central

    Xu, Jun; Brearley, Charles A.; Lin, Wen-Hui; Wang, Yuan; Ye, Rui; Mueller-Roeber, Bernd; Xu, Zhi-Hong; Xue, Hong-Wei

    2005-01-01

    Inositol polyphosphates, such as inositol trisphosphate, are pivotal intracellular signaling molecules in eukaryotic cells. In higher plants the mechanism for the regulation of the type and the level of these signaling molecules is poorly understood. In this study we investigate the physiological function of an Arabidopsis (Arabidopsis thaliana) gene encoding inositol polyphosphate kinase (AtIPK2α), which phosphorylates inositol 1,4,5-trisphosphate successively at the D-6 and D-3 positions, and inositol 1,3,4,5-tetrakisphosphate at D-6, resulting in the generation of inositol 1,3,4,5,6-pentakisphosphate. Semiquantitative reverse transcription-PCR and promoter-β-glucuronidase reporter gene analyses showed that AtIPK2α is expressed in various tissues, including roots and root hairs, stem, leaf, pollen grains, pollen tubes, the flower stigma, and siliques. Transgenic Arabidopsis plants expressing the AtIPK2α antisense gene under its own promoter were generated. Analysis of several independent transformants exhibiting strong reduction in AtIPK2α transcript levels showed that both pollen germination and pollen tube growth were enhanced in the antisense lines compared to wild-type plants, especially in the presence of nonoptimal low Ca2+ concentrations in the culture medium. Furthermore, root growth and root hair development were also stimulated in the antisense lines, in the presence of elevated external Ca2+ concentration or upon the addition of EGTA. In addition, seed germination and early seedling growth was stimulated in the antisense lines. These observations suggest a general and important role of AtIPK2α, and hence inositol polyphosphate metabolism, in the regulation of plant growth most likely through the regulation of calcium signaling, consistent with the well-known function of inositol trisphosphate in the mobilization of intracellular calcium stores. PMID:15618435

  3. The RpiR-Like Repressor IolR Regulates Inositol Catabolism in Sinorhizobium meliloti▿†

    PubMed Central

    Kohler, Petra R. A.; Choong, Ee-Leng; Rossbach, Silvia

    2011-01-01

    Sinorhizobium meliloti, the nitrogen-fixing symbiont of alfalfa, has the ability to catabolize myo-, scyllo-, and d-chiro-inositol. Functional inositol catabolism (iol) genes are required for growth on these inositol isomers, and they play a role during plant-bacterium interactions. The inositol catabolism genes comprise the chromosomally encoded iolA (mmsA) and the iolY(smc01163)RCDEB genes, as well as the idhA gene located on the pSymB plasmid. Reverse transcriptase assays showed that the iolYRCDEB genes are transcribed as one operon. The iol genes were weakly expressed without induction, but their expression was strongly induced by myo-inositol. The putative transcriptional regulator of the iol genes, IolR, belongs to the RpiR-like repressor family. Electrophoretic mobility shift assays demonstrated that IolR recognized a conserved palindromic sequence (5′-GGAA-N6-TTCC-3′) in the upstream regions of the idhA, iolY, iolR, and iolC genes. Complementation assays found IolR to be required for the repression of its own gene and for the downregulation of the idhA-encoded myo-inositol dehydrogenase activity in the presence and absence of inositol. Further expression studies indicated that the late pathway intermediate 2-keto-5-deoxy-d-gluconic acid 6-phosphate (KDGP) functions as the true inducer of the iol genes. The iolA (mmsA) gene encoding methylmalonate semialdehyde dehydrogenase was not regulated by IolR. The S. meliloti iolA (mmsA) gene product seems to be involved in more than only the inositol catabolic pathway, since it was also found to be essential for valine catabolism, supporting its more recent annotation as mmsA. PMID:21784930

  4. Determination of Myo-Inositol in Infant, Pediatric, and Adult Formulas by Liquid Chromatography-Pulsed Amperometric Detection with Column Switching: Collaborative Study, Final Action 2011.18.

    PubMed

    Butler-Thompson, Linda D; Jacobs, Wesley A; Schimpf, Karen J

    2015-01-01

    AOAC First Action Method 2011.18, Myo-Inositol (Free and Bound as Phosphatidylinositol) in Infant and Pediatric Formulas and Adult Nutritionals, was collaboratively studied. With this method free myo-inositol and phosphatidylinositol bound myo-inositol are extracted using two different sample preparation procedures, separated by ion chromatography using a combination of Dionex Carbo Pac PA1 and MA1 columns with column switching, and detected with pulsed amperometry using a gold electrode. Free myo-inositol is extracted from samples with dilute hydrochloric acid and water. Phosphatidylinositol is extracted from samples with chloroform and separated from other fats with silica SPE cartridges. Myo-inositol is then released from the glycerol backbone with concentrated acetic and hydrochloric acids at 120°C. During this collaborative study, nine laboratories from five different countries analyzed blind duplicates of nine infant and pediatric nutritional formulas for both free and phosphatidylinositol bound myo-inositol, and one additional laboratory only completed the free myo-inositol analyses. The method demonstrated acceptable repeatability and reproducibility and met the AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN) Standard Method Performance Requirements (SMPRs®) for free myo-inositol plus phosphatidylinositol bound myo-inositol for all the matrixes analyzed. SMPRs for repeatability were ≤5% RSD at myo-inositol concentrations of 2-68 mg/100 g ready-to-feed (RTF) liquid. SMPRs for reproducibility were ≤8% RSD in products with myo-inositol concentrations ranging from 2 to 68 mg/100 g RTF liquid. During this collaborative study, repeatability RSDs ranged from 0.51 to 3.22%, and RSDs ranged from 2.66 to 7.55% for free myo-inositol plus phosphatidylinositol bound myo-inositol. PMID:26651580

  5. Modulation of HIV-like particle assembly in vitro by inositol phosphates

    PubMed Central

    Campbell, Stephen; Fisher, Robert J.; Towler, Eric M.; Fox, Stephen; Issaq, Haleem J.; Wolfe, Tracy; Phillips, Lawrence R.; Rein, Alan

    2001-01-01

    HIV-1 Gag protein assembles into 100- to 120-nm diameter particles in mammalian cells. Recombinant HIV-1 Gag protein assembles in a fully defined system in vitro into particles that are only 25–30 nm in diameter and that differ significantly in other respects from authentic particles. However, particles with the size and other properties of authentic virions were obtained in vitro by addition of inositol phosphates or phosphatidylinsitol phosphates to the assembly system. Thus, the interactions between HIV-1 Gag protein molecules are altered by binding of inositol derivatives; this binding is apparently essential for normal HIV-1 particle assembly. This requirement is not seen in a deleted Gag protein lacking residues 16–99 within the matrix domain. PMID:11526217

  6. Binding of zinc and calcium to inositol phosphates (phytate) in vitro.

    PubMed

    Simpson, C J; Wise, A

    1990-07-01

    Inositol compounds with three to five phosphate groups (IP3-IP5) were produced by hydrolysis of phytate (inositol hexaphosphate, IP6) and their binding affinities for calcium and zinc investigated at neutral pH with relative concentrations that had been found in a range of students' meals. Zn solubility was negligible at many of these concentrations, with less Zn bound to precipitates of Ca-IP6 than Ca-IP5. The capacity to precipitate Zn at these ratios fell between IP5 and IP3. Zn was partially desorbed by soluble chelators (histidine and picolinate), especially when it had been adsorbed to preformed Ca-IP precipitates. A lower proportion of Zn was accessible to soluble chelators from Ca-IP4 than the other compounds. IP3-IP4 were hydrolysed by phytase more readily than IP5-IP6. PMID:2400762

  7. Purification, crystallization and preliminary X-ray analysis of inositol dehydrogenase (IDH) from Bacillus subtilis

    SciTech Connect

    Van Straaten, K. E.; Hoffort, A.; Palmer, D. R. J.; Sanders, D. A. R.

    2008-02-01

    Selenomethionine-substituted IDH was crystallized using the microbatch method. The crystals diffracted to beyond 2.0 Å resolution using synchrotron radiation. Inositol dehydrogenase (IDH) is an enzyme that catalyses the NAD{sup +}-dependent oxidation of myo-inositol to scyllo-inosose. The enzyme has been purified to homogeneity by means of Ni{sup 2+}-affinity chromatography and was crystallized in both native and selenomethionine (SeMet) labelled forms using the microbatch method. SAD X-ray diffraction data were collected to 2.0 Å resolution from a SeMet-labelled crystal at the Advanced Photon Source (APS) and a MAD data set was collected to 1.75 Å resolution at the Canadian Light Source (CLS); this is the first reported anomalous diffraction experiment from the CLS. The crystals belong to space group I222 and contain one molecule per asymmetric unit.

  8. Huntington’s disease: Neural dysfunction linked to inositol polyphosphate multikinase

    PubMed Central

    Ahmed, Ishrat; Sbodio, Juan I.; Harraz, Maged M.; Tyagi, Richa; Grima, Jonathan C.; Albacarys, Lauren K.; Hubbi, Maimon E.; Xu, Risheng; Kim, Seyun; Paul, Bindu D.; Snyder, Solomon H.

    2015-01-01

    Huntington’s disease (HD) is a progressive neurodegenerative disease caused by a glutamine repeat expansion in mutant huntingtin (mHtt). Despite the known genetic cause of HD, the pathophysiology of this disease remains to be elucidated. Inositol polyphosphate multikinase (IPMK) is an enzyme that displays soluble inositol phosphate kinase activity, lipid kinase activity, and various noncatalytic interactions. We report a severe loss of IPMK in the striatum of HD patients and in several cellular and animal models of the disease. This depletion reflects mHtt-induced impairment of COUP-TF-interacting protein 2 (Ctip2), a striatal-enriched transcription factor for IPMK, as well as alterations in IPMK protein stability. IPMK overexpression reverses the metabolic activity deficit in a cell model of HD. IPMK depletion appears to mediate neural dysfunction, because intrastriatal delivery of IPMK abates the progression of motor abnormalities and rescues striatal pathology in transgenic murine models of HD. PMID:26195796

  9. Efficacy of IP6 + inositol in the treatment of breast cancer patients receiving chemotherapy: prospective, randomized, pilot clinical study

    PubMed Central

    2010-01-01

    Background Prospective, randomized, pilot clinical study was conducted to evaluate the beneficial effects of inositol hexaphosphate (IP6) + Inositol in breast cancer patients treated with adjuvant therapy. Patients and methods Patients with invasive ductal breast cancer where polychemotherapy was indicated were monitored in the period from 2005-2007. Fourteen patients in the same stage of ductal invasive breast cancer were involved in the study, divided in two randomized groups. One group was subjected to take IP6 + Inositol while the other group was taking placebo. In both groups of patients the same laboratory parameters were monitored. When the treatment was finished, all patients have filled questionnaires QLQ C30 and QLQ-BR23 to determine the quality of life. Results Patients receiving chemotherapy, along with IP6 + Inositol did not have cytopenia, drop in leukocyte and platelet counts. Red blood cell counts and tumor markers were unaltered in both groups. However, patients who took IP6 + Inositol had significantly better quality of life (p = 0.05) and functional status (p = 0.0003) and were able to perform their daily activities. Conclusion IP6 + Inositol as an adjunctive therapy is valuable help in ameliorating the side effects and preserving quality of life among the patients treated with chemotherapy. PMID:20152024

  10. Characterization of a ligand-gated cation channel based on an inositol receptor in the silkworm, Bombyx mori.

    PubMed

    Kikuta, Shingo; Endo, Haruka; Tomita, Natsuo; Takada, Tomoyuki; Morita, Chiharu; Asaoka, Kiyoshi; Sato, Ryoichi

    2016-07-01

    Insect herbivores recognize non-volatile compounds in plants to direct their feeding behavior. Gustatory receptors (Gr) appear to be required for nutrient recognition by gustatory organs in the mouthparts of insects. Gr10 is expressed in Bombyx mori (BmGr10) mouthparts such as maxillary galea, maxillary palp, and labrum. BmGr10 is predicted to function in sugar recognition; however, the precise biochemical function remains obscure. Larvae of B. mori are monophagous feeders able to find and feed on mulberry leaves. Soluble mulberry leaf extract contains sucrose, glucose, fructose, and myo-inositol. In this study, we identified BmGr10 as an inositol receptor using electrophysiological analysis with the Xenopus oocyte expression system and Ca(2+) imaging techniques using mammalian cells. These results demonstrated that Xenopus oocytes or HEK293T cells expressing BmGr10 specifically respond to myo-inositol and epi-inositol but do not respond to any mono-, di-, or tri-saccharides or to some sugar alcohols. These inositols caused Ca(2+) and Na(+) influxes into the cytoplasm independently of a G protein-mediated signaling cascade, indicating that BmGr10 is a ligand-gated cation channel. Overall, BmGr10 plays an important role in the myo-inositol recognition required for B. mori larval feeding behavior. PMID:27132146

  11. Pharmacokinetics and Safety of a Single Intravenous Dose of myo-Inositol in Preterm Infants of 23 to 29 weeks

    PubMed Central

    Phelps, Dale L.; Ward, Robert M.; Williams, Rick L.; Watterberg, Kristi L.; Laptook, Abbot R.; Wrage, Lisa A.; Nolen, Tracy L.; Fennell, Timothy R.; Ehrenkranz, Richard A.; Poindexter, Brenda B.; Cotten, C. Michael; Hallman, Mikko K.; Frantz, Ivan D.; Faix, Roger G.; Zaterka-Baxter, Kristin M.; Das, Abhik; Ball, M. Bethany; O’Shea, T. Michael; Lacy, Conra Backstrom; Walsh, Michele C.; Shankaran, Seetha; Sánchez, Pablo J.; Bell, Edward F.; Higgins, Rosemary D.

    2014-01-01

    Background Myo-inositol given to preterm infants with respiratory distress has reduced death, increased survival without bronchopulmonary dysplasia (BPD) and reduced severe retinopathy of prematurity (ROP) in 2 randomized trials. Pharmacokinetic (PK) studies in extremely preterm infants are needed prior to efficacy trials. Methods Infants of 23–29 weeks gestation were randomized to a single intravenous (IV) dose of inositol at 60 or 120 mg/kg or placebo. Over 96 h, serum levels (sparse sampling population PK) and urine inositol excretion were determined. Population PK models were fit using a nonlinear mixed effects approach. Safety outcomes were recorded. Results A 1-compartment model that included factors for endogenous inositol production, allometric size based on weight, gestational age (GA) strata and creatinine clearance fit the data best. The central volume of distribution was 0.5115 l/kg, the clearance 0.0679 l/kg/h, endogenous production 2.67 mg/kg/h and the half life 5.22 h when modeled without the covariates. During the first 12 h renal inositol excretion quadrupled in the 120 mg/kg group, returning to near baseline after 48 h. There was no diuretic side-effect. No significant differences in adverse events occurred between the 3 groups (p > 0.05). Conclusions A single compartment model accounting for endogenous production satisfactorily described the PK of IV inositol. PMID:24067395

  12. Characterization of glycosyl inositol phosphoryl ceramides from plants and fungi by mass spectrometry.

    PubMed

    Buré, Corinne; Cacas, Jean-Luc; Mongrand, Sébastien; Schmitter, Jean-Marie

    2014-02-01

    Although glycosyl inositol phosphoryl ceramides (GIPCs) represent the most abundant class of sphingolipids in plants, they still remain poorly characterized in terms of structure and biodiversity. More than 50 years after their discovery, little is known about their subcellular distribution and their exact roles in membrane structure and biological functions. This review is focused on extraction and characterization methods of GIPCs occurring in plants and fungi. Global methods for characterizing ceramide moieties of GIPCs revealed the structures of long-chain bases (LCBs) and fatty acids (FAs): LCBs are dominated by tri-hydroxylated molecules such as monounsaturated and saturated phytosphingosine (t18:1 and t18:0, respectively) in plants and mainly phytosphingosine (t18:0 and t20:0) in fungi; FA are generally 14-26 carbon atoms long in plants and 16-26 carbon atoms long in fungi, these chains being often hydroxylated in position 2. Mass spectrometry plays a pivotal role in the assessment of GIPC diversity and the characterization of their structures. Indeed, it allowed to determine that the core structure of GIPC polar heads in plants is Hex(R1)-HexA-IPC, with R1 being a hydroxyl, an amine, or a N-acetylamine group, whereas the core structure in fungi is Man-IPC. Notably, information gained from tandem mass spectrometry spectra was most useful to describe the huge variety of structures encountered in plants and fungi and reveal GIPCs with yet uncharacterized polar head structures, such as hexose-inositol phosphoceramide in Chondracanthus acicularis and (hexuronic acid)4-inositol phosphoceramide and hexose-(hexuronic acid)3-inositol phosphoceramide in Ulva lactuca. PMID:23887274

  13. Reduced myo-inositol and total choline measured with cerebral MRS in acute thyrotoxic Graves' disease.

    PubMed

    Elberling, T V; Danielsen, E R; Rasmussen, A K; Feldt-Rasmussen, U; Waldemar, G; Thomsen, C

    2003-01-14

    Neuropsychiatric symptoms in the acute thyrotoxic phase of Graves' disease suggest involvement of brain processes. Short-echo-time proton MRS was used to measure the cerebral metabolite profile in newly diagnosed and untreated Graves' disease. Sixteen patients with Graves' disease and 18 age- and sex-matched healthy volunteers were studied. The patients had significantly reduced total choline and myo-inositol in the acute phase of Graves' thyrotoxicosis compared with the healthy volunteers. PMID:12525741

  14. An Uncharacterized Member of the Ribokinase Family in Thermococcus kodakarensis Exhibits myo-Inositol Kinase Activity*

    PubMed Central

    Sato, Takaaki; Fujihashi, Masahiro; Miyamoto, Yukika; Kuwata, Keiko; Kusaka, Eriko; Fujita, Haruo; Miki, Kunio; Atomi, Haruyuki

    2013-01-01

    Here we performed structural and biochemical analyses on the TK2285 gene product, an uncharacterized protein annotated as a member of the ribokinase family, from the hyperthermophilic archaeon Thermococcus kodakarensis. The three-dimensional structure of the TK2285 protein resembled those of previously characterized members of the ribokinase family including ribokinase, adenosine kinase, and phosphofructokinase. Conserved residues characteristic of this protein family were located in a cleft of the TK2285 protein as in other members whose structures have been determined. We thus examined the kinase activity of the TK2285 protein toward various sugars recognized by well characterized ribokinase family members. Although activity with sugar phosphates and nucleosides was not detected, kinase activity was observed toward d-allose, d-lyxose, d-tagatose, d-talose, d-xylose, and d-xylulose. Kinetic analyses with the six sugar substrates revealed high Km values, suggesting that they were not the true physiological substrates. By examining activity toward amino sugars, sugar alcohols, and disaccharides, we found that the TK2285 protein exhibited prominent kinase activity toward myo-inositol. Kinetic analyses with myo-inositol revealed a greater kcat and much lower Km value than those obtained with the monosaccharides, resulting in over a 2,000-fold increase in kcat/Km values. TK2285 homologs are distributed among members of Thermococcales, and in most species, the gene is positioned close to a myo-inositol monophosphate synthase gene. Our results suggest the presence of a novel subfamily of the ribokinase family whose members are present in Archaea and recognize myo-inositol as a substrate. PMID:23737529

  15. Conformational Changes in Inositol 1,3,4,5,6-Pentakisphosphate 2-Kinase upon Substrate Binding

    PubMed Central

    Baños-Sanz, José Ignacio; Sanz-Aparicio, Julia; Whitfield, Hayley; Hamilton, Chris; Brearley, Charles A.; González, Beatriz

    2012-01-01

    Inositol 1,3,4,5,6-pentakisphosphate 2-kinase (IP5 2-K) catalyzes the synthesis of inositol 1,2,3,4,5,6-hexakisphosphate from ATP and IP5. Inositol 1,2,3,4,5,6-hexakisphosphate is implicated in crucial processes such as mRNA export, DNA editing, and phosphorus storage in plants. We previously solved the first structure of an IP5 2-K, which shed light on aspects of substrate recognition. However, failure of IP5 2-K to crystallize in the absence of inositide prompted us to study putative conformational changes upon substrate binding. We have made mutations to residues on a region of the protein that produces a clasp over the active site. A W129A mutant allowed us to capture IP5 2-K in its different conformations by crystallography. Thus, the IP5 2-K apo-form structure displays an open conformation, whereas the nucleotide-bound form shows a half-closed conformation, in contrast to the inositide-bound form obtained previously in a closed conformation. Both nucleotide and inositide binding produce large conformational changes that can be understood as two rigid domain movements, although local changes were also observed. Changes in intrinsic fluorescence upon nucleotide and inositide binding are in agreement with the crystallographic findings. Our work suggests that the clasp might be involved in enzyme kinetics, with the N-terminal lobe being essential for inositide binding and subsequent conformational changes. We also show how IP5 2-K discriminates between inositol 1,3,4,5-tetrakisphosphate and 3,4,5,6-tetrakisphosphate enantiomers and that substrate preference can be manipulated by Arg130 mutation. Altogether, these results provide a framework for rational design of specific inhibitors with potential applications as biological tools for in vivo studies, which could assist in the identification of novel roles for IP5 2-K in mammals. PMID:22745128

  16. Depletion of arachidonic acid from GH3 cells. Effects on inositol phospholipid turnover and cellular activation.

    PubMed Central

    Dudley, D T; Macfarlane, D E; Spector, A A

    1987-01-01

    We have adapted rat pituitary GH3 cells to grow in delipidated culture medium. In response, esterfied linoleic acid and arachidonic acid become essentially undetectable, whereas eicosa-5,8,11-trienoic acid accumulates and oleic acid increases markedly. These changes occur in all phospholipid classes, but are particularly pronounced in inositol phospholipids, where the usual stearate/arachidonate profile is replaced with oleate/eicosatrienoate (n - 9) and stearate/eicosatrienoate (n - 9). Incubation of arachidonate-depleted cells with 10 microM-arachidonic acid for only 24 h results in extensive remodelling of phospholipid fatty acids, such that close-to-normal compositions and arachidonic acid content are achieved for the inositol phospholipids. In comparison studies with arachidonic acid-depleted or -repleted cells, it was found that the arachidonate content does not affect thyrotropin-releasing-hormone (TRH)-stimulated responses measured at long time points, including [32P]Pi labelling of phosphatidylinositol and phosphatidic acid, stimulation of protein phosphorylation, and basal or TRH-stimulated prolactin release. However, transient events such as stimulated breakdown of inositol phospholipids and an initial rise in diacylglycerol are enhanced by the presence of arachidonate. These results show that arachidonic acid itself is not required for operation of the phosphatidylinositol cycle and is not an obligatory intermediate in TRH-mediated GH3 cell activation. It is possible that any structural or functional role of arachidonic acid in these processes is largely met by replacement with eicosatrienoate (n - 9). However, since arachidonate in inositol phospholipids facilitates their hydrolysis upon stimulation by TRH, arachidonic acid apparently may have a specific role in the recognition of these lipids by phospholipase C. Images Fig. 4. PMID:3120699

  17. A Combined Therapy with Myo-Inositol and D-Chiro-Inositol Improves Endocrine Parameters and Insulin Resistance in PCOS Young Overweight Women.

    PubMed

    Benelli, Elena; Del Ghianda, Scilla; Di Cosmo, Caterina; Tonacchera, Massimo

    2016-01-01

    Introduction. We evaluated the effects of a therapy that combines myo-inositol (MI) and D-chiro-inositol (DCI) in young overweight women affected by polycystic ovary syndrome (PCOS), characterized by oligo- or anovulation and hyperandrogenism, correlated to insulin resistance. Methods. We enrolled 46 patients affected by PCOS and, randomly, we assigned them to two groups, A and B, treated, respectively, with the association of MI plus DCI, in a 40 : 1 ratio, or with placebo (folic acid) for six months. Thus, we analyzed pretreatment and posttreatment FSH, LH, 17-beta-Estradiol, Sex Hormone Binding Globulin, androstenedione, free testosterone, dehydroepiandrosterone sulphate, HOMA index, and fasting glucose and insulin. Results. We recorded a statistically significant reduction of LH, free testosterone, fasting insulin, and HOMA index only in the group treated with the combined therapy of MI plus DCI; in the same patients, we observed a statistically significant increase of 17-beta-Estradiol levels. Conclusions. The combined therapy of MI plus DCI is effective in improving endocrine and metabolic parameters in young obese PCOS affected women. PMID:27493664

  18. A Combined Therapy with Myo-Inositol and D-Chiro-Inositol Improves Endocrine Parameters and Insulin Resistance in PCOS Young Overweight Women

    PubMed Central

    Benelli, Elena; Del Ghianda, Scilla

    2016-01-01

    Introduction. We evaluated the effects of a therapy that combines myo-inositol (MI) and D-chiro-inositol (DCI) in young overweight women affected by polycystic ovary syndrome (PCOS), characterized by oligo- or anovulation and hyperandrogenism, correlated to insulin resistance. Methods. We enrolled 46 patients affected by PCOS and, randomly, we assigned them to two groups, A and B, treated, respectively, with the association of MI plus DCI, in a 40 : 1 ratio, or with placebo (folic acid) for six months. Thus, we analyzed pretreatment and posttreatment FSH, LH, 17-beta-Estradiol, Sex Hormone Binding Globulin, androstenedione, free testosterone, dehydroepiandrosterone sulphate, HOMA index, and fasting glucose and insulin. Results. We recorded a statistically significant reduction of LH, free testosterone, fasting insulin, and HOMA index only in the group treated with the combined therapy of MI plus DCI; in the same patients, we observed a statistically significant increase of 17-beta-Estradiol levels. Conclusions. The combined therapy of MI plus DCI is effective in improving endocrine and metabolic parameters in young obese PCOS affected women. PMID:27493664

  19. Activation of Ca2+ entry into acinar cells by a non-phosphorylatable inositol trisphosphate

    NASA Astrophysics Data System (ADS)

    Bird, G. S. J.; Rossier, M. F.; Hughes, A. R.; Shears, S. B.; Armstrong, D. L.; , J. W. Putney, Jr.

    1991-07-01

    IN many cell types, receptor activation of phosphoinositidase C results in an initial release of intracellular Ca2+ stores followed by sustained Ca2+ entry across the plasma membrane. Inositol 1,4,5-trisphosphate is the mediator of the initial Ca2+ release1, although its role in the mechanism underlying Ca2+ entry remains controversial1-6. We have now used two techniques to introduce inositol phosphates into mouse lacrimal acinar cells and measure their effects on Ca2+ entry: microinjection into cells loaded with Fura-2, a fluorescent dye which allows the measurement of intra-cellular free calcium concentration by microspectrofluorimetry, and perfusion of patch clamp pipettes in the whole-cell configuration while monitoring the activity of Ca2+-activated K+ channels as an indicator of intracellular Ca2+. We report here that inositol 1,4,5-trisphosphate serves as a signal that is both necessary and sufficient for receptor activation of Ca2+ entry across the plasma membrane in these cells.

  20. Relationship Between Myo-Inositol Supplementary and Gestational Diabetes Mellitus: A Meta-Analysis.

    PubMed

    Zheng, Xiangqin; Liu, Zhaozhen; Zhang, Yulong; Lin, Yuan; Song, Jianrong; Zheng, Lianghui; Lin, Sheng

    2015-10-01

    To determine whether myo-inositol supplement will increase the action of endogenous insulin, which is mainly measured by markers of insulin resistance such as homeostasis model assessment of insulin resistance.PubMed, Cochrane Library, Embase, and web of science were comprehensively searched using "gestational diabetes mellitus" and "myo-inositol" to identify relevant studies. Both subject headings and free texts were adopted. The methodological quality of the included studies were assessed and pooled analyzed by the methods recommended by the Cochrane collaboration.A total of 5 trials containing 513 participants were included. There was a significant reduction in aspects of gestational diabetes incidence (risk ratio [RR], 0.29; 95% confidence interval (95% CI), 0.19-0.44), birth weight (mean difference [MD], -116.98; 95% CI, -208.87 to -25.09), fasting glucose oral glucose tolerance test (OGTT) (MD, -0.36; 95% CI, -0.51 to -0.21), 1-h glucose OGTT (MD, -0.63; 95% CI, -1.01 to -0.26), 2-h glucose OGTT (MD, -0.45; 95% CI, -0.75 to -0.16), and related complications (odds ratio [OR], 0.28; 95% CI 0.14-0.58).On the basis of current evidence, myo-inositol supplementation reduces the development of gestational diabetes mellitus (GDM), although this conclusion requires further evaluation in large-scale, multicenter, blinded randomized controlled trials. PMID:26496267

  1. Amino acid depletion activates TonEBP and sodium-coupled inositol transport.

    PubMed

    Franchi-Gazzola, R; Visigalli, R; Dall'Asta, V; Sala, R; Woo, S K; Kwon, H M; Gazzola, G C; Bussolati, O

    2001-06-01

    The expression of the osmosensitive sodium/myo-inositol cotransporter (SMIT) is regulated by multiple tonicity-responsive enhancers (TonEs) in the 5'-flanking region of the gene. In response to hypertonicity, the nuclear abundance of the transcription factor TonE-binding protein (TonEBP) is increased, and the transcription of the SMIT gene is induced. Transport system A for neutral amino acids, another osmosensitive mechanism, is progressively stimulated if amino acid substrates are not present in the extracellular compartment. Under this condition, as in hypertonicity, cells shrink and mitogen-activated protein kinases are activated. We demonstrate here that a clear-cut nuclear redistribution of TonEBP, followed by SMIT expression increase and inositol transport activation, is observed after incubation of cultured human fibroblasts in Earle's balanced salts (EBSS), an isotonic, amino acid-free saline. EBSS-induced SMIT stimulation is prevented by substrates of system A, although these compounds do not compete with inositol for transport through SMIT. We conclude that the incubation in isotonic, amino acid-free saline triggers an osmotic stimulus and elicits TonEBP-dependent responses like hypertonic treatment. PMID:11350742

  2. Effect of a myo-Inositol Antagonist, 2-O, C-Methylene-myo-Inositol, on the Metabolism of myo-Inositol-2-3H and d-Glucose-1-14C in Lilium longiflorum Pollen 1

    PubMed Central

    Chen, Minshen; Loewus, Mary Walz; Loewus, Frank A.

    1977-01-01

    2-O,C-Methylene-myo-inositol (MMO), a myo-inositol (MI) antagonist, inhibits germination and tube elongation of pollen from Lilium longiflorum cv. Ace or 44. The presence of 5 mm MMO in Dickinson's pentaerythritol medium (Plant Physiol. 43:1-8) partially blocks germination. The tubes produced are short and fail to elongate. In the presence of MI, MMO's toxic effect is blocked. As little as 0.56 mm MI will maintain normal germination in the presence of 43 mm MMO, and pollen tubes continue to elongate for 2 to 3 hr. Eventually, the toxic action of MMO prevents further growth. MMO does not inhibit UDP-d-glucose dehydrogenase from lily pollen. Uptake of MI-2-3H and incorporation of tritium into galacturonic acid and pentose units of tube wall pectin are blocked by MMO. The site of this inhibition is undertermined. Uptake of d-glucose-1-14C and incorporation of 14C into 70% ethyl alcohol-insoluble polysaccharides of germinated pollen are not blocked by MMO, but distribution of label into polysaccharide product is altered. In MMO-treated pollen, very little 14C is found in uronic acid or pentose units. At 30 mm MMO, about two-thirds of the carbon flow from d-glucose to these pectic components is interrupted. MMO also alters d-glucose metabolism in the 70% ethyl alcohol-soluble fraction, but the compound involved must still be identified. These results offer fresh evidence of an intermediary role for MI during UDP-d-glucuronate biosynthesis in germinated pollen. PMID:16659913

  3. 4.1N binding regions of inositol 1,4,5-trisphosphate receptor type 1.

    PubMed

    Fukatsu, Kazumi; Bannai, Hiroko; Inoue, Takafumi; Mikoshiba, Katsuhiko

    2006-04-01

    Zhang et al. and Maximov et al. [S. Zhang, A. Mizutani, C. Hisatsune, T. Higo, H. Bannai, T. Nakayama, M. Hattori, and K. Mikoshiba, Protein 4.1N is required for translocation of inositol 1,4,5-trisphosphate receptor type 1 to the basolateral membrane domain in polarized Madin-Darby canine kidney cells, J. Biol. Chem. 278 (2003) 4048-4056; A. Maximov, T. S. Tang, and I. Bezprozvanny, Association of the type 1 inositol (1,4,5)-trisphosphate receptor with 4.1N protein in neurons, Mol. Cell. Neurosci. 22 (2003) 271-283.] reported that 4.1N is a binding partner of inositol 1,4,5-trisphosphate receptor type 1 (IP(3)R1), however the binding site of IP(3)R1 differed: the former determined the C-terminal 14 amino acids of the cytoplasmic tail (CTT14aa) as the binding site, while the latter assigned another segment, cytoplasmic tail middle 1 (CTM1). To solve this discrepancy, we performed immunoprecipitation and found that both the segments had binding activity to 4.1N. Both segments also interfered the 4.1N-regulated IP(3)R1 diffusion in neuronal dendrites. However, IP(3)R1 lacking the CTT14aa (IP(3)R1-DeltaCTT14aa) does not bind to 4.1N [S. Zhang, A. Mizutani, C. Hisatsune, T. Higo, H. Bannai, T. Nakayama, M. Hattori, and K. Mikoshiba, Protein 4.1N is required for translocation of inositol 1,4,5-trisphosphate receptor type 1 to the basolateral membrane domain in polarized Madin-Darby canine kidney cells, J. Biol. Chem. 278 (2003) 4048-4056.] and its diffusion constant is larger than that of IP(3)R1 full-length in neuronal dendrites [K. Fukatsu, H. Bannai, S. Zhang, H. Nakamura, T. Inoue, and K. Mikoshiba, Lateral diffusion of inositol 1,4,5-trisphosphate receptor type 1 is regulated by actin filaments and 4.1N in neuronal dendrites, J. Biol. Chem. 279 (2004) 48976-48982.]. We conclude that both the CTT14aa and CTM1 sequences can bind to 4.1N in peptide fragment forms. However, we propose that the responsible binding site for 4.1N binding in full-length tetramer form of IP

  4. Anion-exchange high-performance liquid chromatography with post-column detection for the analysis of phytic acid and other inositol phosphates

    NASA Technical Reports Server (NTRS)

    Rounds, M. A.; Nielsen, S. S.; Mitchell, C. A. (Principal Investigator)

    1993-01-01

    The use of gradient anion-exchange HPLC, with a simple post-column detection system, is described for the separation of myo-inositol phosphates, including "phytic acid" (myo-inositol hexaphosphate). Hexa-, penta-, tetra-, tri- and diphosphate members of this homologous series are clearly resolved within 30 min. This method should facilitate analysis and quantitation of "phytic acid" and other inositol phosphates in plant, food, and soil samples.

  5. Variations in myo-inositol in fronto-limbic regions and clinical response to electroconvulsive therapy in major depression.

    PubMed

    Njau, Stephanie; Joshi, Shantanu H; Leaver, Amber M; Vasavada, Megha; Van Fleet, Jessica; Espinoza, Randall; Narr, Katherine L

    2016-09-01

    Though electroconvulsive therapy (ECT) is an established treatment for severe depression, the neurobiological factors accounting for the clinical effects of ECT are largely unknown. Myo-inositol, a neurometabolite linked with glial activity, is reported as reduced in fronto-limbic regions in patients with depression. Whether changes in myo-inositol relate to the antidepressant effects of ECT is unknown. Using magnetic resonance spectroscopy ((1)H-MRS), we measured dorsomedial anterior cingulate cortex (dmACC) and left and right hippocampal myo-inositol in 50 ECT patients (mean age: 43.78, 14 SD) and 33 controls (mean age: 39.33, 12 SD) to determine cross sectional effects of diagnosis and longitudinal effects of ECT. Patients were scanned prior to treatment, after the second ECT and at completion of the ECT index series. Controls were scanned twice at intervals corresponding to patients' baseline and end of treatment scans. Myo-inositol increased over the course of ECT in the dmACC (p = 0.042). A significant hemisphere by clinical response effect was observed for the hippocampus (p = 0.003) where decreased myo-inositol related to symptom improvement in the left hippocampus. Cross-sectional differences between patients and controls at baseline were not detected. Changes in myo-inositol observed in the dmACC in association with ECT and in the hippocampus in association with ECT-related clinical response suggest the mechanisms of ECT could include gliogenesis or a reversal of gliosis that differentially affect dorsal and ventral limbic regions. Change in dmACC myo-inositol diverged from control values with ECT suggesting compensation, while hippocampal change suggested normalization. PMID:27285661

  6. Specific isoforms of protein kinase C are essential for prevention of folate-resistant neural tube defects by inositol.

    PubMed

    Cogram, Patricia; Hynes, Andrew; Dunlevy, Louisa P E; Greene, Nicholas D E; Copp, Andrew J

    2004-01-01

    A proportion of neural tube defects (NTDs) can be prevented by maternal folic acid supplementation, although some cases are unresponsive. The curly tail mutant mouse provides a model of folate-resistant NTDs, in which defects can be prevented by inositol therapy in early pregnancy. Hence, inositol represents a possible novel adjunct therapy to prevent human NTDs. The present study investigated the molecular mechanism by which inositol prevents mouse NTDs. Activation of protein kinase C (PKC) is known to be essential, and we examined neurulation-stage embryos for PKC expression and applied PKC inhibitors to curly tail embryos developing in culture. Although all known PKC isoforms were detected in the closing neural tube, use of chemical PKC inhibitors identified a particular requirement for 'conventional' PKC isoforms. Peptide inhibitors offer selective inhibition of individual PKCs, and we demonstrated isoform-specific inhibition of PKC in embryonic cell cultures. Application of peptide inhibitors to neurulation-stage embryos revealed an absolute dependence on the activity of PKCbetaI and gamma for prevention of NTDs by inositol, and partial dependence on PKCzeta, whereas other PKCs (alpha, betaII delta, and epsilon) were dispensable. To investigate the cellular action of inositol and PKCs in NTD prevention, we examined cell proliferation in curly tail embryos. Defective proliferation of hindgut cells is a key component of the pathogenic sequence leading to NTDs in curly tail. Hindgut cell proliferation was stimulated specifically by inositol, an effect that required activation of PKCbetaI. Our findings reveal an essential role of specific PKC isoforms in mediating the prevention of mouse NTDs by inositol. PMID:14613966

  7. Possible evolutionary relationships between streptomycin and bluensomycin biosynthetic pathways: detection of novel inositol kinase and O-carbamoyltransferase activities.

    PubMed Central

    Walker, J B

    1990-01-01

    Bluensomycin (glebomycin) is an aminocyclitol antibiotic that differs structurally from dihydrostreptomycin in having bluensidine (1D-1-O-carbamoyl-3-guanidinodeoxy-scyllo-inositol) rather than streptidine (1,3-diguanidino-1,3-dideoxy-scyllo-inositol) as its aminocyclitol moiety. Extracts of the bluensomycin producer Streptomyces hygroscopicus form glebosus ATCC 14607 (S. glebosus) were found to have aminodeoxy-scyllo-inositol kinase activity but to lack 1D-1-guanidino-3-amino-1,3-dideoxy-scyllo-inositol kinase activity, showing for the first time that these two reactions in streptomycin producers must be catalyzed by different enzymes. S. glebosus extracts therefore possess the same five enzymes required for synthesis of guanidinodeoxy-scyllo-inositol from myo-inositol that are found in streptomycin producers but lack the next three of the four enzymes found in streptomycin producers that are required to synthesize the second guanidino group of streptidine-P. In place of a second guanidino group, S. glebosus extracts were found to catalyze a Mg2(+)-dependent carbamoylation of guanidinodeoxy-scyllo-inositol to form bluensidine, followed by a phosphorylation to form bluensidine-P. The novel carbamoyl-P:guanidinodeoxy-scyllo-inositol O-carbamoyltransferase and ATP:bluensidine phosphotransferase activities were not detected in streptomycin producers or in S. glebosus during its early rapid growth phase. Free bluensidine appears to be a normal intermediate in bluensomycin biosynthesis, in contrast to the case of streptomycin biosynthesis; in the latter, although exogenous streptidine can enter the pathway via streptidine-P, free streptidine is not an intermediate in the endogenous biosynthetic pathway. Comparison of the streptomycin and bluensomycin biosynthetic pathways provides a unique opportunity to evaluate those proposed mechanisms for the evolutionary acquisition of new biosynthetic capabilities that involve gene duplication and subsequent mutational changes in

  8. Certain Malvaceae Plants Have a Unique Accumulation of myo-Inositol 1,2,4,5,6-Pentakisphosphate.

    PubMed

    Phillippy, Brian Q; Perera, Imara Y; Donahue, Janet L; Gillaspy, Glenda E

    2015-01-01

    Methods used to quantify inositol phosphates in seeds lack the sensitivity and specificity necessary to accurately detect the lower concentrations of these compounds contained in the leaves of many plants. In order to measure inositol hexakisphosphate (InsP₆) and inositol pentakisphosphate (InsP₅) levels in leaves of different plants, a method was developed to concentrate and pre-purify these compounds prior to analysis. Inositol phosphates were extracted from leaves with diluted HCl and concentrated on small anion exchange columns. Reversed-phase solid phase extraction cartridges were used to remove compounds that give peaks that sometimes interfere during HPLC. The method permitted the determination of InsP₆ and InsP₅ concentrations in leaves as low as 10 µM and 2 µM, respectively. Most plants analyzed contained a high ratio of InsP₆ to InsP₅. In contrast, certain members of the Malvaceae family, such as cotton (Gossypium) and some hibiscus (Hibiscus) species, had a preponderance of InsP₅. Radiolabeling of cotton seedlings also showed increased amounts of InsP₅ relative to InsP₆. Why some Malvaceae species exhibit a reversal of the typical ratios of these inositol phosphates is an intriguing question for future research. PMID:27135328

  9. Arabidopsis inositol 1,3,4-trisphosphate 5/6 kinase 2 is required for seed coat development.

    PubMed

    Tang, Yong; Tan, Shutang; Xue, Hongwei

    2013-07-01

    Inositol 1,3,4-trisphosphate 5/6 kinase (ITPK) phosphorylates inositol 1,3,4-trisphosphate to form inositol 1,3,4,5-tetrakisphosphate and inositol 1,3,4,6-tetrakisphosphate which can be finally transferred to inositol hexaphosphate (IP₆) and play important roles during plant growth and development. There are 4 putative ITPK members in Arabidopsis. Expression pattern analysis showed that ITPK2 is constitutively expressed in various tissues. A T-DNA knockout mutant of ITPK2 was identified and scanning electron microscopy (SEM) analysis showed that the epidermis structure of seed coat was irregularly formed in seeds of itpk2-1 mutant, resulting in the increased permeability of seed coat to tetrazolium salts. Further analysis by gas chromatography coupled with mass spectrometry of lipid polyester monomers in cell wall confirmed a dramatic decrease in composition of suberin and cutin, which relate to the permeability of seed coat and the formation of which is accompanied with seed coat development. These results indicate that ITPK2 plays an essential role in seed coat development and lipid polyester barrier formation. PMID:23595027

  10. Certain Malvaceae Plants Have a Unique Accumulation of myo-Inositol 1,2,4,5,6-Pentakisphosphate

    PubMed Central

    Phillippy, Brian Q.; Perera, Imara Y.; Donahue, Janet L.; Gillaspy, Glenda E.

    2015-01-01

    Methods used to quantify inositol phosphates in seeds lack the sensitivity and specificity necessary to accurately detect the lower concentrations of these compounds contained in the leaves of many plants. In order to measure inositol hexakisphosphate (InsP6) and inositol pentakisphosphate (InsP5) levels in leaves of different plants, a method was developed to concentrate and pre-purify these compounds prior to analysis. Inositol phosphates were extracted from leaves with diluted HCl and concentrated on small anion exchange columns. Reversed-phase solid phase extraction cartridges were used to remove compounds that give peaks that sometimes interfere during HPLC. The method permitted the determination of InsP6 and InsP5 concentrations in leaves as low as 10 µM and 2 µM, respectively. Most plants analyzed contained a high ratio of InsP6 to InsP5. In contrast, certain members of the Malvaceae family, such as cotton (Gossypium) and some hibiscus (Hibiscus) species, had a preponderance of InsP5. Radiolabeling of cotton seedlings also showed increased amounts of InsP5 relative to InsP6. Why some Malvaceae species exhibit a reversal of the typical ratios of these inositol phosphates is an intriguing question for future research. PMID:27135328

  11. [3H]Indole-3-acetyl-myo-inositol hydrolysis by extracts of Zea mays L. vegetative tissue

    NASA Technical Reports Server (NTRS)

    Hall, P. J.; Bandurski, R. S.

    1986-01-01

    [3H]Indole-3-acetyl-myo-inositol was hydrolyzed by buffered extracts of acetone powders prepared from 4 day shoots of dark grown Zea mays L. seedlings. The hydrolytic activity was proportional to the amount of extract added and was linear for up to 6 hours at 37 degrees C. Boiled or alcohol denatured extracts were inactive. Analysis of reaction mixtures by high performance liquid chromatography demonstrated that not all isomers of indole-3-acetyl-myo-inositol were hydrolyzed at the same rate. Buffered extracts of acetone powders were prepared from coleoptiles and mesocotyls. The rates of hydrolysis observed with coleoptile extracts were greater than those observed with mesocotyl extracts. Active extracts also catalyzed the hydrolysis of esterase substrates such as alpha-naphthyl acetate and the methyl esters of indoleacetic acid and naphthyleneacetic acid. Attempts to purify the indole-3-acetyl-myo-inositol hydrolyzing activity by chromatographic procedures resulted in only slight purification with large losses of activity. Chromatography over hydroxylapatite allowed separation of two enzymically active fractions, one of which catalyzed the hydrolysis of both indole-3-acetyl-myo-inositol and esterase substrates. With the other enzymic hydrolysis of esterase substrates was readily demonstrated, but no hydrolysis of indole-3-acetyl-myo-inositol was ever detected.

  12. Formation of inositol 1,3,4,6-tetrakisphosphate during angiotensin II action in bovine adrenal glomerulosa cells

    SciTech Connect

    Balla, T.; Guillemette, G.; Baukal, A.J.; Catt, K.J.

    1987-10-14

    Angiotensin II stimulates the formation of several inositol polyphosphates in cultured bovine adrenal glomerulosa cells prelabelled with (/sup 3/H) inositol. Analysis by high performance anion exchange chromatography of the inositol-phosphate compounds revealed the existence of two additional inositol tetrakisphosphate (InsP4) isomers in proximity to Ins-1,3,4,5-P4, the known phosphorylation product of Ins-1,4,5-trisphosphate and precursor of Ins-1,3,4-trisphosphate. Both of these new compounds showed a slow increase after stimulation with angiotensin II. The structure of one of these new InsP4 isomers, which is a phosphorylation product of Ins-1,3,4-P3, was deduced by its resistance to periodate oxidation to be Ins-1,3,4,6-P4. The existence of multiple cycles of phosphorylation-dephosphorylation reactions for the processing of Ins-1,4,5-P4 may represent a new aspect of the inositol-lipid related signalling mechanism in agonist-activated target cells.

  13. Asymmetric Distribution of Glucose and Indole-3-Acetyl-myo-Inositol in Geostimulated Zea mays Seedlings 1

    PubMed Central

    Momonoki, Yoshie S.

    1988-01-01

    Indole-3-acetyl-myo-inositol occurs in both the kernel and vegetative shoot of germinating Zea mays seedlings. The effect of a gravitational stimulus on the transport of [3H]-5-indole-3-acetyl-myo-inositol and [U-14C]-d-glucose from the kernel to the seedling shoot was studied. Both labeled glucose and labeled indole-3-acetyl-myo-inositol become asymmetrically distributed in the mesocotyl cortex of the shoot with more radioactivity occurring in the bottom half of a horizontally placed seedling. Asymmetric distribution of [3H]indole-3-acetic acid, derived from the applied [3H]indole-3-acetyl-myo-inositol, occurred more rapidly than distribution of total 3H-radioactivity. These findings demonstrate that the gravitational stimulus can induce an asymmetric distribution of substances being transported from kernel to shoot. They also indicate that, in addition to the transport asymmetry, gravity affects the steady state amount of indole-3-acetic acid derived from indole-3-acetyl-myo-inositol. PMID:11537873

  14. Effect of atropine and gammahydroxybutyrate on ischemically induced changes in the level of radioactivity in (/sup 3/H)inositol phosphates in gerbil brain in vivo

    SciTech Connect

    Wikiel, H.; Halat, G.; Strosznajder, J.

    1988-05-01

    Brain ischemia in gerbils was induced by ligation of both common carotid arteries for 1 min or 10 min. Sham-operated animals served as controls. Intracerebral injection of (3H)inositol into gerbil brain 16 hr before ischemic insult resulted in equilibration of the label between inositol lipids and water-soluble inositol phosphate. A short ischemic period (1 min) resulted in a statistically significant increase in the radioactivity of inositol triphosphate (IP3) and inositol monophosphate (IP), by about 48% and 79%, respectively, with little change in that of the intermediate inositol biphosphate (IP2), which increased by about 16%. When the ischemic period was prolonged (10 min), an increase in the radioactivity of inositol monophosphate exclusively, by about 84%, was observed. The level of radioactivity in inositol phosphates IP2 and IP3 decreased by about 50%, probably as a consequence of phosphatase activation by the ischemic insult. The agonist of the cholinergic receptor, carbachol, injected intracerebrally (40 micrograms per animal) increased accumulation of radioactivity in all inositol phosphates. The level of radioactivity in IP3, IP2, and IP was elevated by about 40, 23, and 147%, respectively. The muscarinic cholinergic antagonist, atropine, injected intraperitoneally in doses of 100 mg/kg body wt. depressed phosphoinositide metabolism in control animals. The level of radioactivity in water-soluble inositol metabolites in the brain of animals pretreated with atropine was evidently about 32% lower than in untreated animals. Pretreatment with atropine decreased the radioactivity of all inositol phosphates in the brain of animals subjected to 1-min ischemia and the radioactivity of IP in the case of 10-min brain ischemia.

  15. Prefrontal inositol levels and implicit decision-making in healthy individuals and depressed patients.

    PubMed

    Jollant, Fabrice; Richard-Devantoy, Stéphane; Ding, Yang; Turecki, Gustavo; Bechara, Antoine; Near, Jamie

    2016-08-01

    Risky decision-making is found in several mental disorders and is associated with deleterious consequences. Current research aims at understanding the biological underpinnings of this complex cognitive function and the basis of individual variability. We used 3T proton Magnetic Resonance Spectroscopy to measure in vivo glutamate, GABA, N-acetyl-aspartate (NAA), and myo-inositol levels at rest in the right dorsal prefrontal cortex of 54 participants, comprising 24 unmedicated depressed patients and 30 healthy individuals. Participants were also tested with the Iowa Gambling Task (IGT), a classical measure of value-based decision-making. No group differences were found in terms of compound levels or decision-making performance. However, high inositol levels were associated with lower decision-making scores independently from group, notably during the initial stage of the task when explicit rules are still unknown and decisions are largely based on implicit processes (whole sample: F=4.0; p=0.02), with a large effect size (Cohen׳s d=0.8, 95% [0.2-1.5]). This effect was stronger when explicit knowledge was taken into account, with explicit knowledge showing an independent effect on performance. There was no association with other compounds. This study suggests, for the first time, a role for the inositol pathway on the implicit learning component of decision-making, without any direct effect on the explicit component. Hypothesized mechanisms implicate intracellular calcium modulation and subsequent synaptic plasticity. These findings represent a first step in the understanding of the biochemical mechanisms underlying decision-making and the identification of therapeutic targets. They also emphasize a dimensional approach in the study of the neurobiological determinants of mental disorders. PMID:27342631

  16. Inositol Metabolism in Plants. IV. Biosynthesis of Apiose in Lemna and Petroselinum

    PubMed Central

    Roberts, R. M.; Shah, R. H.; Loewus, F.

    1967-01-01

    The biosynthesis of apiose was investigated in cell wall polysaccharide of Lemna gibba G3 (duckweed) and in detached leaves of Petroselinum crispum (parsley). Lemna grown either in short days or in continuous light incorporated 14C from a medium containing myo-inositol-2-14C into d-apiosyl and d-xylosyl units of cell wall polysaccharides. Labeled d-apiose was characterized by paper chromatography, by formation of labeled crystalline di-O-isopropylidene d-apiose, and by gas chromatography of trimethylsilyl derivatives of apiose and of its sodium borohydride reduction product, apiitol. Periodate oxidation of labeled apiose revealed 86 to 94% of the 14C was located in formaldehyde fragments corresponding to C3′ and C4. Comparison of this result with work reported by Grisebach and Doebereiner and by Beck and Kandler supports the conclusion that myo-inositol-2-14C was converted to d-apiose labeled specifically at C4. When l-arabinose-l-14C was supplied to Lemna, both l-arabinosyl and d-xylosyl units of cell wall polysaccharides became labeled, but no 14C was found in d-apiose. Analysis of the medium external to the plants revealed the presence of a polysaccharide-like polymer that also contained labeled xylose and arabinose. Petroselinum leaves utilized myo-inositol-2-3H for the synthesis of apiose in apiin. These results provide direct evidence for a pathway of apiose biosynthesis involving d-glucuronic acid metabolism. PMID:16656551

  17. Production of inositol trisphosphates upon. cap alpha. -adrenergic stimulation in BC3H-1 muscle cells

    SciTech Connect

    Ambler, S.K.; Thompson, B.; Brown, J.H.; Taylor, P.

    1986-05-01

    Activation of ..cap alpha../sub 1/-adrenergic receptors in BC3H-1 muscle cells rapidly mobilizes intracellular and results in a paradoxically slower accumulation of inositol trisphosphate. A possible explanation for this discrepancy may be provided by the recent findings of Irvine et al. of additional Ins P3 isomers besides the Ca/sup + +/-mobilizing isomer, Ins 1,4,5-P3. They have eluted and separated the inositol phosphates of BC3H-1 cells with an NH/sub 4//sup +/ x HCO/sub 2//sup -//H/sub 3/PO/sub 4/ gradient on a Whatman Partisil 10SAX column using Hewlett-Packard HPLC. Commercial (/sup 3/H)Ins 1,4,5-P3 and (/sup 3/H)inositol phosphates from carbachol-stimulated parotid glands were used as standards. Little or no Ins 1,3,4-P3 could be detected in control or phenylephrine-treated BC3H-1 cells. Ins 1,4,5-P3 followed the pattern of agonist stimulation observed previously. As a positive control, Ins P3 isomers were also measured in 1321N1 astrocytoma cells. Muscarinic stimulation of 1321N1 cells results in both the rapid accumulation of Ins P3 and Ca/sup + +/ mobilization. There is no detectable basal Ins 1,3,4-P3, but carbachol stimulates a rapid production of this compound in 1321N1 cells. Agonist activation also results in a rapid increase in Ins 1,4,5-P3 above basal values. These studies indicate that Ins 1,3,4-P3 does not contribute to the InsP3 signal in BC3H-1 cells and multiple mechanisms may exist for the coupling of receptors to PI turnover.

  18. Fungal Inositol Pyrophosphate IP7 Is Crucial for Metabolic Adaptation to the Host Environment and Pathogenicity

    PubMed Central

    Lev, Sophie; Li, Cecilia; Desmarini, Desmarini; Saiardi, Adolfo; Fewings, Nicole L.; Schibeci, Stephen D.; Sharma, Raghwa; Sorrell, Tania C.

    2015-01-01

    ABSTRACT Inositol pyrophosphates (PP-IPs) comprising inositol, phosphate, and pyrophosphate (PP) are essential for multiple functions in eukaryotes. Their role in fungal pathogens has never been addressed. Cryptococcus neoformans is a model pathogenic fungus causing life-threatening meningoencephalitis. We investigate the cryptococcal kinases responsible for the production of PP-IPs (IP7/IP8) and the hierarchy of PP-IP importance in pathogenicity. Using gene deletion and inositol polyphosphate profiling, we identified Kcs1 as the major IP6 kinase (producing IP7) and Asp1 as an IP7 kinase (producing IP8). We show that Kcs1-derived IP7 is the most crucial PP-IP for cryptococcal drug susceptibility and the production of virulence determinants. In particular, Kcs1 kinase activity is essential for cryptococcal infection of mouse lungs, as reduced fungal burdens were observed in the absence of Kcs1 or when Kcs1 was catalytically inactive. Transcriptome and carbon source utilization analysis suggested that compromised growth of the KCS1 deletion strain (Δkcs1 mutant) in the low-glucose environment of the host lung is due to its inability to utilize alternative carbon sources. Despite this metabolic defect, the Δkcs1 mutant established persistent, low-level asymptomatic pulmonary infection but failed to elicit a strong immune response in vivo and in vitro and was not readily phagocytosed by primary or immortalized monocytes. Reduced recognition of the Δkcs1 cells by monocytes correlated with reduced exposure of mannoproteins on the Δkcs1 mutant cell surface. We conclude that IP7 is essential for fungal metabolic adaptation to the host environment, immune recognition, and pathogenicity. PMID:26037119

  19. Is inositol (1,3,4,5)-tetrakisphosphate a new second messenger

    SciTech Connect

    Hansen, C.A.; Williamson, J.R.

    1986-05-01

    Hormone-stimulated hydrolysis of inositol (Ins) lipids results in the rapid formation of Ins(1,4,5)P/sub 3/, the second messenger for intracellular Ca/sup 2 +/ mobilization. Recently, a more polar inositol phosphate, Ins(1,3,4,5)P/sub 4/ as well as its probable hydrolysis product Ins(1,3,4)P/sub 3/ have been reported to accumulate in carbachol-stimulated brain slices. Vasopressin addition to hepatocytes prelabeled with (/sup 3/H)-Ins also showed a rapid increase of Ins(1,3,4,5)P/sub 4/, which was similar to that of Ins(1,4,5)P/sub 3/, while the accumulation of Ins(1,3,4)P/sub 3/ was slower. In order to examine whether Ins(1,3,4,5)P/sub 4/ has any functional effects on Ca/sup 2 +/ homeostasis, it was synthesized enzymatically from (/sup 3/H)-Ins(1,4,5)P/sub 3/ using a partially purified phosphoinositol kinase activity from rat brain cortex. (/sup 3/H)-labeled inositol phosphates were separated by anion exchange chromatography and analyzed by HPLC using ammonium formate/phosphoric acid gradient elution. Preliminary experiments indicate that Ins(1,3,4,5)P/sub 4/ up to 10 ..mu..M does not release Ca/sup 2 +/ from vesicular pools in saponin-permeabilized hepatocytes. It has a slight inhibitory effect on Ins(1,4,5)P/sub 3/-induced Ca/sup 2 +/ release. The effect of Ins(1,3,4,5)P/sub 4/ on plasma membrane Ca/sup 2 +/ fluxes are presently being investigated.

  20. Inositol-trisphosphate reduces alveolar apoptosis and pulmonary edema in neonatal lung injury.

    PubMed

    Preuss, Stefanie; Stadelmann, Sabrina; Omam, Friede D; Scheiermann, Julia; Winoto-Morbach, Supandi; von Bismarck, Philipp; Knerlich-Lukoschus, Friederike; Lex, Dennis; Adam-Klages, Sabine; Wesch, Daniela; Held-Feindt, Janka; Uhlig, Stefan; Schütze, Stefan; Krause, Martin F

    2012-08-01

    D-myo-inositol-1,2,6-trisphosphate (IP3) is an isomer of the naturally occurring second messenger D-myo-inositol-1,4,5-trisphosphate, and exerts anti-inflammatory and antiedematous effects in the lung. Myo-inositol (Inos) is a component of IP3, and is thought to play an important role in the prevention of neonatal pulmonary diseases such as bronchopulmonary dysplasia and neonatal acute lung injury (nALI). Inflammatory lung diseases are characterized by augmented acid sphingomyelinase (aSMase) activity leading to ceramide production, a pathway that promotes increased vascular permeability, apoptosis, and surfactant alterations. A novel, clinically relevant triple-hit model of nALI was developed, consisting of repeated airway lavage, injurious ventilation, and lipopolysaccharide instillation into the airways, every 24 hours. Thirty-five piglets were randomized to one of four treatment protocols: control (no intervention), surfactant alone, surfactant + Inos, and surfactant + IP3. After 72 hours of mechanical ventilation, lungs were excised from the thorax for subsequent analyses. Clinically, oxygenation and ventilation improved, and extravascular lung water decreased significantly with the S + IP3 intervention. In pulmonary tissue, we observed decreased aSMase activity and ceramide concentrations, decreased caspase-8 concentrations, reduced alveolar epithelial apoptosis, the reduced expression of interleukin-6, transforming growth factor-β1, and amphiregulin (an epithelial growth factor), reduced migration of blood-borne cells and particularly of CD14(+)/18(+) cells (macrophages) into the airspaces, and lower surfactant surface tensions in S + IP3-treated but not in S + Inos-treated piglets. We conclude that the admixture of IP3 to surfactant, but not of Inos, improves gas exchange and edema in our nALI model by the suppression of the governing enzyme aSMase, and that this treatment deserves clinical evaluation. PMID:22403805

  1. Genome-wide screen for inositol auxotrophy in Saccharomyces cerevisiae implicates lipid metabolism in stress response signaling

    PubMed Central

    Villa-García, Manuel J.; Choi, Myung Sun; Hinz, Flora I.; Gaspar, María L.; Jesch, Stephen A.

    2011-01-01

    Inositol auxotrophy (Ino− phenotype) in budding yeast has classically been associated with misregulation of INO1 and other genes involved in lipid metabolism. To identify all non-essential yeast genes that are necessary for growth in the absence of inositol, we carried out a genome-wide phenotypic screening for deletion mutants exhibiting Ino− phenotypes under one or more growth conditions. We report the identification of 419 genes, including 385 genes not previously reported, which exhibit this phenotype when deleted. The identified genes are involved in a wide range of cellular processes, but are particularly enriched in those affecting transcription, protein modification, membrane trafficking, diverse stress responses, and lipid metabolism. Among the Ino− mutants involved in stress response, many exhibited phenotypes that are strengthened at elevated temperature and/or when choline is present in the medium. The role of inositol in regulation of lipid metabolism and stress response signaling is discussed. PMID:21136082

  2. Myo-inositol esters of indole-3-acetic acid are endogenous components of Zea mays L. shoot tissue

    NASA Technical Reports Server (NTRS)

    Chisnell, J. R.

    1984-01-01

    Indole-3-acetyl-myo-inositol esters have been demonstrated to be endogenous components of etiolated Zea mays shoots tissue. This was accomplished by comparison of the putative compounds with authentic, synthetic esters. The properties compared were liquid and gas-liquid chromatographic retention times and the 70-ev mass spectral fragmentation pattern of the pentaacetyl derivative. The amount of indole-3-acetyl-myo-inositol esters in the shoots was determined to be 74 nanomoles per kilogram fresh weight as measured by isotope dilution, accounting for 19% of the ester indole-3-acetic acid of the shoot. This work is the first characterization of an ester conjugate of indole-3-acetate acid from vegetative shoot tissue using multiple chromatographic properties and mass spectral identification. The kernel and the seedling shoot both contain indole-3-acetyl-myo-inositol esters, and these esters comprise approximately the same percentage of the total ester content of the kernel and of the shoot.

  3. A cold-induced myo-inositol transporter-like gene confers tolerance to multiple abiotic stresses in transgenic tobacco plants.

    PubMed

    Sambe, Mame Abdou Nahr; He, Xueying; Tu, Qinghua; Guo, Zhenfei

    2015-03-01

    A full length cDNA encoding a myo-inositol transporter-like protein, named as MfINT-like, was cloned from Medicago sativa subsp. falcata (herein falcata), a species with greater cold tolerance than alfalfa (M. sativa subsp. sativa). MfINT-like is located on plasma membranes. MfINT-like transcript was induced 2-4 h after exogenous myo-inositol treatment, 24-96 h with cold, and 96 h by salinity. Given that myo-inositol accumulates higher in falcata after 24 h of cold treatment, myo-inositol is proposed to be involved in cold-induced expression of MfINT-like. Higher levels of myo-inositol was observed in leaves of transgenic tobacco plants overexpressing MfINT-like than the wild-type but not in the roots of plants grown on myo-inositol containing medium, suggesting that transgenic plants had higher myo-inositol transport activity than the wild-type. Transgenic plants survived better to freezing temperature, and had lower ion leakage and higher maximal photochemical efficiency of photosystem II (Fv /Fm ) after chilling treatment. In addition, greater plant fresh weight was observed in transgenic plants as compared with the wild-type when plants were grown under drought or salinity stress. The results suggest that MfINT-like mediated transport of myo-inositol is associated with plant tolerance to abiotic stresses. PMID:25131886

  4. Rv2131c gene product: An unconventional enzyme that is both inositol monophosphatase and fructose-1,6-bisphosphatase

    SciTech Connect

    Gu Xiaoling; Chen Mao; Shen Hongbo; Jiang Xin; Huang Yishu; Wang Honghai . E-mail: hhwang@fudan.edu.cn

    2006-01-20

    Inositol monophosphatase is an enzyme in the biosynthesis of myo-inostiol, a crucial substrate for the synthesis of phosphatidylinositol, which has been demonstrated to be an essential component of mycobacteria. In this study, the Rv2131c gene from Mycobacterium tuberculosis H37Rv was cloned into the pET28a vector and the recombinant plasmid was transformed into Escherichia coli BL21 (DE3) strain, allowing the expression of the enzyme in fusion with a histidine-rich peptide on the N-terminal. The fusion protein was purified from the soluble fraction of the lysed cells under native conditions by immobilized metal affinity chromatography (IMAC). The purified Rv2131c gene product showed inositol monophosphatase activity but with substrate specificity that was broader than those of several bacterial and eukaryotic inositol monophosphatases, and it also acted as fructose-1,6-bisphosphatase. The dimeric enzyme exhibited dual activities of IMPase and FBPase, with K {sub m} of 0.22 {+-} 0.03 mM for inositol-1-phosphate and K {sub m} of 0.45 {+-} 0.05 mM for fructose-1,6-bisphosphatase. To better understand the relationship between the function and structure of the Rv2131c enzyme, we constructed D40N, L71A, and D94N mutants and purified these corresponding proteins. Mutations of D40N and D94N caused the proteins to almost completely lose both the inositol monophosphatase and fructose-1,6-bisphosphatase activities. However, L71A mutant did not cause loss either of the activities, but the activity toward the inositol was 12-fold more resistant to inhibition by lithium (IC{sub 5} {approx} 60 mM). Based on the substrate specificity and presence of conserved sequence motifs of the M. tuberculosis Rv2131c, we proposed that the enzyme belonged to class IV fructose-1,6-bisphosphatase (FBPase IV)

  5. Inositol 1,3,4,5,6-Pentakisphosphate 2-Kinase from Maize: Molecular and Biochemical Characterization[OA

    PubMed Central

    Sun, Yuejin; Thompson, Mark; Lin, Gaofeng; Butler, Holly; Gao, Zhifang; Thornburgh, Scott; Yau, Kerrm; Smith, Doug A.; Shukla, Vipula K.

    2007-01-01

    Inositol 1,3,4,5,6-pentakisphosphate 2-kinase, an enzyme encoded by the gene IPK1, catalyzes the terminal step in the phytic acid biosynthetic pathway. We report here the isolation and characterization of IPK1 cDNA and genomic clones from maize (Zea mays). DNA Southern-blot analysis revealed that ZmIPK1 in the maize genome constitutes a small gene family with two members. Two nearly identical ZmIPK1 paralogs, designated as ZmIPK1A and ZmIPK1B, were identified. The transcripts of ZmIPK1A were detected in various maize tissues, including leaves, silks, immature ears, seeds at 12 d after pollination, midstage endosperm, and maturing embryos. However, the transcripts of ZmIPK1B were exclusively detected in roots. A variety of alternative splicing products of ZmIPK1A were discovered in maize leaves and seeds. These products are derived from alternative acceptor sites, alternative donor sites, and retained introns in the transcripts. Consequently, up to 50% of the ZmIPK1A transcripts in maize seeds and leaves have an interrupted open reading frame. In contrast, only one type of splicing product of ZmIPK1B was detected in roots. When expressed in Escherichia coli and subsequently purified, the ZmIPK1 enzyme catalyzes the conversion of myo-inositol 1,3,4,5,6-pentakisphosphate to phytic acid. In addition, it is also capable of catalyzing the phosphorylation of myo-inositol 1,4,6-trisphosphate, myo-inositol 1,4,5,6-tetrakisphosphate, and myo-inositol 3,4,5,6-tetrakisphosphate. Nuclear magnetic resonance spectroscopy analysis indicates that the phosphorylation product of myo-inositol 1,4,6-trisphosphate is inositol 1,2,4,6-tetrakisphosphate. Kinetic studies showed that the Km for ZmIPK1 using myo-inositol 1,3,4,5,6-pentakisphosphate as a substrate is 119 μm with a Vmax at 625 nmol/min/mg. These data describing the tissue-specific accumulation and alternative splicing of the transcripts from two nearly identical ZmIPK1 paralogs suggest that maize has a highly sophisticated

  6. Diastereoselective synthesis of new O-alkylated and C-branched inositols and their corresponding fluoro analogues

    PubMed Central

    Chrétien, Françoise; Chapleur, Yves

    2016-01-01

    Summary Efficient routes were developed for the diastereoselective synthesis of new O-alkylated and C-branched inositols and their corresponding fluoro analogues. The key steps of the synthesis were the easy accessibility of different types of arms in term of configuration (myo and scyllo), the linking method and length, which could modulate the biological properties. These inositol derivatives, bearing an arm terminated either with a hydroxy group or a fluorine atom, could be interesting candidates for diastereoisomeric intermediates and biological evaluations, especially for PET imaging experiments. PMID:26977196

  7. Data on the optimization of a GC-MS procedure for the determination of total plasma myo-inositol.

    PubMed

    Guo, Jin; Shi, Yingfei; Xu, Chengbao; Zhong, Rugang; Zhang, Feng; Niu, Bo; Wang, Jianhua; Zhang, Ting

    2016-09-01

    Myo-inositol (MI) is one of the stereoisomers of hexahydroxycyclohexane, which plays an important role in intracellular signal pathway. Derivatization is an indispensable step in both external and internal standard method during the chromatography-mass spectrometer (GC-MS) detection, as MI can't be ionized directly. It is valuable to optimize the derivative process and the detection volume for clinical detection. This article contains optimization data related to research publication "Quantification of plasma myo-inositol using gas chromatography-mass spectrometry" [1]. Here we introduce the data on the optimized derivatization volume, temperature, duration and the detection volume. PMID:27508261

  8. CHARMM Additive All-Atom Force Field for Acyclic Polyalcohols, Acyclic Carbohydrates and Inositol

    PubMed Central

    Hatcher, Elizabeth; Guvench, Olgun; MacKerell, Alexander D.

    2009-01-01

    Parametrization of the additive all-atom CHARMM force field for acyclic polyalcohols, acyclic carbohydrates and inositol is conducted. Initial parameters were transferred from the alkanes and hexopyranose carbohydrates, with subsequent development and optimization of parameters unique to the molecules considered in this study. Using the model compounds acetone and acetaldehyde, nonbonded parameters for carbonyls were optimized targeting quantum mechanical interaction data for solute-water pairs and pure solvent thermodynamic data. Bond and angle parameters were adjusted by comparing optimized geometries to small molecule crystal survey data and by performing vibrational analyses on acetone, acetaldehyde and glycerol. C-C-C-C, C-C-C-O, C-C-OH and O-C-C-O torsional parameters for polyol chains were fit to quantum mechanical dihedral potential energy scans comprising over 1500 RIMP2/cc-pVTZ//MP2/6-31G(d) conformations using an automated Monte Carlo simulated annealing procedure. Comparison of computed condensed-phase data, including crystal lattice parameters and densities, NMR proton-proton couplings, densities and diffusion coefficients of aqueous solutions, to experimental data validated the optimized parameters. Parameter development for these compounds proved particularly challenging because of the flexibility of the acyclic sugars and polyalcohols as well as the intramolecular hydrogen bonding between vicinal hydroxyls for all of the compounds. The newly optimized additive CHARMM force field parameters are anticipated to be of utility for atomic level of detail simulations of acyclic polyalcohols, acyclic carbohydrates and inositol in solution. PMID:20160980

  9. INOSITOL HEXAKISPHOSPHATE MEDIATES APOPTOSIS IN HUMAN BREAST ADENOCARCINOMA MCF-7 CELL LINE VIA INTRINSIC PATHWAY

    SciTech Connect

    Agarwal, Rakhee; Ali, Nawab

    2010-04-12

    Inositol polyphosphates (InsP{sub s}) are naturally occurring compounds ubiquitously present in plants and animals. Inositol hexakisphosphate (InsP{sub 6}) is the most abundant among all InsP{sub s} and constitutes the major portion of dietary fiber in most cereals, legumes and nuts. Certain derivatives of InsP{sub s} also regulate cellular signaling mechanisms. InsP{sub s} have also been shown to reduce tumor formation and induce apoptosis in cancerous cells. Therefore, in this study, the effects of InsPs on apoptosis were studied in an attempt to investigate their potential anti-cancer therapeutic application and understand their mechanism of action. Acridine orange and ethidium bromide staining suggested that InsP{sub 6} dose dependently induced apoptosis in human breast adenocarcinoma MCF-7 cells. Among InsP{sub s} tested (InsP{sub 3}, InsP{sub 4}, InsP{sub 5}, and InsP{sub 6}), InsP{sub 6} was found to be the most effective in inducing apoptosis. Furthermore, effects of InsP{sub 6} were found most potent inducing apoptosis. Etoposide, the drug known to induce apoptosis in both in vivo and in vitro, was used as a positive control. Western blotting experiments using specific antibodies against known apoptotic markers suggested that InsP{sub 6} induced apoptotic changes were mediated via an intrinsic apoptotic pathway.

  10. The role of the inositol phosphate cascade in visual excitation of invertebrate microvillar photoreceptors

    PubMed Central

    1991-01-01

    The identity of the transmitter(s) involved in visual transduction in invertebrate microvillar photoreceptors remains unresolved. In this study, the role of inositol 1,4,5-trisphosphate (IP3) was examined in Limulus ventral photoreceptors by studying the effects on the light response of heparin and neomycin, agents that inhibit the production or action of IP3. Both heparin and neomycin reduce responses to brief flashes of light and the transient component of responses to steps of light, and also inhibit IP3-induced calcium release, indicating that IP3 plays a direct role in invertebrate visual excitation. The effects of BAPTA, a calcium buffer, were also examined and shown to be consistent with a role for IP3-mediated calcium release in visual excitation. However, all three agents fail to block the plateau component of the response to a step of light, indicating that a single pathway involving IP3 and calcium cannot solely be responsible for visual excitation in invertebrates. We suggest that the inositol phosphate cascade and a second parallel process that is not dependent on IP3 are involved in the production of the light response. PMID:1905344

  11. Sac2/INPP5F is an inositol 4-phosphatase that functions in the endocytic pathway

    PubMed Central

    Nakatsu, Fubito; Messa, Mirko; Nández, Ramiro; Czapla, Heather; Zou, Yixiao; Strittmatter, Stephen M.

    2015-01-01

    The recruitment of inositol phosphatases to endocytic membranes mediates dephosphorylation of PI(4,5)P2, a phosphoinositide concentrated in the plasma membrane, and prevents its accumulation on endosomes. The importance of the conversion of PI(4,5)P2 to PtdIns during endocytosis is demonstrated by the presence of both a 5-phosphatase and a 4-phosphatase (Sac domain) module in the synaptojanins, endocytic PI(4,5)P2 phosphatases conserved from yeast to humans and the only PI(4,5)P2 phosphatases in yeast. OCRL, another 5-phosphatase that couples endocytosis to PI(4,5)P2 dephosphorylation, lacks a Sac domain. Here we show that Sac2/INPP5F is a PI4P phosphatase that colocalizes with OCRL on endocytic membranes, including vesicles formed by clathrin-mediated endocytosis, macropinosomes, and Rab5 endosomes. An OCRL–Sac2/INPP5F interaction could be demonstrated by coimmunoprecipitation and was potentiated by Rab5, whose activity is required to recruit Sac2/INPP5F to endosomes. Sac2/INPP5F and OCRL may cooperate in the sequential dephosphorylation of PI(4,5)P2 at the 5 and 4 position of inositol in a partnership that mimics that of the two phosphatase modules of synaptojanin. PMID:25869668

  12. Inositol phosphate kinase 2 is required for imaginal disc development in Drosophila.

    PubMed

    Seeds, Andrew M; Tsui, Marco M; Sunu, Christine; Spana, Eric P; York, John D

    2015-12-22

    Inositol phosphate kinase 2 (Ipk2), also known as IP multikinase IPMK, is an evolutionarily conserved protein that initiates production of inositol phosphate intracellular messengers (IPs), which are critical for regulating nuclear and cytoplasmic processes. Here we report that Ipk2 kinase activity is required for the development of the adult fruit fly epidermis. Ipk2 mutants show impaired development of their imaginal discs, the primordial tissues that form the adult epidermis. Although disk tissue seems to specify normally during early embryogenesis, loss of Ipk2 activity results in increased apoptosis and impairment of proliferation during larval and pupal development. The proliferation defect is in part attributed to a reduction in JAK/STAT signaling, possibly by controlling production or secretion of the pathway's activating ligand, Unpaired. Constitutive activation of the JAK/STAT pathway downstream of Unpaired partially rescues the disk growth defects in Ipk2 mutants. Thus, IP production is essential for proliferation of the imaginal discs, in part, by regulating JAK/STAT signaling. Our work demonstrates an essential role for Ipk2 in producing inositide messengers required for imaginal disk tissue maturation and subsequent formation of adult body structures and provides molecular insights to signaling pathways involved in tissue growth and stability during development. PMID:26647185

  13. Inositol hexakisphosphate kinase-1 interacts with perilipin1 to modulate lipolysis.

    PubMed

    Ghoshal, Sarbani; Tyagi, Richa; Zhu, Qingzhang; Chakraborty, Anutosh

    2016-09-01

    Lipolysis leads to the breakdown of stored triglycerides (TAG) to release free fatty acids (FFA) and glycerol which is utilized by energy expenditure pathways to generate energy. Therefore, a decrease in lipolysis augments fat accumulation in adipocytes which promotes weight gain. Conversely, if lipolysis is not complemented by energy expenditure, it leads to FFA induced insulin resistance and type-2 diabetes. Thus, lipolysis is under stringent physiological regulation, although the precise mechanism of the regulation is not known. Deletion of inositol hexakisphosphate kinase-1 (IP6K1), the major inositol pyrophosphate biosynthetic enzyme, protects mice from high fat diet (HFD) induced obesity and insulin resistance. IP6K1-KO mice are lean due to enhanced energy expenditure. Therefore, IP6K1 is a target in obesity and type-2 diabetes. However, the mechanism/s by which IP6K1 regulates adipose tissue lipid metabolism is yet to be understood. Here, we demonstrate that IP6K1-KO mice display enhanced basal lipolysis. IP6K1 modulates lipolysis via its interaction with the lipolytic regulator protein perilipin1 (PLIN1). Furthermore, phosphorylation of IP6K1 at a PKC/PKA motif modulates its interaction with PLIN1 and lipolysis. Thus, IP6K1 is a novel regulator of PLIN1 mediated lipolysis. PMID:27373682

  14. Inositol phosphate kinase 2 is required for imaginal disc development in Drosophila

    PubMed Central

    Seeds, Andrew M.; Tsui, Marco M.; Sunu, Christine; Spana, Eric P.; York, John D.

    2015-01-01

    Inositol phosphate kinase 2 (Ipk2), also known as IP multikinase IPMK, is an evolutionarily conserved protein that initiates production of inositol phosphate intracellular messengers (IPs), which are critical for regulating nuclear and cytoplasmic processes. Here we report that Ipk2 kinase activity is required for the development of the adult fruit fly epidermis. Ipk2 mutants show impaired development of their imaginal discs, the primordial tissues that form the adult epidermis. Although disk tissue seems to specify normally during early embryogenesis, loss of Ipk2 activity results in increased apoptosis and impairment of proliferation during larval and pupal development. The proliferation defect is in part attributed to a reduction in JAK/STAT signaling, possibly by controlling production or secretion of the pathway’s activating ligand, Unpaired. Constitutive activation of the JAK/STAT pathway downstream of Unpaired partially rescues the disk growth defects in Ipk2 mutants. Thus, IP production is essential for proliferation of the imaginal discs, in part, by regulating JAK/STAT signaling. Our work demonstrates an essential role for Ipk2 in producing inositide messengers required for imaginal disk tissue maturation and subsequent formation of adult body structures and provides molecular insights to signaling pathways involved in tissue growth and stability during development. PMID:26647185

  15. Inositol lipids: from an archaeal origin to phosphatidylinositol 3,5-bisphosphate faults in human disease.

    PubMed

    Michell, Robert H

    2013-12-01

    The last couple of decades have seen an extraordinary transformation in our knowledge and understanding of the multifarious biological roles of inositol phospholipids. Herein, I briefly consider two topics. The first is the role that recently acquired biochemical and genomic information - especially from archaeons - has played in illuminating the possible evolutionary origins of the biological employment of inositol in lipids, and some questions that these studies raise about the 'classical' biosynthetic route to phosphatidylinositol. The second is the growing recognition of the importance in eukaryotic cells of phosphatidylinositol 3,5-bisphosphate. Phosphatidylinositol 3,5-bisphosphate only entered our phosphoinositide consciousness quite recently, but it is speedily gathering a plethora of roles in diverse cellular processes and diseases thereof. These include: control of endolysosomal vesicular trafficking and of the activity of ion channels and pumps in the endolysosomal compartment; control of constitutive and stimulated protein traffic to and from plasma membrane subdomains; control of the nutrient and stress-sensing target of rapamycin complex 1 pathway (TORC1); and regulation of key genes in some central metabolic pathways. PMID:23902363

  16. Chemogenetic Characterization of Inositol Phosphate Metabolic Pathway Reveals Druggable Enzymes for Targeting Kinetoplastid Parasites.

    PubMed

    Cestari, Igor; Haas, Paige; Moretti, Nilmar Silvio; Schenkman, Sergio; Stuart, Ken

    2016-05-19

    Kinetoplastids cause Chagas disease, human African trypanosomiasis, and leishmaniases. Current treatments for these diseases are toxic and inefficient, and our limited knowledge of drug targets and inhibitors has dramatically hindered the development of new drugs. Here we used a chemogenetic approach to identify new kinetoplastid drug targets and inhibitors. We conditionally knocked down Trypanosoma brucei inositol phosphate (IP) pathway genes and showed that almost every pathway step is essential for parasite growth and infection. Using a genetic and chemical screen, we identified inhibitors that target IP pathway enzymes and are selective against T. brucei. Two series of these inhibitors acted on T. brucei inositol polyphosphate multikinase (IPMK) preventing Ins(1,4,5)P3 and Ins(1,3,4,5)P4 phosphorylation. We show that IPMK is functionally conserved among kinetoplastids and that its inhibition is also lethal for Trypanosoma cruzi. Hence, IP enzymes are viable drug targets in kinetoplastids, and IPMK inhibitors may aid the development of new drugs. PMID:27133314

  17. Potentiometric and spectroscopic study of the interaction of 3d transition metal ions with inositol hexakisphosphate

    NASA Astrophysics Data System (ADS)

    Veiga, Nicolás; Macho, Israel; Gómez, Kerman; González, Gabriel; Kremer, Carlos; Torres, Julia

    2015-10-01

    Among myo-inositol phosphates, the most abundant in nature is the myo-inositol hexakisphosphate, InsP6. Although it is known to be vital to cell functioning, the biochemical research into its metabolism needs chemical and structural analysis of all the protonation, complexation and precipitation processes that it undergoes in the biological media. In view of its high negative charge at physiological level, our group has been leading a thorough research into the InsP6 chemical and structural behavior in the presence of the alkali and alkaline earth metal ions essential for life. The aim of this article is to extend these studies, dealing with the chemical and structural features of the InsP6 interaction with biologically relevant 3d transition metal ions (Fe(II), Fe(III), Mn(II), Co(II), Ni(II), Cu(II) and Zn(II)), in a non-interacting medium and under simulated physiological conditions. The metal-complex stability constants were determined by potentiometry, showing under ligand-excess conditions the formation of mononuclear species in different protonation states. Under metal ion excess, polymetallic species were detected for Fe(II), Fe(III), Zn(II) and Cu(II). Additionally, the 31P NMR and UV-vis spectroscopic studies provided interesting structural aspects of the strong metal ion-InsP6 interaction.

  18. Inositol Hexakisphosphate Mediates Apoptosis in Human Breast Adenocarcinoma MCF-7 Cell Line via Intrinsic Pathway

    NASA Astrophysics Data System (ADS)

    Agarwal, Rakhee; Ali, Nawab

    2010-04-01

    Inositol polyphosphates (InsPs) are naturally occurring compounds ubiquitously present in plants and animals. Inositol hexakisphosphate (InsP6) is the most abundant among all InsPs and constitutes the major portion of dietary fiber in most cereals, legumes and nuts. Certain derivatives of InsPs also regulate cellular signaling mechanisms. InsPs have also been shown to reduce tumor formation and induce apoptosis in cancerous cells. Therefore, in this study, the effects of InsPs on apoptosis were studied in an attempt to investigate their potential anti-cancer therapeutic application and understand their mechanism of action. Acridine orange and ethidium bromide staining suggested that InsP6 dose dependently induced apoptosis in human breast adenocarcinoma MCF-7 cells. Among InsPs tested (InsP3, InsP4, InsP5, and InsP6), InsP6 was found to be the most effective in inducing apoptosis. Furthermore, effects of InsP6 were found most potent inducing apoptosis. Etoposide, the drug known to induce apoptosis in both in vivo and in vitro, was used as a positive control. Western blotting experiments using specific antibodies against known apoptotic markers suggested that InsP6 induced apoptotic changes were mediated via an intrinsic apoptotic pathway.

  19. Myo-Inositol in the Treatment of Teenagers Affected by PCOS

    PubMed Central

    Barbakadze, Ludmila; Kvashilava, Nana

    2016-01-01

    Objective. To compare the effectiveness of myo-inositol (MI) and oral contraceptive pills (OCPs) in monotherapy and MI in combination with OCPs in the treatment of teenagers affected by polycystic ovary syndrome (PCOS). Methods. 61 adolescent girls aged 13–19 years, with PCOS, were involved in the prospective, open-label study. Patients were randomized into three groups: I group, 20 patients receiving drospirenone 3 mg/ethinyl estradiol 30 μg; II group, 20 patients receiving 4 g myo-inositol plus 400 mg folic acid; III group, 21 patients receiving both medications. Results. After receiving MI significant reduction in weight, BMI, glucose, C-peptide, insulin, HOMA-IR, FT, and LH was detected. The levels of SHBG, TT, FAI, DHEA-S, and AMH did not change statistically significantly. After receiving OCPs weight and BMI slightly increased, but metabolic parameters did not change. Combination of MI and OCPs did not change weight and BMI, but reduction in C-peptide, insulin, and HOMA-IR was detected. TT, FT, FAI, DHEA-S, LH, and AMH levels decreased and SHBG increased. Conclusions. Administration of MI is a safe and effective method to prevent and correct metabolic disorders in teenagers affected by PCOS. With combination of MI and OCPs antiandrogenic effects are enhanced, negative impact of OCPs on weight gain is balanced, and metabolic profile is improved.

  20. Brevis plant1, a putative inositol polyphosphate 5-phosphatase, is required for internode elongation in maize

    PubMed Central

    Avila, Luis M.; Cerrudo, Diego; Swanton, Clarence

    2016-01-01

    In maize (Zea mays L.), as in other grass species, stem elongation occurs during growth and most noticeably upon the transition to flowering. Genes that reduce stem elongation have been important to reduce stem breakage, or lodging. Stem elongation has been mediated by dwarf and brachytic/brevis plant mutants that affect giberellic acid and auxin pathways, respectively. Maize brevis plant1 (bv1) mutants, first identified over 80 years ago, strongly resemble brachytic2 mutants that have shortened internodes, short internode cells, and are deficient in auxin transport. Here, we characterized two novel bv1 maize mutants. We found that an inositol polyphosphate 5-phosphatase orthologue of the rice gene dwarf50 was the molecular basis for the bv1 phenotype, implicating auxin-mediated inositol polyphosphate and/or phosphoinositide signalling in stem elongation. We suggest that auxin-mediated internode elongation involves processes that also contribute to stem gravitropism. Genes misregulated in bv1 mutants included genes important for cell wall synthesis, transmembrane transport, and cytoskeletal function. Mutant and wild-type plants were indistinguishable early in development, responded similarly to changes in light quality, had unaltered flowering times, and had normal flower development. These attributes suggest that breeding could utilize bv1 alleles to increase crop grain yields. PMID:26767748

  1. Kinetic crystallization separation process of the inositol isomers by controlling metastable zones

    NASA Astrophysics Data System (ADS)

    Konuki, Kaname; Hirasawa, Izumi

    2013-06-01

    D-chiro-inositol (DCI) is prepared by the immobilized enzyme reaction which uses myo-inositol (MI) as the substrate and the conversion rate is about 13%. The aim of this study was to develop a separation method for high purity DCI crystals from a reaction solution including low purity DCI only by the crystallization process. We succeeded in separating DCI crystals of 96% purity by water cooling crystallization, but it was presumed that scale-up was difficult. Although we tried anti-solvent crystallization similar to water cooling crystallization, high purity DCI crystals were not obtained. Therefore, we proposed the crystallization separation process by controlling metastable zones. The purity of a desired compound is controlled by this process, because solid-liquid separation is achieved before crystallization of compound in metastable zone. By the crystallization using this method, the DCI crystals of 97% purity were obtained. Although the yield per batch is about 50%, the actual yield is improved as the last mother liquor returns into the process of the following batch. When this process was repeated, the purity and the yield of DCI were reproduced and the robustness of this process was proved. It is expected that scale-up of this process will be successful, and this purification method could be applicable to similar systems such as separation of isomers and analogs.

  2. Brevis plant1, a putative inositol polyphosphate 5-phosphatase, is required for internode elongation in maize.

    PubMed

    Avila, Luis M; Cerrudo, Diego; Swanton, Clarence; Lukens, Lewis

    2016-03-01

    In maize (Zea mays L.), as in other grass species, stem elongation occurs during growth and most noticeably upon the transition to flowering. Genes that reduce stem elongation have been important to reduce stem breakage, or lodging. Stem elongation has been mediated by dwarf and brachytic/brevis plant mutants that affect giberellic acid and auxin pathways, respectively. Maize brevis plant1 (bv1) mutants, first identified over 80 years ago, strongly resemble brachytic2 mutants that have shortened internodes, short internode cells, and are deficient in auxin transport. Here, we characterized two novel bv1 maize mutants. We found that an inositol polyphosphate 5-phosphatase orthologue of the rice gene dwarf50 was the molecular basis for the bv1 phenotype, implicating auxin-mediated inositol polyphosphate and/or phosphoinositide signalling in stem elongation. We suggest that auxin-mediated internode elongation involves processes that also contribute to stem gravitropism. Genes misregulated in bv1 mutants included genes important for cell wall synthesis, transmembrane transport, and cytoskeletal function. Mutant and wild-type plants were indistinguishable early in development, responded similarly to changes in light quality, had unaltered flowering times, and had normal flower development. These attributes suggest that breeding could utilize bv1 alleles to increase crop grain yields. PMID:26767748

  3. Microtubule-Mediated Inositol Lipid Signaling Plays Critical Roles in Regulation of Blebbing

    PubMed Central

    Sugiyama, Tatsuroh; Pramanik, Md. Kamruzzaman; Yumura, Shigehiko

    2015-01-01

    Cells migrate by extending pseudopods such as lamellipodia and blebs. Although the signals leading to lamellipodia extension have been extensively investigated, those for bleb extension remain unclear. Here, we investigated signals for blebbing in Dictyostelium cells using a newly developed assay to induce blebbing. When cells were cut into two pieces with a microneedle, the anucleate fragments vigorously extended blebs. This assay enabled us to induce blebbing reproducibly, and analyses of knockout mutants and specific inhibitors identified candidate molecules that regulate blebbing. Blebs were also induced in anucleate fragments of leukocytes, indicating that this assay is generally applicable to animal cells. After cutting, microtubules in the anucleate fragments promptly depolymerized, followed by the extension of blebs. Furthermore, when intact cells were treated with a microtubule inhibitor, they frequently extended blebs. The depolymerization of microtubules induced the delocalization of inositol lipid phosphatidylinositol 3,4,5-trisphosphate from the cell membrane. PI3 kinase-null cells frequently extended blebs, whereas PTEN-null cells extended fewer blebs. From these observations, we propose a model in which microtubules play a critical role in bleb regulation via inositol lipid metabolism. PMID:26317626

  4. Quantification of myo-inositol, 1,5-anhydro- D-sorbitol, and D-chiro-inositol using high-performance liquid chromatography with electrochemical detection in very small volume clinical samples.

    PubMed

    Schimpf, Karen J; Meek, Claudia C; Leff, Richard D; Phelps, Dale L; Schmitz, Daniel J; Cordle, Christopher T

    2015-11-01

    Inositol is a six-carbon sugar alcohol and is one of nine biologically significant isomers of hexahydroxycyclohexane. Myo-inositol is the primary biologically active form and is present in higher concentrations in the fetus and newborn than in adults. It is currently being examined for the prevention of retinopathy of prematurity in newborn preterm infants. A robust method for quantifying myo-inositol (MI), D-chiro-inositol (DCI) and 1,5-anhydro- D-sorbitol (ADS) in very small-volume (25 μL) urine, blood serum and/or plasma samples was developed. Using a multiple-column, multiple mobile phase liquid chromatographic system with electrochemical detection, the method was validated with respect to (a) selectivity, (b) accuracy/recovery, (c) precision/reproducibility, (d) sensitivity, (e) stability and (f) ruggedness. The standard curve was linear and ranged from 0.5 to 30 mg/L for each of the three analytes. Above-mentioned performance measures were within acceptable limits described in the Food and Drug Administration's Guidance for Industry: Bioanalytical Method Validation. The method was validated using blood serum and plasma collected using four common anticoagulants, and also by quantifying the accuracy and sensitivity of MI measured in simulated urine samples recovered from preterm infant diaper systems. The method performs satisfactorily measuring the three most common inositol isomers on 25 μL clinical samples of serum, plasma, milk, and/or urine. Similar performance is seen testing larger volume samples of infant formulas and infant formula ingredients. MI, ADS and DCI may be accurately tested in urine samples collected from five different preterm infant diapers if the urine volume is greater than 2-5 mL. PMID:26010453

  5. Comparison of muscarine- and vasopressin-stimulated inositol phospholipid metabolism in the superior cervical ganglion of the rat

    SciTech Connect

    Horwitz, J.; Anderson, C.; Perlman, R.L.

    1986-03-05

    Both muscarine and vasopressin have previously been shown to increase the accumulation of /sup 3/H-inositol phosphates (/sup 3/H-IP) in superior cervical ganglia in which the phospholipids were labeled with /sup 3/H-inositol. They have compared the effects of muscarine and vasopressin on phospholipid metabolism in the ganglion. The effects of these agents on /sup 3/H-IP accumulation are additive. The response to muscarine plateaus after approximately 10 min whereas the response to vasopressin increases for at least 30 min. Decentralization and maintenance in organ culture appear to potentiate the effect of muscarine on /sup 3/H-IP accumulation but do not effect the response of the ganglia to vasopressin. Muscarine and vasopressin also increase the incorporation of /sup 3/H-inositol into phospholipids in the ganglion. Autoradiographic techniques were used to localize the inositol-containing phospholipids in the ganglion. Muscarine increases phospholipid labeling primarily in the cell bodies of the principal ganglionic neurons, whereas vasopressin increases phospholipid labeling primarily in the neuropil. These data are consistent with the hypothesis that muscarine and vasopressin stimulate the metabolism of different pools of phospholipids.

  6. Identification and Quantitation of Various Inositols and O-methylinositols Present in Plant Roots Using Gas Chromatograpghy/Mass Spectrometry

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Many inositols and O-methylinositols serve important roles in medicine and plant biology. A simple method was developed for the identification of these compounds in plant roots by extracting with 80% ethanol, derivatizing with trimethylsilyl imidazole, and analyzing by gas chromatography/mass spect...

  7. Role of Inositol Phosphosphingolipid Phospholipase C1, the Yeast Homolog of Neutral Sphingomyelinases in DNA Damage Response and Diseases.

    PubMed

    Tripathi, Kaushlendra

    2015-01-01

    Sphingolipids play a very crucial role in many diseases and are well-known as signaling mediators in many pathways. Sphingolipids are produced during the de novo process in the ER (endoplasmic reticulum) from the nonsphingolipid precursor and comprise both structural and bioactive lipids. Ceramide is the central core of the sphingolipid pathway, and its production has been observed following various treatments that can induce several different cellular effects including growth arrest, DNA damage, apoptosis, differentiation, and senescence. Ceramides are generally produced through the sphingomyelin hydrolysis and catalyzed by the enzyme sphingomyelinase (SMase) in mammals. Presently, there are many known SMases and they are categorized into three groups acid SMases (aSMases), alkaline SMases (alk-SMASES), and neutral SMases (nSMases). The yeast homolog of mammalians neutral SMases is inositol phosphosphingolipid phospholipase C. Yeasts generally have inositol phosphosphingolipids instead of sphingomyelin, which may act as a homolog of mammalian sphingomyelin. In this review, we shall explain the structure and function of inositol phosphosphingolipid phospholipase C1, its localization inside the cells, mechanisms, and its roles in various cell responses during replication stresses and diseases. This review will also give a new basis for our understanding for the mechanisms and nature of the inositol phosphosphingolipid phospholipase C1/nSMase. PMID:26346287

  8. Raman spectra of organic (myo-inositol hexakis phosphate) and inorganic P sepctra show pH dependence

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Understanding phosphorous fate and transport is in part limited by technical difficulties and/or access to expensive equipment associated with differentiating ortho-phosphate (P) from organic phosphate in complex environmental samples. Myo-inositol hexakis phosphate (IHP) is the most prevalent form...

  9. An improved Bacillus subtilis cell factory for producing scyllo-inositol, a promising therapeutic agent for Alzheimer’s disease

    PubMed Central

    2013-01-01

    Background Bacillus subtilis 168 possesses an efficient pathway to metabolize some of the stereoisomers of inositol, including myo-inositol (MI) and scyllo-inositol (SI). Previously we reported a prototype of a B. subtilis cell factory with modified inositol metabolism that converts MI into SI in the culture medium. However, it wasted half of initial 1.0% (w/v) MI, and the conversion was limited to produce only 0.4% (w/v) SI. To achieve a more efficient SI production, we attempted additional modifications. Results All “useless” genes involved in MI and SI metabolism were deleted. Although no elevation in SI production was observed in the deletion strain, it did result in no wastage of MI anymore. Thus additionally, overexpression of the key enzymes, IolG and IolW, was appended to demonstrate that simultaneous overexpression of them enabled complete conversion of all MI into SI. Conclusions The B. subtilis cell factory was improved to yield an SI production rate of 10 g/L/48 h at least. The improved conversion was achieved only in the presence of enriched nutrition in the form of 2% (w/v) Bacto soytone in the medium, which may be due to the increasing demand for regeneration of cofactors. PMID:24325193

  10. Nomenclature Committee of the International Union of Biochemistry (NC-IUB). Numbering of atoms in myo-inositol. Recommendations 1988.

    PubMed

    1989-04-01

    A relaxation of previous recommendations on the numbering of the atoms of myo-inositol is suggested: substituents need not necessarily be numbered so that the smallest possible locant is used. This allows an alternative designation to be used, when authors wish, to bring out structural relationships. PMID:2714267

  11. Numbering of atoms in myo-inositol. Recommendations 1988. Nomenclature Committee of the International Union of Biochemistry.

    PubMed

    1989-02-15

    A relaxation of previous recommendations on the numbering of the atoms of myo-inositol is suggested, so that substituents are not necessarily numbered so that the smallest possible locant is used. This allows an alternative designation to be used, when authors wish, to bring out structural relationships. PMID:2930499

  12. A limitation of the continuous spectrophotometric assay for the measurement of myo-inositol-1-phosphate synthase activity.

    PubMed

    Huang, Xinyi; Hernick, Marcy

    2011-10-15

    Myo-inositol-1-phosphate synthase (MIPS) catalyzes the conversion of glucose-6-phosphate to myo-inositol-1-phosphate. The reaction catalyzed by MIPS is the first step in the biosynthesis of inositol and inositol-containing molecules that serve important roles in both eukaryotes and prokaryotes. Consequently, MIPS is a target for the development of therapeutic agents for the treatment of infectious diseases and bipolar disorder. We recently reported a continuous spectrophotometric method for measuring MIPS activity using a coupled assay that allows the rapid characterization of MIPS in a multiwell plate format. Here we validate the continuous assay as a high-throughput alternative for measuring MIPS activity and report on one limitation of this assay-the inability to examine the effect of divalent metal ions (at high concentrations) on MIPS activity. In addition, we demonstrate that the activity of MIPS from Arabidopsis thaliana is moderately enhanced by the addition Mg(2+) and is not enhanced by other divalent metal ions (Zn(2+) and Mn(2+)), consistent with what has been observed for other eukaryotic MIPS enzymes. Our findings suggest that the continuous assay is better suited for characterizing eukaryotic MIPS enzymes that require monovalent cations as cofactors than for characterizing bacterial or archeal MIPS enzymes that require divalent metal ions as cofactors. PMID:21729692

  13. Supplementation of plant-based diets for rainbow trout, Oncorhynchus mykiss with macro-minerals and inositol.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Replacement of fish meal with plant products in aquafeeds results in the elimination of dietary compounds which may be important for optimal growth and physiology. A study was conducted to determine if supplementation with macro-minerals and/or inositol would improve performance of rainbow trout fe...

  14. Involvement of Inositol Biosynthesis and Nitric Oxide in the Mediation of UV-B Induced Oxidative Stress

    PubMed Central

    Lytvyn, Dmytro I.; Raynaud, Cécile; Yemets, Alla I.; Bergounioux, Catherine; Blume, Yaroslav B.

    2016-01-01

    The involvement of NO-signaling in ultraviolet B (UV-B) induced oxidative stress (OS) in plants is an open question. Inositol biosynthesis contributes to numerous cellular functions, including the regulation of plants tolerance to stress. This work reveals the involvement of inositol-3-phosphate synthase 1 (IPS1), a key enzyme for biosynthesis of myo-inositol and its derivatives, in the response to NO-dependent OS in Arabidopsis. Homozygous mutants deficient for IPS1 (atips1) and wild-type plants were transformed with a reduction- grx1-rogfp2 and used for the dynamic measurement of UV-B-induced and SNP (sodium nitroprusside)-mediated oxidative stresses by confocal microscopy. atips1 mutants displayed greater tissue-specific resistance to the action of UV-B than the wild type. SNP can act both as an oxidant or repairer depending on the applied concentration, but mutant plants were more tolerant than the wild type to nitrosative effects of high concentration of SNP. Additionally, pretreatment with low concentrations of SNP (10, 100 μM) before UV-B irradiation resulted in a tissue-specific protective effect that was enhanced in atips1. We conclude that the interplay between nitric oxide and inositol signaling can be involved in the mediation of UV-B-initiated oxidative stress in the plant cell. PMID:27148278

  15. Combination of inositol and alpha lipoic acid in metabolic syndrome-affected women: a randomized placebo-controlled trial

    PubMed Central

    2013-01-01

    Background Inositol has been reported to improve insulin sensitivity since it works as a second messenger achieving insulin-like effects on metabolic enzymes. The aim of this study was to evaluate the inositol and alpha lipoic acid combination effectiveness on metabolic syndrome features in postmenopausal women at risk of breast cancer. Methods A six-month prospective, randomized placebo-controlled trial was carried out on a total of 155 postmenopausal women affected by metabolic syndrome at risk of breast cancer, the INOSIDEX trial. All women were asked to follow a low-calorie diet and were assigned randomly to daily consumption of a combination of inositol and alpha lipoic acid (77 pts) or placebo (78 pts) for six months. Primary outcomes we wanted to achieve were both reduction of more than 20% of the HOMA-IR index and of triglycerides serum levels. Secondary outcomes expected were both the improvement of high-density lipoprotein cholesterol levels and the reduction of anthropometric features such as body mass index and waist-hip ratio. Results A significant HOMA-IR reduction of more than 20% was evidenced in 66.7% (P <0.0001) of patients, associated with a serum insulin level decrease in 89.3% (P <0.0000). A decrease in triglycerides was evidenced in 43.2% of patients consuming the supplement (P <0.0001). An increase in HDL cholesterol (48.6%) was found in the group consuming inositol with respect to the placebo group. A reduction in waist circumference and waist-hip ratio was found in the treated group with respect to the placebo group. Conclusions Inositol combined with alpha lipoic acid can be used as a dietary supplement in insulin-resistant patients in order to increase their insulin sensitiveness. Daily consumption of inositol combined with alpha lipoic acid has a significant bearing on metabolic syndrome. As metabolic syndrome is considered a modifiable risk factor of breast tumorigenesis, further studies are required to assess whether inositol combined

  16. Inositol trisphosphate 3-kinase B is increased in human Alzheimer brain and exacerbates mouse Alzheimer pathology.

    PubMed

    Stygelbout, Virginie; Leroy, Karelle; Pouillon, Valérie; Ando, Kunie; D'Amico, Eva; Jia, Yonghui; Luo, H Robert; Duyckaerts, Charles; Erneux, Christophe; Schurmans, Stéphane; Brion, Jean-Pierre

    2014-02-01

    ITPKB phosphorylates inositol 1,4,5-trisphosphate into inositol 1,3,4,5-tetrakisphosphate and controls signal transduction in various hematopoietic cells. Surprisingly, it has been reported that the ITPKB messenger RNA level is significantly increased in the cerebral cortex of patients with Alzheimer's disease, compared with control subjects. As extracellular signal-regulated kinases 1/2 activation is increased in the Alzheimer brain and as ITPKB is a regulator of extracellular signal-regulated kinases 1/2 activation in some hematopoietic cells, we tested whether this increased activation in Alzheimer's disease might be related to an increased activity of ITPKB. We show here that ITPKB protein level was increased 3-fold in the cerebral cortex of most patients with Alzheimer's disease compared with control subjects, and accumulated in dystrophic neurites associated to amyloid plaques. In mouse Neuro-2a neuroblastoma cells, Itpkb overexpression was associated with increased cell apoptosis and increased β-secretase 1 activity leading to overproduction of amyloid-β peptides. In this cellular model, an inhibitor of mitogen-activated kinase kinases 1/2 completely prevented overproduction of amyloid-β peptides. Transgenic overexpression of ITPKB in mouse forebrain neurons was not sufficient to induce amyloid plaque formation or tau hyperphosphorylation. However, in the 5X familial Alzheimer's disease mouse model, neuronal ITPKB overexpression significantly increased extracellular signal-regulated kinases 1/2 activation and β-secretase 1 activity, resulting in exacerbated Alzheimer's disease pathology as shown by increased astrogliosis, amyloid-β40 peptide production and tau hyperphosphorylation. No impact on pathology was observed in the 5X familial Alzheimer's disease mouse model when a catalytically inactive ITPKB protein was overexpressed. Together, our results point to the ITPKB/inositol 1,3,4,5-tetrakisphosphate/extracellular signal-regulated kinases 1

  17. Phosphotransferase system-independent glucose utilization in corynebacterium glutamicum by inositol permeases and glucokinases.

    PubMed

    Lindner, Steffen N; Seibold, Gerd M; Henrich, Alexander; Krämer, Reinhard; Wendisch, Volker F

    2011-06-01

    Phosphoenolpyruvate-dependent glucose phosphorylation via the phosphotransferase system (PTS) is the major path of glucose uptake in Corynebacterium glutamicum, but some growth from glucose is retained in the absence of the PTS. The growth defect of a deletion mutant lacking the general PTS component HPr in glucose medium could be overcome by suppressor mutations leading to the high expression of inositol utilization genes or by the addition of inositol to the growth medium if a glucokinase is overproduced simultaneously. PTS-independent glucose uptake was shown to require at least one of the inositol transporters IolT1 and IolT2 as a mutant lacking IolT1, IolT2, and the PTS component HPr could not grow with glucose as the sole carbon source. Efficient glucose utilization in the absence of the PTS necessitated the overexpression of a glucokinase gene in addition to either iolT1 or iolT2. IolT1 and IolT2 are low-affinity glucose permeases with K(s) values of 2.8 and 1.9 mM, respectively. As glucose uptake and phosphorylation via the PTS differs from glucose uptake via IolT1 or IolT2 and phosphorylation via glucokinase by the requirement for phosphoenolpyruvate, the roles of the two pathways for l-lysine production were tested. The l-lysine yield by C. glutamicum DM1729, a rationally engineered l-lysine-producing strain, was lower than that by its PTS-deficient derivate DM1729Δhpr, which, however, showed low production rates. The combined overexpression of iolT1 or iolT2 with ppgK, the gene for PolyP/ATP-dependent glucokinase, in DM1729Δhpr enabled l-lysine production as fast as that by the parent strain DM1729 but with 10 to 20% higher l-lysine yield. PMID:21478323

  18. Effect of co-solutes and process variables on crystallinity and the crystal form of freeze-dried myo-inositol.

    PubMed

    Izutsu, Ken-Ichi; Kusano, Riho; Arai, Ryoko; Yoshida, Hiroyuki; Ito, Masataka; Shibata, Hiroko; Sugano, Kiyohiko; Goda, Yukihiro; Terada, Katsuhide

    2016-07-25

    The purpose of this study was to elucidate how co-solutes affect the crystallization of small solute molecules during freeze-drying and subsequent storage. Crystallization profiles of myo-inositol and its mixture with dextran 40k in frozen solutions and dried solids were assessed by thermal analysis (DSC), powder-X-ray diffraction, and simultaneous DSC and PXRD analysis. Higher mass ratios of dextran maintained myo-inositol in the non-crystalline mixture state, in frozen solutions, during freeze-drying process, and exposure of dried solids to higher temperatures. Co-lyophilization with a lower mass ratio of dextran resulted in solids containing a variety of myo-inositol crystal forms and crystallinity depending on the composition and thermal history of the process. Heating of some inositol-rich amorphous solids showed crystallization of myo-inositol in the metastable form and its transition to stable form before melting. Heat-treatment of inositol-rich frozen solutions resulted in high crystallinity stable-form inositol solids, leaving dextran in the amorphous state. Sufficient direct molecular interactions (e.g., hydrogen bonding) should explain the stability of dextran-rich amorphous solids. Optimizing solute composition and processes should be a potent way to control crystal form and crystallinity of components in freeze-dried formulations. PMID:27282535

  19. The separation of ( sup 32 P)inositol phosphates by ion-pair chromatography: Optimization of the method and biological applications

    SciTech Connect

    Sulpice, J.C.; Gascard, P.; Journet, E.; Rendu, F.; Renard, D.; Poggioli, J.; Giraud, F. )

    1989-05-15

    We have developed an ion-pair reverse-phase HPLC method to measure inositol phosphates in {sup 32}P-labeled cells. The different chromatographic parameters were analyzed to optimize the resolution of the {sup 32}P-labeled metabolites. Analysis of inositol phosphates in biological samples was improved by a single charcoal pretreatment which eliminated interfering nucleotides without removing inositol phosphates. The kinetics of production of inositol phosphates in calcium-activated erythrocytes, vasopressin-stimulated hepatocytes, and thrombin-activated platelets were analyzed. Original data on the activation of phosphoinositide phospholipase C were obtained in intact erythrocytes by direct measurement of inositol (1,4,5)P3. Data from agonist-stimulated hepatocytes and platelets were consistent with those from previous studies. In conclusion, this technique offers many advantages over the methodologies currently employed involving anion-exchange chromatography and ({sup 3}H)inositol labeling: (i) {sup 32}P labeling is less expensive and more efficient than {sup 3}H labeling and can be used with all types of cells without permeabilization treatments and (ii) ion-pair HPLC gives good resolution of inositol phosphates from nucleotides with shorter retention times, and long reequilibration periods are not required.

  20. A Novel Inositol Pyrophosphate Phosphatase in Saccharomyces cerevisiae: Siw14 PROTEIN SELECTIVELY CLEAVES THE β-PHOSPHATE FROM 5-DIPHOSPHOINOSITOL PENTAKISPHOSPHATE (5PP-IP5).

    PubMed

    Steidle, Elizabeth A; Chong, Lucy S; Wu, Mingxuan; Crooke, Elliott; Fiedler, Dorothea; Resnick, Adam C; Rolfes, Ronda J

    2016-03-25

    Inositol pyrophosphates are high energy signaling molecules involved in cellular processes, such as energetic metabolism, telomere maintenance, stress responses, and vesicle trafficking, and can mediate protein phosphorylation. Although the inositol kinases underlying inositol pyrophosphate biosynthesis are well characterized, the phosphatases that selectively regulate their cellular pools are not fully described. The diphosphoinositol phosphate phosphohydrolase enzymes of the Nudix protein family have been demonstrated to dephosphorylate inositol pyrophosphates; however, theSaccharomyces cerevisiaehomolog Ddp1 prefers inorganic polyphosphate over inositol pyrophosphates. We identified a novel phosphatase of the recently discovered atypical dual specificity phosphatase family as a physiological inositol pyrophosphate phosphatase. Purified recombinant Siw14 hydrolyzes the β-phosphate from 5-diphosphoinositol pentakisphosphate (5PP-IP5or IP7)in vitro. In vivo,siw14Δ yeast mutants possess increased IP7levels, whereas heterologousSIW14overexpression eliminates IP7from cells. IP7levels increased proportionately whensiw14Δ was combined withddp1Δ orvip1Δ, indicating independent activity by the enzymes encoded by these genes. We conclude that Siw14 is a physiological phosphatase that modulates inositol pyrophosphate metabolism by dephosphorylating the IP7isoform 5PP-IP5to IP6. PMID:26828065

  1. A defect in sodium-dependent amino acid uptake in diabetic rabbit peripheral nerve. Correction by an aldose reductase inhibitor or myo-inositol administration.

    PubMed Central

    Greene, D A; Lattimer, S A; Carroll, P B; Fernstrom, J D; Finegold, D N

    1990-01-01

    A myo-inositol-related defect in nerve sodium-potassium ATPase activity in experimental diabetes has been suggested as a possible pathogenetic factor in diabetic neuropathy. Because the sodium-potassium ATPase is essential for other sodium-cotransport systems, and because myo-inositol-derived phosphoinositide metabolites regulate multiple membrane transport processes, sodium gradient-dependent amino acid uptake was examined in vitro in endoneurial preparations derived from nondiabetic and 14-d alloxan diabetic rabbits. Untreated alloxan diabetes reduced endoneurial sodium-gradient dependent uptake of the nonmetabolized amino acid 2-aminoisobutyric acid by greater than 50%. Administration of an aldose reductase inhibitor prevented reductions in both nerve myo-inositol content and endoneurial sodium-dependent 2-aminoisobutyric acid uptake. Myo-inositol supplementation that produced a transient pharmacological elevation in plasma myo-inositol concentration, but did not raise nerve myo-inositol content, reproduced the effect of the aldose reductase inhibitor on endoneurial sodium-dependent 2-aminoisobutyric acid uptake. Phorbol myristate acetate, which acutely normalizes sodium-potassium ATPase activity in diabetic nerve, did not acutely correct 2-aminoisobutyric uptake when added in vitro. These data suggest that depletion of a small myo-inositol pool may be implicated in the pathogenesis of defects in amino acid uptake in diabetic nerve and that rapid correction of sodium-potassium ATPase activity with protein kinase C agonists in vitro does not acutely normalize sodium-dependent 2-aminoisobutyric acid uptake. PMID:2185278

  2. The Phosphatidyl-myo-Inositol Mannosyltransferase PimA Is Essential for Mycobacterium tuberculosis Growth In Vitro and In Vivo

    PubMed Central

    Boldrin, Francesca; Ventura, Marcello; Degiacomi, Giulia; Ravishankar, Sudha; Sala, Claudia; Svetlikova, Zuzana; Ambady, Anisha; Dhar, Neeraj; Kordulakova, Jana; Zhang, Ming; Serafini, Agnese; Vishwas, V. G.; Kolly, Gaëlle S.; Kumar, Naveen; Palù, Giorgio; Guerin, Marcelo E.; Mikusova, Katarina; Cole, Stewart T.

    2014-01-01

    The cell envelope of Mycobacterium tuberculosis contains glycans and lipids of peculiar structure that play prominent roles in the biology and pathogenesis of tuberculosis. Consequently, the chemical structure and biosynthesis of the cell wall have been intensively investigated in order to identify novel drug targets. Here, we validate that the function of phosphatidyl-myo-inositol mannosyltransferase PimA is vital for M. tuberculosis in vitro and in vivo. PimA initiates the biosynthesis of phosphatidyl-myo-inositol mannosides by transferring a mannosyl residue from GDP-Man to phosphatidyl-myo-inositol on the cytoplasmic side of the plasma membrane. To prove the essential nature of pimA in M. tuberculosis, we constructed a pimA conditional mutant by using the TetR-Pip off system and showed that downregulation of PimA expression causes bactericidality in batch cultures. Consistent with the biochemical reaction catalyzed by PimA, this phenotype was associated with markedly reduced levels of phosphatidyl-myo-inositol dimannosides, essential structural components of the mycobacterial cell envelope. In addition, the requirement of PimA for viability was clearly demonstrated during macrophage infection and in two different mouse models of infection, where a dramatic decrease in viable counts was observed upon silencing of the gene. Notably, depletion of PimA resulted in complete clearance of the mouse lungs during both the acute and chronic phases of infection. Altogether, the experimental data highlight the importance of the phosphatidyl-myo-inositol mannoside biosynthetic pathway for M. tuberculosis and confirm that PimA is a novel target for future drug discovery programs. PMID:25049093

  3. Osmotic regulation and tissue localization of the myo-inositol biosynthesis pathway in tilapia (Oreochromis mossambicus) larvae.

    PubMed

    Sacchi, Romina; Gardell, Alison M; Chang, Nicole; Kültz, Dietmar

    2014-10-01

    The myo-inositol biosynthesis (MIB) pathway converts glucose-6-phosphate to the compatible osmolyte myo-inositol, which protects cells from salinity stress. We exposed tilapia larvae just after yolk sac resorption to various hypersaline environments and recorded robust induction of the enzymes that constitute the MIB pathway, myo-inositol-phosphate synthase (MIPS), and inositol monophosphatase 1 (IMPA1). Strong up-regulation of these enzymes is evident at both mRNA (quantitative real-time PCR) and protein (densitometric analysis of Western blots) levels. The highest level of induction of these enzymes occurs at the highest salinity that larvae were exposed to (90 ppt). Less severe salinity stress causes a proportionately reduced induction of the MIB pathway. Two distinct MIPS mRNA variants are present in tilapia larvae and both are induced at comparable levels for all the salinity challenges tested (34, 70, and 90 ppt). Immunohistochemical localization of IMPA1 protein in sagittal sections of salinity stressed and control larvae identified tissues that are particularly potent in inducing the MIB pathway. These tissues include the skin (epidermis), gills, eye (ciliary epithelium) and heart. In particular, the epidermis directly facing the external milieu showed a very strong induction of IMPA1 immunoreactivity. IMPA1 induction in response to salinity stress was not observed in other tissues suggesting that tilapia larvae may also utilize compatible organic osmolytes other than solely myo-inositol for osmoprotection. We conclude that the MIB pathway plays an important role in protecting multiple (but not all) tissues of tilapia larvae from hyperosmotic salinity stress. PMID:25045088

  4. Mechanism of myo-inositol hexakisphosphate sorption on amorphous aluminum hydroxide: spectroscopic evidence for rapid surface precipitation.

    PubMed

    Yan, Yupeng; Li, Wei; Yang, Jun; Zheng, Anmin; Liu, Fan; Feng, Xionghan; Sparks, Donald L

    2014-06-17

    Inositol hexakisphosphates are the most abundant organic phosphates (OPs) in most soils and sediments. Adsorption, desorption, and precipitation reactions at environmental interfaces govern the reactivity, speciation, mobility, and bioavailability of inositol hexakisphosphates in terrestrial and aquatic environments. However, surface complexation and precipitation reactions of inositol hexakisphosphates on soil minerals have not been well understood. Here we investigate the surface complexation-precipitation process and mechanism of myo-inositol hexakisphosphate (IHP, phytate) on amorphous aluminum hydroxide (AAH) using macroscopic sorption experiments and multiple spectroscopic tools. The AAH (16.01 μmol m(-2)) exhibits much higher sorption density than boehmite (0.73 μmol m(-2)) and α-Al2O3 (1.13 μmol m(-2)). Kinetics of IHP sorption and accompanying OH(-) release, as well as zeta potential measurements, indicate that IHP is initially adsorbed on AAH through inner-sphere complexation via ligand exchange, followed by AAH dissolution and ternary complex formation; last, the ternary complexes rapidly transform to surface precipitates and bulk phase analogous to aluminum phytate (Al-IHP). The pH level, reaction time, and initial IHP loading evidently affect the interaction of IHP on AAH. In situ ATR-FTIR and solid-state NMR spectra further demonstrate that IHP sorbs on AAH and transforms to surface precipitates analogous to Al-IHP, consistent with the results of XRD analysis. This study indicates that active metal oxides such as AAH strongly mediate the speciation and behavior of IHP via rapid surface complexation-precipitation reactions, thus controlling the mobility and bioavailability of inositol phosphates in the environment. PMID:24871399

  5. Fagopyritol B1, O-alpha-D-galactopyranosyl-(1-->2)-D-chiro-inositol, a galactosyl cyclitol in maturing buckwheat seeds associated with desiccation tolerance.

    PubMed

    Horbowicz, M; Brenac, P; Obendorf, R L

    1998-05-01

    O-alpha-D-Galactopyranosyl-(1-->2)-D-chiro-inositol, herein named fagopyritol B1, was identified as a major soluble carbohydrate (40% of total) in buckwheat (Fagopyrum esculentum Moench, Polygonaceae) embryos. Analysis of hydrolysis products of purified compounds and of the crude extract led to the conclusion that buckwheat embryos have five alpha-galactosyl D-chiro-inositols: fagopyritol A1 and fagopyritol B1 (mono-galactosyl D-chiro-inositol isomers), fagopyritol A2 and fagopyritol B2 (di-galactosyl D-chiro-inositol isomers), and fagopyritol B3 (tri-galactosyl D-chiro-inositol). Other soluble carbohydrates analyzed by high-resolution gas chromatography included sucrose (42% of total), D-chiro-inositol, myo-inositol, galactinol, raffinose and stachyose (1% of total), but no reducing sugars. All fagopyritols were readily hydrolyzed by alpha-galactosidase (EC 3.2.1.22) from green coffee bean, demonstrating alpha-galactosyl linkage. Retention time of fagopyritol B1 was identical to the retention time of O-alpha-D-galactopyranosyl-(1-->2)-D-chiro-inositol from soybean (Glycine max (L.) Merrill, Leguminosae), suggesting that the alpha-galactosyl linkage is to the 2-position of D-chiro-inositol. Accumulation of fagopyritol B1 was associated with acquisition of desiccation tolerance during seed development and maturation in planta, and loss of fagopyritol B1 correlated with loss of desiccation tolerance during germination. Embryos of seeds grown at 18 degrees C, a condition that favors enhanced seed vigor and storability, had a sucrose-to-fagopyritol B1 ratio of 0.8 compared to a ratio of 2.46 for seeds grown at 25 degrees C. We propose that fagopyritol B1 facilitates desiccation tolerance and storability of buckwheat seeds. PMID:9599801

  6. Uncoupling of attenuated myo-(3H)inositol uptake and dysfunction in Na(+)-K(+)-ATPase pumping activity in hypergalactosemic cultured bovine lens epithelial cells

    SciTech Connect

    Cammarata, P.R.; Tse, D.; Yorio, T. )

    1991-06-01

    Attenuation of both the active transport of myo-inositol and Na(+)-K(+)-ATPase pumping activity has been implicated in the onset of sugar cataract and other diabetic complications in cell culture and animal models of the disease. Cultured bovine lens epithelial cells (BLECs) maintained in galactose-free Eagle's minimal essential medium (MEM) or 40 mM galactose with and without sorbinil for up to 5 days were examined to determine the temporal effects of hypergalactosemia on Na(+)-K(+)-ATPase and myo-inositol uptake. The Na(+)-K(+)-ATPase pumping activity after 5 days of continuous exposure to galactose did not change, as demonstrated by 86Rb uptake. The uptake of myo-(3H)inositol was lowered after 20 h of incubation in galactose and remained below that of the control throughout the 5-day exposure period. The coadministration of sorbinil to the galactose medium normalized the myo-(3H)inositol uptake. No significant difference in the rates of passive efflux of myo-(3H)inositol or 86Rb from preloaded galactose-treated and control cultures was observed. Culture-media reversal studies were also carried out to determine whether the galactose-induced dysfunction in myo-inositol uptake could be corrected. BLECs were incubated in galactose for 5 days, then changed to galactose-free physiological medium with and without sorbinil for a 1-day recovery period. myo-Inositol uptake was reduced to 34% of control after 6 days of continuous exposure to galactose. Within 24 h of media reversal, myo-inositol uptake returned to or exceeded control values in BLECs switched to either MEM or MEM with sorbinil.2+ reversible and occurred independently of changes in Na(+)-K(+)-ATPase pumping activity in cultured lens epithelium, indicating that the two parameters are not strictly associated and that the deficit in myo-inositol uptake occurs rapidly during hypergalactosemia.

  7. Effect of myo-inositol and alpha-lipoic acid on oocyte quality in polycystic ovary syndrome non-obese women undergoing in vitro fertilization: a pilot study.

    PubMed

    Rago, R; Marcucci, I; Leto, G; Caponecchia, L; Salacone, P; Bonanni, P; Fiori, C; Sorrenti, G; Sebastianelli, A

    2015-01-01

    The aim of the present study was to evaluate the effectiveness of the combined administration of myo-inositol and α-lipoic acid in polycystic ovary syndrome (PCOS) patients with normal body mass index (BMI), who had previously undergone intracytoplasmic sperm injection (ICSI) and received myo-inositol alone. Thirty-six of 65 normal-weight patients affected by PCOS who did not achieve pregnancy and one patient who had a spontaneous abortion were re-enrolled and given a cycle of treatment with myo-inositol and α-lipoic acid. For all female partners of the treated couples, the endocrine-metabolic and ultrasound parameters, ovarian volume, oocyte and embryo quality, and pregnancy rates were assessed before and after three months of treatment and compared with those of previous in vitro fertilization (IVF) cycle(s). After supplementation of myo-inositol with α-lipoic acid, insulin levels, BMI and ovarian volume were significantly reduced compared with myo-inositol alone. No differences were found in the fertilization and cleavage rate or in the mean number of transferred embryos between the two different treatments, whereas the number of grade 1 embryos was significantly increased, with a significant reduction in the number of grade 2 embryos treated with myo-inositol plus α-lipoic acid. Clinical pregnancy was not significantly different with a trend for a higher percentage for of myo-inositol and α-lipoic acid compared to the myo-inositol alone group. Our preliminary data suggest that the supplementation of myo-inositol and α-lipoic acid in PCOS patients undergoing an IVF cycle can help to improve their reproductive outcome and also their metabolic profiles, opening potential for their use in long-term prevention of PCOS. PMID:26753656

  8. Isolation of 1-monomethylphosphoinositol 4,5-bisphosphate (a product of methanolysis of inositol 1,2-(cyclic)-4,5-trisphosphate) from Swiss mouse 3T3 cells

    SciTech Connect

    Lips, D.L.; Bross, T.E.; Majerus, P.W.

    1988-01-01

    We have noted two previously undescribed inositol polyphosphates in neutral methanol extracts from Swiss mouse 3T3 cells that were grown in (/sup 3/H)inositol and stimulated with platelet-derived growth factor. They have been identified as 1-monomethylphosphoinositol 4,5-bisphosphate and 1-monomethylphosphoinositol 4-phosphate by comparison to a synthesized standard using HPLC chromatography, paper electrophoresis, and enzymatic dephosphorylation with inositol polyphosphate 5-phosphomonoesterase and intestinal alkaline phosphatase. We propose that these compounds are formed by methanolysis of inositol 1,2-(cyclic)-4,5-trisphosphate and inositol 1,2-(cyclic)-4-bisphosphate present in the cells. Inositol cyclic phosphates did not react with neutral methanol in the absence of the cells, which are required for the methanolysis reaction. These findings suggest a role for inositol cyclic phosphates as reactive compounds that are added to as yet unidentified cellular acceptors.

  9. Studying isoform-specific inositol 1,4,5-trisphosphate receptor function and regulation

    PubMed Central

    Betzenhauser, Matthew J.; Wagner, Larry E.; Won, Jong-Hak; Yule, David I.

    2011-01-01

    Inositol 1,4,5-trisphosphate receptors (InsP3R) are a family of ubiquitously expressed intracellular Ca2+ channels. Isoform-specific properties of the three family members may play a prominent role in defining the rich diversity of the spatial and temporal characteristics of intracellular Ca2+ signals. Studying the properties of the particular family members is complicated because individual receptor isoforms are typically never expressed in isolation. In this article, we discuss strategies for studying Ca2+ release through individual InsP3R family members with particular reference to methods applicable following expression of recombinant InsP3R and mutant constructs in the DT40-3KO cell line, an unambiguously null InsP3R expression system. PMID:18929664

  10. Myo-Inositol trisphosphate mobilizes calcium from fusogenic carrot (Daucus carota L. ) protoplasts

    SciTech Connect

    Rincon, M.; Boss, W.F.

    1987-02-01

    To determine whether or not inositol trisphosphate (IP/sub 3/) mobilizes calcium in higher plant cells; they investigated the effect of IP/sub 3/ on Ca/sup 2 +/ fluxes in fusogenic carrot (Daucus carota L.) protoplasts. The protoplasts were incubated in /sup 45/Ca/sup 2 +/-containing medium and the /sup 45/Ca/sup 2 +/ associated with the protoplasts was monitored with time. Addition of IP/sub 3/ (20 micromolar) caused a 17% net loss of the accumulated /sup 45/Ca/sup 2 +/ within 4 minutes. There was a reuptake of /sup 45/Ca/sup 2 +/ and the protoplasts recovered to their initial value by 10 minutes. Phytic acid (IP/sub 6/), also stimulated /sup 45/Ca/sup 2 +/ efflux from the protoplasts. Both the IP/sub 3/- and the IP/sub 6/-induced /sup 45/Ca/sup 2 +/ efflux were inhibited by the calmodulin antagonist, trifluoperazine.