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Between spring 1982 and autumn 1984 the physiological role of Ins(1,4,5)P-3 as a calcium-mobilizing second messenger was first suggested and then experimentally established. At the same time the unexpected complexity of inositide ...
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Between spring 1982 and autumn 1984 the physiological role of Ins(1,4,5)P-3 as a calcium-mobilizing second messenger was first suggested and then experimentally established. At the same time the unexpected complexity of inositide metabolism began to be exposed by the discovery of Ins(1,3,4)P-3. This article recalls my entanglement with these two inositol phosphates.
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A critical evaluation of a recent attempt to measure inositol hexakisphosphate (IP6) in mammalian plasma by mass spectroscopy leads to the conclusion that as yet there is no unambiguous evidence that plasma contains any IP6.
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Inositol pyrophosphates (PP-InsPs) are an emerging class of "high-energy" intracellular signaling molecules, containing one or two diphosphate groups attached to an inositol ring, that are connected with phosphate sensing, jasmona...
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Inositol pyrophosphates (PP-InsPs) are an emerging class of "high-energy" intracellular signaling molecules, containing one or two diphosphate groups attached to an inositol ring, that are connected with phosphate sensing, jasmonate signaling, and inositol hexakisphosphate (InsP(6)) storage in plants. While information regarding this new class of signaling molecules in plants is scarce, the enzymes responsible for their synthesis have recently been elucidated. This review focuses on InsP(6) synthesis and its conversion into PP-InsPs, containing seven and eight phosphate groups (InsP(7) and InsP(8)). These steps involve two types of enzymes: the ITPKs that phosphorylate InsP(6) to InsP(7), and the PPIP5Ks that phosphorylate InsP(7) to InsP(8). This review also considers the potential roles of PP-InsPs in plant hormone and inorganic phosphate (Pi) signaling, along with an emerging role in bioenergetic homeostasis. PP-InsP synthesis and signaling are important for plant breeders to consider when developing strategies that reduce InsP(6) in plants, as this will likely also reduce PP-InsPs. Thus, this review is primarily intended to bridge the gap between the basic science aspects of PP-InsP synthesis/signaling and breeding/engineering strategies to fortify foods by reducing InsP(6).
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Inositol phosphate (IP) kinases constitute an emerging class of cellular kinases linked to multiple cellular activities. Here, we report a previously uncharacterized cellular function in Hedgehog (Hh) signaling for the IP kinase d...
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Inositol phosphate (IP) kinases constitute an emerging class of cellular kinases linked to multiple cellular activities. Here, we report a previously uncharacterized cellular function in Hedgehog (Hh) signaling for the IP kinase designated inositol hexakisphosphate kinase-2 (IP6K2) that produces diphosphoryl inositol phosphates (PP-IPs). In zebrafish embryos, IP6K2 activity was required for normal development of craniofacial structures, somites, and neural crest cells. ip6k2 depletion in both zebrafish and mammalian cells also inhibited Hh target gene expression. Inhibiting IP_6 kinase activity using N(2)-(m-(trifluoromethy)lbenzyl) N(6)-(p-nitrobenzyl) purine (TNP) resulted in altered Hh signal transduction. In zebrafish, restoring IP6K2 levels with exogenous ip6k2 mRNA reversed the effects of IP6K2 depletion. Furthermore, overexpression of ip6k2 in mammalian cells enhanced the Hh pathway response, suggesting IP6K2 is a positive regulator of Hh signaling. Perturbations from IP6K2 depletion or TNP were reversed by overexpressing smoM2, gli1, or ip6k2. Moreover, the inhibitory effect of cyclopamine was reversed by overexpressing ip6k2. This identified roles for the inositol kinase pathway in early vertebrate development and tissue morphogenesis, and in Hh signaling. We propose that IP6K2 activity is required at the level or downstream of Smooth-ened but upstream of the transcription activator Gli1.
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The parasitic protozoon Trichomonas vaginalis is the pathogen of trichomoniasis, the most common non-viral, sexually transmitted disease in humans. Inositol phosphates function in the pathomechanisms of a number of human pathogeni...
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The parasitic protozoon Trichomonas vaginalis is the pathogen of trichomoniasis, the most common non-viral, sexually transmitted disease in humans. Inositol phosphates function in the pathomechanisms of a number of human pathogenic protozoa. Recent findings point to a role of inositol phosphates in T. vaginalis' adaption to oxygen exposure during change of host. Six inositol phosphate kinase genes (tvip6k1-4, tvipk1-2) were identified in the T. vaginalis genome by us all coding for proteins containing canonical sequence motifs of the major group of animal inositol phosphate kinases (PDKG, SSLL, DFG/A). When characterizing the purified protein product of tvip6k1, we discovered that the major activity of the highly active enzyme ((similar to)2 mu mol/min/mg) is a conversion of InsP(6) to 6PP-InsP(5) and not 5PP-InsP(5) as by animal isoforms. Thus TvIP6K1 is a novel IP6-6K. The enzyme also converts Ins(1,3,4,5,6)P-5 to products pyrophosphorylated both at 6- and 4-phosphate still having a free 5-hydroxyl. In addition, the enzyme has a minor selectivity to phosphorylate the 3-OH in Ins(1,2,4,5)P-4 and Ins (1,2,4,5,6)P-5 . To present knowledge this novel enzyme is restricted to protozoa. Since its structure is predicted to be distinctly different from animal IP6K (IP6-5K) forms, TvIP6-6K may become a promising target to search for novel trichomoniasis specific drugs.
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Four inositol phosphate kinases catalyze phosphorylation of the second messenger inositol 1,4,5-trisphosphate [Ins(1,4,5)P-3] to inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P-4]: these enzymes comprise three isoenzymes of inos...
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Four inositol phosphate kinases catalyze phosphorylation of the second messenger inositol 1,4,5-trisphosphate [Ins(1,4,5)P-3] to inositol 1,3,4,5-tetrakisphosphate [Ins(1,3,4,5)P-4]: these enzymes comprise three isoenzymes of inositol 1,4,5-trisphosphate 3-kinase (Itpk), referred to as Itpka, Itpkb and Itpkc, and the inositol polyphosphate multikinase (IPMK). The four enzymes that act on Ins(1,4,5)P-3 are all expressed in rat pheochromocytoma PC12 cells, a model that is used to study neurite outgrowth induced by nerve growth factor (NGF). We compared the effect of over-expression of the four GFP-tagged kinases on NGF-induced neurite outgrowth. Our data show that over-expression of the Itpka and Itpkb isoforms inhibits NGF-induced neurite outgrowth, but over-expression of Itpkc and IPMK does not. Surprisingly, over-expression of the N-terminal F-actin binding domain of Itpka, which lacks catalytic activity, was as effective at inhibiting neurite outgrowth as the full-length enzyme. Neurite length was also significantly decreased in cells over-expressing Itpka and Itpkb but not Itpkc or IPMK. This result did not depend on the over-expression level of any of the kinases. PC12 cells over-expressing GFP-tagged kinase-dead mutants Itpka/b have shorter neurites than GFP control cells. The decrease in neurite length was never as pronounced as observed with wild-type GFP-tagged Itpka/b. Finally, the percentage of neurite-bearing cells was increased in cells over-expressing the membranous typeI Ins(1,4,5)P-3 5-phosphatase. We conclude that Itpka and Itpkb inhibit neurite outgrowth through both F-actin binding and localized Ins(1,4,5)P-3 3-kinase activity. Itpkc and IPMK do not influence neurite outgrowth or neurite length in this model.
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Inositols (cyclohexanehexols) comprise nine isomeric cyclic sugar alcohols, several of which occur in all domains of life with various functions. Many bacteria can utilize inositols as carbon and energy sources via a specific path...
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Inositols (cyclohexanehexols) comprise nine isomeric cyclic sugar alcohols, several of which occur in all domains of life with various functions. Many bacteria can utilize inositols as carbon and energy sources via a specific pathway involving inositol dehydrogenases (IDHs) as the first step of catabolism. The microbial cell factory Corynebacterium glutamicum can grow with myo -inositol as a sole carbon source. Interestingly, this species encodes seven potential IDHs, raising the question of the reason for this multiplicity. We therefore investigated the seven IDHs to determine their function, activity, and selectivity toward the biologically most important isomers myo -, scyllo -, and d - chiro -inositol. We created an ΔIDH strain lacking all seven IDH genes, which could not grow on the three inositols. scyllo - and d -chiro -inositol were identified as novel growth substrates of C. glutamicum . Complementation experiments showed that only four of the seven IDHs (IolG, OxiB, OxiD, and OxiE) enabled growth of the ΔIDH strain on two of the three inositols. The kinetics of the four purified enzymes agreed with the complementation results. IolG and OxiD are NAD + -dependent IDHs accepting myo - and d -chiro -inositol but not scyllo -inositol. OxiB is an NAD + -dependent myo -IDH with a weak activity also for scyllo -inositol but not for d - chiro -inositol. OxiE on the other hand is an NAD + -dependent scyllo -IDH showing also good activity for myo -inositol and a very weak activity for d -chiro -inositol. Structural models, molecular docking experiments, and sequence alignments enabled the identification of the substrate binding sites of the active IDHs and of residues allowing predictions on the substrate specificity. IMPORTANCE myo -, scyllo -, and d - chiro -inositol are C 6 cyclic sugar alcohols with various biological functions, which also serve as carbon sources for microbes. Inositol catabolism starts with an oxidation to keto-inositols catalyzed by inositol dehydrogenases (IDHs). The soil bacterium C. glutamicum encodes seven potential IDHs. Using a combination of microbiological, biochemical, and modeling approaches, we analyzed the function of these enzymes and identified four IDHs involved in the catabolism of inositols. They possess distinct substrate preferences for the three isomers, and modeling and sequence alignments allowed the identification of residues important for substrate specificity. Our results expand the knowledge of bacterial inositol metabolism and provide an important basis for the rational development of producer strains for these valuable inositols, which show pharmacological activities against, e.g., Alzheimer’s disease, polycystic ovarian syndrome, or type II diabetes.
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The enzyme inositol 1,3,4-trisphosphate 5/6-kinase (ITPK1) catalyzes the rate-limiting step in the formation of higher phosphory-lated forms of inositol in mammalian cells. Because it sits at a key regulatory point in the inositol...
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The enzyme inositol 1,3,4-trisphosphate 5/6-kinase (ITPK1) catalyzes the rate-limiting step in the formation of higher phosphory-lated forms of inositol in mammalian cells. Because it sits at a key regulatory point in the inositol metabolic pathway, its activity is likely to be regulated. We have previously shown that ITPK1 is phosphorylated, a posttranslational modification used by cells to regulate enzyme activity. We show here that ITPK1 is modified by acetylation of internal lysine residues. The acetylation sites, as determined by mass spectrometry, were found to be lysines 340, 383, and 410, which are all located on the surface of this protein. Overexpression of the acetyltransferases CREB-binding protein or p300 resulted in the acetylation of ITPK1, whereas overexpression of mammalian silent information regulator 2 resulted in the dea-cetylation of ITPK1. Functionally, ITPK1 acetylation regulates its stability. CREB-binding protein dramatically decreased the half-life of ITPK1. We further found that ITPK1 acetylation down-regulated its enzyme activity. HEK293 cells stably expressing acetylated ITPK1 had reduced levels of the higher phosphorylated forms of inositol, compared with the levels seen in cells expressing unacetylated ITPK1. These results demonstrate that lysine acetylation alters both the stability as well as the activity of ITPK1 in cells.
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We recently reported that inositol dehydrogenase(EC 1.1.1.18)from Bacillus subtilis can catalyze the highly stereoselective oxidation of 1L-4-O-substituted myo-inositol derivatives,as well as disaccharides melibiose and isomaltose...
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We recently reported that inositol dehydrogenase(EC 1.1.1.18)from Bacillus subtilis can catalyze the highly stereoselective oxidation of 1L-4-O-substituted myo-inositol derivatives,as well as disaccharides melibiose and isomaltose,but not gentiobiose or maltose,consistent with the requirement of an alpha-(1->6)linkage.We believed that the enzyme might therefore catalyze efficient stereoselective oxidation of the appropriate alpha-linked glycosyl inositols.We have synthesized alpha-D-glucopyranosyl-(l->4)-(Dh)-myo-inositol and alpha-D-galactopyranosyl-(1->4)-(DL)-myo-inositol using the Appel-Lee protocol to couple benzyl-protected glycosyl donors to protected inositols.This method failed in our hands using glycosyl donors derived from D-mannose and 2-azido-2-deoxy-D-glucose.When myo-inositol 1,3,5-monoorthoformate is used as the acceptor,the reaction is regiospecific for the 4/6-posi-tion.We report here the mildest conditions known for the removal of the orthoformate group.2-Azido-2-deoxy-alpha-D-glucopyrano-syl-(1->4)-(DL)-myo-inositol was synthesized using the trichloroacetimidate derivative as the donor,and all three pseudo-disaccharides were substrates for inositol dehydrogenase.The glucopyranosyl and galactopyranosyl derivatives displayed apparent second-order rate constants comparable to that of myo-inositol.
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