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Biochimica Et Biophysica Acta Jun 2015Phosphatidylinositol is a metabolic precursor of phosphoinositides and soluble inositol phosphates. Both sets of molecules represent versatile intracellular chemical... (Review)
Review
Phosphatidylinositol is a metabolic precursor of phosphoinositides and soluble inositol phosphates. Both sets of molecules represent versatile intracellular chemical signals in eukaryotes. While much effort has been invested in understanding the enzymes that produce and consume these molecules, central aspects for how phosphoinositide production is controlled and functionally partitioned remain unresolved and largely unappreciated. It is in this regard that phosphatidylinositol (PtdIns) transfer proteins (PITPs) are emerging as central regulators of the functional channeling of phosphoinositide pools produced on demand for specific signaling purposes. The physiological significance of these proteins is amply demonstrated by the consequences that accompany deficits in individual PITPs. Although the biological problem is fascinating, and of direct relevance to disease, PITPs remain largely uncharacterized. Herein, we discuss our perspectives regarding what is known about how PITPs work as molecules, and highlight progress in our understanding of how PITPs are integrated into cellular physiology. This article is part of a Special Issue entitled Phosphoinositides.
Topics: Biological Transport; CDP-Diacylglycerol-Inositol 3-Phosphatidyltransferase; Gene Expression Regulation; Humans; Lipid Metabolism; Models, Molecular; Phosphatidylinositols; Phospholipid Transfer Proteins; Phosphotransferases (Alcohol Group Acceptor); Saccharomyces cerevisiae; Signal Transduction; Type C Phospholipases
PubMed: 25592381
DOI: 10.1016/j.bbalip.2014.12.011 -
ELife Dec 2022How cellular metabolic state impacts cellular programs is a fundamental, unresolved question. Here, we investigated how glycolytic flux impacts embryonic development,...
How cellular metabolic state impacts cellular programs is a fundamental, unresolved question. Here, we investigated how glycolytic flux impacts embryonic development, using presomitic mesoderm (PSM) patterning as the experimental model. First, we identified fructose 1,6-bisphosphate (FBP) as an in vivo sentinel metabolite that mirrors glycolytic flux within PSM cells of post-implantation mouse embryos. We found that medium-supplementation with FBP, but not with other glycolytic metabolites, such as fructose 6-phosphate and 3-phosphoglycerate, impaired mesoderm segmentation. To genetically manipulate glycolytic flux and FBP levels, we generated a mouse model enabling the conditional overexpression of dominant active, cytoplasmic PFKFB3 (cytoPFKFB3). Overexpression of cytoPFKFB3 indeed led to increased glycolytic flux/FBP levels and caused an impairment of mesoderm segmentation, paralleled by the downregulation of Wnt-signaling, reminiscent of the effects seen upon FBP-supplementation. To probe for mechanisms underlying glycolytic flux-signaling, we performed subcellular proteome analysis and revealed that cytoPFKFB3 overexpression altered subcellular localization of certain proteins, including glycolytic enzymes, in PSM cells. Specifically, we revealed that FBP supplementation caused depletion of Pfkl and Aldoa from the nuclear-soluble fraction. Combined, we propose that FBP functions as a flux-signaling metabolite connecting glycolysis and PSM patterning, potentially through modulating subcellular protein localization.
Topics: Animals; Mice; Mesoderm; Glycolysis; Embryonic Development; Embryo, Mammalian; Wnt Signaling Pathway; Phosphotransferases
PubMed: 36469462
DOI: 10.7554/eLife.83299 -
Chemical Communications (Cambridge,... Nov 2019DNA-encoded library (DEL) screening has emerged as an important method for early stage drug and probe molecule discovery. The vast majority of screens using DELs have... (Review)
Review
DNA-encoded library (DEL) screening has emerged as an important method for early stage drug and probe molecule discovery. The vast majority of screens using DELs have been relatively simple binding assays. The library is incubated with a target molecule, which is almost always a protein, and the DNAs that remain associated with the target after thorough washing are amplified and deep sequenced to reveal the chemical structures of the ligands they encode. Recently however, a number of different screening formats have been introduced that demand more than simple binding. These include a format that demands hits exhibit high selectivity for target vs. off-targets, a protocol to screen for enzyme inhibitors and another to identify organocatalysts in a DEL. These and other novel assay formats are reviewed in this article. We also consider some of the most significant remaining challenges in DEL assay development.
Topics: Drug Discovery; Enzyme Inhibitors; Gene Library; Humans; Ligands; Membrane Proteins; Peptide Library; Phosphotransferases; Protein Binding
PubMed: 31633708
DOI: 10.1039/c9cc06256d -
Journal of Experimental Botany Jul 2016The directional transport of auxin, known as polar auxin transport (PAT), allows asymmetric distribution of this hormone in different cells and tissues. This system... (Review)
Review
The directional transport of auxin, known as polar auxin transport (PAT), allows asymmetric distribution of this hormone in different cells and tissues. This system creates local auxin maxima, minima, and gradients that are instrumental in both organ initiation and shape determination. As such, PAT is crucial for all aspects of plant development but also for environmental interaction, notably in shaping plant architecture to its environment. Cell to cell auxin transport is mediated by a network of auxin carriers that are regulated at the transcriptional and post-translational levels. Here we review our current knowledge on some aspects of the 'non-genomic' regulation of auxin transport, placing an emphasis on how phosphorylation by protein and lipid kinases controls the polarity, intracellular trafficking, stability, and activity of auxin carriers. We describe the role of several AGC kinases, including PINOID, D6PK, and the blue light photoreceptor phot1, in phosphorylating auxin carriers from the PIN and ABCB families. We also highlight the function of some receptor-like kinases (RLKs) and two-component histidine kinase receptors in PAT, noting that there are probably RLKs involved in co-ordinating auxin distribution yet to be discovered. In addition, we describe the emerging role of phospholipid phosphorylation in polarity establishment and intracellular trafficking of PIN proteins. We outline these various phosphorylation mechanisms in the context of primary and lateral root development, leaf cell shape acquisition, as well as root gravitropism and shoot phototropism.
Topics: Biological Transport; Body Patterning; Indoleacetic Acids; Lipids; Phosphorylation; Phosphotransferases; Plant Development; Plant Growth Regulators; Protein Kinases
PubMed: 27242371
DOI: 10.1093/jxb/erw216 -
Journal of Lipid Research 2021Choline phospholipids (PLs) such as phosphatidylcholine (PC) and 1-alkyl-2-acyl-sn-glycerophosphocholine are important components for cell membranes and also serve as a...
Choline phospholipids (PLs) such as phosphatidylcholine (PC) and 1-alkyl-2-acyl-sn-glycerophosphocholine are important components for cell membranes and also serve as a source of several lipid mediators. These lipids are biosynthesized in mammals in the final step of the CDP-choline pathway by the choline phosphotransferases choline phosphotransferase 1 (CPT1) and choline/ethanolamine phosphotransferase 1 (CEPT1). However, the contributions of these enzymes to the de novo biosynthesis of lipids remain unknown. Here, we established and characterized CPT1- and CEPT1-deficient human embryonic kidney 293 cells. Immunohistochemical analyses revealed that CPT1 localizes to the trans-Golgi network and CEPT1 to the endoplasmic reticulum. Enzyme assays and metabolic labeling with radiolabeled choline demonstrated that loss of CEPT1 dramatically decreases choline PL biosynthesis. Quantitative PCR and reintroduction of CPT1 and CEPT1 revealed that the specific activity of CEPT1 was much higher than that of CPT1. LC-MS/MS analysis of newly synthesized lipid molecular species from deuterium-labeled choline also showed that these enzymes have similar preference for the synthesis of PC molecular species, but that CPT1 had higher preference for 1-alkyl-2-acyl-sn-glycerophosphocholine with PUFA than did CEPT1. The endogenous level of PC was not reduced by the loss of these enzymes. However, several 1-alkyl-2-acyl-sn-glycerophosphocholine molecular species were reduced in CPT1-deficient cells and increased in CEPT1-deficient cells when cultured in 0.1% FBS medium. These results suggest that CEPT1 accounts for most choline PL biosynthesis activity, and that both enzymes are responsible for the production of different lipid molecular species in distinct organelles.
Topics: Cells, Cultured; Choline; Diacylglycerol Cholinephosphotransferase; HEK293 Cells; Humans; Phospholipids; Transferases (Other Substituted Phosphate Groups)
PubMed: 34331935
DOI: 10.1016/j.jlr.2021.100100 -
Structural Basis of the Mechanisms of Action and Immunity of Lactococcin A, a Class IId Bacteriocin.Applied and Environmental Microbiology Mar 2023Lactococcin A (LcnA), a class IId bacteriocin, induces membrane leakage and cell death by specifically binding to the membrane receptor-mannose phosphotransferase system...
Lactococcin A (LcnA), a class IId bacteriocin, induces membrane leakage and cell death by specifically binding to the membrane receptor-mannose phosphotransferase system (man-PTS), as is the case for pediocin-like (class IIa) bacteriocins. The cognate immunity protein of bacteriocins, which protects the producer cell from its own bacteriocin, recognizes and binds to the bacteriocin-man-PTS complex, consequently blocking membrane leakage. We previously deciphered the mode of action and immunity of class IIa bacteriocins. Here, we determined the structure of the ternary complex of LcnA, LciA (, the immunity protein), and its receptor, , the man-PTS of Lactococcus lactis (ll-man-PTS). An external loop on the membrane-located component IIC of ll-man-PTS was found to prevent specific binding of the N-terminal region of LcnA to the site recognized by pediocin-like bacteriocins. Thus, the N-terminal β-sheet region of LcnA recognized an adjacent site on the extracellular side of ll-man-PTS, with the LcnA C-terminal hydrophobic helix penetrating into the membrane. The cytoplasmic cleft formed within the man-PTS Core and Vmotif domains induced by embedded LcnA from the periplasmic side is adopted by the appropriate angle between helices H3 and H4 of the N terminus of LciA. The flexible C terminus of LciA then blocks membrane leakage. To summarize, our findings reveal the molecular mechanisms of action and immunity of LcnA and LciA, laying a foundation for further design of class IId bacteriocins. Class IId (lactococcin-like) bacteriocins and class IIa (pediocin-like) bacteriocins share a few similarities: (i) both induce membrane leakage and cell death by specifically binding the mannose phosphotransferase system (man-PTS) on their target cells, and (ii) cognate immunity proteins recognize and bind to the bacteriocin-man-PTS complex to block membrane leakage. However, class IId bacteriocins lack the "pediocin box" motif, which is typical of class IIa bacteriocins, and basically target only lactococcal cells; in contrast, class IIa bacteriocins target diverse bacterial cells, but not lactococcal cells. We previously solved the structure of class IIa bacteriocin-receptor-immunity ternary complex from Lactobacillus sakei. Here, we determined the structure of the ternary complex of class IId bacteriocin LcnA, its cognate immunity protein LciA, and its receptor, the man-PTS of Lactococcus lactis. By comparing the interactions between man-PTS and class IIa and class IId bacteriocins, this study affords some clues to better understand the specificity of bacteriocins targeting the mannose phosphotransferase system.
Topics: Pediocins; Mannose; Bacteriocins; Lactococcus lactis; Phosphotransferases
PubMed: 36840592
DOI: 10.1128/aem.00066-23 -
Biomolecules Aug 2021Nucleic acid derivatives are involved in cell growth and replication, but they are also particularly important as building blocks for RNA and DNA synthesis [...].
Nucleic acid derivatives are involved in cell growth and replication, but they are also particularly important as building blocks for RNA and DNA synthesis [...].
Topics: Animals; Bacteria; Biocatalysis; Biotechnology; DNA; Fungi; Gene Expression; Glycosyltransferases; Humans; Nucleosides; Nucleotides; Pentosyltransferases; Phosphotransferases; Phosphotransferases (Phosphate Group Acceptor); RNA; Ribonucleotide Reductases
PubMed: 34439813
DOI: 10.3390/biom11081147 -
Cell Death and Differentiation Jan 2021Tauopathies are a group of neurodegenerative diseases characterized by hyperphosphorylation of the microtubule-binding protein, tau, and typically feature axon...
Tauopathies are a group of neurodegenerative diseases characterized by hyperphosphorylation of the microtubule-binding protein, tau, and typically feature axon impairment and synaptic dysfunction. Cyclin-dependent kinase5 (Cdk5) is a major tau kinase and its activity requires p35 or p25 regulatory subunits. P35 is subjected to rapid proteasomal degradation in its membrane-bound form and is cleaved by calpain under stress to a stable p25 form, leading to aberrant Cdk5 activation and tau hyperphosphorylation. The type Ib transmembrane protein RPS23RG1 has been implicated in Alzheimer's disease (AD). However, physiological and pathological roles for RPS23RG1 in AD and other tauopathies are largely unclear. Herein, we observed retarded axon outgrowth, elevated p35 and p25 protein levels, and increased tau phosphorylation at major Cdk5 phosphorylation sites in Rps23rg1 knockout (KO) mice. Both downregulation of p35 and the Cdk5 inhibitor roscovitine attenuated tau hyperphosphorylation and axon outgrowth impairment in Rps23rg1 KO neurons. Interestingly, interactions between the RPS23RG1 carboxyl-terminus and p35 amino-terminus promoted p35 membrane distribution and proteasomal degradation. Moreover, P301L tau transgenic (Tg) mice showed increased tau hyperphosphorylation with reduced RPS23RG1 levels and impaired axon outgrowth. Overexpression of RPS23RG1 markedly attenuated tau hyperphosphorylation and axon outgrowth defects in P301L tau Tg neurons. Our results demonstrate the involvement of RPS23RG1 in tauopathy disorders, and implicate a role for RPS23RG1 in inhibiting tau hyperphosphorylation through homeostatic p35 degradation and suppression of Cdk5 activation. Reduced RPS23RG1 levels in tauopathy trigger aberrant Cdk5-p35 activation, consequent tau hyperphosphorylation, and axon outgrowth impairment, suggesting that RPS23RG1 may be a potential therapeutic target in tauopathy disorders.
Topics: Alzheimer Disease; Animals; Mice; Mice, Inbred C57BL; Mice, Knockout; Neuronal Outgrowth; Neurons; Phosphorylation; Phosphotransferases; Ribosomal Proteins; tau Proteins
PubMed: 32908202
DOI: 10.1038/s41418-020-00620-y -
Advances in Biological Regulation Jan 2017The inositol pyrophosphates (diphosphoinositol polyphosphates), which include 1-InsP, 5-InsP, and InsP, are highly 'energetic' signaling molecules that play important... (Review)
Review
The inositol pyrophosphates (diphosphoinositol polyphosphates), which include 1-InsP, 5-InsP, and InsP, are highly 'energetic' signaling molecules that play important roles in many cellular processes, particularly with regards to phosphate and bioenergetic homeostasis. Two classes of kinases synthesize the PP-InsPs: IP6Ks and PPIP5Ks. The significance of the IP6Ks - and their 5-InsP product - has been widely reported. However, relatively little is known about the biological significance of the PPIP5Ks. The purpose of this review is to provide an update on developments in our understanding of key features of the PPIP5Ks, which we believe strengthens the hypothesis that their catalytic activities serve important cellular functions. Central to this discussion is the recent discovery that the PPIP5K is a rare example of a single protein that catalyzes a kinase/phosphatase futile cycle.
Topics: Acid Anhydride Hydrolases; Amino Acid Motifs; Energy Metabolism; Gene Expression Regulation; Humans; Inositol Phosphates; Phosphotransferases (Phosphate Group Acceptor); Phytic Acid; Protein Domains; Signal Transduction
PubMed: 27776974
DOI: 10.1016/j.jbior.2016.10.003 -
Molecules (Basel, Switzerland) Mar 2020In mammals, a family of three inositol hexakisphosphate kinases (IP6Ks) synthesizes the inositol pyrophosphate 5-IP7 from IP6. Genetic deletion of protects mice from... (Review)
Review
In mammals, a family of three inositol hexakisphosphate kinases (IP6Ks) synthesizes the inositol pyrophosphate 5-IP7 from IP6. Genetic deletion of protects mice from high fat diet induced obesity, insulin resistance and fatty liver. IP6K1 generated 5-IP7 promotes insulin secretion from pancreatic β-cells, whereas it reduces insulin signaling in metabolic tissues by inhibiting the protein kinase Akt. Thus, IP6K1 promotes high fat diet induced hyperinsulinemia and insulin resistance in mice while its deletion has the opposite effects. IP6K1 also promotes fat accumulation in the adipose tissue by inhibiting the protein kinase AMPK mediated energy expenditure. Genetic deletion of protects mice from age induced fat accumulation and insulin resistance. Accordingly, the pan IP6K inhibitor TNP [N2-(-trifluorobenzyl), N6-(-nitrobenzyl)purine] ameliorates obesity, insulin resistance and fatty liver in diet induced obese mice by improving Akt and AMPK mediated insulin sensitivity and energy expenditure. TNP also protects mice from bone loss, myocardial infarction and ischemia reperfusion injury. Thus, the IP6K pathway is a potential target in obesity and other metabolic diseases. Here, we summarize the studies that established IP6Ks as a potential target in metabolic diseases. Further studies will reveal whether inhibition of this pathway has similar pleiotropic benefits on metabolic health of humans.
Topics: Animals; Energy Metabolism; Enzyme Inhibitors; Humans; Inositol Phosphates; Metabolic Diseases; Mice; Molecular Targeted Therapy; Phosphotransferases (Phosphate Group Acceptor); Phytic Acid
PubMed: 32204420
DOI: 10.3390/molecules25061403