Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0011849 (diabetes)
 277,896 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

High intracellular glucose concentrations increase flux though the hexosamine biosynthetic pathway, resulting in elevated UDP-N-acetylglucosamine (GlcNAc) concentrations. The nucleocytoplasmic enzyme O-linked N-acetylglucosaminyltransferase (OGT) uses UDP-GlcNAc as a donor to modify numerous critical substrates, including nuclear pore proteins and transcription factors. Here, we document (a) the overwhelming enrichment of pancreatic OGT transcripts in the beta-cells of the islets of Langerhans, (b) the physiologically significant increase in the level of O-GlcNAc residues present in beta-cells, and (c) the action of streptozotocin, a close analogue of GlcNAc, to selectively inhibit O-GlcNAcase, an enzyme involved in the removal of O-GlcNAc residues. Taken together, these findings suggest that pancreatic beta cells maintain a highly elevated O-GlcNAc metabolism and that the diabetes inducing drug streptozotocin inhibits O-GlcNAcase.
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PMID:Elevated O-linked N-acetylglucosamine metabolism in pancreatic beta-cells. 991 27

Dynamic modification of cytoplasmic and nuclear proteins by O-linked N-acetylglucosamine (O-GlcNAc) on Ser/Thr residues is ubiquitous in higher eukaryotes and is analogous to protein phosphorylation. The enzyme for the addition of this modification, O-GlcNAc transferase, has been cloned from several species. Here, we have cloned a human brain O-GlcNAcase that cleaves O-GlcNAc off proteins. The cloned cDNA encodes a polypeptide of 916 amino acids with a predicted molecular mass of 103 kDa and a pI value of 4.63, but the protein migrates as a 130-kDa band on SDS-polyacrylamide gel electrophoresis. The cloned O-GlcNAcase has a pH optimum of 5.5-7.0 and is inhibited by GlcNAc but not by GalNAc. p-Nitrophenyl (pNP)-beta-GlcNAc, but not pNP-beta-GalNAc or pNP-alpha-GlcNAc, is a substrate. The cloned enzyme cleaves GlcNAc, but not GalNAc, from glycopeptides. Cell fractionation suggests that the overexpressed protein is mostly localized in the cytoplasm. It therefore has all the expected characteristics of O-GlcNAcase and is distinct from lysosomal hexosaminidases. Northern blots show that the transcript is expressed in every human tissue examined but is the highest in the brain, placenta, and pancreas. An understanding of O-GlcNAc dynamics and O-GlcNAcase may be key to elucidating the relationships between O-phosphate and O-GlcNAc and to the understanding of the molecular mechanisms of diseases such as diabetes, cancer, and neurodegeneration.
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PMID:Dynamic O-glycosylation of nuclear and cytosolic proteins: cloning and characterization of a neutral, cytosolic beta-N-acetylglucosaminidase from human brain. 1114 10

It is known well that activation of the hexosamine pathway causes insulin resistance, but how this activation influences pancreatic beta-cell function remains unclear. In this study, we found that in isolated rat islets adenovirus-mediated overexpression of glutamine:fructose-6-phosphate amidotransferase (GFAT), the first and rate-limiting enzyme of the hexosamine pathway, leads to deterioration of beta-cell function, which is similar to that found in diabetes. Overexpression of GFAT or treatment with glucosamine results in impaired glucose-stimulated insulin secretion and reduction in the expression levels of several beta-cell specific genes (insulin, GLUT2, and glucokinase). Additionally, the DNA binding activity of PDX-1, an important transcription factor for these three genes, was markedly reduced. These phenomena were not mimicked by the induction of O-linked glycosylation with an inhibitor of O-GlcNAcase, PUGNAc. It was also found that glucosamine increases hydrogen peroxide levels and that several hexosamine pathway-mediated changes were suppressed by treatment with the antioxidant N-acetyl-l-cysteine. In conclusion, activation of the hexosamine pathway leads to deterioration of beta-cell function through the induction of oxidative stress rather than O-linked glycosylation. Thus, the hexosamine pathway may contribute to the deterioration of beta-cell function found in diabetes.
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PMID:Activation of the hexosamine pathway leads to deterioration of pancreatic beta-cell function through the induction of oxidative stress. 1139 Apr 7

The molecular complexity that defines different cell types and their biological responses occurs at the level of the cell's proteome. The recent increase in availability of genomic sequence information is a valuable tool for the field of proteomics. While most proteomic studies focus on differential expression levels, post-translational modifications such as phosphorylation, glycosylation, and acetylation, provide additional levels of functional complexity to the cell's proteome. The reversible post-translational modification O-linked beta-N-acetylglucosamine (O-GlcNAc) is found on serines and threonines of nuclear and cytoplasmic proteins. It appears to be as widespread as phosphorylation. While phosphorylation is recognized as a fundamental mechanism for controlling protein function, less is known about the specific roles of O-GlcNAc modification. However, evidence is building that O-GlcNAc may compete with phosphate at some sites of attachment. Aberrant O-GlcNAc modification has been linked to several disease states, including diabetes and Alzheimer's disease. Regulated enzymes catalyzing the addition (O-GlcNAc transferase, OGT) and removal (O-GlcNAcase) of the modification have been cloned and OGT is required for life at the single cell level. Here we review the properties of O-GlcNAc that suggest it is a regulatory modification analogous to phosphorylation. We also discuss the use of comparative functional proteomics to elucidate functions for this ubiquitous intracellular carbohydrate modification.
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PMID:Nucleocytoplasmic O-glycosylation: O-GlcNAc and functional proteomics. 1152 85

The addition of O-linked N-acetylglucosamine (O-GlcNAc) to target proteins may serve as a signaling modification analogous to protein phosphorylation. Like phosphorylation, O-GlcNAc is a dynamic modification occurring in the nucleus and cytoplasm. Various analytical methods have been developed to detect O-GlcNAc and distinguish it from glycosylation in the endomembrane system. Many target molecules have been identified; these targets are typically components of supramolecular complexes such as transcription factors, nuclear pore proteins, or cytoskeletal components. The enzymes responsible for O-GlcNAc addition and removal are highly conserved molecules having molecular features consistent with a signaling role. The O-GlcNAc transferase and O-GlcNAcase are likely to act in consort with kinases and phosphatases generating various isoforms of physiological substrates. These isoforms may differ in such properties as protein-protein interactions, protein stability, and enzymatic activity. Since O-GlcNAc plays a critical role in the regulation of signaling pathways of higher plants, the glycan modification is likely to perform similar signaling functions in mammalian cells. Glucose and amino acid metabolism generates hexosamine precursors that may be key regulators of a nutrient sensing pathway involving O-GlcNAc signaling. Altered O-linked GlcNAc metabolism may also occur in human diseases including neurodegenerative disorders, diabetes mellitus and cancer.
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PMID:Glycan-dependent signaling: O-linked N-acetylglucosamine. 1153 66

Increased flux of glucose through the hexosamine biosynthetic pathway (HSP) is believed to mediate hyperglycemia-induced insulin resistance in diabetes. The end product of the HSP, UDP beta-N-acetylglucosamine (GlcNAc), is a donor sugar nucleotide for complex glycosylation in the secretory pathway and for O-linked GlcNAc (O-GlcNAc) addition to nucleocytoplasmic proteins. Cycling of the O-GlcNAc posttranslational modification was blocked by pharmacological inhibition of O-GlcNAcase, the enzyme that catalyzes O-GlcNAc removal from proteins, with O-(2-acetamido-2-deoxy-d-glucopyranosylidene)amino-N-phenylcarbamate (PUGNAc). PUGNAc treatment increased levels of O-GlcNAc and caused insulin resistance in 3T3-L1 adipocytes. Insulin resistance induced through the HSP by glucosamine and chronic insulin treatment correlated with increased O-GlcNAc levels on nucleocytoplasmic proteins. Whereas insulin receptor autophosphorylation and insulin receptor substrate 2 tyrosine phosphorylation were not affected by PUGNAc inhibition of O-GlcNAcase, downstream phosphorylation of Akt at Thr-308 and glycogen synthase kinase 3 beta at Ser-9 was inhibited. PUGNAc-induced insulin resistance was associated with increased O-GlcNAc modification of several proteins including insulin receptor substrate 1 and beta-catenin, two important effectors of insulin signaling. These results suggest that elevation of O-GlcNAc levels attenuate insulin signaling and contribute to the mechanism by which increased flux through the HSP leads to insulin resistance in adipocytes.
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PMID:Elevated nucleocytoplasmic glycosylation by O-GlcNAc results in insulin resistance associated with defects in Akt activation in 3T3-L1 adipocytes. 1195 83

Increased flux through the hexosamine biosynthetic pathway and increased O-linked glycosylation (N-acetylglucosamine [O-GlcNAc]) of proteins have been implicated in insulin resistance. Previous research in 3T3-L1 adipocytes indicated that insulin-stimulated glucose uptake and phosphorylation of Akt were reduced after incubation with O-(2-acetamido-2-deoxy-D-glucopyranosylidene)amino-N-phenylcarbamate (PUGNAc; 100 micromol/l), an inhibitor of the O-GlcNAcase that catalyzes removal of O-GlcNAc from proteins. Therefore, in this study, we tested the effects of PUGNAc on skeletal muscle. Incubation of rat epitrochlearis muscles for 19 h with 100 micromol/l PUGNAc resulted in a marked increase in O-GlcNAcylation of multiple proteins. Incubation with PUGNAc reduced glucose transport with a physiologic insulin concentration without affecting glucose transport without insulin or with supraphysiologic insulin. PUGNAc did not significantly alter insulin-stimulated phosphorylation of Akt (serine and threonine) or its substrates glycogen synthase kinase (GSK)3 alpha and GSK3 beta. Insulin stimulated a dose-dependent (12.0 > 0.6 > 0 nmol/l) increase in the phosphorylation of a 160-kDa protein detected using an antibody against an Akt substrate phosphomotif. PUGNAc treatment did not alter phosphorylation of this protein. These results indicate that PUGNAc is an effective inhibitor of O-GlcNAcase in skeletal muscle and suggest that O-GlcNAc modification of proteins can induce insulin resistance in skeletal muscle independent of attenuated phosphorylation of Akt, GSK 3 alpha, GSK3 beta, and a 160-kDa protein with an Akt phosphomotif.
Diabetes 2004 Apr
PMID:Prolonged incubation in PUGNAc results in increased protein O-Linked glycosylation and insulin resistance in rat skeletal muscle. 1504 6

Signal transduction from the insulin receptor to downstream effectors is attenuated by phosphorylation at a number of Ser/Thr residues of insulin receptor substrate-1 (IRS-1) resulting in resistance to insulin action, the hallmark of type II diabetes. Ser/Thr residues can also be reversibly glycosylated by O-linked beta-N-acetylglucosamine (O-GlcNAc) monosaccharide, a dynamic posttranslational modification that offers an alternative means of protein regulation to phosphorylation. To identify sites of O-GlcNAc modification in IRS-1, recombinant rat IRS-1 isolated from HEK293 cells was analyzed by two complementary mass spectrometric methods. Using data-dependent neutral loss MS3 mass spectrometry, MS/MS data were scanned for peptides that exhibited a neutral loss corresponding to the mass of N-acetylglucosamine upon dissociation in an ion trap. This methodology provided sequence coverage of 84% of the protein, permitted identification of a novel site of phosphorylation at Thr-1045, and facilitated the detection of an O-GlcNAc-modified peptide of IRS-1 at residues 1027-1073. The level of O-GlcNAc modification of this peptide increased when cells were grown under conditions of high glucose with or without chronic insulin stimulation or in the presence of an inhibitor of the O-GlcNAcase enzyme. To map the exact site of O-GlcNAc modification, IRS-1 peptides were chemically derivatized with dithiothreitol following beta-elimination and Michael addition prior to LC-MS/MS. This approach revealed Ser-1036 as the site of O-GlcNAc modification. Site-directed mutagenesis and Western blotting with an anti-O-GlcNAc antibody suggested that Ser-1036 is the major site of O-GlcNAc modification of IRS-1. Identification of this site will facilitate exploring the biological significance of the O-GlcNAc modification.
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PMID:Identification of the major site of O-linked beta-N-acetylglucosamine modification in the C terminus of insulin receptor substrate-1. 1624 61

O-linked N-acetylglucosaminyltransferase (OGT) catalyzes the transfer of O-linked GlcNAc to serine or threonine residues of a variety of substrate proteins, including nuclear pore proteins, transcription factors, and proteins implicated in diabetes and neurodegenerative disorders. We have identified two nucleocytoplasmic isoforms of OGT (ncOGT and sOGT) and one isoform that localizes to the mitochondria (mOGT). These three isoforms contain identical catalytic regions but differ in the number of tetratricopeptide repeat motifs found at the N-terminus of each enzyme. We expressed each of these OGT isoforms in a soluble form in Escherichia coli and have used them to identify novel targets including the Src-family tyrosine kinase yes and O-GlcNAc-ase. We demonstrate that some substrate proteins, such as Nup62 and casein kinase II, are glycosylated by both ncOGT and mOGT, while others such as O-GlcNAcase and tau are specifically modified by ncOGT. The yes kinase was specifically modified by mOGT. The short isoform of OGT (sOGT) did not glycosylate any of the substrates tested, although it retains a potentially active catalytic domain. Our findings demonstrate the potential utility of recombinant OGT in identifying new targets and illustrate the necessity to examine all active isoforms of the enzyme. The identification of a tyrosine kinase and O-GlcNAcase as OGT targets suggests the potential for OGT participation in numerous signal transduction cascades.
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PMID:Recombinant O-GlcNAc transferase isoforms: identification of O-GlcNAcase, yes tyrosine kinase, and tau as isoform-specific substrates. 1643 89

A dynamic cycle of O-linked N-acetylglucosamine (O-GlcNAc) addition and removal acts on nuclear pore proteins, transcription factors, and kinases to modulate cellular signaling cascades. Two highly conserved enzymes (O-GlcNAc transferase and O-GlcNAcase) catalyze the final steps in this nutrient-driven "hexosamine-signaling pathway." A single nucleotide polymorphism in the human O-GlcNAcase gene is linked to type 2 diabetes. Here, we show that Caenorhabditis elegans oga-1 encodes an active O-GlcNAcase. We also describe a knockout allele, oga-1(ok1207), that is viable and fertile yet accumulates O-GlcNAc on nuclear pores and other cellular proteins. Interfering with O-GlcNAc cycling with either oga-1(ok1207) or the O-GlcNAc transferase-null ogt-1(ok430) altered Ser- and Thr-phosphoprotein profiles and increased glycogen synthase kinase 3beta (GSK-3beta) levels. Both the oga-1(ok1207) and ogt-1(ok430) strains showed elevated stores of glycogen and trehalose, and decreased lipid storage. These striking metabolic changes prompted us to examine the insulin-like signaling pathway controlling nutrient storage, longevity, and dauer formation in the C. elegans O-GlcNAc cycling mutants. Indeed, we found that the oga-1(ok1207) knockout augmented dauer formation induced by a temperature sensitive insulin-like receptor (daf-2) mutant under conditions in which the ogt-1(ok430)-null diminished dauer formation. Our findings suggest that the enzymes of O-GlcNAc cycling "fine-tune" insulin-like signaling in response to nutrient flux. The knockout of O-GlcNAcase (oga-1) in C. elegans mimics many of the metabolic and signaling changes associated with human insulin resistance and provides a genetically amenable model of non-insulin-dependent diabetes.
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PMID:Caenorhabditis elegans ortholog of a diabetes susceptibility locus: oga-1 (O-GlcNAcase) knockout impacts O-GlcNAc cycling, metabolism, and dauer. 1688 29


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