Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01275 (glucagon)
 26,492 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The intracellular distribution and molecular heterogeneity of carboxypeptidase H was studied in rat insulinoma tissue and isolated islets of Langerhans by a combination of immunohistochemical, ultrastructural, subcellular fractionation, and immunoblotting analyses. Immunofluorescence microscopy of islets demonstrated the presence of carboxypeptidase H in both insulin-containing B cells and glucagon-containing A cells. Quantitative ultrastructural analyses of islet B cells indicated that the enzyme was concentrated in mature insulin secretory granules, clathrin-coated condensing granules, and to a lesser extent the Golgi apparatus. Carboxypeptidase H activity was localized principally to secretory granule subfractions of insulinoma tissue, where it was present for the major part (70%) as a form which is readily solubilizable at pH values prevailing in the granule interior (5.5). This species migrated as a diffuse band of 53-57 kilodaltons (kDa) on immunoblot analysis using antisera raised against the purified native enzyme. In contrast, the insoluble form which was associated with the granule membrane at pH 5.5, migrated as a relatively compact band of 55-57 kDa. Carboxypeptidase H activity was also present in subcellular fractions which contained Golgi membranes together with elements of the endoplasmic reticulum, and in a low density secretory granule fraction which may represent immature granules. The enzyme in these compartments, like the granule membrane species, migrated as a compact 55-57 kDa band on immunoblots. Two-dimensional electrophoretic immunoblot analysis of secretory granules suggested that both membrane and soluble forms of the enzyme were glycoproteins and that the terminal glycosylation was similar in both instances. Antiserum raised against the deduced C-terminal 11 amino acids of the cloned carboxypeptidase H sequence recognized the 55-57 kDa membrane component in granules but did not react with the 53-57 kDa soluble species. A major difference between the soluble and membrane forms therefore appears to be a structural modification or proteolytic removal of the C-terminal domain in the trans-Golgi or early secretory granule compartment. The concept that proteolysis is involved is further supported by the observation that the relative proportion of the high and low mol wt forms of the enzyme in different subcellular fractions correlated with that of proinsulin and insulin, respectively. The membrane association of the 55-57 kDa form of carboxypeptidase H is disrupted at pH values of 9 and is dependent on ionic strength. This further suggests that the C-terminus of the protein may have an important role in the sorting or concentration of the enzyme in vesicular elements of the regulated pathway of secretion.
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PMID:Molecular heterogeneity and cellular localization of carboxypeptidase H in the islets of Langerhans. 185 71

Developmental patterns for rat pancreatic opioid peptides and islet hormones were studied from gestational day 20 through adulthood. Fetal tissue was obtained as well as pancreas at birth (day 0), and postnatal days 3, 7, 14, and 21, and 7 weeks. The hormones measured included insulin, glucagon, and somatostatin. The opioids measured were beta-endorphin, Met- and Leu-enkephalins, and the high molecular weight enkephalin precursors. Pancreata were pooled as necessary and extracted (acid alcohol, or hot acetic acid), and opioids were further purified on reversed-phase C-18 (Sep-pak) cartridges. In all instances measurements were made by radioimmunoassays. Precursor peptides were first digested (with trypsin and carboxypeptidase B) prior to immunoassay. All opioids and hormones except the precursors for enkephalins showed a well-defined surge in pancreatic concentration during the first postnatal week. In contrast, the precursors had the highest concentration in the fetus, and by the seventh day of life had decreased by greater than 50%. This progressive decrease may represent maturation of the enkephalin convertase and trypsin-like enzymes in the islets. The opioid and hormonal surges that we have described are similar to the surge in islet concentration of thyroid-releasing hormone (TRH) previously described in neonatal rat islets. It is suggested that these postnatal alterations in opioid and hormone concentration relate to a specific function in the development of the endocrine pancreas.
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PMID:Developmental patterns for pancreatic opioids in the rat. 253 May 76

Enkephalin convertase (carboxypeptidase E,H; EC 3.4.17.10) is a carboxypeptidase B-like enzyme which appears to be physiologically associated with the biosynthesis of the enkephalins and certain other peptides. We have localized enkephalin convertase in the brain and other tissues autoradiographically by labeling studies with [3H]guanidinoethylmercaptosuccinic acid ([3H]GEMSA). In the brain, [3H]GEMSA localizations parallel enkephalin distribution but with certain exceptions, suggesting a role in relation to other peptides. In the pancreas, [3H]GEMSA binding sites are localized to the islets suggesting an involvement in insulin, glucagon, or somatostatin formation. The selective concentration of [3H]GEMSA grains in cardiac atria suggests a link to atrial natriuretic factor.
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PMID:Enkephalin convertase: characterization and localization using [3H]guanidinoethylmercaptosuccinic acid. 313 43

The hormone insulin remains the cornerstone of diabetic therapy since it is required for almost all cases of Type 1 and many cases of Type 2 diabetes. Since the discovery of insulin in 1921, much has been learned about its chemistry, structure and action as well as its production in the beta cell. Insulin is formed through a series of precursors, beginning with preproinsulin, the protein encoded in the insulin gene. These precursors direct the prohormone into the secretory pathway and ultimately into the secretory granules where it is converted into insulin and C-peptide. These products are stored and secreted together in a highly regulated manner in response to glucose and other stimuli. This review focuses on the recently discovered prohormone convertases, PC2 and PC3 (PC1), the enzymes responsible for the endoproteolytic processing of proinsulin to insulin and C-peptide in the beta cell as well as for the selective processing of proglucagon to glucagon in the alpha cell or GLP1 in intestinal L-cells. PC2 and PC3 are calcium-dependent serine proteases related to the bacterial enzyme subtilisin. They cleave selectively at Lys-Arg or Arg-Arg sites in precursors, generating products with C-terminal basic residues that are then removed by carboxypeptidase E, an exopeptidase. All 3 enzymes are expressed mainly in secretory granules of neuroendocrine cells throughout the body and in the brain. Inherited defects affecting the prohormone-processing enzymes have recently been found in association with unusual syndromes of obesity and other metabolic disorders.
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PMID:The role of prohormone convertases in insulin biosynthesis: evidence for inherited defects in their action in man and experimental animals. 879 89

Proglucagon is alternatively processed to glucagon in pancreatic alpha-cells, or to glucagon-like peptide-1 in intestinal L cells. Here, the specificity of PC2, the major prohormone convertase of alpha-cells, was examined both in vivo and in vitro. Adenovirus-mediated co-expression of proglucagon and PC2 in GH4C1 cells resulted in a pattern of processing products very similar to that observed in alpha-cells. Oxyntomodulin, an intermediate in the processing of proglucagon, was quantitatively converted to glucagon in vitro by purified recombinant PC2, in combination with carboxypeptidase E. It is concluded that PC2 is able to act alone in the pancreatic pathway of proglucagon processing.
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PMID:Role of the prohormone convertase PC2 in the processing of proglucagon to glucagon. 928 28

Studies on the developing mammalian pancreas have suggested that insulin and glucagon co-exist in a transient cell population and that peptide YY (PYY) marks the earliest developing endocrine cells. We have investigated this in the embryonic avian pancreas, which is characterised by anatomical separation of insulin and glucagon islets. Moreover, we have compared the development of the endocrine cells to that of processing enzymes involved in pancreatic hormone biosynthesis. PYY-like immunoreactivity occurred in islet cells from the youngest stages examined: it increased in amount from approximately 5 days of incubation and was co-localised with glucagon and to a lesser extent with insulin. Insulin and glucagon cells were numerous: co-existence of the two peptides in the same cells was but rarely observed. From the youngest stages examined, prohormone convertase (PC) 1/3-like immunoreactivity was detected in insulin cells and PC2-, 7B2- and carboxypeptidase E-like immunoreactivity in both glucagon and insulin cells. We conclude that: (1) PYY-like immunoreactivity occurs in avian islet cells but generally in lesser amounts than in mammals at the earlier stages, (2) the paucity of cells co-expressing insulin and glucagon indicate that all avian insulin cells do not pass through a stage where they co-express glucagon and (3) the early expression of the enzymes responsible for the processing of prohormones suggests that this process is initiated soon after islet cells first differentiate.
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PMID:Development of hormonal peptides and processing enzymes in the embryonic avian pancreas with special reference to co-localisation. 1105 59

The maturation of many peptide hormones is attenuated in carboxypeptidase E (CPE)-deficient fat/fat mice, leading to a slowly developing, adult-onset obesity with mild diabetes. To determine the contribution of the hormones generated from the proglucagon precursor to this phenotype, we studied the tissue-specific processing of glucagon and glucagon-like peptide-1 (GLP-1) in these mice. In all tissues examined there was a great reduction in mature amidated GLP-1. Furthermore, a lack of CPE attenuates prohormone convertase processing of proglucagon in both the pancreas and the intestine. These findings suggest that defects in proglucagon processing together with other endocrine malfunctions could contribute to the diabetic and obesity phenotype in fat/fat mice.
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PMID:Attenuated processing of proglucagon and glucagon-like peptide-1 in carboxypeptidase E-deficient mice. 1137 30

We have reinvestigated the stability and intracellular routing of mutant carboxypeptidase E in NIT3 cells, a pancreatic beta-cell line derived from the Cpe(fat)/Cpe(fat) mouse. Pulse-chase experiments demonstrated that this protein has a half-life of approximately 3 h in these cells and that up to 45% of the proCPE(202) can escape degradation by the proteosome. In double-label immunofluorescence microscopy, a portion of the mutant CPE did not colocalize with calnexin, an endoplasmic reticulum marker, but was found in prohormone convertase 2-containing secretory granules, demonstrating that it had escaped degradation and arrived at a post-Golgi compartment. The mutant CPE as well as prohormone convertase 2 were secreted into the medium in a stimulated manner by treatment with the physiological secretagogue, glucagon-like peptide-1, consistent with its presence in granules of the regulated secretory pathway. The presence of mutant carboxypeptidase E in granules supports a potential role for its involvement as a sorting/retention receptor in the trafficking of proinsulin to the regulated secretory pathway.
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PMID:Trafficking of mutant carboxypeptidase E to secretory granules in a beta-cell line derived from Cpe(fat)/Cpe(fat) mice. 1248 57

Pluripotent embryonic stem cells hold a great promise as an unlimited source of tissue for treatment of chronic diseases such as Type 1 diabetes. Herein, we describe a protocol using all-trans-retinoic acid, basic fibroblast growth factor and dibutyryl cAMP (DBcAMP) in the absence of embryoid body formation, for differentiation of murine embryonic stem cells into definitive endoderm that may serve as pancreatic precursors. The produced cells were analyzed by quantitative PCR, immunohistochemistry and static insulin release assay for markers of trilaminar embryo, and pancreas. Differentiated cells displayed increased Sox17 and Foxa2 expression consistent with definitive endoderm production. There was minimal production of Sox7, an extraembryonic endoderm marker, and Oct4, a marker of pluripotency. There was minimal mesoderm or neuroectoderm formation based on expression levels of the markers brachyury and Sox1, respectively. Various assays revealed that the cell clusters generated by this protocol express markers of the pancreatic lineage including insulin I, insulin II, C-peptide, PDX-1, carboxypeptidase E, pan-cytokeratin, amylase, glucagon, PAX6, Ngn3 and Nkx6.1. This protocol using all-trans-retinoic acid, DBcAMP, in the absence of embryoid bodies, generated cells that have features of definitive endoderm that may serve as pancreatic endocrine precursors.
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PMID:Differentiation of mouse embryonic stem cells into endoderm without embryoid body formation. 2115 87

Proglucagon is expressed in pancreatic alpha cells, intestinal L cells and brainstem neurons. Tissue-specific processing of proglucagon yields the peptide hormones glucagon in the alpha cell and glucagon-like peptide (GLP)-1 and GLP-2 in L cells. Both glucagon and GLP-1 are secreted in response to nutritional status and are critical for regulating glycaemia. The sorting of proglucagon to the dense-core secretory granules of the regulated secretory pathway is essential for the appropriate secretion of glucagon and GLP-1. We examined the roles of carboxypeptidase E (CPE), a prohormone sorting receptor, the processing enzymes PC1/3 and PC2 and putative intrinsic sorting signals in proglucagon sorting. In Neuro 2a cells that lacked CPE, PC1/3 and PC2, proglucagon co-localised with the Golgi marker p115 as determined by quantitative immunofluorescence microscopy. Expression of CPE, but not of PC1/3 or PC2, enhanced proglucagon sorting to granules. siRNA-mediated knockdown of CPE disrupted regulated secretion of glucagon from pancreatic-derived alphaTC1-6 cells, but not of GLP-1 from intestinal cell-derived GLUTag cells. Mutation of the PC cleavage site K70R71, the dibasic R17R18 site within glucagon or the alpha-helix of glucagon, all significantly affected the sub-cellular localisation of proglucagon. Protein modelling revealed that alpha helices corresponding to glucagon, GLP-1 and GLP-2, are arranged within a disordered structure, suggesting some flexibility in the sorting mechanism. We conclude that there are multiple mechanisms for sorting proglucagon to the regulated secretory pathway, including a role for CPE in pancreatic alpha cells, initial cleavage at K70R71 and multiple sorting signals.
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PMID:The sorting of proglucagon to secretory granules is mediated by carboxypeptidase E and intrinsic sorting signals. 2341 62


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