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

Antidiabetic effects of CS-045 were evaluated in 5-month-old C57BL/KsJ-db/db mice (db/db). CS-045 administered for 3 weeks to diabetic db/db mice as a 0.2% food admixture improved hyperglycemia (855 +/- 25 v 298 +/- 62 mg/dL, P less than .01) and glucose intolerance, and lowered plasma triglyceride (299.6 +/- 28.7 v 76.3 +/- 20.7 mg/dL, P less than .01) and free fatty acid (FFA) levels (1.16 +/- 0.14 v 0.57 +/- 0.07 mEq/L, P less than .01). Food intake was not changed, while a small but significant increase in body weight was observed in CS-045-treated mice. Plasma insulin levels gradually increased after 5 days of CS-045 treatment, and a nonsignificant increase was observed in plasma insulin levels after 3 weeks (1.85 +/- 0.50 v 4.54 +/- 1.47 mg/mL). In contrast, the plasma glucagon levels decreased after 3 weeks of CS-045 treatment. Histological examination by aldehyde-fucshin staining demonstrated that pancreatic beta cells in CS-045-treated db/db mice were heavily regranulated, whereas most of the beta cells were extensively degranulated in nontreated db/db mice. The heavily regranulated state of beta cells was also compatible with an increase in pancreatic insulin content in CS-045-treated db/db mice. Electron microscopic analysis showed a well-developed endoplasmic reticulum and the accumulation of much amorphous structural material in the intracisternal space of beta cells from CS-045-treated db/db mice, which were suggestive of an increase in insulin synthesis. Moreover, CS-045 treatment decreased exocrine-containing islets, which was associated with the islets' degeneration process. Immunohistochemical staining of islets showed that CS-045 treatment normalized the distribution pattern of endocrine cells in the islets of db/db mice, reflected by a predominantly peripheral location of alpha and delta cells.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of CS-045, a new oral antidiabetic agent, II. Effects on glycemic control and pancreatic islet structure at a late stage of the diabetic syndrome in C57BL/KsJ-db/db mice. 194 50

Daily cyclosporine doses of 10 mg/kg body weight for 21 days in Wistar rats cause impairment in glucose homeostasis and changes in the amount of immunostainable hormones and in the ultrastructure of the cells of the pancreatic islets. CsA induces hyperglycemia and reduced glucose tolerance, and causes a decrease in immunoreactive insulin and an increase of somatostatin and pancreatic polypeptide (PP) immunoreactivities, leaving glucagon immunoreactivity unaffected. Ultrastructurally, different degrees of dilation of rough endoplasmic reticulum cisternae and enlargement of Golgi apparatus can be observed in B cells, together with a pronounced reduction in the number of secretory granules. Nevertheless, there were no apparent morphological changes of the other cytoplasmic organelles, suggesting that the drug, besides a depression of protein synthesis, as previously stated, also induces a substantial defect in granulogenesis, probably due to impairment in the intracellular transport of the hormone from the sites of synthesis to the secretory granules. The B cell alterations are not accompanied by any sign of B cell degeneration or death. Non-B cells did not show any of the ultrastructural changes found in B cells and were similar to those of the control rats. The above findings indicate that CsA at immunotherapeutic doses causes impairment in the secretory processes of B cells specifically. An hypothesis on the mode of action of CsA on B cells is drawn.
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PMID:Immunocytochemical and ultrastructural changes of islet cells in rats treated long-term with cyclosporine at immunotherapeutic doses. 218 26

Ultrastructural and autoradiographic observations of cultured chick hepatocytes under the following conditions are described: Induction of glycogen synthesis with glucose alone and glucose plus insulin, and glucagon-induced glycogen breakdown. Profiles of hepatocytes cultured in medium containing 10 mM glucose showed typical cellular organelles and occasionally a few glycogen granules. After incubation of hepatocytes with 3H-glucose, silver grains were found over these sparse glycogen granules, indicating a low level of glycogen synthesis by a few cells. After addition of 75 mM glucose for 1 hr about 3% of the profiles of cells showed glycogen, and by 24 hr half of the hepatocytes had glycogen. Addition of insulin plus glucose induced glycogen synthesis in 82% of the cells after 6 hr, and by 24 hr almost every cellular profile showed glycogen particles. Morphologically, glycogen accumulation was similar whether the cells were stimulated by high glucose or by glucose plus insulin: glycogen granules appeared in restricted regions of the cytoplasm, which were rich in smooth endoplasmic reticulum (SER), and peroxisomes were found close to the newly deposited glycogen particles. At maximum glycogen accumulation the association of SER and peroxisomes with glycogen was less obvious. Glycogenolysis induced by incubation of glycogen-rich hepatocytes with glucagon resulted in proliferation of SER in the glycogen regions of the cells. These observations are compatible with the concept of regions in the hepatocyte cytoplasm specialized for glycogen metabolism. Possible roles for SER and peroxisomes found near glycogen particles and other organelles in hepatic glycogen metabolism are discussed.
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PMID:Glycogen metabolism in cultured chick hepatocytes: a morphological study. 219 38

The control of RNA degradation by amino acids, insulin, and glucagon was investigated in perfused livers of fed rats previously labeled in vivo with [6-14C] orotic acid; rates were determined from the release of [14C]cytidine in the presence of 0.5 mM cytidine to suppress reutilization. Studies with cyclically perfused livers showed that plasma amino acids at 10 times (10X) normal concentrations inhibited RNA breakdown by 85%. Similar inhibition was obtained with a known regulatory amino acid mixture (Leu, Met, Pro, Trp, and His), whereas leucine alone (0.8 mM) decreased degradation by 47%. Perfusions carried out in the single-pass mode with graded levels of plasma amino acids revealed that the acceleration of RNA degradation over the full range of amino acid deprivation (0 to 10X normal levels) was the same as that for protein breakdown (3.19 and 3.15% h-1, respectively), and both were equally suppressed by insulin (2.4 micrograms h-1). Glucagon (10 micrograms h-1), though, was far less effective in stimulating RNA than protein turnover. A direct comparison of the two dose responses revealed a strong dissociation at 1 and 2 times normal amino acid levels. These findings support the notion that RNA and protein are degraded within a single macroautophagic compartment during amino acid and insulin deprivation. Glucagon, however, appeared to induce a second pathway in which the proportion of sequestered RNA to protein was selectively reduced. Electron micrographs showed that the ratio of vacuoles containing rough as compared with smooth endoplasmic reticulum was decreased by nearly 80% under these conditions.
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PMID:Amino acid and hormonal control of macromolecular turnover in perfused rat liver. Evidence for selective autophagy. 244 87

Treatment of intact adipocytes with either or both insulin and adrenaline stimulated membrane cyclic AMP phosphodiesterase activity only in the endoplasmic reticulum subfraction. The cyclic GMP-inhibited cyclic AMP phosphodiesterase activity was also found in this fraction. Quantitative Western blotting using a specific polyclonal antibody, raised against the homogeneous 'dense-vesicle' cyclic AMP phosphodiesterase from rat liver, identified a single 63 kDa species which was localized in the adipocyte endoplasmic reticulum fraction. The ability of adrenaline to stimulate adipocyte membrane cyclic AMP phosphodiesterase was shown to be mediated via beta-adrenoceptors and not alpha 1-adrenoceptors. Membrane cyclic AMP phosphodiesterase was stimulated by glucagon but not by vasopressin, A23187 or 12-O-tetradecanoylphorbol 13-acetate (TPA). Treatment of adipocytes with either chloroquine or dansyl cadaverine failed to affect the ability of insulin to stimulate cyclic AMP phosphodiesterase activity. Treatment of an isolated adipocyte endoplasmic reticulum membrane fraction with purified protein kinase A increased its cyclic AMP phosphodiesterase activity some 2-fold. When this fraction was treated with purified protein kinase A and [32P]ATP, label was incorporated into a 63 kDa protein which was specifically immunoprecipitated with the antiserum against the liver 'dense-vesicle' cyclic AMP phosphodiesterase.
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PMID:Subcellular localization and hormone sensitivity of adipocyte cyclic AMP phosphodiesterase. 255 12

3-Hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase is the limiting enzyme step in cholesterol formation in mammalian liver and other tissues. It is a glycoprotein of 97,000 daltons embedded in the endoplasmic reticulum with a long cytoplasmic extension that is the site of catalytic conversion of HMG CoA to mevalonate. The enzyme is subject to both long-term (induction/repression; degradation) and short-term control (reversible phosphorylation) mediated by endocrine signaling (insulin, glucagon) and through negative feedback by metabolic products of mevalonate (e.g., cholesterol). The catalytic capacity of microsomal reductase falls rapidly in the presence of several protein kinases (reductase kinase, protein kinase-C, calmodulin-dependent protein kinase). Activity is restored with various protein phosphatases. Increased phosphorylation of reductase in intact cells after addition of glucagon or mevalonate is followed by enhanced degradation of the enzyme. In an in vitro model system, phosphorylated, native microsomal reductase is more rapidly cleaved by the calcium-dependent, neutral protease calpain than the dephosphorylated from of reductase. Our present research which centers on the mechanism of the in vitro model system is reviewed. Calpain in the presence of Ca2+ cleaves the cytosolic domain of phosphorylated 97 kDa reductase at two points giving rise to two fragments of nearly the same size that appear as a 52-56,000 dalton doublet by electrophoresis and immunoblotting. In the same system native reductase labeled with [gamma-32P]ATP generates a doublet with 32P solely in the upper (heavier) band. This indicates that serine phosphorylation sites lie between the two calpain cleavage loci. These are positioned in the "linker" region of the long carboxy-terminal cytosolic domain near the membrane. This segment possesses five invariant serine residues and two PEST sequences (constellations of proline, glutamate, serine and threonine) that are characteristic of proteins with short half-lives. If phosphorylation of HMG CoA reductase is confined to the linker region, we must look to this domain in order to interpret the resulting conformational changes that markedly influence reductase catalytic activity and prepare the enzyme for degradation.
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PMID:Phosphorylation and degradation of HMG CoA reductase. 262 76

The degradation of intracellular protein and other cytoplasmic macromolecules in liver is an ongoing process that regulates cytoplasmic mass and provides amino acids for energy and other metabolic uses early in starvation. Cellular proteins are conveniently divided into two general classes according to readily discernable differences in average rates of turnover. A short-lived class, having a half-life of approximately 10 min, comprises about 0.6% of total protein. Its degradation is not physiologically controlled, and the mechanism is probably nonlysosomal in nature. The second or long-lived group, with an average half-life 250 times greater, constitutes more than 99% of the cell's protein. By contrast, its breakdown is strongly regulated, and the site of catabolism is believed to be the vacuolar-lysosomal system. Cytoplasmic sequestration by lysosomes can be divided into two categories; macro- and microautophagy. The first is induced by amino acid and/or insulin deprivation. Amino acids are considered to be primary regulators, since they can control this process over the full range of induced proteolysis in the absence of hormones. Glucagon, cyclic AMP, and beta-agonists also stimulate macroautophagy in hepatocytes but have opposite effects in myocytes. Micrautophagy differs from the former in that the cytoplasmic "bite" is smaller and the uptake process is not acutely regulated. However, the latter does decrease during starvation in parallel with basal proteolysis, effects that might be linked to the loss of endoplasmic reticulum. The primary control of macroautophagy is accomplished through a small group of direct regulators (Leu, Tyr/Phe, Gln, Pro, Met, His, and Trp) and a specific coregulatory action of alanine. As a group, regulatory amino acids produce direct inhibitory responses in the perfused rat liver that are identical to those of the complete amino acid mixture at 0.5x and 4x (times) normal plasma concentrations. However, they lose effectiveness almost completely within a narrow zone centered at normal levels, a loss that can be abolished by the addition of alanine at its normal plasma concentration (0.5 mM). At this level, alanine does not inhibit directly. Interestingly, this zonal loss is also eliminated by insulin. Glucagon, though, specifically blocks the initial inhibition evoked by 0.5x amino acid mixtures and thus induces maximal rates of protein degradation at normal amino acid concentrations.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Mechanism and regulation of protein degradation in liver. 264 36

In order to clarify the regulatory mechanism of BAT function, the effects of noradrenaline (NA) (6mM), insulin (I) (40nM), glucagon (G) (400nM) and nerve growth factor (NGF) (2-10nM), alone or in combination, were investigated directly in BAT from neonatal rats (ca. 3 days old), cultured in 10% fetal bovine serum-medium 199 in 95% air-5% CO2 gas phase at 33 degrees C for 1 to 2 weeks. I stimulated lipid accumulation and enlarged outgrown cell size, but mitochondria in the cells of tissue block were smaller and their cristae less distinct. I + G enlarged nucleus and cytoplasm, and suppressed the lipid accumulation induced by I, but mitochondria in the cells of tissue block were larger and their cristae became more prominent than those of I-added cells. G induced the similar changes to those by I + G. I + NA also induced the similar effects to those by I + G, but their mitochondria size did not differ from that of I-added cells. NGF caused the similar effects of those by G, inducing the development of mitochondria, rough endoplasmic reticulum and Golgi complex. These results suggest that multiple factors such as NA, I, G and NGF regulate differentiation and functional development of BAT.
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PMID:[Studies on regulation of the function of thermogenic tissue, brown adipose tissue (BAT) by means of tissue culture method]. 265 88

Four endocrine cell types were identified using peroxidase-antiperoxidase (PAP) technique and ultrastructurally characterized in the pancreas of Mauremys caspica in both winter and summer. In winter, insulin-immunoreactive cells were more abundant and the cell groups larger in the splenic than in the duodenal region, whereas in summer, medium or small cell groups were evenly distributed. Glucagon- and somatostatin-immunoreactive cells were found throughout the gland; they were more numerous in the splenic than in the duodenal region. Polypeptide pancreatic (PP)-immunoreactive cells were found only in the duodenal region. Somatostatin-immunoreactive cells were mainly isolated in winter and grouped in summer. Glucagon- and PP-immunoreactive cells had a similar arrangement in both seasons. Somatostatin- and PP-containing cells showed cytoplasmic processes and could be found next to the pancreatic ducts; the latter were also observed near insulin-immunoreactive cells. Some large secretory granules and numerous, isolated and long rough endoplasmic reticulum (RER) cisternae were seen in winter B cells; in summer B cells numerous lysosomes and few, dilated RER cisternae were found. Summer A cells showed well-developed, dilated RER cisternae and numerous vacuoles; secretory granules were more numerous in winter A cells. In winter B cells and summer A cells some nuclear filamentous inclusions were observed. Few RER cisternae were observed in winter D cells and many in summer D cells; secretory granules were found, the shape and electron density of which differed with the season. PP cells were characterized by their small secretory granules, which were less numerous in winter than in summer, being clustered at the cell pole or dispersed in the cytoplasm, respectively; in winter, the well-developed RER cisternae were dilated and irregularly distributed.
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PMID:Comparative study on the endocrine cells in the pancreas of Mauremys caspica (chelonia) in summer and winter. 267 1


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