Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01189 (beta-endorphin)
 21,003 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two variant cell lines were recently established from parent AtT-20 cells. Whereas HYA.15.10.T.2 have a reduced level of secretory granules, HYA.15.6.T.3 are completely devoid of both the regulated pathway of secretion and of dense-core secretory granules. AtT-20 cells normally express the processing enzymes PC1, PC2, furin, carboxypeptidase E, and peptidylglycine alpha-amidating monooxygenase, as well as proopiomelanocortin, chromogranin B, and 7B2. We measured the expression of these mRNAs in both variant cell lines. Although some differences in mRNA level were noted, HYA.15.10.T.2 and HYA.15.6.T.3 cell lines maintained their expression of the processing enzymes and of 7B2. Furthermore, PC1 and PC2 were shown to be functionally active in the HYA.15.6.T.3 cells. In contrast, proopiomelanocortin and chromogranin B mRNA levels were no longer detectable in HYA.15.6.T.3 cells. Interestingly, stimulation of the HYA.15.6.T.3 cells with cAMP restored proopiomelanocortin mRNA, beta-endorphin immunoreactivity, and dense-core granules. Furthermore, at the ultrastructural level, beta-lipotropin immunoreactivity was detected in granules of cAMP-induced HYA.15.6.T.3 cells. Finally, depolarization of cAMP-induced HYA.15.6.T.3 cells with 56 mM potassium chloride resulted in a marked increase in the release of beta-endorphin immunoreactivity. These observations demonstrate that cAMP restores the regulated pathway of secretion in HYA.15.6.T.3 cells, which under untreated conditions do not demonstrate regulated release. These variant cell lines are unique models to understand better the relationship of the regulated pathway and the expression of the processing enzymes.
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PMID:Maintained PC1 and PC2 expression in the AtT-20 variant cell line 6T3 lacking regulated secretion and POMC: restored POMC expression and regulated secretion after cAMP treatment. 786 35

POMC processing is frequently altered in ACTH-secreting nonpituitary tumors in which intermediate lobe-like peptides such as corticotropin-like intermediate lobe peptide (CLIP) are occasionally generated. In rodent pituitaries, the exclusive presence of prohormone convertase PC2 in the melanotrophs of the intermediate lobe is responsible for the specific conversion of ACTH to alpha MSH and CLIP, by contrast with corticotrophs of the anterior lobe, which do not contain PC2 and, therefore, only produce ACTH. The goal of our study was to look for PC2 expression in ACTH-secreting nonpituitary tumors in man. Using Northern blot analysis, PC2 transcripts were detected in five nonpituitary tumors that contained large proportions of CLIP (from 40-95% of the total C-terminal immunoreactive ACTH). A predominant PC2 messenger ribonucleic acid migrated with an apparent mol wt of 5 kilobases, and a minor signal at 3 kilobases was also detected. No PC2 messenger ribonucleic acid could be detected in the small cell carcinoma of the lung-derived DMS-79 human cell line, which produces unprocessed POMC, or in three pituitary tumors responsible for Cushing's disease or Nelson's syndrome, which produced intact ACTH, but no CLIP. These data strongly suggest that, as in rodents, PC2 is responsible for the production of smaller POMC end products, such as CLIP, frequently observed in ACTH-secreting nonpituitary tumors in man.
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PMID:Expression of the prohormone convertase PC2 correlates with the presence of corticotropin-like intermediate lobe peptide in human adrenocorticotropin-secreting tumors. 796 50

The prohormone convertases, PC1 (SPC3) and PC2, are subtilisin-like serine proteases capable of processing neuropeptide precursors. In cotransfection experiments, other investigators have found that PC1 and PC2 can process POMC to appropriate peptide products. In this study, recombinant rat PC1 was stably expressed in a mouse L-cell line and partially purified. Mouse POMC was cleaved by recombinant PC1 to generate ACTH intermediates, ACTH, ACTH linked to joining peptide, joining peptide, 16-kilodalton N-POMC, N-POMC-(1-74), and beta-lipotropin. Recombinant PC1 was also found to cleave ACTH to ACTH-(1-15) and bovine N-POMC-(1-77) to gamma 3 MSH. The pH optimum of the cleavages was 6.0. We conclude that recombinant PC1 is capable of processing POMC in vitro at all of the paired basic residues, with the exception of Lys-Arg and Lys-Lys in beta-lipotropin and beta-endorphin, respectively. This in vitro study showed a more general specificity of recombinant PC1 for paired and tetrabasic residues of POMC than was previously found in cotransfection experiments. Other cellular regulatory mechanisms probably play a role in limiting the processing of POMC in vivo in the anterior pituitary, where gamma 3 MSH and alpha MSH are not found in significant amounts.
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PMID:In vitro processing of proopiomelanocortin by recombinant PC1 (SPC3). 807 Mar 78

Mice harboring a transgene composed of proopiomelanocortin (POMC) gene promoter sequences (nucleotides -706 to +64) ligated to the simian virus (SV) 40 early gene encoding large T antigen developed large POMC-expressing pituitary tumors. Histologically the tumors arose from the intermediate lobe, contained nuclear SV40 T antigen and POMC peptides, but did not express other pituitary hormones. POMC processing in the pituitary tumors was indistinguishable from normal mouse intermediate lobe melanotrophs and was characterized by high proportions of acetylated and carboxyl-terminal shortened beta-endorphins, and amino-terminal acetylated alpha-melanocyte-stimulating hormone, and virtually no adrenocorticotropic hormone (ACTH)(1-39), beta-lipotropin, or POMC. The tumors contained abundant levels of mRNA for the prohormone convertase PC2 and undetectable levels of PC1. Normal mouse neurointermediate lobe also has a high ratio of PC2/PC1 expression that is distinct from the relative abundance of PC1 in anterior lobe and AtT-20 corticotroph cells. In contrast, extracts from tumors transplanted subcutaneously in nude mice contained predominantly nonacetylated forms of beta-endorphin(1-31) and -(1-27), very little ACTH(1-39), almost no corticotropin-like intermediate peptide or alpha-melanocyte-stimulating hormone, and higher proportions of intact POMC. Surprisingly, despite the less efficient proteolytic cleavage, a transplanted tumor expressed both PC1 and PC2. These studies are the first biochemical documentation of a melanotroph pituitary tumor in a rodent species and provide a new model for the investigation of pituitary oncogenesis and the molecular basis of tissue-specific prohormone post-translational processing.
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PMID:Post-translational processing of proopiomelanocortin (POMC) in mouse pituitary melanotroph tumors induced by a POMC-simian virus 40 large T antigen transgene. 822 58

We have investigated processing of monkey pro-opiomelanocortin (POMC) following transfection into heterologous neuronal Neuro 2A (N2A) cells. In several separately transfected stable cell lines (termed N2A/POMC2-like; n = 4), POMC was processed to beta E only, by direct cleavage from the precursor. Thus, these cell lines did not produce beta E in the orderly manner observed in the pituitary, that is, via the intermediate peptide beta LPH. Analysis of one representative N2A/POMC2 cell line revealed that the extent of processing to beta E appeared to be negatively correlated with precursor expression level, suggesting that the processing enzyme(s) in these cells was present in limiting amounts. Northern analysis of PC1 and PC2, two recently cloned processing enzymes, showed that N2A/POMC2 cells expressed low levels of PC2 mRNA, but no detectable PC1 mRNA. These data suggest that (1) the order of processing observed in the pituitary is not exclusively determined by tertiary folding of the precursor, but rather by the complement of processing enzymes in a particular cell, and (2) if PC2 is responsible for POMC processing in N2A/POMC2 cells, this enzyme, expressed in limiting amounts, appeared to show selectivity for the beta E amino terminal processing site.
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PMID:Characterization of pro-opiomelanocortin processing in heterologous neuronal cells that express PC2 mRNA. 832 69

By using reverse transcriptase/PCR and oligonucleotide sequences derived from conserved segments (including the conserved RRGDL sequence) of the known proprotein convertases (PCs) PC1, PC2, furin, and PC4, we identified a subtilisin/kexin-like PC called PC5 in both mouse and rat tissues. The composite structure (2.85 kb) was deduced from the analysis of the reverse transcription/PCR products combined with the sequence from a clone isolated from a cDNA library made from corticotropin-activated mouse adrenocortical Y1 cells. The deduced cDNA structures of mouse PC5 and rat PC5 showed that the closest homologue is PACE4. Furthermore, like furin, Drosophila melanogaster (d) dfurin2, and PACE4, PC5 shows the presence of a C-terminal Cys-rich domain containing either 5 (PC5 and PACE4) or 10 (dfurin2) repeats of the consensus motif Cys-Xaa2-Cys-Xaa3-Cys-Xaa(5-7)-Cys-Xaa2-Cys-Xaa (8-15)-Cys-Xaa3-Cys-Xaa(9-16). The richest sources of rat PC5 mRNA (3.8 kb) are the adrenal and gut, but it can also be detected in many endocrine and nonendocrine tissues. Corticotropin-stimulated adrenocortical Y1 cells showed an increased expression of PC5 mRNA, suggesting an upregulation by cAMP. In situ hybridization of rat brain sections demonstrated a unique distribution of PC5 compared to PC1, PC2, and furin.
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PMID:cDNA structure of the mouse and rat subtilisin/kexin-like PC5: a candidate proprotein convertase expressed in endocrine and nonendocrine cells. 834 87

A lysed preparation of isolated insulin secretory granules efficiently cleaved murine proopiomelanocortin (mPOMC) at physiologically important Lys-Arg processing sites. This processing was mostly attributed to an activity that co-eluted with the proinsulin processing type-II endopeptidase from anion exchange chromatography (Lys-Arg-directed; Davidson, H. W., Rhodes, C. J., and Hutton, J. C. (1988) Nature 333, 93-96). The principal peptide hormone products generated by the insulin secretory granule lysate were identified by specific radioimmunoassay and NH2-terminal microsequencing analysis of high performance liquid chromatography-separated products as alpha-melanocyte-stimulating hormone, corticotropin-like intermediate, gamma-lipotropin, beta-endorphin-(1-31), 18-kDa NH2-terminal fragment and, to a lesser extent, adrenocorticotrophin and beta-lipotropin. This processing had an acidic pH optimum (pH 5-5.5) and was Ca(2+)-dependent (K0.5 activation = 5-80 microM). With increasing Ca2+ concentrations there was an increase in the extent to which mPOMC was processed. The in vitro processing of mPOMC by the insulin secretory granule endopeptidase activity reported here is in excellent agreement with the in vivo processing of this prohormone by a combination of PC2 and PC3, candidates of prohormone endpeptidase, in gene transfer studies with cells that express the regulated secretory pathway (Thomas, L., Leduc, R., Thorne, B. A., Smeekens, S. S., Steiner, D. F., and Thomas, G. (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 5297-5301).
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PMID:Processing of proopiomelanocortin by insulin secretory granule proinsulin processing endopeptidases. 838 98

In the adult pituitary, anterior lobe corticotrophs and intermediate lobe melanotrophs differentially process proopiomelanocortin (POMC). Within the corticotrophs, POMC is processed mainly to corticotropin (ACTH) and beta-lipotropin, while alpha-melanotropin (alpha MSH) and beta-endorphin are the major end products in the melanotrophs. The observed transient presence of alpha MSH-like immunoreactivity during ontogeny suggested an age-dependent variation in POMC processing in the adenohypophysis. In this tissue, cell-specific POMC products are likely the result of differential expression of the two known prohormone convertases PC1 and PC2. In the present ontogeny study done in the mouse intermediate and anterior pituitary, we examined how the expression pattern of PC1 and PC2 mRNA transcripts correlates with that of ACTH and alpha MSH-like immunoreactivities. Our data demonstrated that both PC1 and PC2 transcripts can be detected in the presumptive adenohypophysis starting on embryonic day 15 (E15). In the intermediate lobe, PC1 and PC2 mRNAs appear on E18 and E16, respectively, and their levels increased during ontogeny, reaching maximal expression in the adult. Similarly, PC1 expression in the anterior pituitary increased from E15 to adulthood. However, PC2 mRNA expression peaked between postnatal days 1 (P1) and 14 (P14) and then decreased to adult levels. The distribution of PC1 and PC2 immunoreactivity is nicely correlated with the in situ hybridization data. In the anterior lobe, during the P1-P14 postnatal period, PC2 immunoreactivity was detected within cells synthesizing an alpha MSH-like peptide(s). This observation substantiates our earlier biochemical data suggesting that PC2 is the important convertase in the processing of POMC into alpha MSH. Furthermore, the demonstrated variation in the relative ratio of PC1/PC2 expression during ontogeny rationalizes the observed plasticity of POMC processing in the adenohypophysis. It is expected that beta-endorphin processing will follow that of alpha MSH.
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PMID:Ontogeny of the prohormone convertases PC1 and PC2 in the mouse hypophysis and their colocalization with corticotropin and alpha-melanotropin. 838 57

Chromogranins A and B and secretogranin II are a family of acidic proteins found in neuroendocrine secretory vesicles; these proteins contain multiple potential cleavage sites for proteolytic processing by the mammalian subtilisin-like serine endoproteases PC1 and PC2 (prohormone convertases 1 and 2), and furin. We explored the role of these endoproteases in chromogranin processing in AtT-20 mouse pituitary corticotropes. Expression of inducible antisense PC1 mRNA virtually abolished PC1 immunoreactivity on immunoblots. Chromogranin A immunoblots revealed chromogranin A processing, from both the NH2 and COOH termini, in both wild-type AtT-20 and AtT-20 antisense PC1 cells. After antisense PC1 induction, an approximately 66-kD chromogranin A NH2-terminal fragment as well as the parent chromogranin A molecule accumulated, while an approximately 50 kD NH2-terminal and an approximately 30 kD COOH-terminal fragment declined in abundance. Chromogranin B and secretogranin II immunoblots showed no change after PC1 reduction. [35S]Methionine/cysteine pulse-chase metabolic labeling in AtT-20 antisense PC1 and antisense furin cells revealed reciprocal changes in secreted chromogranin A COOH-terminal fragments (increased approximately 82 kD and decreased approximately 74 kD forms, as compared with wild-type AtT-20 cells) indicating decreased cleavage, while AtT-20 cells overexpressing PC2 showed increased processing to and secretion of approximately 71 and approximately 27 kD NH2-terminal chromogranin A fragments. Antisense PC1 specifically abolished regulated secretion of both chromogranin A and beta-endorphin in response to the usual secretagogue, corticotropin-releasing hormone. Moreover, immunocytochemistry demonstrated a relative decrease of chromogranin A in processes (where regulated secretory vesicles accumulate) of AtT-20 cells overexpressing either PC1 or PC2. These results demonstrate that chromogranin A is a substrate for the endogenous endoproteases PC1 and furin in vivo, and that such processing influences its trafficking into the regulated secretory pathway; furthermore, lack of change in chromogranin B and secretogranin II cleavage after diminution of PCl suggests that the action of PC1 on chromogranin A may be specific within the chromogranin/secretogranin protein family.
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PMID:Chromogranin A processing and secretion: specific role of endogenous and exogenous prohormone convertases in the regulated secretory pathway. 869 Jul 87

Pro-opiomelanocortin (POMC) is a polyhormone precursor produced predominantly in the pars distalis and pars intermedia of the pituitary gland where it undergoes tissue specific processing to produce a whole array of peptides. We have shown previously that peptides derived from the N-terminal region of POMC are involved in adrenal growth in rats. Using specific two site immunoradiometric assays we have found that the plasma of 17 week old fetal sheep contain a 50 fold excess of pro-gamma-MSH over ACTH. As term approached, the levels of pro-gamma-MSH fell and ACTH rose with evidence of fragmentation of pro-gamma-MSH, suggesting that these peptides act in concert in the development of the fetal adrenal cortex and also provide the necessary drive to bring about parturition. In an attempt to explore the pathophysiology of adrenal function we have cloned human POMC cDNA which led to the discovery of a 9bp addition/deletion mutation in the C-terminus of gamma 3-MSH between positions 67-73. Chinese hamster ovary cells (CHO) cells stably transfected with constructs containing the variant POMC cDNAs have shown a degree of partial processing. Work is currently underway to further investigate the effects of these mutations on the processing by the prohormone converting enzymes PC1 and PC2.
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PMID:Pro-opiomelanocortin and adrenal function. 896 87


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