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Identification of a cDNA encoding a second putative prohormone convertase related to PC2 in AtT20 cells and islets of Langerhans.

ABSTRACT: PC2 and furin are two recently identified members of a class of mammalian proteins homologous to the yeast precursor processing protease kex2 and the bacterial subtillisins. We have used the polymerase chain reaction to identify and clone a cDNA (PC3) from the mouse AtT20 anterior pituitary cell line that represents an additional member of this growing family of mammalian proteases. PC3 encodes a 753-residue protein that begins with a signal peptide and contains a 292-residue domain closely related to the catalytic modules of PC2, furin, and kex2. Within this region 58%, 65%, and 50% of the amino acids of PC3 are identical to those of the aligned PC2, furin, and kex2 sequences, respectively, and the catalytically important Asp, His, and Ser residues are all conserved. On Northern blots, PC3 hybridizes to two transcripts of 3 and 5 kilobases. Tissue distribution studies indicate that both PC2 and PC3 are expressed in a variety of neuroendocrine tissues, including pancreatic islets and brain, but are not expressed in liver, kidney, skeletal muscle, and spleen. The high degree of similarity of PC3, PC2, and furin suggests that they are all members of a superfamily of mammalian proteases that are involved in the processing of prohormones and/or other protein precursors. In contrast to furin, PC3, like PC2, lacks a hydrophobic transmembrane anchor, but it has a potential C-terminal amphipathic helical segment similar to the putative membrane anchor of carboxypeptidase H. These and other differences suggest that these proteins carry out compartmentalized proteolysis within cells, such as processing within regulated versus constitutive secretory pathways.

SUBMITTER: Smeekens SP

PROVIDER: S-EPMC50806 | biostudies-other | 1991 Jan

REPOSITORIES: biostudies-other

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Identification of a cDNA encoding a second putative prohormone convertase related to PC2 in AtT20 cells and islets of Langerhans.

Smeekens S P SP Avruch A S AS LaMendola J J Chan S J SJ Steiner D F DF

Proceedings of the National Academy of Sciences of the United States of America 19910101 2

PC2 and furin are two recently identified members of a class of mammalian proteins homologous to the yeast precursor processing protease kex2 and the bacterial subtillisins. We have used the polymerase chain reaction to identify and clone a cDNA (PC3) from the mouse AtT20 anterior pituitary cell line that represents an additional member of this growing family of mammalian proteases. PC3 encodes a 753-residue protein that begins with a signal peptide and contains a 292-residue domain closely rela ...[more]

PMID: 1988934

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Project description:Neuropeptides regulate cell-cell signaling and influence many biological processes in vertebrates, including development, growth, and reproduction. The complex processing of neuropeptides from prohormone proteins by prohormone convertases, combined with the evolutionary distance between the chicken and mammalian species that have experienced extensive neuropeptide research, has led to the empirical confirmation of only 18 chicken prohormone proteins. To expand our knowledge of the neuropeptide and prohormone convertase gene complement, we performed an exhaustive survey of the chicken genomic, EST, and proteomic databases using a list of 95 neuropeptide and 7 prohormone convertase genes known in other species. Analysis of the EST resources and 22 microarray studies offered a comprehensive portrait of gene expression across multiple conditions. Five neuropeptide genes (apelin, cocaine-and amphetamine-regulated transcript protein, insulin-like 5, neuropeptide S, and neuropeptide B) previously unknown in chicken were identified and 62 genes were confirmed. Although most neuropeptide gene families known in human are present in chicken, there are several gene not present in the chicken. Conversely, several chicken neuropeptide genes are absent from mammalian species, including C-RF amide, c-type natriuretic peptide 1 precursor, and renal natriuretic peptide. The prohormone convertases, with one exception, were found in the chicken genome. Bioinformatic models used to predict prohormone cleavages confirm that the processing of prohormone proteins into neuropeptides is similar between species. Neuropeptide genes are most frequently expressed in the brain and head, followed by the ovary and small intestine. Microarray analyses revealed that the expression of adrenomedullin, chromogranin-A, augurin, neuromedin-U, platelet-derived growth factor A and D, proenkephalin, relaxin-3, prepronociceptin, and insulin-like growth factor I was most susceptible (P-value<0.005) to changes in developmental stage, gender, and genetic line among other conditions studied. Our complete survey and characterization facilitates understanding of neuropeptides genes in the chicken, an animal of importance to biomedical and agricultural research.

Project description:BackgroundThe pig is a biomedical model to study human and livestock traits. Many of these traits are controlled by neuropeptides that result from the cleavage of prohormones by prohormone convertases. Only 45 prohormones have been confirmed in the pig. Sequence homology can be ineffective to annotate prohormone genes in sequenced species like the pig due to the multifactorial nature of the prohormone processing. The goal of this study is to undertake the first complete survey of prohormone and prohormone convertases genes in the pig genome. These genes were functionally annotated based on 35 gene expression microarray experiments. The cleavage sites of prohormone sequences into potentially active neuropeptides were predicted.ResultsWe identified 95 unique prohormone genes, 2 alternative calcitonin-related sequences, 8 prohormone convertases and 1 cleavage facilitator in the pig genome 10.2 assembly and trace archives. Of these, 11 pig prohormone genes have not been reported in the UniProt, UniGene or Gene databases. These genes are intermedin, cortistatin, insulin-like 5, orexigenic neuropeptide QRFP, prokineticin 2, prolactin-releasing peptide, parathyroid hormone 2, urocortin, urocortin 2, urocortin 3, and urotensin 2-related peptide. In addition, a novel neuropeptide S was identified in the pig genome correcting the previously reported pig sequence that is identical to the rabbit sequence. Most differentially expressed prohormone genes were under-expressed in pigs experiencing immune challenge relative to the un-challenged controls, in non-pregnant relative to pregnant sows, in old relative to young embryos, and in non-neural relative to neural tissues. The cleavage prediction based on human sequences had the best performance with a correct classification rate of cleaved and non-cleaved sites of 92% suggesting that the processing of prohormones in pigs is similar to humans. The cleavage prediction models did not find conclusive evidence supporting the production of the bioactive neuropeptides urocortin 2, urocortin 3, torsin family 2 member A, tachykinin 4, islet amyloid polypeptide, and calcitonin receptor-stimulating peptide 2 in the pig.ConclusionsThe present genomic and functional characterization supports the use of the pig as an effective animal model to gain a deeper understanding of prohormones, prohormone convertases and neuropeptides in biomedical and agricultural research.

Dataset Information

Identification of a cDNA encoding a second putative prohormone convertase related to PC2 in AtT20 cells and islets of Langerhans.

Publications

Identification of a cDNA encoding a second putative prohormone convertase related to PC2 in AtT20 cells and islets of Langerhans.

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