title: A unique combination of plasma membrane Ca2+-ATPase isoforms is expressed in islets of Langerhans and pancreatic beta-cell lines.
abstract: Changes in free intracellular Ca2+ concentration regulate insulin secretion from pancreatic beta-cells. The existence of steep Ca2+ gradients within the beta-cell requires the presence of specialized Ca2+ exclusion systems. In this study we have characterized the plasma membrane Ca2+-ATPases (PMCAs) which extrude Ca2+ from the cytoplasm. PMCA isoform- and subtype-specific mRNA expression was investigated in rodent pancreatic alpha- and beta-cell lines, and in human and rat islets of Langerhans using reverse-transcription PCR with primers flanking the calmodulin-binding region of rat PMCA. The expression pattern of PMCA 1 and 2 was conserved in different species and islet-cell types since both rat and human islets of Langerhans and all cell lines tested contained the 1b and 2b forms. PMCA 4 isoform subtypes, however, were expressed in a cell-type-specific manner since beta-cells expressed PMCA 4b only, whereas in islets of Langerhans, which contain alpha, beta, delta and polypeptide-secreting cells, PMCA 4a and 4b were simultaneously present. No evidence was obtained for the expression of PMCA 3. Characterization of the beta-cell Ca2+-pump protein showed that it shared several similarities with the erythrocyte PMCA. It is a P-type ATPase; its phosphorylated intermediate was stabilized by La3+; it reacted with a PMCA-specific antibody; and it was not N-glycosylate. However, the beta-cell PMCA had a higher molecular mass than that of the erythrocyte; this difference could be explained by either predominant translation of the PMCA2 form, which has a molecular mass 3-8 kDa higher than the erythrocyte PMCA 1 and 4 proteins, or by a possible sequence insertion. Thus a unique combination of functionally distinct PMCA isoforms (1b, 2b, 4b) participates in Ca2+ homoeostasis in the beta-cell.
authors.affiliation: University of Oxford, Nuffield Department of Clinical Biochemistry, John Radcliffe Hospital, Headington, Oxford, U.K.
authors.forename: A
authors.initials: A
authors.lastname: Váradi
authors.forename: E
authors.initials: E
authors.lastname: Molnár
authors.forename: S J
authors.initials: SJ
authors.lastname: Ashcroft
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D000252:Calcium-Transporting ATPases / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D017931:DNA Primers; D005260:Female; D006801:Humans; D007515:Islets of Langerhans / Q000201:enzymology*; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar
chemical_list: D017931:DNA Primers; D007527:Isoenzymes; D012333:RNA, Messenger; D000252:Calcium-Transporting ATPases