title: Expression of the Slc12a1 gene in pancreatic β-cells: molecular characterization and in silico analysis.
abstract: The solute carrier protein family 12 group A, member one (Slc12a1) and two (Slc12a2) encode several splice variants of the kidney-specific and the ubiquitous isoforms, respectively, of the bumetanide (BTD)-sensitive Na-dependent K2Cl co-transporter. The Slc12a2 co-transporter is involved in the maintenance of a high intracellular chloride concentration [Cl(-)](i) in β-cells and its inhibition with BTD blocks glucose-induced insulin secretion. In β-cells, [Cl(-)](i) plays an important role in glucose-induced depolarization and insulin secretion. Glucose promotes electrogenic efflux of Cl(-) contributing to β-cell's electrical and secretory activity. To identify the expression pattern of Slc12a1 and Slc12a2 genes in β-cells we have used RT-PCR, Western blotting and immunolocalization studies in mouse pancreatic islets, β-cell lines and rat tissues. Our results demonstrate expression of specific splice variants of Slc12a1 and Slc12a2 transcripts in β-cells i.e., variants 1 of Slc12a1 (NKCC2A) and Slc12a2 (NKCC1a). Molecular cloning and characterization of Slc12a1 variant 1 transcripts from β-cells revealed an alternative splicing event involving the 5'-UTR region. NKCC2A expression at the protein level in islets and β-cells was confirmed by immunoblotting and immunolocalization. Further, NKCC2A, NKCC1a and pro-insulin co-localized in β-cells but not in the exocrine pancreas. Therefore, our results provide for the first time evidence of NKCC2A expression in pancreatic β-cells where it may play a role in insulin secretion.
authors: Department of Pharmacology and Toxicology, Wright State University, School of Medicine, Dayton, OH 45435, USA.
authors: Saeed
authors: S
authors: Alshahrani
authors: Francisco J
authors: FJ
authors: Alvarez-Leefmans
authors: Mauricio
authors: M
authors: Di Fulvio
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D020224:Expressed Sequence Tags; D015870:Gene Expression; D007328:Insulin / Q000235:genetics / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D009928:Organ Specificity; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D051381:Rats; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D028021:Sodium-Potassium-Chloride Symporters / Q000235:genetics / Q000378:metabolism*; D064466:Solute Carrier Family 12, Member 1; D064506:Solute Carrier Family 12, Member 2
chemical_list: D007328:Insulin; D020033:Protein Isoforms; C578715:Slc12a1 protein, mouse; C578716:Slc12a1 protein, rat; C578878:Slc12a2 protein, mouse; C578879:Slc12a2 protein, rat; D028021:Sodium-Potassium-Chloride Symporters; D064466:Solute Carrier Family 12, Member 1; D064506:Solute Carrier Family 12, Member 2