Ca 2+/calmodulin-dependent protein kinase II δ 2 regulates gene expression of insulin in INS-1 rat insulinoma cells

Abstract

Ca 2+/calmodulin-dependent protein kinase II is a member of a broad family of ubiquitously expressed Ca 2+ sensing serine/threonine-kinases. Ca 2+/calmodulin-dependent protein kinase II is highly expressed in insulin secreting cells and is associated with insulin secretory granules and has been proposed to play an important role in exocytosis or in insulin granule transport to release sites. To elucidate its function the antisense sequence of the major β-cell subtype, Ca 2+/calmodulin-dependent protein kinase II δ 2, was stably expressed in INS-1 rat insulinoma cells. This caused a loss of Ca 2+/calmodulin-dependent protein kinase II δ 2 expression at the mRNA and protein level, while the expression of the 95% homologous Ca 2+/calmodulin-dependent protein kinase II γ and of β-cell specific proteins such as the homeodomain factor pancreatic–duodenal homeobox factor-1 (PDX-1, also referred to as islet/duodenum homeobox-1, IDX-1, insulin promoter factor-1, IPF-1 and somatostatin transactivating factor-1, STF-1), the glucagon-like peptide-1 (GLP-1) receptor and K ATP-channels K IR6.2/SUR-1 (sulfonylurea receptor-1) was not altered. Unexpectedly, the cells showed a large reduction of insulin gene expression, which was due to reduced insulin gene transcription. Electrophoretic mobility shift assays of PDX-1 binding to the insulin promoter A1 and E2/A3A4 elements showed additional bands indicating alterations of PDX-1 complex formation. Stable over expression of Ca 2+/calmodulin-dependent protein kinase II δ 2, by contrast, was associated with elevated expression of insulin mRNA. Therefore, we conclude that Ca 2+/calmodulin-dependent protein kinase II δ 2 links fuel-dependent increases in intracellular Ca 2+ concentrations to transcriptional regulation of genes related to the metabolic control of insulin secretion.