RESUMO
Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is expressed in insulin-secreting ß cells. However, the effects of CaMKII on insulin synthesis are unknown. Although Ser133 phosphorylation of cyclic AMP-responsive element-binding protein (CREB) typically increases CREB transcriptional activity, CaMKII phosphorylates CREB at Ser142 and at Ser133 to exert a dominant inhibitory effect. Our objective was to characterize the role of CaMKII in insulin gene expression. In MIN6 cells, insulin gene promoter activity was significantly down-regulated by wild-type (WT) CaMKIIδ2, but was significantly upregulated after small interfering RNA (siRNA) knockdown of CaMKIIδ expression. These results were independent of glucose concentrations and membrane depolarization. Insulin mRNA levels were also decreased by WT CaMKIIδ2 and increased by CaMKIIδ siRNA. Downregulation of insulin gene promoter activity by WT CaMKIIδ2 was partly mediated via cyclic AMP-responsive element 2 (CRE2). WT CaMKIIδ2 significantly increased CREB phosphorylation at Ser142 and significantly decreased binding to CREB binding protein (CBP), whereas kinase dead CaMKIIδ2 did not. Our results indicate that CaMKIIδ2 downregulates insulin gene expression by Ser142 phosphorylation of CREB and reducing binding of CREB to CBP.
Assuntos
Proteína de Ligação a CREB/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Regulação da Expressão Gênica , Insulina/genética , Animais , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Linhagem Celular Tumoral , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Regulação para Baixo , Técnicas de Silenciamento de Genes , Camundongos , Fosforilação , Regiões Promotoras Genéticas , Serina/genética , Serina/metabolismoRESUMO
Hepatic gluconeogenesis is crucial for glucose homeostasis. Although sirtuin 1 (Sirt1) is implicated in the regulation of gluconeogenesis in the liver, the effects of other histone deacetylases (HDAC) on gluconeogenesis are unclear. The aim of this study was to identify the role of class I HDACs in hepatic gluconeogenesis. In HepG2 cells and the liver of mice, the expressions of phosphoenol pyruvate carboxykinase (PEPCK) and hepatocyte nuclear factor 4α (HNF4α) were significantly decreased by treatment with a newly designed class I HDAC inhibitor, Ky-2. SiRNA knockdown of HDAC1 expression, but not of HDAC2 or HDAC3, in HepG2 cells decreased PEPCK and HNF4α expression. In HepG2 cells, insulin-stimulated phosphorylation of Akt and forkhead box O 1 (FoxO1) was increased by Ky-2. Pyruvate tolerance tests in Ky-2-treated high-fat-diet (HFD)-fed mice showed a marked reduction in blood glucose compared with vehicle-treated HFD mice. These data suggest that class I HDACs increase HNF4α protein expression and the transcriptional activity of FoxO1, followed by the induction of PEPCK mRNA expression and gluconeogenesis in liver.