Abstract

Objective: Prolonged exposure of pancreatic beta cells to simultaneously elevated levels of fatty acids and glucose (glucolipotoxicity) impairs insulin gene transcription. However, the intracellular signalling pathways mediating these effects are mostly unknown. This study was aimed to ascertain the role of ERK1/2, PKB, and PASK in palmitate inhibition of insulin gene expression in pancreatic beta cells.

Research Design and Methods: MIN6 cells and isolated rat islets were cultured in the presence of elevated glucose, with or without palmitate or ceramide. ERK1/2 phosphorylation, PKB phosphorylation, and PASK expression were examined by immunoblotting and real-time PCR. The role of these kinases in insulin gene expression was assessed using pharmacological and molecular approaches.

Results: Exposure of MIN6 cells and islets to elevated glucose induced ERK1/2 and PKB phosphorylation, which was further enhanced by palmitate. Inhibition of ERK1/2, but not of PKB, partially prevented the inhibition of insulin gene expression in the presence of palmitate or ceramide. Glucose-induced expression of PASK mRNA and protein levels was reduced in the presence of palmitate. Overexpression of wild-type PASK increased insulin and PDX-1 gene expression in MIN6 cells and rat islets incubated with glucose and palmitate, while overexpression of a kinase-dead PASK mutant overexpressed in rat islets decreased expression of insulin and PDX-1 and increased C/EBPβ expression.

Conclusions: Both the PASK and ERK1/2 signalling pathways mediate palmitate inhibition of insulin gene expression. These findings identify PASK as a novel mediator of glucolipotoxicity on the insulin gene in pancreatic beta cells.