MEK/ERK pathway mediates insulin-promoted degradation of MKP-3 protein in liver cells

► In this study, we examined the mechanism of insulin promoted MKP-3 protein degradation in cultured liver cells. ► We demonstrated that the MEK/ERK pathway mediates the effect of insulin on degrading MKP-3 protein. ► We further confirmed that ERK phosphorylation deficient MKP-3 mutant is resistant to insulin promoted protein degradation. ► We also showed that the MEK/ERK pathway affects glucose output, possibly through regulating MKP-3 protein level. MAP kinase phosphatase 3 (MKP-3) was recently identified as an important regulator of glucose homeostasis in the liver and its expression can be repressed by insulin at post transcriptional level. In this study, the mechanism underlying insulin promoted decrease of MKP-3 protein was investigated by studying MKP-3 protein stability via immunoblot analysis in the presence of cycloheximide using cultured liver cells. Several pathways were examined and activation of the MEK/ERK pathway was found to mediate reduction of MKP-3 protein expression in response to insulin. MEK inhibitor markedly slowed down MKP-3 protein degradation. Mutation of two ERK phosphorylation sites on MKP-3 rendered it resistant to insulin and constitutively active MEK-induced MKP-3 protein degradation. To understand the biological effect of MKP-3 protein stability on liver cell glucose output, expression level of G6Pase gene, which encodes the key enzyme controlling the last step of de novo glucose synthesis in liver cells, was examined by real time PCR analysis upon manipulation of MEK signaling. Activation of MEK pathway in Fao cells resulted in decreased expression of G6Pase gene and lowered glucose output. Consistent with this result, MEK inhibitor increased expression of G6Pase gene and glucose output in Fao cells. In conclusion, insulin likely promotes MKP-3 protein degradation through activation of MEK/ERK pathway in liver cells and MKP-3 protein level affects the capability of Fao cells to output glucose.