Pyruvate carboxylase is a downstream target of p53 in regulating insulin secretion

Abstract

Pyruvate carboxylase (PC), converting pyruvate to oxaloacetate (OAA), is a critical contributor to anaplerosis in pancreatic β-cell, the process that can replenish the intermediates in Krebs cycle. The level of PC is markedly high in pancreatic β-cell, about 7-fold higher than that in α-cell. PC activity is reduced in the islets of animals with type 2 diabetes. Moreover, the rate of pyruvate carboxylation catalyzed by PC is well correlated with glucose-stimulated insulin secretion (GSIS), further supporting the important role of PC in insulin secretion. Tumor protein p53 or p53, best known for its role as a tumor suppressor, has been studied extensively for its broad influence and complex regulation. p53 suppresses tumor progression by responding to a wide variety of intrinsic and extrinsic cellular stress signals. Recent studies have revealed novel roles of p53 in response to metabolic stress, such as oxidative or nutrient stress. MDM2 is the major E3 ubiquitin ligases to control p53 activity negatively. p53 and MDM2 form a negative-feedback loop, where p53 stimulates the expression of MDM2, in turn MDM2 inhibits not only the stability but also the transcriptional activity of p53. In the previous study, mice lacking Mdm2 specifically in pancreatic β-cells have been generated, and display impaired insulin secretion and glucose metabolism. Further study has suggested that the impaired insulin secretion is caused by downregulation of pyruvate carboxylase (PC). To explore the interaction of PC with MDM2-p53 pathway in regulating insulin secretion, we inserted human PC DNA into a shuttle vector pShuttle-CMV to generate a recombinant adenovirus containing human PC gene, that is then used to restore the level of PC in Mdm2 KO islet. The recovery of insulin secretion confirmed that the downregulation of PC leads to β-cell dysfunction. Given that MDM2 is the main E3 ubiquitin protein ligase to restrain p53 activity, abnormal high level of p53 is suggested to suppress the activity of PC in Mdm2 β-cell KO mice. The putative p53 response element is found in a sequence of PC intron gene, indicating that PC can be a downstream target of p53. Using pGL3 Luciferase Reporter Vector, it is verified that p53 suppresses the activity of PC by directly targeting PC.