Chronic ethanol intake impairs insulin signaling in rats by disrupting Akt association with the cell membrane. Role of TRB3 in inhibition of Akt/protein kinase B activation.

Chronic and excessive alcohol consumption is an important and modifiable risk factor for type 2 diabetes. We previously reported elevations in hepatic Class 1 alcohol dehydrogenase (ADH) expression in ethanol-fed rats correspondent with reduced levels of mature, nuclear sterol-regulatory element-binding protein-1 (SREBP-1), an insulin-induced transcriptional repressor of the ADH gene. In this report, we have studied the effects of insulin and ethanol on ADH gene expression in a highly differentiated rat hepatoma cell line (FGC-4), as well as the in vivo effects of chronic intake of an ethanol-containing diet on hepatic insulin signaling. Insulin inhibited ADH gene expression, and this was abolished by LY294002 (a phosphatidylinositol 3-kinase inhibitor) and small interfering RNA knockdown of SREBP-1. Chronic ethanol intake led to decreased phosphorylation of Akt (protein kinase B) at Thr308, increased phosphorylation of Akt at Ser473, and decreased phosphorylation of glycogen synthase kinase-3beta (a downstream effector of Akt). Hepatic membrane-associated Akt content was decreased and cytosolic Akt content was increased in rats fed an ethanol-containing diet. Thus, disruptive effects of ethanol on insulin signaling occurred via impaired phosphorylation of Akt at Thr308. TRB3, a negative regulator of Akt, was induced in liver of ethanol-fed rats. In ethanol-treated FGC-4 cells, small interfering RNA knockdown of TRB3 increased membrane-associated Akt and the phosphorylation of Akt at Thr308. Our results suggest that ethanol induces TRB3, which, through binding to the pleckstrin homology domain of Akt, prevents its plasma membrane association, Akt-Thr308 phosphorylation, and subsequent Akt-mediated signaling. Ethanol inhibition of insulin signaling reduces nuclear SREBP accumulation and results in disinhibition of Class 1 ADH transcription.