Elucidating the role of ubiquitination of insulin signaling molecules in HepG2 cells insulin resistance model

Abstract ID 99316 Poster Board 359 Prior studies have investigated the impact of palmitate treatment on insulin resistance and posttranslational modification of the key insulin signaling molecules. Exposure to sodium palmitate can lead to the alteration of insulin-stimulated phosphorylation of key insulin signaling molecules like insulin receptor (IR), insulin receptor substrate-1 (IRS1), insulin receptor substrate-2 (IRS2), and Protein kinase B (Akt). Our previous study demonstrated that the global ubiquitin linkage profile is altered upon insulin and glucagon treatment. We hypothesized that ubiquitination of key insulin signaling molecules is a potential mechanism by which palmitate induces insulin resistance, and the inhibition of ubiquitination can be a potential strategy for preventing and treating insulin resistance and diabetes. We investigated the IRS1 and IRS2 upon palmitate treatment in HepG2 cells. We treated HepG2 cells with or without sodium palmitate for 24 hours and probed the cell lysates for the levels of IR, IRS1, IRS2, and Akt, and their respective phosphoprotein levels. In addition, we studied the ubiquitination profile of endogenous and transiently overexpressed proteins. Linkage-specific K63, K48, and total ubiquitin antibodies showed significant alterations in the linkage profile of insulin-signaling proteins. The effect of palmitate on the ubiquitination of IRS1/2 is implicated in the development of insulin resistance; thus, revealing a connection between impaired ubiquitination equilibrium and insulin resistance. Therefore, characterizing the molecular mechanisms of impaired insulin action may reveal new therapeutic targets for diabetes. Keywords: HepG2, insulin resistance, ubiquitination, ubiquitin linkages, IRS Akt, diabetes