Abstract
The mechanistic target of rapamycin (mTOR) pathway is the master regulator of cell growth and proliferation. mTOR is a serine/threonine kinase that forms two distinct protein complexes in mammals, mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). Hyperactivation of mTORC1 is seen in certain cancers and neurodegenerative diseases, making mTORC1 inhibition a promising strategy for treating these diseases. Our laboratory recently discovered a novel small molecule mTORC1 inhibitor, CB3A. Preliminary results suggest that CB3A action requires TSC2, the main negative regulator of mTORC1, and also increases ubiquitination of TSC2. Previous studies have implicated the E3 ubiquitin ligase RNF152 as a critical negative regulator of mTORC1. Here I show that the overexpression of RNF152 causes an increase in ubiquitination of endogenous TSC2. Ubiquitination decreased with K63R- ubiquitin, suggesting that the increase in ubiquitination is K63-linked. Additionally, immunoprecipitation of overexpressed RNF152 showed CB3A causes an increase in the association between overexpressed RNF152 and endogenous TSC2. These findings suggest a possible model for the mechanism of CB3A-induced inhibition of mTORC1, where CB3A inhibits mTORC1 by increasing the K63-linked ubiquitination of TSC2 through an increase in the association between RNF152 and TSC2.