Regulation of Epidermal Growth Factor Receptor Degradation by Heterotrimeric Gαs Protein

Abstract

Heterotrimeric G proteins have been implicated in the regulation of membrane trafficking, but the mechanisms involved are not well understood. Here, we report that overexpression of the stimulatory G protein subunit (Gαs) promotes ligand-dependent degradation of epidermal growth factor (EGF) receptors and Texas Red EGF, and knock-down of Gαs expression by RNA interference (RNAi) delays receptor degradation. We also show that Gαs and its GTPase activating protein (GAP), RGS-PX1, interact with hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs), a critical component of the endosomal sorting machinery. Gαs coimmunoprecipitates with Hrs and binds Hrs in pull-down assays. By immunofluorescence, exogenously expressed Gαs colocalizes with myc-Hrs and GFP-RGS-PX1 on early endosomes, and expression of either Hrs or RGS-PX1 increases the localization of Gαs on endosomes. Furthermore, knock-down of both Hrs and Gαs by double RNAi causes greater inhibition of EGF receptor degradation than knock-down of either protein alone, suggesting that Gαs and Hrs have cooperative effects on regulating EGF receptor degradation. These observations define a novel regulatory role for Gαs in EGF receptor degradation and provide mechanistic insights into the function of Gαs in endocytic sorting.