Regulation of epidermal growth factor receptor degradation by heterotrimeric Galphas protein.
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 (Galphas) promotes ligand-dependent degradation of epidermal growth factor (EGF) receptors and Texas Red EGF, and knock-down of Galphas expression by RNA interference (RNAi) delays receptor degradation. We also show that Galphas 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. Galphas coimmunoprecipitates with Hrs and binds Hrs in pull-down assays. By immunofluorescence, exogenously expressed Galphas colocalizes with myc-Hrs and GFP-RGS-PX1 on early endosomes, and expression of either Hrs or RGS-PX1 increases the localization of Galphas on endosomes. Furthermore, knock-down of both Hrs and Galphas by double RNAi causes greater inhibition of EGF receptor degradation than knock-down of either protein alone, suggesting that Galphas and Hrs have cooperative effects on regulating EGF receptor degradation. These observations define a novel regulatory role for Galphas in EGF receptor degradation and provide mechanistic insights into the function of Galphas in endocytic sorting.
Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093-0651, USA.
Bin
B
Zheng
Christine
C
Lavoie
Ting-Dong
TD
Tang
Phuong
P
Ma
Timo
T
Meerloo
Anthony
A
Beas
Marilyn G
MG
Farquhar
D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D056827:Endosomal Sorting Complexes Required for Transport; D011992:Endosomes / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism*; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D020690:GTPase-Activating Proteins / Q000378:metabolism; D006801:Humans; D010750:Phosphoproteins / Q000378:metabolism; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D020710:RGS Proteins / Q000378:metabolism; D034622:RNA Interference; D051381:Rats; D015398:Signal Transduction; D013997:Time Factors; D014966:Xanthenes / Q000494:pharmacology
D056827:Endosomal Sorting Complexes Required for Transport; D020690:GTPase-Activating Proteins; D010750:Phosphoproteins; D020710:RGS Proteins; D014966:Xanthenes; C096164:hepatocyte growth factor-regulated tyrosine kinase substrate; D004815:Epidermal Growth Factor; C034657:Texas red; D066246:ErbB Receptors; D019205:GTP-Binding Protein alpha Subunits, Gs