Endocytosis of epidermal growth factor receptor regulated by Grb2-mediated recruitment of the Rab5 GTPase-activating protein RN-tre.
The Grb2 adaptor protein is best known for its role in signaling to the small GTPase p21(ras), mediated through its interaction with the SOS guanine nucleotide exchange factor. Here, we demonstrate that Grb2 also signals to Rab5, a small GTPase that plays a key role in early endocytic trafficking. Grb2 functions through association with RN-tre, a GTPase-activating protein for Rab5. Grb2 and RN-tre associate both in vitro and in vivo, with interaction mediated by both SH3 domains of Grb2 and extended proline-rich sequences in RN-tre. Association between Grb2 and RN-tre is constitutive and occurs independently of Eps8, a previously identified binding partner of RN-tre. Epidermal growth factor (EGF) stimulates recruitment of RN-tre to the EGF receptor (EGFR) in a Grb2-dependent manner. Grb2 and the EGFR are internalized and co-localized in endocytic vesicles in response to EGF. Overexpression of RN-tre blocks the internalization of both proteins, consistent with its function as a negative regulator of Rab5 and endocytosis. Strikingly, RN-tre does not block EGF-induced internalization of a Grb2 mutant deficient in RN-tre binding. These results 1) suggest that the ability of RN-tre to inhibit internalization of the EGFR requires Grb2-mediated binding to the receptor and 2) identify Grb2 as a critical regulator of Rab5 and EGFR endocytosis.
Martinu
Lenka
L
Department of Cell and Developmental Biology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA.
Santiago-Walker
Ademi
A
Qi
Hongwei
H
Chou
Margaret M
MM
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D019556:COS Cells; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D004705:Endocytosis / Q000502:physiology*; D004789:Enzyme Activation; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000502:physiology*; D005347:Fibroblasts; D051380:GRB2 Adaptor Protein; D020690:GTPase-Activating Proteins; D006367:HeLa Cells; D006801:Humans; D015514:Oncogene Proteins, Fusion / Q000378:metabolism*; D017354:Point Mutation; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D020696:rab5 GTP-Binding Proteins / Q000378:metabolism*; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D020690:GTPase-Activating Proteins; D015514:Oncogene Proteins, Fusion; D011506:Proteins; D011994:Recombinant Proteins; C100665:USP6NL protein, human; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D020696:rab5 GTP-Binding Proteins