title: Regulation of Arf6 and ACAP1 signaling by the PTB-domain-containing adaptor protein GULP.
abstract: The GTPase Arf6 regulates multiple cellular processes, including endocytosis, secretion, phagocytosis, cell adhesion, and cell migration [1, 2]. The Arf6-specific GAP ACAP1 is a negative regulator of Arf6-mediated signaling [3-7]. However, regulation of ACAP1- and Arf6-mediated signaling by other cellular proteins is not well understood. GULP/CED-6 is a phosphotyrosine binding (PTB)-domain-containing adaptor protein linked to engulfment of apoptotic cells [8-13] and to cholesterol homeostasis [14]. Here, we identify a novel role for GULP as a positive regulator of Arf6. Knockdown of GULP decreased cellular Arf6-GTP, whereas GULP overexpression increased cellular Arf6-GTP. At the mechanistic level, GULP influenced Arf6 at four levels. First, GULP bound directly to GDP-bound Arf6 via its PTB domain. Second, GULP associated with the Arf6-GAP ACAP1 at endogenous levels. Third, GULP reversed the Arf6-GTP decrease induced by ACAP1, and countered the ACAP1-mediated inhibition of cell migration. Fourth, GULP, ACAP1, and GDP-bound Arf6 were part of a tripartite complex, suggesting sequestration of ACAP1 as one mechanism of GULP action. Taken together, these data identify GULP as a modifier of cellular Arf6-GTP through regulation of ACAP1. Because PTB-domain-containing adaptor proteins influence endocytosis and trafficking of membrane proteins and cell migration [15, 16], our data support a model wherein PTB-domain-containing adaptor proteins regulate Arf family proteins.
authors.affiliation: Carter Immunology Center, Department of Microbiology, University of Virginia, Charlottesville, Virginia 22908, USA.
authors.forename: Zhong
authors.initials: Z
authors.lastname: Ma
authors.forename: Zhongzhen
authors.initials: Z
authors.lastname: Nie
authors.forename: Ruibai
authors.initials: R
authors.lastname: Luo
authors.forename: James E
authors.initials: JE
authors.lastname: Casanova
authors.forename: Kodi S
authors.initials: KS
authors.lastname: Ravichandran
mesh_terms: D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D020690:GTPase-Activating Proteins / Q000378:metabolism*; D006153:Guanosine Diphosphate / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D017434:Protein Structure, Tertiary
chemical_list: D000089982:ADP-Ribosylation Factor 6; D048868:Adaptor Proteins, Signal Transducing; D020690:GTPase-Activating Proteins; D006153:Guanosine Diphosphate; D020727:ADP-Ribosylation Factors; C000715348:ARF6 protein, human; C000715727:Arf6 protein, mouse