title: EFA6 controls Arf1 and Arf6 activation through a negative feedback loop.
abstract: Guanine nucleotide exchange factors (GEFs) of the exchange factor for Arf6 (EFA6), brefeldin A-resistant Arf guanine nucleotide exchange factor (BRAG), and cytohesin subfamilies activate small GTPases of the Arf family in endocytic events. These ArfGEFs carry a pleckstrin homology (PH) domain in tandem with their catalytic Sec7 domain, which is autoinhibitory and supports a positive feedback loop in cytohesins but not in BRAGs, and has an as-yet unknown role in EFA6 regulation. In this study, we analyzed how EFA6A is regulated by its PH and C terminus (Ct) domains by reconstituting its GDP/GTP exchange activity on membranes. We found that EFA6 has a previously unappreciated high efficiency toward Arf1 on membranes and that, similar to BRAGs, its PH domain is not autoinhibitory and strongly potentiates nucleotide exchange on anionic liposomes. However, in striking contrast to both cytohesins and BRAGs, EFA6 is regulated by a negative feedback loop, which is mediated by an allosteric interaction of Arf6-GTP with the PH-Ct domain of EFA6 and monitors the activation of Arf1 and Arf6 differentially. These observations reveal that EFA6, BRAG, and cytohesins have unanticipated commonalities associated with divergent regulatory regimes. An important implication is that EFA6 and cytohesins may combine in a mixed negative-positive feedback loop. By allowing EFA6 to sustain a pool of dormant Arf6-GTP, such a circuit would fulfill the absolute requirement of cytohesins for activation by Arf-GTP before amplification of their GEF activity by their positive feedback loop.
authors.affiliation: Laboratoire d'Enzymologie et Biochimie Structurales, Centre National de la Recherche Scientifique, Centre de Recherche de Gif, 91198 Gif-sur-Yvette, France and dominique.padovani@parisdescartes.fr cherfils@lebs.cnrs-gif.fr.
authors.forename: Dominique
authors.initials: D
authors.lastname: Padovani
authors.affiliation: Laboratoire d'Enzymologie et Biochimie Structurales, Centre National de la Recherche Scientifique, Centre de Recherche de Gif, 91198 Gif-sur-Yvette, France and.
authors.forename: Marcia
authors.initials: M
authors.lastname: Folly-Klan
authors.affiliation: Laboratoire d'Enzymologie et Biochimie Structurales, Centre National de la Recherche Scientifique, Centre de Recherche de Gif, 91198 Gif-sur-Yvette, France and.
authors.forename: Audrey
authors.initials: A
authors.lastname: Labarde
authors.affiliation: Institut de Pharmacologie Moléculaire et Cellulaire, Centre National de la Recherche Scientifique and University of Nice-Sophia Antipolis, 06560 Sophia-Antipolis, France.
authors.forename: Sonia
authors.initials: S
authors.lastname: Boulakirba
authors.affiliation: Laboratoire d'Enzymologie et Biochimie Structurales, Centre National de la Recherche Scientifique, Centre de Recherche de Gif, 91198 Gif-sur-Yvette, France and.
authors.forename: Valérie
authors.initials: V
authors.lastname: Campanacci
authors.affiliation: Institut de Pharmacologie Moléculaire et Cellulaire, Centre National de la Recherche Scientifique and University of Nice-Sophia Antipolis, 06560 Sophia-Antipolis, France.
authors.forename: Michel
authors.initials: M
authors.lastname: Franco
authors.affiliation: Laboratoire d'Enzymologie et Biochimie Structurales, Centre National de la Recherche Scientifique, Centre de Recherche de Gif, 91198 Gif-sur-Yvette, France and.
authors.forename: Mahel
authors.initials: M
authors.lastname: Zeghouf
authors.affiliation: Laboratoire d'Enzymologie et Biochimie Structurales, Centre National de la Recherche Scientifique, Centre de Recherche de Gif, 91198 Gif-sur-Yvette, France and dominique.padovani@parisdescartes.fr cherfils@lebs.cnrs-gif.fr.
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Cherfils
mesh_terms: D020823:ADP-Ribosylation Factor 1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution; D000818:Animals; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D006224:Cricetinae; D004789:Enzyme Activation; D025461:Feedback, Physiological; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008081:Liposomes; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015238:Sulfotransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D000089982:ADP-Ribosylation Factor 6; C411874:CHST15 protein, human; D015815:Cell Adhesion Molecules; D020662:Guanine Nucleotide Exchange Factors; D008081:Liposomes; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; C109970:PSD protein, human; D011994:Recombinant Proteins; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D015238:Sulfotransferases; D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors; C000715348:ARF6 protein, human
keywords: autoinhibition; endocytosis; kinetics; membrane traffic
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