title: Endocytic membrane turnover at the leading edge is driven by a transient interaction between Cdc42 and GRAF1.
abstract: Changes in cell morphology require coordination of plasma membrane turnover and cytoskeleton dynamics, processes that are regulated by Rho GTPases. Here, we describe how a direct interaction between the Rho GTPase Cdc42 and the GTPase-activating protein (GAP) GRAF1 (also known as ARHGAP26), facilitates rapid cell surface turnover at the leading edge. Both Cdc42 and GRAF1 were required for fluid-phase uptake and regulated the generation of transient GRAF1-coated endocytic carriers, which were distinct from clathrin-coated vesicles. GRAF1 was found to transiently assemble at discrete Cdc42-enriched punctae at the plasma membrane, resulting in a corresponding decrease in the microdomain association of Cdc42. However, Cdc42 captured in its active state was, through a GAP-domain-mediated interaction, localised together with GRAF1 on accumulated internal structures derived from the cell surface. Correlative fluorescence and electron tomography microscopy revealed that these structures were clusters of small membrane carriers with defective endosomal processing. We conclude that a transient interaction between Cdc42 and GRAF1 drives endocytic turnover and controls the transition essential for endosomal maturation of plasma membrane internalised by this mechanism.
authors.affiliation: Integrative Medical Biology, Umeå University, Umeå 901 87, Sweden Medical Biochemistry and Biophysics, Umeå University, Umeå 901 87, Sweden.
authors.forename: Monika K
authors.initials: MK
authors.lastname: Francis
authors.affiliation: Integrative Medical Biology, Umeå University, Umeå 901 87, Sweden.
authors.forename: Mikkel R
authors.initials: MR
authors.lastname: Holst
authors.affiliation: Integrative Medical Biology, Umeå University, Umeå 901 87, Sweden.
authors.forename: Maite
authors.initials: M
authors.lastname: Vidal-Quadras
authors.affiliation: Medical Biochemistry and Biophysics, Umeå University, Umeå 901 87, Sweden Molecular Biology, Umeå University, Umeå 901 87, Sweden.
authors.forename: Sara
authors.initials: S
authors.lastname: Henriksson
authors.affiliation: European Molecular Biology Laboratory, Meyerhofstr. 1, Heidelberg 69 117, Germany.
authors.forename: Rachel
authors.initials: R
authors.lastname: Santarella-Mellwig
authors.affiliation: Molecular Biology, Umeå University, Umeå 901 87, Sweden.
authors.forename: Linda
authors.initials: L
authors.lastname: Sandblad
authors.affiliation: Integrative Medical Biology, Umeå University, Umeå 901 87, Sweden Medical Biochemistry and Biophysics, Umeå University, Umeå 901 87, Sweden richard.lundmark@umu.se.
authors.forename: Richard
authors.initials: R
authors.lastname: Lundmark
mesh_terms: D000199:Actins / Q000378:metabolism*; D022163:Clathrin-Coated Vesicles / Q000378:metabolism; D004705:Endocytosis; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D020764:cdc42 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
chemical_list: C115971:ARHGAP26 protein, human; D000199:Actins; D020690:GTPase-Activating Proteins; D020764:cdc42 GTP-Binding Protein
keywords: Actin; Cdc42; Cell surface; Clathrin-independent endocytosis; GRAF1