title: Negative charges in the flexible N-terminal domain of Rho GDP-dissociation inhibitors (RhoGDIs) regulate the targeting of the RhoGDI-Rac1 complex to membranes.
abstract: In its resting state, Rho GDP-dissociation inhibitor (RhoGDI) α forms a soluble cytoplasmic heterodimer with the GDP-bound form of Rac. Upon stimulation, the dissociation of RhoGDIα from the RhoGDIα-Rac complex is a mandatory step for Rac activation; however, this mechanism is poorly understood. In this study, we examined how the cytoplasm/membrane cycles of the RhoGDI-Rac complex are regulated, as well as where RhoGDI dissociates from the RhoGDI-Rac complex, during FcγR-mediated phagocytosis. The negatively charged and flexible N terminus (25 residues) of RhoGDIα, particularly its second negative amino acid cluster possessing five negatively charged amino acids, was a pivotal regulator in the cytoplasm/membrane cycles of the RhoGDI-Rac complex. We also found that RhoGDIα translocated to the phagosomes as a RhoGDIα-Rac1 complex, and this translocation was mediated by an interaction between the polybasic motif in the C terminus of Rac1 and anionic phospholipids produced on phagosomes, such as phosphatidic acid, that is, by a phagosome-targeting mechanism of Rac1. Thus, we demonstrated that the targeting/accumulation of the RhoGDIα-Rac1 complex to phagosomes is regulated by a balance between three factors: 1) the negatively charged and flexible N-terminal of RhoGDIα, 2) the binding affinity of RhoGDIα for Rac1, and 3) anionic phospholipids produced on phagosomes. Moreover, we demonstrated that the mechanism of targeting/accumulation of the RhoGDIα-Rac1 complex is also applicable for the RhoGDIβ-Rac1 complex.
authors.affiliation: Laboratory of Molecular Pharmacology, Biosignal Research Center, Kobe University, Kobe 657-8501, Japan. tueyama@kobe-u.ac.jp
authors.forename: Takehiko
authors.initials: T
authors.lastname: Ueyama
authors.forename: Jeonghyun
authors.initials: J
authors.lastname: Son
authors.forename: Takeshi
authors.initials: T
authors.lastname: Kobayashi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Hamada
authors.forename: Takashi
authors.initials: T
authors.lastname: Nakamura
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Sakaguchi
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Shirafuji
authors.forename: Naoaki
authors.initials: N
authors.lastname: Saito
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D051379:Mice; D018613:Microscopy, Confocal; D010588:Phagosomes; D021381:Protein Transport / Q000502:physiology; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors / Q000737:chemistry* / Q000378:metabolism*
chemical_list: C107953:RAC1 protein, human; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors; D020830:rac1 GTP-Binding Protein