The yeast Arf-GAP Glo3p is required for the endocytic recycling of cell surface proteins Journal Article

Author(s): Kawada, Daiki; Kobayashi, Hiromu; Tomita, Tsuyoshi; Nakata, Eisuke; Nagano, Makoto; Siekhaus, Daria E; Toshima, Junko Y; Toshimaa, Jiro
Article Title: The yeast Arf-GAP Glo3p is required for the endocytic recycling of cell surface proteins
Affiliation IST Austria
Abstract: Small GTP-binding proteins of the Ras superfamily play diverse roles in intracellular trafficking. Among them, the Rab, Arf, and Rho families function in successive steps of vesicle transport, in forming vesicles from donor membranes, directing vesicle trafficking toward target membranes and docking vesicles onto target membranes. These proteins act as molecular switches that are controlled by a cycle of GTP binding and hydrolysis regulated by guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs). In this study we explored the role of GAPs in the regulation of the endocytic pathway using fluorescently labeled yeast mating pheromone α-factor. Among 25 non-essential GAP mutants, we found that deletion of the GLO3 gene, encoding Arf-GAP protein, caused defective internalization of fluorescently labeled α-factor. Quantitative analysis revealed that glo3Δ cells show defective α-factor binding to the cell surface. Interestingly, Ste2p, the α-factor receptor, was mis-localized from the plasma membrane to the vacuole in glo3Δ cells. Domain deletion mutants of Glo3p revealed that a GAP-independent function, as well as the GAP activity, of Glo3p is important for both α-factor binding and Ste2p localization at the cell surface. Additionally, we found that deletion of the GLO3 gene affects the size and number of Arf1p-residing Golgi compartments and causes a defect in transport from the TGN to the plasma membrane. Furthermore, we demonstrated that glo3Δ cells were defective in the late endosome-to-TGN transport pathway, but not in the early endosome-to-TGN transport pathway. These findings suggest novel roles for Arf-GAP Glo3p in endocytic recycling of cell surface proteins.
Keywords: Endocytosis; ARF-GAP; Small GTPase; Vesicular trafficking; Intracellular trafficking
Journal Title: Biochimica et Biophysica Acta - Molecular Cell Research
Volume: 1853
Issue 1
ISSN: 0167-4889
Publisher: Elsevier  
Date Published: 2015-01-01
Start Page: 144
End Page: 156
DOI: 10.1016/j.bbamcr.2014.10.009
Notes: This work was supported by the Japan Society for the Promotion of Science for J.Y.T., and the Novartis Foundation (Japan), the Mitsubishi Foundation, the Naito Foundation, the Mochida Memorial Foundation for Medical and Pharmaceutical Research, the Futaba Electronics Memorial Foundation, the Astellas Foundation for Research on Metabolic Disorders, the Hamaguchi Foundation for the Advancement of Biochemistry, the Takeda Science Foundation, and the Kurata Memorial Hitachi Science and Technology Foundation, to J.T.
Open access: yes (repository)