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ArfGAP1 promotes COPI vesicle formation by facilitating coatomer polymerization.

ABSTRACT: The role of ArfGAP1 in COPI vesicle biogenesis has been controversial. In work using isolated Golgi membranes, ArfGAP1 was found to promote COPI vesicle formation. In contrast, in studies using large unilamellar vesicles (LUVs) as model membranes, ArfGAP1 functioned as an uncoating factor inhibiting COPI vesicle formation. We set out to discriminate between these models. First, we reexamined the effect of ArfGAP1 on LUVs. We found that ArfGAP1 increased the efficiency of coatomer-induced deformation of LUVs. Second, ArfGAP1 and peptides from cargo facilitated self-assembly of coatomer into spherical structures in the absence of membranes, reminiscent of clathrin self-assembly. Third, in vivo, ArfGAP1 overexpression induced the accumulation of vesicles and allowed normal trafficking of a COPI cargo. Taken together, these data support the model in which ArfGAP1 promotes COPI vesicle formation and membrane traffic and identify a function for ArfGAP1 in the assembly of coatomer into COPI.

SUBMITTER: Shiba Y

PROVIDER: S-EPMC3265926 | biostudies-literature |

REPOSITORIES: biostudies-literature

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A major transmembrane protein of Golgi-derived COPI-coated vesicles involved in coatomer binding.

Project description:Formation of non-clathrin-coated vesicles requires the recruitment of several cytosolic factors to the Golgi membrane. To identify membrane proteins involved in this budding process, a highly abundant type I transmembrane protein (p23) was isolated from mammalian Golgi-derived COPI-coated vesicles, and its cDNA was cloned and sequenced. It belongs to the p24 family of proteins involved in the budding of transport vesicles (Stamnes, M.A., M.W. Craighead, M.H. Hoe, N. Lampen, S. Geromanos, P. Tempst, and J.E. Rothman. 1995. Proc. Natl. Acad. Sci. USA. 92:8011-8015). p23 consists of a large NH2-terminal luminal domain and a short COOH-terminal cytoplasmic tail (-LRRFFKAKKLIE-CO2-) that shows similarity, but not identity, with the sequence motif-KKXX-CO2-, known as a signal for retrieval of escaped ER-resident membrane proteins (Jackson, M.R., T. Nilsson, and P.A. Peterson. 1990. EMBO (Eur. Mol. Biol. Organ.) J. 9:3153-3162; Nilsson, T., M. Jackson, and P.A. Peterson. 1989. Cell. 58:707-718). The cytoplasmic tail of p23 binds to coatomer with similar efficiency as known KKXX motifs. However, the p23 tail differs from the KKXX motif in having an additional motif needed for binding of coatomer. p23 is localized to Golgi cisternae and, during vesicle formation, it concentrates into COPI-coated buds and vesicles. Biochemical analysis revealed that p23 is enriched in vesicles by a factor of approximately 20, as compared with the donor Golgi fraction, and is present in amounts stoichiometric to the small GTP-binding protein ADP-ribosylation factor (ARF) and coatomer. From these data we conclude that p23 represents a Golgi-specific receptor for coatomer involved in the formation of COPI-coated vesicles.

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