Budding from Golgi membranes requires the coatomer complex of non-clathrin coat proteins

Abstract

Do the coats on vesicles budded from the Golgi apparatus actually cause the budding, or do they simply coat buds (Fig. 1)? One view (the membrane-mediated budding hypothesis) is that budding is an intrinsic property of Golgi membranes not requiring extrinsic coat proteins. Assembly of coats from dispersed subunits is super-imposed upon the intrinsic budding process and is proposed to convert the tips of tubules into vesicles. The alternative view (the coat-mediated budding hypothesis) is that coat formation provides the essential driving force for budding. The membrane-mediated budding hypothesis was inspired by the microtubule-dependent extension of apparently uncoated, 90-nm-diameter membrane tubules from the Golgi apparatus and other organelles in vivo after treatment with brefeldin A, a drug that inhibits the assembly of coat proteins onto Golgi membranes. This hypothesis predicts that tubules will be extended when coat proteins are unavailable to convert tubule-derived membrane into vesicles. Here we use a cell-free system in which coated vesicles are formed from Golgi cisternae to show that, on the contrary, when budding diminishes as a result of immunodepletion of coat protein pools, tubules are not formed at the expense of vesicles. We conclude that coat proteins are required for budding from Golgi membranes.

Cloning and functional characterization of mammalian homologues of the COPII component Sec23.

“…COPI was identified by the isolation of proteins that mediate vesicle budding within the mammalian Golgi apparatus (Balch et al, 1984;Orci et al, 1986Orci et al, , 1989Orci et al, , 1993b. Upon the binding of ARF-GTP to a putative receptor present on the Golgi membrane coatomer, a 700-kDa protein complex is recruited to form a coated t Corresponding author. bud that separates from the donor membrane in a process requiring acyl-CoA (Orci et al, 1993a;Ostermann et al, 1993).…”

Biochemical Heterogeneity and Phosphorylation of Coatomer Subunits

“…The role of COPI in transport between the ER and the Golgi complex, as well as in endosome function, has been established by inhibition of transport by microinjected anti-␤-COP antibody (10) as well as by the isolation of COPI mutants in yeast and Chinese hamster ovary cells (8,11). There is also considerable evidence from in vitro reconstitution experiments that at least COPI coats are involved in transport through the Golgi stack (12).…”

“…It is becoming increasingly apparent that one or more COPI components bind to and/or function at multiple intracellular sites in both the secretory and endocytic pathways (4,11,12,20). Recruitment of COPI subunits to these various sites of action may, at least in part, be controlled by the phosphorylation state of ␤-COP or ␦-COP.…”

COPI Budding within the Golgi Stack

“…The two cytosolic components essential for COPI vesicle formation in vitro are Arf1 and the heptameric coat complex coatomer (Serafini et al 1991a;Orci et al 1993).…”