ARNO3, A SEC7-DOMAIN GUANINE-NUCLEOTIDE EXCHANGE FACTOR FOR ADP-RIBOSYLATION FACTOR 1, IS INVOLVED IN THE CONTROL OF GOLGI STRUCTURE AND FUNCTION

Budding of transport vesicles in the Golgi apparatus requires the recr uitment of coat proteins and is regulated by ADP ribosylation factor ( ARF) 1. ARF1 activation is promoted by guanine nucleotide exchange fac tors (GEFs), which catalyze the transition to GTP-bound ARF1. We recen tly have identified a human protein, ARNO (ARF nucleotide-binding-site opener), as an ARF1-GEF that shares a conserved domain with the yeast Sec7 protein, We now describe a human Sec7 domain-containing GEF refe rred to as ARNO3. ARNO and ARNO3, as well as a third GEF called cytohe sin-1, form a family of highly related proteins with identical structu ral organization that consists of a central Sec7 domain and a carboxy- terminal pleckstrin homology domain. We show that all three proteins a ct as ARF1 GEF in vitro, whereas they have no effect on ARF6, an ARF p rotein implicated in the early endocytic pathway. Substrate specificit y of ARNO-like GEFs for ARF1 depends solely on the Sec7 domain, Overex pression of ARNO3 in mammalian cells results in (i) fragmentation of t he Golgi apparatus, (ii) redistribution of Golgi resident proteins as well as the coat component beta-COP, and (iii) inhibition of SEAP tran sport (secreted form of alkaline phosphatase). In contrast, the distri bution of endocytic markers is not affected. This study indicates that Sec7 domain-containing GEFs control intracellular membrane compartmen t structure and function through the regulation of specific ARF protei ns in mammalian cells.