GAIP, a G alpha(i-3)-binding protein, is associated with Golgi-derived vesicles and protein trafficking

Proteins of the regulators of G protein signaling (RGS) family bind to G al pha subunits to downregulate their signaling in a variety of systems. G alp ha-interacting protein (GAIP) is a mammalian RGS protein that shows high af finity for the activated state of G alpha(i-3), a protein known to regulate post-Golgi trafficking of secreted proteins in kidney epithelial cells. Th is study aimed to localize GAIP in epithelial cells and to investigate its potential role in the regulation of membrane trafficking. LLC-PK1 cells wer e stably transfected with a c-myc-tagged GAIP cDNA. in the transfected and untransfected cells, GAIP was found in the cytosol and on cell membranes. I mmunogold labeling showed that membrane-bound GAIP was localized on budding vesicles around Golgi stacks. When an in vitro assay was used to generate vesicles from isolated rat liver and Madin-Darby canine kidney cell Golgi m embranes, GAIP was found to be concentrated in fractions of newly budded Go lgi vesicles. Finally, the constitutive trafficking and secretion of sulfat ed proteoglycans was measured in cell Lines overexpressing GAIP. We show ev idence for GAIP regulation of secretory trafficking before the level of the trans-Golgi network, but not in post-Golgi secretion. The location and fun ctional effects of GAIP overlap only partially with those of G alpha(i-3) a nd suggest multiple roles for GAIP in epithelial cells.