Proteins of the regulators of G protein signaling (RGS) family bind to G subunits to downregulate their signaling in a variety of systems. G-interacting protein (GAIP) is a mammalian RGS protein that shows high affinity for the activated state of G_i-3, a protein known to regulate post-Golgi trafficking of secreted proteins in kidney epithelial cells. This study aimed to localize GAIP in epithelial cells and to investigate its potential role in the regulation of membrane trafficking. LLC-PK₁ cells were 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. Immunogold 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 membranes, GAIP was found to be concentrated in fractions of newly budded Golgi vesicles. Finally, the constitutive trafficking and secretion of sulfated proteoglycans was measured in cell lines overexpressing GAIP. We show evidence for GAIP regulation of secretory trafficking before the level of the trans-Golgi network but not in post-Golgi secretion. The location and functional effects of GAIP overlap only partially with those of G_i-3 and suggest multiple roles for GAIP in epithelial cells.

regulators of G protein signaling family of proteins; vesicle trafficking; heterotrimeric G protein regulation

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