RGS4 and RGS2 bind coatomer and inhibit COPI association with Golgi membranes and intracellular transport

COPI, a protein complex consisting of coatomer and the small GTPase ARF1, i s an integral component of some intracellular transport carriers. The assoc iation of COPI with secretory membranes has been implicated in the maintena nce of Golgi integrity and the normal functioning of intracellular transpor t in eukaryotes. The regulator of G protein signaling, RGS4, interacted wit h the COPI subunit beta'-COP in a yeast two-hybrid screen. Both recombinant RGS4 and RGS2 bound purified recombinant beta'-COP in vitro. Endogenous cy tosolic RGS4 from NG108 cells and RGS2 from HEK293T cells cofractionated wi th the COPI complex by gel filtration. Binding of beta'-COP to RGS4 occurre d through two dilysine motifs in RGS4, similar to those contained in some a minoglycoside antibiotics that are known to bind coatomer. RGS4 inhibited C OPI binding to Golgi membranes independently of its GTPase-accelerating act ivity on G(i alpha). In RCS4-transfected LLC-PK1 cells, the amount of COPI in the Golgi region was considerably reduced compared with that in wild-typ e cells, but there was no detectable difference in the amount of either Gol gi-associated ARF1 or the integral Golgi membrane protein giantin, indicati ng that Golgi integrity was preserved. In addition, RGS4 expression inhibit ed trafficking of aquaporin 1 to the plasma membrane in LLC-PK1 cells and i mpaired secretion of placental alkaline phosphatase from HEK293T cells. The inhibitory effect of RGS4 in these assays was independent of GTPase-accele rating activity but correlated with its ability to bind COPI. Thus, these d ata support the hypothesis that these RGS proteins sequester coatomer in th e cytoplasm and inhibit its recruitment onto Golgi membranes, which may in turn modulate Golgi-plasma membrane or intra-Golgi transport.