The X-linked retinitis pigmentosa protein RP2 facilitates G protein traffic

The X-linked retinitis pigmentosa protein RP2 is a GTPase activating protein (GAP) for the small GTPase Arl3 and both proteins are implicated in the traffic of proteins to the primary cilia. Here, we show that RP2 can facilitate the traffic of the G beta subunit of transducin (G beta 1). Glutathione S-transferase (GST)-RP2 pulled down G beta from retinal lysates and the interaction was specific to G beta 1, as G beta 3 or G beta 5L did not bind RP2. RP2 did not appear to interact with the G beta:G gamma heterodimer, in contrast G gamma 1 competed with RP2 for G beta binding. Overexpression of G beta 1 in SK-N-SH cells led to a cytoplasmic accumulation of G beta 1, while co-expression of RP2 or G gamma 1 with G beta 1 restored membrane association of G beta 1. Furthermore, RP2 small interfering RNA in ARPE19 cells resulted in a reduction in G beta 1 membrane association that was rescued by G gamma 1 overexpression. The interaction of RP2 with G beta 1 required RP2 N-terminal myristolyation and the co-factor C (TBCC) homology domain. The interaction was also disrupted by the pathogenic mutation R118H, which blocks Arl3 GAP activity. Interestingly, Arl3-Q71L competed with G beta 1 for RP2 binding, suggesting that Arl3-GTP binding by RP2 would release G beta 1. RP2 also stimulated the association of G beta 1 with Rab11 vesicles. Collectively, the data support a role for RP2 in facilitating the membrane association and traffic of G beta 1, potentially prior to the formation of the obligate G beta:G gamma heterodimer. Combined with other recent evidence, this suggests that RP2 may co-operate with Arl3 and its effectors in the cilia-associated traffic of G proteins.