Regulator of G protein signaling domain-containing Rho guanine-nucleotide exchange factors (RGS-RhoGEFs) directly links activated forms of the G12 family of heterotrimeric G protein alpha subunits to the small GTPase Rho. Stimulation of G(12/13)-coupled GPCRs or expression of constitutively activated forms of alpha(12) and alpha(13) has been shown to induce the translocation of the RGS-RhoGEF, p115-RhoGEF, from the cytoplasm to the plasma membrane (PM). However, little is known regarding the functional importance and mechanisms of this regulated PM recruitment, and thus PM recruitment of p115-RhoGEF is the focus of this report. A constitutively PM-localized mutant of p115-RhoGEF shows a much greater activity compared to wild type p115-RhoGEF in promoting Rho-dependent neurite retraction of NGF-differentiated PC12 cells, providing the first evidence that PM localization can activate p115-RhoGEF signaling. Next, we uncovered the unexpected finding that Rho is required for alpha(13)-induced PM translocation of p115-RhoGEF. However, inhibition of Rho did not prevent alpha(12)-induced PM translocation of p115-RhoGEF. Additional differences between alpha(13) and alpha(12) in promoting PM recruitment of p115-RhoGEF were revealed by analyzing RGS domain mutants of p115-RhoGEF. Activated alpha(12) effectively recruits the isolated RGS domain of p115-RhoGEF to the PM, whereas alpha(13) only weakly does. On the other hand, alpha(13) strongly recruits to the PM a p115-RhoGEF mutant containing amino acid substitutions in an acidic region at the N-terminus of the RGS domain; however, alpha(12) is unable to recruit this p115-RhoGEF mutant to the PM. These studies provide new insight into the function and mechanisms of alpha(12/13)-mediated PM recruitment of p115-RhoGEF.