Specificity and structural requirements of phospholipase C-β stimulation by Rho GTPases versus G protein βγ dimers

Phospholipase C-beta(2) (PLCbeta(2)) is activated both by heterotrimeric G protein alpha- and betagamma- subunits and by Rho GTPases. In this study, activated Rho GTPases are shown to stimulate PLCbeta isozymes with the rank order of PLCbeta(2) > PLCbeta(3) greater than or equal to PLCbeta(1). The sensitivity of PLCbeta isozymes to Rho GTPases was clearly different from that observed for G protein betagamma dimers, which decreased in the following order: PLCbeta(3) > PLCbeta(2) > PLCbeta(1), for beta(1)gamma(1/2) and PLCbeta(2) > PLCbeta(1) >>> PLCbeta(3) for beta(5)gamma(2). Rac1 and Rac2 were found to be more potent and efficacious activators of PLCbeta(2) than was Cdc42Hs. The stimulation of PLCbeta(2) by Rho GTPases and G protein betagamma dimers was additive, suggesting that PLCbeta(2) activation can be augmented by independent regulation of the enzyme by the two stimuli. Using chimeric PLCbeta(1)-PLCbeta(2) enzymes, betagamma dimers, and Rho GTPases are shown to require different regions of PLCbeta(2) to mediate efficient stimulation of the enzyme. Although the catalytic subdomains X and Y of PLCbeta(2) were sufficient for efficient stimulation by betagamma, the presence of the putative pleckstrin homology domain of PLCbeta(2) was absolutely required for the stimulation of the enzyme by Rho GTPases. Taken together, these results identify Rho GTPases as novel PLCbeta regulators, which mediate PLCbeta isozyme-specific stimulation and are potentially involved in coordinating the activation of PLCbeta(2) by extracellular mediators in intact cells.