Rac Regulates Its Effector Phospholipase Cγ2 through Interaction with a Split Pleckstrin Homology Domain

Several isoforms of phospholipase C (PLC) are regulated through interactions with Ras superfamily GTPases, including Rac proteins. Interestingly, of two closely related PLC gamma isoforms, only PLC gamma(2) has previously been shown to be activated by Rac. Here, we explore the molecular basis of this interaction as well as the structural properties of PLC gamma(2) required for activation. Based on reconstitution experiments with isolated PLC gamma variants and Rac2, we show that an unusual pleckstrin homology (PH) domain, designated as the split PH domain (spPH), is both necessary and sufficient to effect activation of PLC gamma(2) by Rac2. We also demonstrate that Rac2 directly binds to PLC gamma(2) as well as to the isolated spPH of this isoform. Furthermore, through the use of NMR spectroscopy and mutational analysis, we determine the structure of spPH, define the structural features of spPH required for Rac interaction, and identify critical amino acid residues at the interaction interface. We further discuss parallels and differences between PLC gamma(1) and PLC gamma(2) and the implications of our findings for their respective signaling roles.