Gβγ engages PLCβ3 at multiple sites to reorient and facilitate its activation

Phospholipase C {beta} (PLC{beta}) enzymes are activated by heterotrimeric G protein subunits, increasing hydrolysis of phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) at the plasma membrane. All four human PLC{beta} isoforms (PLC{beta}1-4) are activated by Gq, while PLC{beta}1-3 are activated to varying extents by G{beta}{gamma}. The binding sites for Gq on PLC{beta} are well-established and much has been learned about its mechanism of activation, but comparatively little is known about G{beta}{gamma}-dependent activation. In this work, we used cryo-electron microscopy (cryo-EM) single particle analysis (SPA), functional assays, and bioluminescence resonance energy transfer (BRET) to investigate how G{beta}{gamma} interacts with PLC{beta}3 in concert with activated Gq to regulate phospholipase activity. G{beta}{gamma} heterodimers bind multiple surfaces of PLC{beta}3 to promote activation but alone do not recruit the enzyme to the plasma membrane. Instead, G{beta}{gamma} facilitates activation by Gq, most likely by reorienting the phospholipase catalytic site at the membrane to maximize PI(4,5)P2 hydrolysis and downstream Ca2+ release. Cell-based functional assays demonstrate that G{beta}{gamma} is required for maximal PLC{beta}3 activation even when Gq heterotrimers are the sole source of G{beta}{gamma}. Together, these findings demonstrate that G{beta}{gamma} acts as a critical positive allosteric modulator that regularly acts in concert with Gq to activate PLC{beta}3 at the plasma membrane.