Anionic lipids regulate PLC

Phospholipase C-β (PLCβ) enzymes are essential effectors of G protein-coupled receptor signaling that hydrolyze phosphatidylinositol 4,5-bisphosphate (PIP2) at the plasma membrane to generate the second messengers inositol trisphosphate and diacylglycerol. PLCβs play essential roles in diverse physiological processes, including cardiac and neuronal function, macrophage activation, and the pathogenesis of diseases such as hypertrophic cardiomyopathy. PLCβ enzymes are unique in that they are aqueous-soluble and must partition onto the membrane surface to access their substrate, making membrane association a critical regulatory step. For example, we recently demonstrated that Gβγ activates PLCβ by membrane recruitment and orientation of the catalytic core on the membrane surface. Although PLCβ membrane recruitment is required for function, the molecular determinants governing this process remain incompletely understood. Using a quantitative membrane partitioning assay, we show that robust membrane association of PLCβ requires anionic phospholipids, whereas polar phospholipids cannot substitute. Membrane partitioning exhibits a steep dependence on anionic lipid abundance, which is mediated by electrostatic interactions between negatively charged lipids and basic residues within the distal C-terminal domain of PLCβ. We further demonstrate that anionic lipids cooperate with Gβγ to regulate PLCβ membrane recruitment, such that the magnitude of Gβγ-dependent recruitment is dictated by membrane anionic lipid content. These findings reconcile previous discrepancies regarding Gβγ-mediated PLCβ membrane recruitment and establish membrane electrostatics as a key regulatory input that integrates lipid composition with G protein signaling to regulate PLCβ activity.