Cryo-EM Structure of Phospholipase Cϵ Defines N-terminal Domains and their Roles in Activity

Phospholipase C{varepsilon} (PLC{varepsilon}) increases intracellular Ca2+ and protein kinase C (PKC) activity in the cardiovascular system in response to stimulation of G protein coupled receptors (GPCRs) and receptor tyrosine kinases (RTKs). The ability of PLC{varepsilon} to respond to these diverse inputs is due, in part, to multiple, conformationally dynamic regulatory domains. However, this heterogeneity has also limited structural studies of the lipase to either individual domains or its catalytic core. Here, we report the 3.9 [A] reconstruction of the largest fragment of PLC{varepsilon} to date in complex with an antigen binding fragment (Fab). The structure reveals that PLC{varepsilon} contains a pleckstrin homology (PH) domain and four tandem EF hands, including subfamily-specific insertions and intramolecular interactions with the catalytic core. The structure, together with a model of the holoenzyme, suggest that part of the N-terminus and PH domain form a continuous surface that could engage cytoplasmic leaflets of the plasma and perinuclear membranes, contributing to activity. Functional characterization of this surface confirm it is critical for maximum basal and G protein-stimulated activities. This study provides new insights into the autoinhibited, basal conformation of PLC{varepsilon} and the first mechanistic insights into how it engages cellular membranes for activity.