Identification of synaptic DGKq interactors that stimulate DGKq activity

Lipids and their metabolic enzymes are a critical point of regulation for the membrane curvature required to induce membrane fusion during synaptic vesicle recycling. One such enzyme is diacylglycerol kinase theta (DGKq), which produces phosphatidic acid (PtdOH) that generates negative membrane curvature. Synapses lacking DGKq have significantly slower rates of endocytosis, implicating DGKq as an endocytic regulator. Importantly, DGKq kinase activity is required for this function. However, protein regulators of DGKq’s kinase activity in neurons have never been identified. In this study, we employed APEX2 proximity labeling and mass spectrometry to identify endogenous interactors of DGKq in neurons and assayed their ability to modulate its kinase activity. Seven endogenous DGKq interactors were identified and notably, synaptotagmin-1 (Syt1) increased DGKq kinase activity 10-fold. This study is the first to validate endogenous DGKq interactors at the mammalian synapse and suggests a coordinated role between DGKq-produced PtdOH and Syt1 in synaptic vesicle recycling.