Fig 2: Model for DGK?’s interaction with Syt1. Syt1 is a transmembrane protein on synaptic vesicles and the presynaptic membrane. Perhaps to aid in the membrane deformation required in the fusion and fission of vesicles, Syt1 binds to DGK? and stimulates its kinase activity, producing local hotspots of PtdOH that generate negative membrane curvature. AZ = active zone, EZ = endocytic zone. Figure made with BioRender.com.

Fig 3: Validation of candidate DGK?-interacting proteins. (A) Streptavidin beads were used to isolate biotinylated proteins from DGK?-GFP-APEX2 labeled cortical neuron lysates (DIV21), and biotinylation was confirmed biochemically for dynamin-1, synaptogyrin-1, Munc18-1, clathrin heavy chain 1, Syt1, CaMKIIa, Hsc70, and syntaxin-7. (B) DGK? overexpressed in HEK cells is immunoprecipitated with a myc antibody. (C) DGK? interacts with Hsc70, CaMKIIa, Syt1, synaptogyrin-1, dynamin-1, syntaxin-7, and Munc18-1 when both are overexpressed in HEK cells. Hsc70 interacts with DGK? at its endogenous expression level. (D) Clathrin heavy chain 1 does not interact with DGK? when overexpressed in HEK cells. All blots are representative of at least three separate experiments.