Phosphorylation regulates SYD-2/Liprin-α’s phase separation and active zone formation.

(A, B) Immunoprecipitation of endogenous GFP-SYD-2 (A) and endogenous mScarlet-SYD-2-λpptase (B). Western blot for GFP or mScarlet and phosphoserine/phosphothreonine. The indicated immunoprecipitation samples were treated with phosphatase before running on the gel (pptase +). (C) FRAP of endogenous mScarlet-SYD-2 at embryonic nerve ring synapses to measure dynamics. Wild-type SYD-2 is present in dynamic phase-separated condensates, while removal of phase separation in SYD-2(IDRΔ) or removal of phosphorylation in SYD-2-λpptase inhibits condensate formation. Scale bars, 5 μm. (D) Quantification of FRAP in (C). (E) Schematic of the C. elegans HSN that makes stereotyped synapses to vulval muscles to control egg laying. (F) HSN synapse formation phenotypes visualized with Airyscan superresolution imaging of endogenous fluorescent tags in the indicated mutants. Scale bar, 5 μm. (G) Quantification of HSN intensity in (F). ****, p S1 Data. FRAP, fluorescence recovery after photobleaching; HSN, hermaphrodite-specific neuron.