Cell Isolation of Transgenic Worms

Day 1 adult neuronally GFP-labeled worms (Punc119::GFP or Pmec-4::GFP) were prepared for cell isolation as previously described^{15 (link)} with modifications (Extended Data Fig. 2). Synchronized adult worms were washed with M9 buffer to remove excess bacteria. The pellet (~250 µl) was washed with 500 µl lysis buffer (200 mM DTT, 0.25% SDS, 20 mM Hepes pH 8.0, 3% sucrose) and resuspended in 1000 µl lysis buffer. Worms were incubated in lysis buffer with gentle rocking for 6.5 minutes at room temperature. The pellet was washed 6× with M9 and resuspended in 20 mg/ml pronase from Streptomyces griseus (Sigma-Aldrich). Worms were incubated at room temperature (<20 minutes) with periodic mechanical disruption by pipetting every 2 min. When most worm bodies were dissociated, leaving only small debris and eggs, ice-cold PBS buffer containing 2% fetal bovine serum (Gibco) was added. RNA from FACS-sorted neurons was prepared for RNA-seq and subsequent analysis (see Extended Data for details).