LAUSR

Mechanisms of neuromodulatory transmission in the brain remain ambiguous. Dopamine is a prototypical neuromodulator, and it was recently found that its secretion relies on active zone-like release site assemblies. Here, we use in vivo biotin-identification (iBioID) proximity proteomics in mouse striatum to isolate dopamine release site proteins enriched over the general dopamine axonal protein content. Using three bait proteins, we identified 527 proteins that fall into several synaptic protein classes, including active zone, Ca2+ regulatory and synaptic vesicle proteins. We also detected many proteins not previously associated with synaptic exocytosis. Knockout of the presynaptic organizer protein RIM profoundly disrupted dopamine release site composition assessed by iBioID, while Synaptotagmin-1 knockout did not. α-Synuclein, a protein linked to Parkinson’s disease, was enriched at release sites, and this enrichment was lost in both tested mutants. We conclude that RIM organizes scaffolded dopamine release sites and we define the protein composition of these sites.