Structural basis for a Munc13-1 homodimer to Munc13-1/RIM heterodimer switch

C₂ domains are well characterized as Ca²⁺/phospholipid-binding modules, but little is known about how they mediate protein–protein interactions. In neurons, a Munc13–1 C₂A-domain/RIM zinc-finger domain (ZF) heterodimer couples synaptic vesicle priming to presynaptic plasticity. We now show that the Munc13–1 C₂A domain homodimerizes, and that homodimerization competes with Munc13–1/RIM heterodimerization. X-ray diffraction studies guided by nuclear magnetic resonance (NMR) experiments reveal the crystal structures of the Munc13–1 C₂A-domain homodimer and the Munc13–1 C₂A-domain/RIM ZF heterodimer at 1.44 Å and 1.78 Å resolution, respectively. The C₂A domain adopts a β-sandwich structure with a four-stranded concave side that mediates homodimerization, leading to the formation of an eight-stranded β-barrel. In contrast, heterodimerization involves the bottom tip of the C₂A-domain β-sandwich and a C-terminal α-helical extension, which wrap around the RIM ZF domain. Our results describe the structural basis for a Munc13–1 homodimer–Munc13–1/RIM heterodimer switch that may be crucial for vesicle priming and presynaptic plasticity, uncovering at the same time an unexpected versatility of C₂ domains as protein–protein interaction modules, and illustrating the power of combining NMR spectroscopy and X-ray crystallography to study protein complexes.