Synapse Formation

To identify neuronal pathways that control synapse formation and synaptic growth, we have conducted forward genetic screens for temperature-sensitive mutants that disrupt the process. Several Ca²⁺-dependent presynaptic proteins have been implicated in synaptic growth, but the source of Ca²⁺ and its regulation are unknown.  In addition to retrograde signaling, we have found that presynaptic Ca²⁺ influx through N-type channels participates in synaptic growth via signaling pathways that are distinct from those that mediate neurotransmitter release.  The opening of presynaptic N-type channels during robust synaptic activity allows presynaptic Ca²⁺ influx to modulate sprouting mechanisms that locally control synaptic remodeling. Linking presynaptic voltage-gated Ca²⁺ entry to downstream Ca²⁺-sensitive synaptic growth regulators provides an efficient activity-dependent mechanism for modifying synaptic strength.

Several cell adhesion proteins and synaptic growth regulators reside adjacent to the active zone where presynaptic Ca²⁺ channels localize, in domains termed peri-active zones. In collaboration with Barry Ganetzky’s lab, we have recently identified one such peri-active zone protein, termed nervous wreck (nwk), in screens for temperature-sensitive paralytic mutants that causes excessive growth of larval neuromuscular junctions.  Mutations in nwk result in increased synaptic bouton number and branch formation.  NWK encodes an FCH and SH3 domain-containing adaptor protein that localizes to the peri-active zone of presynaptic terminals and binds to the Drosophila ortholog of Wasp (Wsp), a key regulator of actin polymerization. wsp null mutants display synaptic overgrowth similar to nwk and enhance the nwk morphological phenotype in a dose-dependent manner. Evolutionarily, NWK belongs to a previously undescribed family of adaptor proteins that includes the human srGAPs, which regulate Rho activity downstream of Robo receptors. We propose that NWK controls synapse morphology by regulating actin dynamics downstream of growth signals in presynaptic terminals. Further genetic analysis of synaptic growth mutants is underway to identify additional molecular pathways that modulate synaptic connectivity and alter behavioral output.