Regulation of synaptic architecture and synaptic vesicle pools by Nervous wreck at Drosophila Type 1b glutamatergic synapses

Nervous wreck (Nwk), a protein that is present at Type 1 glutamatergic synapses that contains an SH3 domain and an FCH motif, is a Drosophila homolog of the human srGAP3/MEGAP protein, which is associated with mental retardation. Confocal microscopy revealed that circles in Nwk reticulum enclosed T-shaped active zones (T-AZs) and partially colocalized with synaptic vesicle (SV) markers and both exocytosis and endocytosis components. Results from an electron microscopic (EM) analysis showed that Nwk proteins localized at synaptic edges and in SV pools. Both the synaptic areas and the number of SVs in the readily releasable (RRPs) and reserve (RPs) SV pools in nwk(2) were significantly reduced. Synergistic, morphological phenotypes observed from eag(1);nwk(2) neuromuscular junctions suggested that Nwk may regulate synaptic plasticity differently from activity-dependent Hebbian plasticity. Although the synaptic areas in eag(1);nwk(2) boutons were not significantly different from those of nwk(2), the number of SVs in the RRPs was similar to those of Canton-S. In addition, three-dimensional, high-voltage EM tomographic analysis demonstrated that significantly fewer enlarged SVs were present in nwk(2) RRPs. Furthermore, Nwk formed protein complexes with Drosophila Synapsin and Synaptotagmin 1 (DSypt1). Taken together, these findings suggest that Nwk is able to maintain synaptic architecture and both SV size and distribution at T-AZs by interacting with Synapsin and DSypt1.