INCEPT —Interdisciplinary Neuroscience Colloquia of Extramural Postdoctoral Talks, is a new talk series organized by the INCEPT committee of Neurobio Postdoc Club to provide exceptional senior postdocs with more opportunities to receive feedback and practice their prospective job talks.

Speaker: Dr. DelSignore
Topic: Organization and regulation of synaptic endocytic machinery

The synaptic endocytic machinery regulates the traffic of cargoes important for synaptic development, activity, and plasticity, and defects in this machinery are associated with a variety of neurological diseases. Unlike non-neuronal cells, in which endocytic machinery assembles transiently at discrete 50-100nm sites of endocytosis, the synaptic endocytic machinery assembles persistently at membranes in a poorly understood micron scale domain called the ‘Periactive Zone (PAZ)’, within which active endocytic events occur sparsely in space and time. This suggests that there are neuron-specific mechanisms to broadly organize and then locally activate this machinery. Problem: How does micron-scale PAZ machinery direct nano-scale endocytic events?

We previously found that the PAZ protein Nervous Wreck couples membrane remodeling to force-generating actin assembly, but is locked in an autoinhibited state even after it associates with membranes. I hypothesized that local relief of autoregulation could promote discrete membrane remodeling events. Using in vitro assays, I found that SH3-mediated interactions among Nervous Wreck, the actin assembly factor WASp, and the scaffold Dap160/Intersectin cooperate to fully relieve autoinhibition of membrane binding and actin assembly. To test how this mechanism regulates synaptic endocytosis, I developed novel methods at the Drosophila neuromuscular junction to quantify PAZ activation by quantitative live imaging of actin assembly, and PAZ organization by super-resolution microscopy. I found that loss of Nervous Wreck, Dap160, or their interaction resulted in short-lived, likely unproductive actin assembly, and compromised synaptic endocytosis. In currently ongoing work, I am further investigating the mechanisms that organize PAZ micron-scale architecture. I find that the same SH3 interactions that promote PAZ activation are also required for its micron-scale architecture at synapses, and drive phase separation in vitro, suggesting a biochemical mechanism for micron-scale assembly. This raises the exciting question of how phase separation and autoregulation might collaboratively organize PAZ architecture and function.
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