View Program - 63rd Annual Drosophila Research Conference

Smooth ER (endoplasmic reticulum) forms a continuous network inside of each motorneuron, dominating the intracellular landscape and extending through each axon from root to tip. Smooth ER acts as a source and sink of Ca⁺² for organelles and tethers itself to other organelles, all lending to the hypothesis that it could be dynamically communicating and trafficking intracellular information, like a “neuron within a neuron”. Recent findings suggest that VAPB (vesicle-associated membrane protein [VAMP]-associated protein B) regulates smooth ER morphology and function in cultured cells. A highly conserved portion of VAPB can also be cleaved to act as an extracellular ligand. Intriguingly, this domain can harbor a point mutation (P58S) causative of amyotrophic lateral sclerosis (ALS). The complete molecular mechanisms linking VAPB to ER and motorneuron development and degeneration remain elusive. We will investigate these interactions using a simple Drosophila Crispr-mediated knockout model that allows us to knock back in both GFP-tagged rescue and P58S VAPB constructs. Excitingly, the VAPB knockout I engineered seems viable. In conjunction with new adult neuromuscular junction (NMJ) dissection techniques, this uniquely poises us to track synaptic development and VAPB localization at the NMJ in vivo from larvae to adult. We will observe gross ER morphology and function by crossing these mutants with existing lines that mark ER, calcium signaling, and autophagosomal trafficking. Flight behavior assays will measure motorneuron activity. We hypothesize, due to prior in vitro studies, that VAPB knockout and VAPB P58S mutant Drosophila will exhibit deficiencies in VAPB localization and ER morphology and function at the NMJ. We further hypothesize that this will correlate with deficits in motorneuron development, and ultimately degeneration and altered flight behaviors.

Investigating the role of VAPB in axonal ER and motorneuron development and degeneration