Biophysical Society Conference | Estes Park 2023

Membrane Budding and Fusion

Poster Abstracts

27-POS Board 9 ELUCIDATING THE COMPLEXIN C-TERMINAL DOMAIN MECHANISM IN SYNAPTIC REGULATION

Justine A Lottermoser 1 ; Jeremy S Dittman 1 ; 1 Weill Cornell Medicine, Biochemistry, New York, NY, USA

The molecular mechanisms of synaptic transmission as well as its plasticity and pathophysiology remain poorly understood. Complexin (Cpx) is a small cytosolic presynaptic protein that acts on neuronal SNARE proteins to control synaptic vesicle (SV) exocytosis. The Cpx C-terminal domain (CTD) interacts with membranes and directs Cpx to SVs. In absence of the CTD, Cpx is mislocalized and fails to inhibit spontaneous SV fusion, severely disrupting nervous system function. Using C. elegans as a model nervous system, we replaced the Cpx CTD with various SV proteins and screened for functional rescue relative to synaptic abundance to address the following questions: Does the CTD have any function besides localization? Is there an essential feature in the CTD required for Cpx inhibitory function? C. elegans transgenes included mGFP to monitor steady-state synaptic Cpx fusion protein abundance in intact living animals using confocal imaging. Functional rescue was assessed through a quantitative behavioral assay that is highly sensitive to Cpx inhibitory function at the synapse. We found that the SV-resident GTPase RAB-3 can functionally replace the CTD when the fusion protein is expressed in the absence of endogenous Cpx. We conclude the CTD has no essential features beyond sequestering Cpx onto vesicles. The ability of RAB-3 to restore truncated Cpx function was further investigated by perturbing RAB-3 GTPase activity. Preliminary data shows that stabilizing the GTP-bound state decreased Cpx function hinting at the possibility that the GDP bound state better positions Cpx for its inhibitory control of synaptic transmission. Future efforts will focus on the GDP-bound state of the RAB-3 anchor as well as potential interactions between RAB-3 and Cpx.

102