Activation in voltage-gated Na channels modified by the tarantula toxin ProTxll

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DESCRIPTION (provided by applicant): Neuronal voltage-gated ion channels allow the nervous system to precisely modulate its excitability according to recent events and changes in the local environment. Individual channels open and close in response to fluctuations in membrane potential. Activation, the process by which channels transition from a closed, non-conducting state to an open conducting state, is mediated by conformational changes in the four voltage sensors of these channels. Previous studies have shown that differences in activation gating properties can be traced to differences in the timing and coordination of voltage sensor movements. Therefore, understanding how the voltage sensors move and coordinate with one another during activation in different types of channels is crucial in designing diagnostic, prophylactic, and therapeutic tools that target specific channels. In voltage-gated sodium (Na) channels voltage sensor movements occur with different timecourses and likely exhibit differences in coupling. ProTxIl, a recently discovered tarantula peptide toxin, modifies activation in voltage-gated sodium (Na) channels. Here, I propose to use this toxin to probe voltage sensor movements within individual domains of the channel during activation gating. Whole cell, single channel, and gating current analyses will first be used to determine the mechanistic basis of the multiple effects of ProTxIl. Fluorescent labeling techniques combined with targeted mutagenesis will then be used to systematically explore the site(s) of toxin interaction with each of the channel's voltage sensor regions. Disruptions in the effect of ProTxIl on gating in these mutants will reveal the domain-specific role of toxin interactions and voltage sensor movements in channel activation. Disruptions in Na channel gating are the cause of many severe neuropathies including epilepsy, sleep disorders, and neurogenic pain. Understanding the way these channels work is the necessary first step towards developing prophylactic and therapeutic tools for the prevention and treatment of these conditions.