RESEARCH ARTICLE Identifying a Kinase Network Regulating FGF14:Nav1.6 Complex Assembly Using Split-

Kinases play fundamental roles in the brain. Through complex signaling pathways, kinases regulate the strength of protein:protein interactions (PPI) influencing cell cycle, signal trans-duction, and electrical activity of neurons. Changes induced by kinases on neuronal excit-ability, synaptic plasticity and brain connectivity are linked to complex brain disorders, but the molecular mechanisms underlying these cellular events remain for the most part elu-sive. To further our understanding of brain disease, new methods for rapidly surveying ki-nase pathways in the cellular context are needed. The bioluminescence-based luciferase complementation assay (LCA) is a powerful, versatile toolkit for the exploration of PPI. LCA relies on the complementation of two firefly luciferase protein fragments that are functionally reconstituted into the full luciferase enzyme by two interacting binding partners. Here, we applied LCA in live cells to assay 12 kinase pathways as regulators of the PPI complex formed by the voltage-gated sodium channel, Nav1.6, a transmembrane ion channel that elicits the action potential in neurons and mediates synaptic transmission, and its multiva-lent accessory protein, the fibroblast growth factor 14 (FGF14). Through extensive dose-de