Identification of novel interaction sites that determine specificity between fibroblast growth factor homologous factors and voltage-gated sodium channels.
Fibroblast growth factor homologous factors (FHFs, FGF11-14) bind to the C termini (CTs) of specific voltage-gated sodium channels (VGSC) and thereby regulate their function. The effect of an individual FHF on a specific VGSC varies greatly depending upon the individual FHF isoform. How individual FHFs impart distinctive effects on specific VGSCs is not known and the specificity of these pairwise interactions is not understood. Using several biochemical approaches combined with functional analysis, we mapped the interaction site for FGF12B on the Na(V)1.5 C terminus and discovered previously unknown determinants necessary for FGF12 interaction. Also, we demonstrated that FGF12B binds to some, but not all Na(V)1 CTs, suggesting specificity of interaction. Exploiting a human single nucleotide polymorphism in the core domain of FGF12 (P149Q), we identified a surface proline that contributes a part of this pairwise specificity. This proline is conserved among all FHFs, and mutation of the homologous residue in FGF13 also leads to loss of interaction with a specific VGSC CT (Na(V)1.1) and loss of modulation of the resultant Na(+) channel function. We hypothesized that some of the specificity mediated by this proline may result from differences in the affinity of the binding partners. Consistent with this hypothesis, surface plasmon resonance data showed that the P149Q mutation decreased the binding affinity between FHFs and VGSC CTs. Moreover, immunocytochemistry revealed that the mutation prevented proper subcellular targeting of FGF12 to the axon initial segment in neurons. Together, these results give new insights into details of the interactions between FHFs and Na(V)1.x CTs, and the consequent regulation of Na(+) channels.
Wang, C; Wang, C; Hoch, EG; Pitt, GS
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