Effects of the terminal structural regions in isoform-specific sodium channel variations

Voltage-gated sodium channels are membrane proteins that initiate action potentials in neurons following membrane depolarization. Nav1.2 and Nav1.6 are two members of this family that differ in their electrophysiological properties and cellular distributions. The functional variations of these isoforms contribute to differences in the membrane excitability where the channels are expressed. Though both isoforms are found in the adult CNS neuron, they appear to have preferences for different subdomains depending on the myelination state of the axon. In this study, we generated chimeras between Nav1.2 and Nav1.6 to test specific structural regions for determinants that might contribute to the isoform-specific differences in channel activities and localization. We showed by voltage clamping in Xenopus oocytes that the N-terminal 202 amino acids of Nav1.2 and the corresponding N-region from Nav1.6 specified isoformic voltage-dependent activation. Exchanging the C-terminal 451 amino acids of Nav1.2 with the corresponding C-region from Nav1.6 altered the voltage-dependence and kinetics of inactivation, but the changes did not reflect a direct transfer of inactivation properties between the isoforms. In addition, the N-region and C-region of Nav1.6 demonstrated a cooperativity that might help to self-modulate the steady-state inactivation properties of the channel. We showed by expressing fluorescence protein-tagged chimeras in both neurons and heterologous MDCK cells that the N-terminal 202 amino acids of Nav1.2 mediated isoform-specific sorting to the apical membrane of the polarized MDCK cell. However, the sequence was necessary but not sufficient for directing isoform-specific axonal localization in unmyelinated neuron. In contrast, the C-terminal 451 amino acids of Nav1.2 affected axonal but not apical localization. Therefore, both terminal regions of Na v1.2 might be involved in targeting the isoform to axon.