Pairwise biosynthesis of ion channels stabilizes excitability and mitigates arrhythmias.

Abstract

Coordinated expression of ion channels is crucial for cardiac rhythms, neural signaling, and cell cycle progression. Perturbation of this balance results in many disorders including cardiac arrhythmias. Prior work revealed association of mRNAs encoding cardiac Na_V1.5 (<i>SCN5A</i>) and hERG1 (<i>KCNH2</i>), but the functional significance of this association was not established. Here, we provide a more comprehensive picture of <i>KCNH2</i>, <i>SCN5A</i>, <i>CACNA1C</i>, and <i>KCNQ1</i> transcripts collectively copurifying with nascent hERG1, Na_V1.5, Ca_V1.2, or KCNQ1 channel proteins. Single-molecule fluorescence in situ hybridization (smFISH) combined with immunofluorescence reveals that the channel proteins are synthesized predominantly as heterotypic pairs from discrete molecules of mRNA, not as larger cotranslational complexes. Puromycin disrupted colocalization of mRNA with its encoded protein, as expected, but remarkably also pairwise mRNA association, suggesting that transcript association relies on intact translational machinery or the presence of the nascent protein. Targeted depletion of <i>KCHN2</i> by specific shRNA resulted in concomitant reduction of all associated mRNAs, with a corresponding reduction in the encoded channel currents. This co-knockdown effect, originally described for <i>KCNH2</i> and <i>SCN5A</i>, thus appears to be a general phenomenon among transcripts encoding functionally related proteins. In multielectrode array recordings, proarrhythmic behavior arose when I_Kr was reduced by the selective blocker dofetilide at IC₅₀ concentrations, but not when equivalent reductions were mediated by shRNA, suggesting that co-knockdown mitigates proarrhythmic behavior expected from the selective reduction of a single channel species. We propose that coordinated, cotranslational association of functionally related ion channel mRNAs confers electrical stability by co-regulating complementary ion channels in macromolecular complexes.