Pairwise biosynthesis of ion channels stabilizes excitability and mitigates arrhythmias.

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 NaV1.5 (SCN5A) and hERG1 (KCNH2), but the functional significance of this association was not established. Here, we provide a more comprehensive picture of KCNH2, SCN5A, CACNA1C, and KCNQ1 transcripts collectively copurifying with nascent hERG1, NaV1.5, CaV1.2, 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 KCHN2 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 KCNH2 and SCN5A, thus appears to be a general phenomenon among transcripts encoding functionally related proteins. In multielectrode array recordings, proarrhythmic behavior arose when IKr 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.

The voltage-gated sodium channel inhibitor, 4,9-anhydrotetrodotoxin, blocks human Nav1.1 in addition to Nav1.6.

Voltage-gated sodium channels (VGSCs) are responsible for the initiation and propagation of action potentials in neurons. The human genome includes ten human VGSC α-subunit genes, SCN(X)A, encoding Nav1.1-1.9 plus Nax. To understand the unique role that each VGSC plays in normal and pathophysiological function in neural networks, compounds with high affinity and selectivity for specific VGSC subtypes are required. Toward that goal, a structural analog of the VGSC pore blocker tetrodotoxin, 4,9-anhydrotetrodotoxin (4,9-ah-TTX), has been reported to be more selective in blocking Na+ current mediated by Nav1.6 than other TTX-sensitive VGSCs, including Nav1.2, Nav1.3, Nav1.4, and Nav1.7. While SCN1A, encoding Nav1.1, has been implicated in several neurological diseases, the effects of 4,9-ah-TTX on Nav1.1-mediated Na+ current have not been tested. Here, we compared the binding of 4,9-ah-TTX for human and mouse brain preparations, and the effects of 4,9-ah-TTX on human Nav1.1-, Nav1.3- and Nav1.6-mediated Na+ currents using the whole-cell patch clamp technique in heterologous cells. We show that, while 4,9-ah-TTX administration results in significant blockade of Nav1.6-mediated Na+ current in the nanomolar range, it also has significant effects on Nav1.1-mediated Na+ current. Thus, 4,9-ah-TTX is not a useful tool in identifying Nav1.6-specific effects in human brain networks.

A microtranslatome coordinately regulates sodium and potassium currents in the human heart.

Catastrophic arrhythmias and sudden cardiac death can occur with even a small imbalance between inward sodium currents and outward potassium currents, but mechanisms establishing this critical balance are not understood. Here, we show that mRNA transcripts encoding INa and IKr channels (SCN5A and hERG, respectively) are associated in defined complexes during protein translation. Using biochemical, electrophysiological and single-molecule fluorescence localization approaches, we find that roughly half the hERG translational complexes contain SCN5A transcripts. Moreover, the transcripts are regulated in a way that alters functional expression of both channels at the membrane. Association and coordinate regulation of transcripts in discrete 'microtranslatomes' represents a new paradigm controlling electrical activity in heart and other excitable tissues.

Scn1b deletion in adult mice results in seizures and SUDEP.

Pathogenic loss-of-function variants in SCN1B are linked to Dravet syndrome (DS). Previous work suggested that neuronal pathfinding defects underlie epileptogenesis and SUDEP in the Scn1b null mouse model of DS. We tested this hypothesis by inducing Scn1b deletion in adult mice that had developed normally. Epilepsy and SUDEP, which occur by postnatal day 21 in Scn1b null animals, were observed within 20 days of induced Scn1b deletion in adult mice, suggesting that epileptogenesis in SCN1B-DS does not result from defective brain development. Thus, the developmental brain defects observed previously in Scn1b null mice may model other co-morbidities of DS.