Genome-wide association studies (GWAS) have identified thousands of genetic variants linked to the risk of human disease. However, GWAS have so far remained largely underpowered in relation to identifying associations in the rare and low-frequency allelic spectrum and have lacked the resolution to trace causal mechanisms to underlying genes¹. Here we combined whole-exome sequencing in 392,814 UK Biobank participants with imputed genotypes from 260,405 FinnGen participants (653,219 total individuals) to conduct association meta-analyses for 744 disease endpoints across the protein-coding allelic frequency spectrum, bridging the gap between common and rare variant studies. We identified 975 associations, with more than one-third being previously unreported. We demonstrate population-level relevance for mutations previously ascribed to causing single-gene disorders, map GWAS associations to likely causal genes, explain disease mechanisms, and systematically relate disease associations to levels of 117 biomarkers and clinical-stage drug targets. Combining sequencing and genotyping in two population biobanks enabled us to benefit from increased power to detect and explain disease associations, validate findings through replication and propose medical actionability for rare genetic variants. Our study provides a compendium of protein-coding variant associations for future insights into disease biology and drug discovery.

Voltage-gated Na⁺channels (VGSC) are transmembrane proteins that are essential for the initiation and propagation of action potentials in neuronal excitability. Because neurons express a mixture of Na⁺channel isoforms and protein kinase C (PKC) isozymes, the nature of which channel is being regulated by which PKC isozyme is not known. We showed that DRG VGSC Na_v1.7 (TTX-sensitive) and Na_v1.8 (TTX-resistant), expressed in Xenopus oocytes were differentially regulated by protein kinase A (PKA) and PKC isozymes using the two-electrode voltage-clamp method. PKA activation resulted in a dose-dependent potentiation of Na_v1.8 currents and an attenuation of Na_v1.7 currents. PKA-induced increases (Na_v1.8) and decreases (Na_v1.7) in peak currents were not associated with shifts in voltage-dependent activation or inactivation. The PKA-mediated increase in Na_v1.8 current amplitude was prevented by chloroquine, suggesting that cell trafficking may contribute to the changes in Na_v1.8 current amplitudes. A dose-dependent decrease in Na_v1.7 and Na_v1.8 currents was observed with the PKC activators phorbol 12-myristate, 13-acetate (PMA) and phorbol 12,13-dibutyrate. PMA induced shifts in the steady-state activation of Na_v1.7 and Na_v1.8 channels by 6.5 and 14 mV, respectively, in the depolarizing direction. The role of individual PKC isozymes in the regulation of Na_v1.7 and Na_v1.8 was determined using PKC-isozyme-specific peptide activators and inhibitors. The decrease in the Na_v1.8 peak current induced by PMA was prevented by a specific ϵPKC isozyme peptide antagonist, whereas the PMA effect on Na_v1.7 was prevented by ϵPKC and βIIPKC peptide inhibitors. The data showed that Na_v1.7 and Na_v1.8 were differentially modulated by PKA and PKC. This is the first report demonstrating a functional role for ϵPKC and βIIPKC in the regulation of Na_v1.7 and Na_v1.8 Na⁺channels. Identification of the particular PKC isozymes(s) that mediate the regulation of Na⁺channels is essential for understanding the molecular mechanism involved in neuronal ion channel regulation in normal and pathological conditions.

Brugada syndrome is an inherited cardiac disorder caused by mutations in the cardiac sodium channel gene, SCN5A, that leads to ventricular fibrillation and sudden death. This study reports the changes in functional expression and cellular localization of an SCN5A double mutant (R1232W/T1620M) recently discovered in patients with Brugada syndrome. Mutant and wild-type (WT) human heart sodium channels (hNa _v 1.5) were expressed in tsA201 cells in the presence of the β ₁ -auxiliary subunit. Patch-clamp experiments in whole-cell configuration were conducted to assess functional expression. Immunohistochemistry and confocal microscopy were used to determine the spatial distribution of either WT or mutant cardiac sodium channels. The results show an abolition of functional sodium channel expression of the hNa _v 1.5/R1232W/T1620M mutant in the tsA201 cells. A conservative positively charged mutant, hNa _v 1.5/R1232K/T1620M, produced functional channels. Immunofluorescent staining showed that the FLAG-tagged hNa _v 1.5/WT transfected into tsA201 cells was localized on the cell surface, whereas the FLAG-tagged hNa _v 1.5/R1232W/T1620M mutant was colocalized with calnexin within the endoplasmic reticulum (ER). These results indicate that a positively charged arginine or lysine residue at position 1232 in the double mutant is required for the proper transport and functional expression of the hNa _v 1.5 protein. These results support the concept that loss of function of the cardiac Na ⁺ channel is responsible for the Brugada syndrome. The full text of this article is available at http://www.circresaha.org.

Cardiac Na⁺ channels encoded by the SCN5A gene are essential for initiating heart beats and maintaining a regular heart rhythm. Mutations in these channels have recently been associated with atrial fibrillation, ventricular arrhythmias, conduction disorders, and dilated cardiomyopathy (DCM). We investigated a young male patient with a mixed phenotype composed of documented conduction disorder, atrial flutter, and ventricular tachycardia associated with DCM. Further family screening revealed DCM in the patient's mother and sister and in three of the mother's sisters. Because of the complex clinical phenotypes, we screened SCN5A and identified a novel mutation, R219H, which is located on a highly conserved region on the fourth helix of the voltage sensor domain of Na_v1.5. Three family members with DCM carried the R219H mutation. The wild-type (WT) and mutant Na⁺ channels were expressed in a heterologous expression system, and intracellular pH (pHi) was measured using a pH-sensitive electrode. The biophysical characterization of the mutant channel revealed an unexpected selective proton leak with no effect on its biophysical properties. The H⁺ leak through the mutated Na_v1.5 channel was not related to the Na⁺ permeation pathway but occurred through an alternative pore, most probably a proton wire on the voltage sensor domain. We propose that acidification of cardiac myocytes and/or downstream events may cause the DCM phenotype and other electrical problems in affected family members. The identification of this clinically significant H⁺ leak may lead to the development of more targeted treatments.

Voltage sensor domain (VSDs) are a feature of voltage gated ion channel (VGICs) and voltage sensitive proteins. They are composed of four transmembrane (TM) segments (S1 to S4). Currents leaking through VSDs are called omega or gating pore currents.Gating pores are caused by mutations of the highly conserved positively charged amino acids in the S4 segment that disrupt interactions between the S4 segment and the gating charge transfer center (GCTC). The GCTC separates the intracellular and extracellular water crevices. The disruption of S4–GCTC interactions allows these crevices to communicate and create a fast activating and non-inactivating alternative cation-selective permeation pathway of low conductance, or a gating pore.Gating pore currents have recently been shown to cause periodic paralysis phenotypes. There is also increasing evidence that gating pores are linked to several other familial diseases. For example, gating pores in Nav1.5 and Kv7.2 channels may underlie mixed arrhythmias associated with dilated cardiomyopathy (DCM) phenotypes and peripheral nerve hyperexcitability (PNH) respectively. There is little evidence for the existence of gating pore blockers. Moreover, it is known that a number of toxins bind to the VSD of a specific domain of Na+ channels. These toxins may thus modulate gating pore currents. This focus on the VSD motif opens up a new area of research centered on developing molecules to treat a number of cell excitability disorders such as epilepsy, cardiac arrhythmias, and pain.The purpose of the present review is to summarize existing knowledge of the pathophysiology, biophysics, and pharmacology of gating pore currents and to serve as a guide for future studies aimed at improving our understanding of gating pores and their pathophysiological roles.