Comparative structural analysis of human Nav1.1 and Nav1.5 reveals mutational hotspots for sodium channelopathies.
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SUBMITTER: Pan X
PROVIDER: S-EPMC7980448 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Background & aimsSCN5A encodes the α-subunit of the voltage-gated sodium channel NaV1.5. Many patients with cardiac arrhythmias caused by mutations in SCN5A also have symptoms of irritable bowel syndrome (IBS). We investigated whether patients with IBS have SCN5A variants that affect the function of NaV1.5.MethodsWe performed genotype analysis of SCN5A in 584 persons with IBS and 1380 without IBS (controls). Mutant forms of SCN5A were expressed in human embryonic kidney-293 cells, and functions were assessed by voltage clamp analysis. A genome-wide association study was analyzed for an association signal for the SCN5A gene, and replicated in 1745 patients in 4 independent cohorts of IBS patients and controls.ResultsMissense mutations were found in SCN5A in 13 of 584 patients (2.2%, probands). Diarrhea-predominant IBS was the most prevalent form of IBS in the overall study population (25%). However, a greater percentage of individuals with SCN5A mutations had constipation-predominant IBS (31%) than diarrhea-predominant IBS (10%; P < .05). Electrophysiologic analysis showed that 10 of 13 detected mutations disrupted NaV1.5 function (9 loss-of-function and 1 gain-of-function function). The p. A997T-NaV1.5 had the greatest effect in reducing NaV1.5 function. Incubation of cells that expressed this variant with mexiletine restored their sodium current and administration of mexiletine to 1 carrier of this mutation (who had constipation-predominant IBS) normalized their bowel habits. In the genome-wide association study and 4 replicated studies, the SCN5A locus was strongly associated with IBS.ConclusionsAbout 2% of patients with IBS carry mutations in SCN5A. Most of these are loss-of-function mutations that disrupt NaV1.5 channel function. These findings provide a new pathogenic mechanism for IBS and possible treatment options.
| S-EPMC4096335 | biostudies-literature
Project description:Abnormalities in the human Nav1.5 (hNav1.5) voltage-gated sodium ion channel (VGSC) are associated with a wide range of cardiac problems and diseases in humans. Current structural models of hNav1.5 are still far from complete and, consequently, their ability to study atomistic interactions of this channel is very limited. Here, we report a comprehensive atomistic model of the hNav1.5 ion channel, constructed using homology modeling technique and refined through long molecular dynamics simulations (680 ns) in the lipid membrane bilayer. Our model was comprehensively validated by using reported mutagenesis data, comparisons with previous models, and binding to a panel of known hNav1.5 blockers. The relatively long classical MD simulation was sufficient to observe a natural sodium permeation event across the channel's selectivity filters to reach the channel's central cavity, together with the identification of a unique role of the lysine residue. Electrostatic potential calculations revealed the existence of two potential binding sites for the sodium ion at the outer selectivity filters. To obtain further mechanistic insight into the permeation event from the central cavity to the intracellular region of the channel, we further employed "state-of-the-art" steered molecular dynamics (SMD) simulations. Our SMD simulations revealed two different pathways through which a sodium ion can be expelled from the channel. Further, the SMD simulations identified the key residues that are likely to control these processes. Finally, we discuss the potential binding modes of a panel of known hNav1.5 blockers to our structural model of hNav1.5. We believe that the data presented here will enhance our understanding of the structure-property relationships of the hNav1.5 ion channel and the underlying molecular mechanisms in sodium ion permeation and drug interactions. The results presented here could be useful for designing safer drugs that do not block the hNav1.5 channel.
| S-EPMC5552146 | biostudies-literature
Project description:Voltage-gated sodium (NaV) channels are transmembrane proteins that initiate and propagate neuronal and cardiac action potentials. NaV channel ? subunits have been widely studied due to their modulatory role. Mice null for Scn1b, which encodes NaV ?1 and ?1b subunits, have defects in neuronal development and excitability, spontaneous generalized seizures, cardiac arrhythmias, and early mortality. A mutation in exon 3 of SCN1B, c.308A>T leading to ?1_p.D103V and ?1b_p.D103V, was previously found in a patient with a history of proarrhythmic conditions with progressive atrial standstill as well as cognitive and motor deficits accompanying structural brain abnormalities. We investigated whether ?1 or ?1b subunits carrying this mutation affect NaV1.5 and/or NaV1.1 currents using a whole cell patch-clamp technique in tsA201 cells. We observed a decrease in sodium current density in cells co-expressing NaV1.5 or NaV1.1 and ?1D103V compared to ?1WT. Interestingly, ?1bD103V did not affect NaV1.1 sodium current density but induced a positive shift in the voltage dependence of inactivation and a faster recovery from inactivation compared to ?1bWT. The ?1bD103V isoform did not affect NaV1.5 current properties. Although the SCN1B_c.308A>T mutation may not be the sole cause of the patient's symptoms, we observed a clear loss of function in both cardiac and brain sodium channels. Our results suggest that the mutant ?1 and ?1b subunits play a fundamental role in the observed electrical dysfunction.
| S-EPMC7550680 | biostudies-literature
Project description:The voltage-gated sodium channel α-subunit genes comprise a highly conserved gene family. Mutations of three of these genes, SCN1A, SCN2A and SCN8A, are responsible for a significant burden of neurological disease. Recent progress in identification and functional characterization of patient variants is generating new insights and novel approaches to therapy for these devastating disorders. Here we review the basic elements of sodium channel function that are used to characterize patient variants. We summarize a large body of work using global and conditional mouse mutants to characterize the in vivo roles of these channels. We provide an overview of the neurological disorders associated with mutations of the human genes and examples of the effects of patient mutations on channel function. Finally, we highlight therapeutic interventions that are emerging from new insights into mechanisms of sodium channelopathies.
| S-EPMC8710247 | biostudies-literature
Project description:Dendrobium is one of the largest genera in Orchidaceae, comprising about 800-1500 species mainly distributed in tropical Asia, Australasia, and Australia. There are 74 species and two varieties of this genus in China. Because of their ornamental and commercial value, Dendrobium orchids have been studied at low taxonomic levels. However, structural changes and effective mutational hotspots of Dendrobium plastomes have rarely been documented. Here, 30 Dendrobium plastomes were compared, comprising 25 newly sequenced in this study and five previously published. Except for their differences in NDH genes, these plastomes shared identical gene content and order. Comparative analyses revealed that the variation in size of Dendroubium plastomes was associated with dramatically changed length of InDels. Furthermore, ten loci were identified as the top-ten mutational hotspots, whose sequence variability was almost unchanged with more than 10 plastomes sampled, suggesting that they may be powerful markers for Dendrobium species. In addition, primer pairs of 47 polymorphic microsatellites were developed. After assessing the mean BS values of all combinations derived from the top-ten hotspots, we recommend that the combination of five hotspots-trnT-trnL, rpl32-trnL, clpP-psbB, trnL intron, and rps16-trnQ-should be used in the phylogenetic and identification studies of Dendrobium.
| S-EPMC5437043 | biostudies-literature
Project description:The SCN5A-encoded voltage-gated mechanosensitive Na+ channel NaV1.5 is expressed in human gastrointestinal smooth muscle cells and interstitial cells of Cajal. NaV1.5 contributes to smooth muscle electrical slow waves and mechanical sensitivity. In predominantly Caucasian irritable bowel syndrome (IBS) patient cohorts, 2-3% of patients have SCN5A missense mutations that alter NaV1.5 function and may contribute to IBS pathophysiology. In this study we examined a racially and ethnically diverse cohort of IBS patients for SCN5A missense mutations, compared them with IBS-negative controls, and determined the resulting NaV1.5 voltage-dependent and mechanosensitive properties. All SCN5A exons were sequenced from somatic DNA of 252 Rome III IBS patients with diverse ethnic and racial backgrounds. Missense mutations were introduced into wild-type SCN5A by site-directed mutagenesis and cotransfected with green fluorescent protein into HEK-293 cells. NaV1.5 voltage-dependent and mechanosensitive functions were studied by whole cell electrophysiology with and without shear force. Five of 252 (2.0%) IBS patients had six rare SCN5A mutations that were absent in 377 IBS-negative controls. Six of six (100%) IBS-associated NaV1.5 mutations had voltage-dependent gating abnormalities [current density reduction (R225W, R433C, R986Q, and F1293S) and altered voltage dependence (R225W, R433C, R986Q, G1037V, and F1293S)], and at least one kinetic parameter was altered in all mutations. Four of six (67%) IBS-associated SCN5A mutations (R225W, R433C, R986Q, and F1293S) resulted in altered NaV1.5 mechanosensitivity. In this racially and ethnically diverse cohort of IBS patients, we show that 2% of IBS patients harbor SCN5A mutations that are absent in IBS-negative controls and result in NaV1.5 channels with abnormal voltage-dependent and mechanosensitive function. NEW & NOTEWORTHY The voltage-gated Na+ channel NaV1.5 contributes to smooth muscle physiology and electrical slow waves. In a racially and ethnically mixed irritable bowel syndrome cohort, 2% had mutations in the NaV1.5 gene SCN5A. These mutations were absent in irritable bowel syndrome-negative controls. Most mutant NaV1.5 channels were loss of function in voltage dependence or mechanosensitivity.
| S-EPMC5966747 | biostudies-literature
Project description:Inherited mutations in voltage-gated sodium channels (VGSCs; or Nav) cause many disorders of excitability, including epilepsy, chronic pain, myotonia, and cardiac arrhythmias. Understanding the functional consequences of the disease-causing mutations is likely to provide invaluable insight into the roles that VGSCs play in normal and abnormal excitability. Here, we sought to test the hypothesis that disease-causing mutations lead to increased resurgent currents, unusual sodium currents that have not previously been implicated in disorders of excitability. We demonstrated that a paroxysmal extreme pain disorder (PEPD) mutation in the human peripheral neuronal sodium channel Nav1.7, a paramyotonia congenita (PMC) mutation in the human skeletal muscle sodium channel Nav1.4, and a long-QT3/SIDS mutation in the human cardiac sodium channel Nav1.5 all substantially increased the amplitude of resurgent sodium currents in an optimized adult rat-derived dorsal root ganglion neuronal expression system. Computer simulations indicated that resurgent currents associated with the Nav1.7 mutation could induce high-frequency action potential firing in nociceptive neurons and that resurgent currents associated with the Nav1.5 mutation could broaden the action potential in cardiac myocytes. These effects are consistent with the pathophysiology associated with the respective channelopathies. Our results indicate that resurgent currents are associated with multiple channelopathies and are likely to be important contributors to neuronal and muscle disorders of excitability.
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Project description:Angiosarcoma is a highly aggressive cancer of blood vessel-forming cells with few effective treatment options and high patient mortality. It is both rare and heterogenous, making large, well-powered genomic studies nearly impossible. Dogs commonly suffer from a similar cancer, called hemangiosarcoma, with breeds like the golden retriever carrying heritable genetic factors that put them at high risk. If the clinical similarity of canine hemangiosarcoma and human angiosarcoma reflects shared genomic etiology, dogs could be a critically needed model for advancing angiosarcoma research. We assessed the genomic landscape of canine hemangiosarcoma via whole-exome sequencing (47 golden retriever hemangiosarcomas) and RNA sequencing (74 hemangiosarcomas from multiple breeds). Somatic coding mutations occurred most frequently in the tumor suppressor TP53 (59.6% of cases) as well as two genes in the PI3K pathway: the oncogene PIK3CA (29.8%) and its regulatory subunit PIK3R1 (8.5%). The predominant mutational signature was the age-associated deamination of cytosine to thymine. As reported in human angiosarcoma, CDKN2A/B was recurrently deleted and VEGFA, KDR, and KIT recurrently gained. We compared the canine data to human data recently released by The Angiosarcoma Project, and found many of the same genes and pathways significantly enriched for somatic mutations, particularly in breast and visceral angiosarcomas. Canine hemangiosarcoma closely models the genomic landscape of human angiosarcoma of the breast and viscera, and is a powerful tool for investigating the pathogenesis of this devastating disease. IMPLICATIONS: We characterize the genomic landscape of canine hemangiosarcoma and demonstrate its similarity to human angiosarcoma.
| S-EPMC7067513 | biostudies-literature
Project description:The discovery of driver genes is a major pursuit of cancer genomics, usually based on observing the same mutation in different patients. But the heterogeneity of cancer pathways plus the high background mutational frequency of tumor cells often cloud the distinction between less frequent drivers and innocent passenger mutations. Here, to overcome these disadvantages, we grouped together mutations from close kinase paralogs under the hypothesis that cognate mutations may functionally favor cancer cells in similar ways. Indeed, we find that kinase paralogs often bear mutations to the same substituted amino acid at the same aligned positions and with a large predicted Evolutionary Action. Functionally, these high Evolutionary Action, non-random mutations affect known kinase motifs, but strikingly, they do so differently among different kinase types and cancers, consistent with differences in selective pressures. Taken together, these results suggest that cancer pathways may flexibly distribute a dependence on a given functional mutation among multiple close kinase paralogs. The recognition of this "mutational delocalization" of cancer drivers among groups of paralogs is a new phenomena that may help better identify relevant mechanisms and therefore eventually guide personalized therapy.
| S-EPMC5157159 | biostudies-literature
Project description:Here an analysis is presented of the roles of recombination and mutation in shaping previously determined haplotype variation in 9.7 kb of genomic DNA sequence from the human lipoprotein lipase gene (LPL), scored in 71 individuals from three populations: 24 African Americans, 24 Finns, and 23 non-Hispanic whites. Recombination and gene-conversion events inferred from data on 88 haplotypes that were defined by 69 variable sites were tested. The analysis revealed 29 statistically significant recombination events and one gene-conversion event. The recombination events were concentrated in a 1.9-kb region, near the middle of the segment, that contains a microsatellite and a pair of tandem and complementary mononucleotide runs; both the microsatellite and the runs show length variation. An analysis of site variation revealed that 9.6% of the nucleotides at CpG sites were variable, as were 3% of the nucleotides found in mononucleotide runs of >/=5 nucleotides, 3% of the nucleotides found </=3 bp from certain putative polymerase alpha-arrest sites, and 0. 5% of the remaining nucleotides. This nonhomogeneous distribution of variation suggests that multiple mutational hits at certain sites are common, an observation that challenges the fundamental assumption of the infinite-sites-mutation model. The nonrandom patterns of recombination and mutation suggest that randomly chosen single-nucleotide polymorphisms may not be optimal for disequilibrium mapping of this gene. Overall, these results indicate that both recombinational and mutational hotspots have played significant roles in shaping the haplotype variation at the LPL locus.
| S-EPMC1288350 | biostudies-literature