Gene variant effects across sodium channelopathies predict function and guide precision therapy

Pathogenic variants in the voltage-gated sodium channel gene family (SCNs) lead to early onset epilepsies, neurodevelopmental disorders, skeletal muscle channelopathies, peripheral neuropathies and cardiac arrhythmias. Disease-associated variants have diverse functional effects ranging from complete loss-of-function to marked gain-of-function. Therapeutic strategy is likely to depend on functional effect. Experimental studies offer important insights into channel function, but are resource intensive and only performed in a minority of cases. Given the evolutionarily conserved nature of the sodium channel genes we investigated whether similarities in biophysical properties between different voltage-gated sodium channels can predict function and inform precision treatment across sodium channelopathies. We performed a systematic literature search identifying functionally assessed variants in any of the nine voltage-gated sodium channel genes until 28 April 2021. We included missense variants that had been electrophysiologically characterised in mammalian cells in whole-cell patch-clamp recordings. We performed an alignment of linear protein sequences of all sodium channel genes and correlated variants by their overall functional effect on biophysical properties. Of 951 identified records, 437 sodium channel-variants met our inclusion criteria and were reviewed for functional properties. Of these, 141 variants were epilepsy-associated (SCN1/2/3/8A), 79 had a neuromuscular phenotype (SCN4/9/10/11A), 149 were associated with a cardiac phenotype (SCN5/10A) and 68 (16%) were considered benign. We detected 38 missense variant pairs with an identical disease-associated variant in a different sodium channel gene. 35 out of 38 of those pairs resulted in similar functional consequences indicating up to 92% biophysical agreement between corresponding sodium channel variants (odds ratio = 11.3; 95% CI = 2.8 to 66.9; P < 0.001). Pathogenic missense variants were clustered in specific functional domains, whereas population variants were significantly more frequent across non conserved domains (odds ratio = 18.6; 95% CI = 10.9 to 34.4; P < 0.001). Pore-loop regions were frequently associated with loss-of-function (LoF) variants, whereas inactivation sites were associated with gain-of-function (GoF; odds ratio = 42.1, 95% CI = 14.5 to 122.4; P < 0.001), whilst variants occurring in voltage-sensing regions comprised a range of gain- and loss-of-function effects. Our findings suggest that biophysical characterisation of variants in one SCN-gene can predict channel function across different SCN-genes where experimental data are not available. The collected data represent the first GoF versus LoF topological map of SCN proteins indicating shared patterns of biophysical effects aiding variant analysis and guiding precision therapy. We integrated our findings into a free online webtool to facilitate functional sodium channel gene variant interpretation (http://SCN-viewer.broadinstitute.org)

Delineation of functionally essential protein regions for 242 neurodevelopmental genes

Neurodevelopmental disorders (NDDs), including severe paediatric epilepsy, autism and intellectual disabilities are heterogeneous conditions in which clinical genetic testing can often identify a pathogenic variant. For many of them, genetic therapies will be tested in this or the coming years in clinical trials. In contrast to first-generation symptomatic treatments, the new disease-modifying precision medicines require a genetic test-informed diagnosis before a patient can be enrolled in a clinical trial. However, even in 2022, most identified genetic variants in NDD genes are "variants of uncertain significance'. To safely enrol patients in precision medicine clinical trials, it is important to increase our knowledge about which regions in NDD-associated proteins can "tolerate' missense variants and which ones are "essential' and will cause a NDD when mutated. In addition, knowledge about functionally indispensable regions in the 3D structure context of proteins can also provide insights into the molecular mechanisms of disease variants. We developed a novel consensus approach that overlays evolutionary, and population based genomic scores to identify 3D essential sites (Essential3D) on protein structures. After extensive benchmarking of AlphaFold predicted and experimentally solved protein structures, we generated the currently largest expert curated protein structure set for 242 NDDs and identified 14 377 Essential3D sites across 189 gene disorders associated proteins. We demonstrate that the consensus annotation of Essential3D sites improves prioritization of disease mutations over single annotations. The identified Essential3D sites were enriched for functional features such as intermembrane regions or active sites and discovered key inter-molecule interactions in protein complexes that were otherwise not annotated. Using the currently largest autism, developmental disorders, and epilepsies exome sequencing studies including > 360 000 NDD patients and population controls, we found that missense variants at Essential3D sites are 8-fold enriched in patients.In summary, we developed a comprehensive protein structure set for 242 NDDs and identified 14377 Essential3D sites in these. All data are available at https://es-ndd.broadinstitute.org for interactive visual inspection to enhance variant interpretation and development of mechanistic hypotheses for 242 NDDs genes. The provided resources will enhance clinical variant interpretation and in silico drug target development for NDD-associated genes and encoded proteins.Peer reviewe

SCN1A variants from bench to bedside-improved clinical prediction from functional characterization

Variants in the SCN1A gene are associated with a wide range of disorders including genetic epilepsy with febrile seizures plus (GEFS+), familial hemiplegic migraine (FHM), and the severe childhood epilepsy Dravet syndrome (DS). Predicting disease outcomes based on variant type remains challenging. Despite thousands of SCN1A variants being reported, only a minority has been functionally assessed. We review the functional SCN1A work performed to date, critically appraise electrophysiological measurements, compare this to in silico predictions, and relate our findings to the clinical phenotype. Our results show, regardless of the underlying phenotype, that conventional in silico software correctly predicted benign from pathogenic variants in nearly 90%, however was unable to differentiate within the disease spectrum (DS vs. GEFS+ vs. FHM). In contrast, patch‐clamp data from mammalian expression systems revealed functional differences among missense variants allowing discrimination between disease severities. Those presenting with milder phenotypes retained a degree of channel function measured as residual whole‐cell current, whereas those without any whole‐cell current were often associated with DS (p = .024). These findings demonstrate that electrophysiological data from mammalian expression systems can serve as useful disease biomarker when evaluating SCN1A variants, particularly in view of new and emerging treatment options in DS

Metal Concentrations in e-Cigarette Liquid and Aerosol Samples:The Contribution of Metallic Coils

Supplemental Material is available online (https://doi.org/10.1289/EHP2175)Electronic cigarettes (e-cigarettes) generate an aerosol by heating a solution (e-liquid) with a metallic coil. Whether metals are transferred from the coil to the aerosol is unknown. Objective: Our goal was to investigate the transfer of metals from the heating coil to the e-liquid in the e-cigarette tank and the generated aerosol. Methods: We sampled 56 e-cigarette devices from daily e-cigarette users and obtained samples from the refilling dispenser, aerosol, and remaining e-liquid in the tank. Aerosol liquid was collected via deposition of aerosol droplets in a series of conical pipette tips. Metals were reported as mass fractions (μg/kg) in liquids and converted to mass concentrations (mg/m3) for aerosols. Results: Median metal concentrations (μg/kg) were higher in samples from the aerosol and tank vs. the dispenser (all p<0.001): 16.3 and 31.2 vs. 10.9 for Al; 8.38 and 55.4 vs. <0.5 for Cr; 68.4 and 233 vs. 2.03 for Ni; 14.8 and 40.2 vs. 0.476 for Pb; and 515 and 426 vs. 13.1 for Zn. Mn, Fe, Cu, Sb, and Sn were detectable in most samples. Cd was detected in 0.0, 30.4, and 55.1% of the dispenser, aerosol, and tank samples respectively. Arsenic was detected in 10.7% of dispenser samples (median 26.7 μg/kg) and these concentrations were similar in aerosol and tank samples. Aerosol mass concentrations (mg/m3) for the detected metals spanned several orders of magnitude and exceeded current health-based limits in close to 50% or more of the samples for Cr, Mn, Ni, and Pb. Conclusions: Our findings indicate that e-cigarettes are a potential source of exposure to toxic metals (Cr, Ni, and Pb), and to metals that are toxic when inhaled (Mn and Zn). Markedly higher concentrations in the aerosol and tank samples versus the dispenser demonstrate that coil contact induced e-liquid contamination.This study is supported by the Cigarette Restitution Fund (State of Maryland; grant PHPA-G2034). P.O. was supported by the Alfonso Martín Escudero Foundation (postdoctoral fellowship 2014). A.A. was supported by the American Heart Association Tobacco Regulation and Addiction Center (grant 1P50HL120163). A.N.A., M.H., and P.O. are supported by the National Institute of Environmental Health Sciences/National Institutes of Health (grant 5P30ES009089)

Targeting qubit states using open-loop control

We present an open-loop (bang-bang) scheme which drives an open two-level quantum system to any target state, while maintaining quantum coherence throughout the process. The control is illustrated by a realistic simulation for both adiabatic and thermal decoherence. In the thermal decoherence regime, the control achieved by the proposed scheme is qualitatively similar, at the ensemble level, to the control realized by the quantum feedback scheme of Wang, Wiseman, and Milburn [Phys. Rev. A 64, #063810 (2001)] for the spontaneous emission of a two-level atom. The performance of the open-loop scheme compares favorably against the quantum feedback scheme with respect to robustness, target fidelity and transition times.Comment: 27 pages, 7 figure

Gene variant effects across sodium channelopathies predict function and guide precision therapy.

Pathogenic variants in the voltage-gated sodium channel gene family lead to early onset epilepsies, neurodevelopmental disorders, skeletal muscle channelopathies, peripheral neuropathies and cardiac arrhythmias. Disease-associated variants have diverse functional effects ranging from complete loss-of-function to marked gain-of-function. Therapeutic strategy is likely to depend on functional effect. Experimental studies offer important insights into channel function but are resource intensive and only performed in a minority of cases. Given the evolutionarily conserved nature of the sodium channel genes, we investigated whether similarities in biophysical properties between different voltage-gated sodium channels can predict function and inform precision treatment across sodium channelopathies. We performed a systematic literature search identifying functionally assessed variants in any of the nine voltage-gated sodium channel genes until 28 April 2021. We included missense variants that had been electrophysiologically characterized in mammalian cells in whole-cell patch-clamp recordings. We performed an alignment of linear protein sequences of all sodium channel genes and correlated variants by their overall functional effect on biophysical properties. Of 951 identified records, 437 sodium channel-variants met our inclusion criteria and were reviewed for functional properties. Of these, 141 variants were epilepsy-associated (SCN1/2/3/8A), 79 had a neuromuscular phenotype (SCN4/9/10/11A), 149 were associated with a cardiac phenotype (SCN5/10A) and 68 (16%) were considered benign. We detected 38 missense variant pairs with an identical disease-associated variant in a different sodium channel gene. Thirty-five out of 38 of those pairs resulted in similar functional consequences, indicating up to 92% biophysical agreement between corresponding sodium channel variants (odds ratio = 11.3; 95% confidence interval = 2.8 to 66.9; P < 0.001). Pathogenic missense variants were clustered in specific functional domains, whereas population variants were significantly more frequent across non-conserved domains (odds ratio = 18.6; 95% confidence interval = 10.9-34.4; P < 0.001). Pore-loop regions were frequently associated with loss-of-function variants, whereas inactivation sites were associated with gain-of-function (odds ratio = 42.1, 95% confidence interval = 14.5-122.4; P < 0.001), whilst variants occurring in voltage-sensing regions comprised a range of gain- and loss-of-function effects. Our findings suggest that biophysical characterisation of variants in one SCN-gene can predict channel function across different SCN-genes where experimental data are not available. The collected data represent the first gain- versus loss-of-function topological map of SCN proteins indicating shared patterns of biophysical effects aiding variant analysis and guiding precision therapy. We integrated our findings into a free online webtool to facilitate functional sodium channel gene variant interpretation (http://SCN-viewer.broadinstitute.org)

Novel spatial domain integral equation formulation for the analysis of rectangular waveguide steps close to arbitrarily shaped dielectric and/or conducting posts

[EN] In this paper, a novel integral equation formulation expressed in the spatial domain is proposed for the analysis of rectangular waveguide step discontinuities. The important novelty of the proposed formulation is that which allows to easily take into account the electrical influence of a given number of arbitrarily shaped conducting and dielectric posts placed close to the waveguide discontinuity. For the sake of simplicity, and without loss of generality, the presented integral equation has been particularized and solved for inductive rectangular waveguide geometry. In this case, the integral equation mixed-potentials kernel is written in terms of parallel plate Green¿s functions with an additional ground plane located on the waveguide step. Therefore, the unknowns of the problem are reduced to an equivalent magnetic surface current on the step aperture and equivalent magnetic and electric surface currents on the dielectric and conducting posts close to the discontinuity. The numerical solution of the final integral equation is efficiently computed after the application of acceleration techniques for the slowly convergent series representing the Green¿s functions of the problem. The numerical method has been validated through several simulation examples of practical microwave devices, including compact size band-pass cavity filters and coupled dielectric resonators filters. The results have been compared to those provided by commercial full-wave electromagnetic simulation software packages, showing in all cases a very good agreement, and with substantially enhanced numerical efficiencies.This research work has been financially supported by the Spanish Ministerio de Economia y Competitividad in the frame of the projects "Demostradores Tecnologicos de Filtros y Multiplexores con Respuestas Selectivas y Sintonizables en Nuevas Guias Compactas para Aplicaciones Espaciales (COMPASSES)" with Ref. TEC2016-75934-C4-1-R, and "Analisis y Diseno de Nuevos Componentes en Microondas y Milimetricas para Comunicaciones por Satelite (MILISAT)" with Ref. TEC2016-75934-C4-4-R. As an additional financial source we thank the regional agency Fundacion Seneca from Region de Murcia under the research project "Desarrollo de Antenas y Componentes Pasivos de Microondas para Sistemas Avanzados de Comunicaciones" with Ref. 19494/PI/14 and Ref. 20147/EE/17, and the PhD scholarship granted by the Spanish national Ministerio de Educacion, Cultura y Deporte with Ref. FPU15/02883. All results of this paper can be reproduced by using the data and information contained in the drawings and in the captions of the figures included in the paper.Quesada Pereira, FD.; Gomez Molina, C.; Alvarez Melcon, A.; Boria Esbert, VE.; Guglielmi, M. (2018). Novel spatial domain integral equation formulation for the analysis of rectangular waveguide steps close to arbitrarily shaped dielectric and/or conducting posts. Radio Science. 53(4):406-419. https://doi.org/10.1002/2017RS006429S406419534Arcioni , P. Bressan , M. Conciauro , G. Perregrini , L. 1997 Generalized Y-matrix of arbitrary H-plane waveguide junctions by the BI-RME method IEEE MTT-S International Microwave Symposium Digest 211 214 DenverCapolino, F., Wilton, D. R., & Johnson, W. A. (2005). Efficient computation of the 2-D Green’s function for 1-D periodic structures using the Ewald method. IEEE Transactions on Antennas and Propagation, 53(9), 2977-2984. doi:10.1109/tap.2005.854556Catina, V., Arndt, F., & Brandt, J. (2005). Hybrid surface integral-equation/mode-matching method for the analysis of dielectric loaded waveguide filters of arbitrary shape. IEEE Transactions on Microwave Theory and Techniques, 53(11), 3562-3567. doi:10.1109/tmtt.2005.857343Fructos, A. L., Boix, R. R., Mesa, F., & Medina, F. (2008). An Efficient Approach for the Computation of 2-D Green’s Functions With 1-D and 2-D Periodicities in Homogeneous Media. IEEE Transactions on Antennas and Propagation, 56(12), 3733-3742. doi:10.1109/tap.2008.2007281Guglielmi, M., & Newport, C. (1990). Rigorous, multimode equivalent network representation of inductive discontinuities. IEEE Transactions on Microwave Theory and Techniques, 38(11), 1651-1659. doi:10.1109/22.60012Hu, Y. L., Li, J., Ding, D. Z., & Chen, R. S. (2016). Analysis of Transient EM Scattering From Penetrable Objects by Time Domain Nonconformal VIE. IEEE Transactions on Antennas and Propagation, 64(1), 360-365. doi:10.1109/tap.2015.2501437Kalantari, M., & Paran, K. (2017). Analysing Metamaterial Layer by Simpler Approach Based on Mode Matching Technique. IET Microwaves, Antennas & Propagation, 11(5), 607-616. doi:10.1049/iet-map.2016.0687Mrvić, M., Potrebić, M., & Tošić, D. (2016). CompactEplane waveguide filter with multiple stopbands. Radio Science, 51(12), 1895-1904. doi:10.1002/2016rs006169Pérez-Soler, F. J., Quesada-Pereira, F. D., Cañete Rebenaque, D., Pascual-García, J., & Alvarez-Melcon, A. (2007). Efficient integral equation formulation for inductive waveguide components with posts touching the waveguide walls. Radio Science, 42(6). doi:10.1029/2006rs003591POGGIO, A. J., & MILLER, E. K. (1973). Integral Equation Solutions of Three-dimensional Scattering Problems. Computer Techniques for Electromagnetics, 159-264. doi:10.1016/b978-0-08-016888-3.50008-8Quesada Pereira , F. Boria , V. E. Gimeno , B. Cañete Rebenaque , D. Pascual Garcia , J. Alvarez Melcon , A. 2006 Investigation of multipaction phenomena in inductively coupled passive waveguide components for space applications IEEE MTT-S International Microwave Symposium Digest 246 249 San Francisco, CAPereira, F. D. Q., Esbert, V. E. B., Garcia, J. P., Ana Vidal Pantaleoni, Melcon, A. A., Tornero, J. L. G., & Gimeno, B. (2007). Efficient Analysis of Arbitrarily Shaped Inductive Obstacles in Rectangular Waveguides Using a Surface Integral-Equation Formulation. IEEE Transactions on Microwave Theory and Techniques, 55(4), 715-721. doi:10.1109/tmtt.2007.893673Quesada Pereira, F. D., Vera Castejón, P., Álvarez Melcón, A., Gimeno, B., & Boria Esbert, V. E. (2011). An efficient integral equation technique for the analysis of arbitrarily shaped capacitive waveguide circuits. Radio Science, 46(2), n/a-n/a. doi:10.1029/2010rs004458Stumpf, M., & Leone, M. (2009). Efficient 2-D Integral Equation Approach for the Analysis of Power Bus Structures With Arbitrary Shape. IEEE Transactions on Electromagnetic Compatibility, 51(1), 38-45. doi:10.1109/temc.2008.2009223Wei, X.-C., Li, E.-P., Liu, E.-X., & Cui, X. (2008). Efficient Modeling of Rerouted Return Currents in Multilayered Power-Ground Planes by Using Integral Equation. IEEE Transactions on Electromagnetic Compatibility, 50(3), 740-743. doi:10.1109/temc.2008.924392Huapeng Zhao, En-Xiao Liu, Jun Hu, & Er-Ping Li. (2014). Fast Contour Integral Equation Method for Wideband Power Integrity Analysis. IEEE Transactions on Components, Packaging and Manufacturing Technology, 4(8), 1317-1324. doi:10.1109/tcpmt.2014.232724

Genome sequence analysis and insecticidal characterization of Bacillus thuringiensis Bt-UNVM_94, a strain showing dual insecticidal activity against lepidopteran and coleopteran pests

Bacillus thuringiensis is a gram-positive and spore-forming bacterium that synthesizes a wide diversity of proteins with insecticidal activity and which has demonstrated its potential and safety as a biocontrol agent for more than four decades. However, several susceptible insect species have been reported for evolving resistance, which demands screening for strains exhibiting novel insecticidal properties. In this work, we performed the genome sequence analysis and the insecticidal characterization of B. thuringiensis strain Bt-UNVM_94 isolated from Argentina. This strain produced quasi symmetric bipyramidal parasporal crystals as shown using Scanning Electron Microscopy. Its genomic sequence harbours one coding sequence showing homology to the crystal toxin Cry7Ga2 and another, with similarity to the Mpp2Aa3 (Mtx2) toxin. Known Cry7A and Cry7B are known to be active against some coleopteran and lepidopteran larvae, respectively. However, bioassays performed with spore-crystal mixtures of strain Bt-UNVM_94 exhibited dual toxicity with 50% and 91% mortality against Cydia pomonella (Lepidoptera: Tortricidae) and Anthonomus grandis (Coleoptera: Curculionidae), respectively. No toxicity was detected against the free-living nematode Panagrellus redivivus (Rhabditidae: Panagrolaimidae). This strain also showed no PCR amplification of the type I b-exotoxin thuE gene, consistent with the absence of mortality in b-exotoxin bioassays with Musca domestica (Diptera: Muscidae). Screenings of novel B. thuringiensis strains may provide toxins with novel insecticidal properties that can be used to suppress insect resistance to the most used B. thuringiensis-crops in the field

Origin and Epidemiological History of HIV-1 CRF14_BG

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Users must also make clear the license terms under which the work was published. CC BY Licence: http://creativecommons.org/licenses/by/4.0/Background: CRF14_BG isolates, originally found in Spain, are characterized by CXCR4 tropism and rapid disease progression. This study aimed to identify the origin of CRF14_BG and reconstruct its epidemiological history based on new isolates from Portugal.Methodology/Principal Findings: C2V3C3 env gene sequences were obtained from 62 samples collected in 1993–1998 from Portuguese HIV-1 patients. Full-length genomic sequences were obtained from three patients. Viral subtypes, diversity, divergence rate and positive selection were investigated by phylogenetic analysis. The molecular structure of the genomes was determined by bootscanning. A relaxed molecular clock model was used to date the origin of CRF14_BG. Geno2pheno was used to predict viral tropism. Subtype B was the most prevalent subtype (45 sequences; 73%) followed by CRF14_BG (8; 13%), G (4; 6%), F1 (2; 3%), C (2; 3%) and CRF02_AG (1; 2%). Three CRF14_BG sequences were derived from 1993 samples. Near full-length genomic sequences were strongly related to the CRF14_BG isolates from Spain. Genetic diversity of the Portuguese isolates was significantly higher than the Spanish isolates (0.044 vs 0.014, P,0.0001). The mean date of origin of the CRF14_BG cluster was estimated to be 1992 (range, 1989 and 1996) based on the subtype G genomic region and 1989 (range, 1984–1993) based on the subtype B genomic region. Most CRF14_BG strains (78.9%) were predicted to be CXCR4. Finally, up to five amino acids were under selective pressure in subtype B V3 loop whereas only one was found in the CRF14_BG cluster.Conclusions: CRF14_BG emerged in Portugal in the early 1990 s soon after the beginning of the HIV-1 epidemics, spread to Spain in late 1990 s as a consequence of IVDUs migration and then to the rest of Europe. CXCR4 tropism is a general characteristic of this CRF that may have been selected for by escape from neutralizing antibody response

Gene family information facilitates variant interpretation and identification of disease-associated genes in neurodevelopmental disorders

Background Classifying pathogenicity of missense variants represents a major challenge in clinical practice during the diagnoses of rare and genetic heterogeneous neurodevelopmental disorders (NDDs). While orthologous gene conservation is commonly employed in variant annotation, approximately 80% of known disease-associated genes belong to gene families. The use of gene family information for disease gene discovery and variant interpretation has not yet been investigated on a genome-wide scale. We empirically evaluate whether paralog-conserved or non-conserved sites in human gene families are important in NDDs. Methods Gene family information was collected from Ensembl. Paralog-conserved sites were defined based on paralog sequence alignments; 10,068 NDD patients and 2078 controls were statistically evaluated for de novo variant burden in gene families. Results We demonstrate that disease-associated missense variants are enriched at paralog-conserved sites across all disease groups and inheritance models tested. We developed a gene family de novo enrichment framework that identified 43 exome-wide enriched gene families including 98 de novo variant carrying genes in NDD patients of which 28 represent novel candidate genes for NDD which are brain expressed and under evolutionary constraint. Conclusion This study represents the first method to incorporate gene family information into a statistical framework to interpret variant data for NDDs and to discover new NDD-associated genes