Impact of functional studies on exome sequence variant interpretation in early-onset cardiac conduction system diseases

Aims The genetic cause of cardiac conduction system disease (CCSD) has not been fully elucidated. Whole-exome sequencing (WES) can detect various genetic variants; however, the identification of pathogenic variants remains a challenge. We aimed to identify pathogenic or likely pathogenic variants in CCSD patients by using WES and 2015 American College of Medical Genetics and Genomics (ACMG) standards and guidelines as well as evaluating the usefulness of functional studies for determining them. Methods and Results We performed WES of 23 probands diagnosed with early-onset (<65 years) CCSD and analyzed 117 genes linked to arrhythmogenic diseases or cardiomyopathies. We focused on rare variants (minor allele frequency < 0.1%) that were absent from population databases. Five probands had protein truncating variants in EMD and LMNA which were classified as “pathogenic” by 2015 ACMG standards and guidelines. To evaluate the functional changes brought about by these variants, we generated a knock-out zebrafish with CRISPR-mediated insertions or deletions of the EMD or LMNA homologs in zebrafish. The mean heart rate and conduction velocities in the CRISPR/Cas9-injected embryos and F2 generation embryos with homozygous deletions were significantly decreased. Twenty-one variants of uncertain significance were identified in 11 probands. Cellular electrophysiological study and in vivo zebrafish cardiac assay showed that 2 variants in KCNH2 and SCN5A, 4 variants in SCN10A, and 1 variant in MYH6 damaged each gene, which resulted in the change of the clinical significance of them from “Uncertain significance” to “Likely pathogenic” in 6 probands. Conclusions Of 23 CCSD probands, we successfully identified pathogenic or likely pathogenic variants in 11 probands (48%). Functional analyses of a cellular electrophysiological study and in vivo zebrafish cardiac assay might be useful for determining the pathogenicity of rare variants in patients with CCSD. SCN10A may be one of the major genes responsible for CCSD. Translational Perspective Whole-exome sequencing (WES) may be helpful in determining the causes of cardiac conduction system disease (CCSD), however, the identification of pathogenic variants remains a challenge. We performed WES of 23 probands diagnosed with early-onset CCSD, and identified 12 pathogenic or likely pathogenic variants in 11 of these probands (48%) according to the 2015 ACMG standards and guidelines. In this context, functional analyses of a cellular electrophysiological study and in vivo zebrafish cardiac assay might be useful for determining the pathogenicity of rare variants, and SCN10A may be one of the major development factors in CCSD

Results of the search for inspiraling compact star binaries from TAMA300's observation in 2000-2004

We analyze the data of TAMA300 detector to search for gravitational waves from inspiraling compact star binaries with masses of the component stars in the range 1-3Msolar. In this analysis, 2705 hours of data, taken during the years 2000-2004, are used for the event search. We combine the results of different observation runs, and obtained a single upper limit on the rate of the coalescence of compact binaries in our Galaxy of 20 per year at a 90% confidence level. In this upper limit, the effect of various systematic errors such like the uncertainty of the background estimation and the calibration of the detector's sensitivity are included.Comment: 8 pages, 4 Postscript figures, uses revtex4.sty The author list was correcte

Observation results by the TAMA300 detector on gravitational wave bursts from stellar-core collapses

We present data-analysis schemes and results of observations with the TAMA300 gravitational-wave detector, targeting burst signals from stellar-core collapse events. In analyses for burst gravitational waves, the detection and fake-reduction schemes are different from well-investigated ones for a chirp-wave analysis, because precise waveform templates are not available. We used an excess-power filter for the extraction of gravitational-wave candidates, and developed two methods for the reduction of fake events caused by non-stationary noises of the detector. These analysis schemes were applied to real data from the TAMA300 interferometric gravitational wave detector. As a result, fake events were reduced by a factor of about 1000 in the best cases. The resultant event candidates were interpreted from an astronomical viewpoint. We set an upper limit of 2.2x10^3 events/sec on the burst gravitational-wave event rate in our Galaxy with a confidence level of 90%. This work sets a milestone and prospects on the search for burst gravitational waves, by establishing an analysis scheme for the observation data from an interferometric gravitational wave detector

Disruption of the CAR1 Gene Encoding Arginase Enhances Freeze Tolerance of the Commercial Baker's Yeast Saccharomyces cerevisiae

The effect of intracellular charged amino acids on freeze tolerance in doughs was determined by constructing homozygous diploid arginase-deficient mutants of commercial baker's yeast. An arginase mutant accumulated higher levels of arginine and/or glutamate and showed increased leavening ability during the frozen-dough baking process, suggesting that disruption of the CAR1 gene enhances freeze tolerance