Gene panel and exome sequencing have revealed a high rate of molecular diagnoses among diseases where the genetic architecture has proven suitable for sequencing approaches, with a large number of distinct and highly penetrant causal variants identified among a growing list of disease genes. The challenge is, given the DNA sequence of a new patient, to distinguish disease-causing from benign variants. Large samples of human standing variation data highlight regional variation in the tolerance to missense variation within the protein-coding sequence of genes. This information is not well captured by existing bioinformatic tools, but is effective in improving variant interpretation. To address this limitation in existing tools, we introduce the missense tolerance ratio (MTR), which summarizes available human standing variation data within genes to encapsulate population level genetic variation. We find that patient-ascertained pathogenic variants preferentially cluster in low MTR regions (P < 0.005) of well-informed genes. By evaluating 20 publicly available predictive tools across genes linked to epilepsy, we also highlight the importance of understanding the empirical null distribution of existing prediction tools, as these vary across genes. Subsequently integrating the MTR with the empirically selected bioinformatic tools in a gene-specific approach demonstrates a clear improvement in the ability to predict pathogenic missense variants from background missense variation in disease genes. Among an independent test sample of case and control missense variants, case variants (0.83 median score) consistently achieve higher pathogenicity prediction probabilities than control variants (0.02 median score; Mann-Whitney U test, P < 1 × 10
MLKL is the essential effector of necroptosis, a form of programmed lytic cell death. We have isolated a mouse strain with a single missense mutation, MlklD139V, that alters the two-helix ‘brace’ that connects the killer four-helix bundle and regulatory pseudokinase domains. This confers constitutive, RIPK3 independent killing activity to MLKL. Homozygous mutant mice develop lethal postnatal inflammation of the salivary glands and mediastinum. The normal embryonic development of MlklD139V homozygotes until birth, and the absence of any overt phenotype in heterozygotes provides important in vivo precedent for the capacity of cells to clear activated MLKL. These observations offer an important insight into the potential disease-modulating roles of three common human MLKL polymorphisms that encode amino acid substitutions within or adjacent to the brace region. Compound heterozygosity of these variants is found at up to 12-fold the expected frequency in patients that suffer from a pediatric autoinflammatory disease, chronic recurrent multifocal osteomyelitis (CRMO).
Advances in genomic sequencing have enormous potential to revolutionize personalized medicine, however distinguishing disease-causing from benign variants remains a challenge. The increasing number of human genome and exome sequences available has revealed areas where unfavourable variation is removed through purifying selection. Here, we present the MTR-Viewer, a web-server enabling easy visualization at the gene or variant level of the Missense Tolerance Ratio (MTR), a measure of regional intolerance to missense variation calculated using variation from 240 000 exome and genome sequences. The MTR-Viewer enables exploration of MTR calculations, using different sliding windows, for over 18 000 human protein-coding genes and 85 000 alternative transcripts. Users can also view MTR scores calculated for specific ethnicities, to enable easy exploration of regions that may be under different selective pressure. The spatial distribution of population and known disease variants is also displayed on the protein's domain structure. Intolerant regions were found to be highly enriched for ClinVar pathogenic and COSMIC somatic missense variants (Mann–Whitney U test P < 2.2 × 10−16). As the MTR is not biased by known domains and protein features, it can highlight functionally important regions within genes overlooked or inaccessible by traditional methods. MTR-Viewer is freely available via a user friendly web-server at http://biosig.unimelb.edu.au/mtr-viewer/.
Summary
EasyVS is a web-based platform built to simplify molecule library selection and virtual screening. With an intuitive interface, the tool allows users to go from selecting a protein target with a known structure and tailoring a purchasable molecule library to performing and visualizing docking in a few clicks. Our system also allows users to filter screening libraries based on molecule properties, cluster molecules by similarity and personalize docking parameters.
Availability and implementation
EasyVS is freely available as an easy-to-use web interface at http://biosig.unimelb.edu.au/easyvs.
Supplementary information
Supplementary data are available at Bioinformatics online.
The identification of disease-causal variants is non-trivial. By mapping population variation from over 448,000 exome and genome sequences to over 81,000 experimental structures and homology models of the human proteome, we have calculated both regional intolerance to missense variation (Missense Tolerance Ratio, MTR), using a sliding window of 21–41 codons, and introduce a new 3D spatial intolerance to missense variation score (3D Missense Tolerance Ratio, MTR3D), using spheres of 5–8 Å. We show that the MTR3D is less biased by regions with limited data and more accurately identifies regions under purifying selection than estimates relying on the sequence alone. Intolerant regions were highly enriched for both ClinVar pathogenic and COSMIC somatic missense variants (Mann–Whitney U test P < 2.2 × 10−16). Further, we combine sequence- and spatial-based scores to generate a consensus score, MTRX, which distinguishes pathogenic from benign variants more accurately than either score separately (AUC = 0.85). The MTR3D server enables easy visualisation of population variation, MTR, MTR3D and MTRX scores across the entire gene and protein structure for >17,000 human genes and >42,000 alternative alternate transcripts, including both Ensembl and RefSeq transcripts. MTR3D is freely available by user-friendly web-interface and API at http://biosig.unimelb.edu.au/mtr3d/.
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