Dataset Information

Prospective functional classification of all possible missense variants in PPARG.

ABSTRACT: Clinical exome sequencing routinely identifies missense variants in disease-related genes, but functional characterization is rarely undertaken, leading to diagnostic uncertainty. For example, mutations in PPARG cause Mendelian lipodystrophy and increase risk of type 2 diabetes (T2D). Although approximately 1 in 500 people harbor missense variants in PPARG, most are of unknown consequence. To prospectively characterize PPAR? variants, we used highly parallel oligonucleotide synthesis to construct a library encoding all 9,595 possible single-amino acid substitutions. We developed a pooled functional assay in human macrophages, experimentally evaluated all protein variants, and used the experimental data to train a variant classifier by supervised machine learning. When applied to 55 new missense variants identified in population-based and clinical sequencing, the classifier annotated 6 variants as pathogenic; these were subsequently validated by single-variant assays. Saturation mutagenesis and prospective experimental characterization can support immediate diagnostic interpretation of newly discovered missense variants in disease-related genes.

SUBMITTER: Majithia AR

PROVIDER: S-EPMC5131844 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

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Prospective functional classification of all possible missense variants in PPARG.

Majithia Amit R AR Tsuda Ben B Agostini Maura M Gnanapradeepan Keerthana K Rice Robert R Peloso Gina G Patel Kashyap A KA Zhang Xiaolan X Broekema Marjoleine F MF Patterson Nick N Duby Marc M Sharpe Ted T Kalkhoven Eric E Rosen Evan D ED Barroso Inês I Ellard Sian S Kathiresan Sekar S O'Rahilly Stephen S Chatterjee Krishna K Florez Jose C JC Mikkelsen Tarjei T Savage David B DB Altshuler David D

Nature genetics 20161017 12

Clinical exome sequencing routinely identifies missense variants in disease-related genes, but functional characterization is rarely undertaken, leading to diagnostic uncertainty. For example, mutations in PPARG cause Mendelian lipodystrophy and increase risk of type 2 diabetes (T2D). Although approximately 1 in 500 people harbor missense variants in PPARG, most are of unknown consequence. To prospectively characterize PPARγ variants, we used highly parallel oligonucleotide synthesis to construc ...[more]

PMID: 27749844

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Project description:BackgroundThe human TET2 gene plays a pivotal role in the epigenetic regulation of normal and malignant hematopoiesis. Somatic TET2 mutations have been repeatedly identified in age-related clonal hematopoiesis and in myeloid neoplasms ranging from acute myeloid leukemia (AML) to myeloproliferative neoplasms. However, there have been no attempts to systematically explore the structural and functional consequences of the hundreds of TET2 missense variants reported to date.MethodsWe have sequenced the TET2 gene in 189 Spanish AML patients using Sanger sequencing and NGS protocols. Next, we performed a thorough bioinformatics analysis of TET2 protein and of the expected impact of all reported TET2 missense variants on protein structure and function, exploiting available structure-and-function information as well as 3D structure prediction tools.ResultsWe have identified 38 TET2 allelic variants in the studied patients, including two frequent SNPs: p.G355D (10 cases) and p.I1762V (28 cases). Four of the detected mutations are reported here for the first time: c.122C>T (p.P41L), c.4535C>G (p.A1512G), c.4760A>G (p.D1587G), and c.5087A>T (p.Y1696F). We predict a complex multidomain architecture for the noncatalytic regions of TET2, and in particular the presence of well-conserved α+β globular domains immediately preceding and following the actual catalytic unit. Further, we provide a rigorous interpretation of over 430 missense SNVs that affect the TET2 catalytic domain, and we hypothesize explanations for ~700 additional variants found within the regulatory regions of the protein. Finally, we propose a systematic classification of all missense mutants and SNPs reported to date into three major categories (severe, moderate, and mild), based on their predicted structural and functional impact.ConclusionsThe proposed classification of missense TET2 variants would help to assess their clinical impact on human neoplasia and may guide future structure-and-function investigations of TET family members.

Dataset Information

Prospective functional classification of all possible missense variants in PPARG.

Publications

Prospective functional classification of all possible missense variants in PPARG.

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