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Multiplex functional assay of variant effect in the retinal transcription factor CRX

视网膜转录因子 CRX 变异效应的多重功能测定

基本信息

批准号：
10555204
负责人：
James Lewis Shepherdson
金额：
$ 3.36万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-01 至 2026-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10555204
关键词：
Address Age of Onset Benign Biological Assay Blindness Catalogs Cells Classification ClinVar Clinical Clinical Management Clinical Research Databases Decision Making Defect Diagnosis Disease Genes Genetic Genetic Transcription Genetic Variation Genotype Homeobox Human Human Genetics Inherited Leber&apos s amaurosis Libraries Location Measures Molecular Mutation Pathogenesis Pathogenicity Patients Phenotype Photoreceptors Proteins Reporter Reporter Genes Reporting Retina Retinal Diseases Retinitis Pigmentosa Severities System Techniques Technology Testing Time Translating United States National Institutes of Health Variant Vertebrate Photoreceptors Vision Work clinical sequencing cone-rod dystrophy disease-causing mutation experimental study gene therapy genetic variant human disease interest loss of function multiplex assay mutation screening patient variability recessive genetic trait transcription factor variant of unknown significance

项目摘要

PROJECT SUMMARY The transcription factor Cone-Rod Homeobox (CRX ) is a master regulator of photoreceptor cell fate. Sequence variants in CRX can cause Retinitis Pigmentosa, Cone-Rod Dystrophy, and Leber Congenital Amaurosis, all inherited causes of vision loss and blindness. CRX is the only gene implicated in the pathogenesis of all three of these diseases, which present with both rod- and cone-centric phenotypes of varying age of onset and severity. Several CRX variants have been reported to cause severe dominant disease through antimorphic genetic interac- tions with wild-type CRX, and yet these mutations are adjacent to variants which are benign or only cause mild, recessive disease. Determining which mutations in CRX are pathogenic and quantifying their effect on functional activity is prerequisite to interpreting patient variation and predicting patient phenotypes. However, most variants in CRX are “Variants of Uncertain Signiﬁcance” (VUS), meaning that insufﬁcient clinical or functional evidence exists to determine their pathogenicity. Without a robust catalog of human genetic variation, advances in patient sequencing cannot be translated into clinical guidance or therapies for patients with uncharacterized variants. One potential solution to this challenge is Deep Mutational Scanning (DMS), which uses massive libraries of variant sequences in multiplexed assays to simultaneously measure the functional consequence of thousands of variants in a gene of interest in a single experiment. In DMS, each gene variant is assigned a quantitative functional pathogenicity score based on its activity in a molecular assay. This proposal will use DMS to simultaneously assay the transcriptional activity and abundance of all point substitutions, truncations, and frameshifts in CRX. The direct product of this work will be a “look-up table” listing the functional consequence of every CRX variant on protein activity and stability, which will be directly applicable to clinical variant classiﬁcation and decision making. Given the retina's privileged location as a facile target for gene therapy, a catalog of human variation in CRX could inform the clinical management of patients afﬂicted with inherited retinopathies in the near term. Furthermore, this work will establish an extensible platform for additional DMS studies of other retinal transcription factors, broadly expanding our understanding of inherited retinal disease.

项目总结