Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies

Autoři: Anya T. Joynt aff001;  Taylor A. Evans aff001;  Matthew J. Pellicore aff001;  Emily F. Davis-Marcisak aff001;  Melis A. Aksit aff001;  Alice C. Eastman aff001;  Shivani U. Patel aff002;  Kathleen C. Paul aff001;  Derek L. Osorio aff001;  Alyssa D. Bowling aff001;  Calvin U. Cotton aff003;  Karen S. Raraigh aff001;  Natalie E. West aff002;  Christian A. Merlo aff002;  Garry R. Cutting aff001;  Neeraj Sharma aff001
Působiště autorů: McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America aff001;  Division of Pulmonary and Critical Care Medicine, Department of Medicine, Johns Hopkins Hospital, Baltimore, Maryland, United States of America aff002;  Departments of Pediatrics, Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio, United States of America aff003
Vyšlo v časopise: Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies. PLoS Genet 16(10): e1009100. doi:10.1371/journal.pgen.1009100
Kategorie: Research Article
doi: 10.1371/journal.pgen.1009100


Elucidating the functional consequence of molecular defects underlying genetic diseases enables appropriate design of therapeutic options. Treatment of cystic fibrosis (CF) is an exemplar of this paradigm as the development of CFTR modulator therapies has allowed for targeted and effective treatment of individuals harboring specific genetic variants. However, the mechanism of these drugs limits effectiveness to particular classes of variants that allow production of CFTR protein. Thus, assessment of the molecular mechanism of individual variants is imperative for proper assignment of these precision therapies. This is particularly important when considering variants that affect pre-mRNA splicing, thus limiting success of the existing protein-targeted therapies. Variants affecting splicing can occur throughout exons and introns and the complexity of the process of splicing lends itself to a variety of outcomes, both at the RNA and protein levels, further complicating assessment of disease liability and modulator response. To investigate the scope of this challenge, we evaluated splicing and downstream effects of 52 naturally occurring CFTR variants (exonic = 15, intronic = 37). Expression of constructs containing select CFTR intronic sequences and complete CFTR exonic sequences in cell line models allowed for assessment of RNA and protein-level effects on an allele by allele basis. Characterization of primary nasal epithelial cells obtained from individuals harboring splice variants corroborated in vitro data. Notably, we identified exonic variants that result in complete missplicing and thus a lack of modulator response (e.g. c.2908G>A, c.523A>G), as well as intronic variants that respond to modulators due to the presence of residual normally spliced transcript (e.g. c.4242+2T>C, c.3717+40A>G). Overall, our data reveals diverse molecular outcomes amongst both exonic and intronic variants emphasizing the need to delineate RNA, protein, and functional effects of each variant in order to accurately assign precision therapies.

Klíčová slova:

Amino acid substitution – Electromyography – Chlorides – Introns – Messenger RNA – Nucleotides – Reverse transcriptase-polymerase chain reaction – RNA splicing


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