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Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies
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...
Autores principales: | , , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7605713/ https://www.ncbi.nlm.nih.gov/pubmed/33085659 http://dx.doi.org/10.1371/journal.pgen.1009100 |
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author | Joynt, Anya T. Evans, Taylor A. Pellicore, Matthew J. Davis-Marcisak, Emily F. Aksit, Melis A. Eastman, Alice C. Patel, Shivani U. Paul, Kathleen C. Osorio, Derek L. Bowling, Alyssa D. Cotton, Calvin U. Raraigh, Karen S. West, Natalie E. Merlo, Christian A. Cutting, Garry R. Sharma, Neeraj |
author_facet | Joynt, Anya T. Evans, Taylor A. Pellicore, Matthew J. Davis-Marcisak, Emily F. Aksit, Melis A. Eastman, Alice C. Patel, Shivani U. Paul, Kathleen C. Osorio, Derek L. Bowling, Alyssa D. Cotton, Calvin U. Raraigh, Karen S. West, Natalie E. Merlo, Christian A. Cutting, Garry R. Sharma, Neeraj |
author_sort | Joynt, Anya T. |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-7605713 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-76057132020-11-05 Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies Joynt, Anya T. Evans, Taylor A. Pellicore, Matthew J. Davis-Marcisak, Emily F. Aksit, Melis A. Eastman, Alice C. Patel, Shivani U. Paul, Kathleen C. Osorio, Derek L. Bowling, Alyssa D. Cotton, Calvin U. Raraigh, Karen S. West, Natalie E. Merlo, Christian A. Cutting, Garry R. Sharma, Neeraj PLoS Genet Research Article 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. Public Library of Science 2020-10-21 /pmc/articles/PMC7605713/ /pubmed/33085659 http://dx.doi.org/10.1371/journal.pgen.1009100 Text en © 2020 Joynt et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Joynt, Anya T. Evans, Taylor A. Pellicore, Matthew J. Davis-Marcisak, Emily F. Aksit, Melis A. Eastman, Alice C. Patel, Shivani U. Paul, Kathleen C. Osorio, Derek L. Bowling, Alyssa D. Cotton, Calvin U. Raraigh, Karen S. West, Natalie E. Merlo, Christian A. Cutting, Garry R. Sharma, Neeraj Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies |
title | Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies |
title_full | Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies |
title_fullStr | Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies |
title_full_unstemmed | Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies |
title_short | Evaluation of both exonic and intronic variants for effects on RNA splicing allows for accurate assessment of the effectiveness of precision therapies |
title_sort | evaluation of both exonic and intronic variants for effects on rna splicing allows for accurate assessment of the effectiveness of precision therapies |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7605713/ https://www.ncbi.nlm.nih.gov/pubmed/33085659 http://dx.doi.org/10.1371/journal.pgen.1009100 |
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