CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts
Collagen VI-related disorders are the second most common congenital muscular dystrophies for which no treatments are presently available. They are mostly caused by dominant-negative pathogenic variants in the genes encoding α chains of collagen VI, a heteromeric network forming collagen; for example...
| Autores: | , , , , , , , , , , , , |
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| Tipo de recurso: | artículo |
| Estado: | Versión publicada |
| Fecha de publicación: | 2022 |
| País: | España |
| Institución: | Universidad de Barcelona |
| Repositorio: | Dipòsit Digital de la UB |
| OAI Identifier: | oai:diposit.ub.edu:2445/186222 |
| Acceso en línea: | https://hdl.handle.net/2445/186222 |
| Access Level: | acceso abierto |
| Palabra clave: | Distròfia muscular Mutació (Biologia) Muscular dystrophy Mutation (Biology) |
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CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblastsLópez Márquez, ArístidesMorín, MatíasFernández-Peñalver, SergioBadosa, CarmenHernández-Delgado, AlejandroNatera-de Benito, DanielOrtez, CarlosNascimento, AndrésGrinberg Vaisman, Daniel RaúlBalcells Comas, SusanaRoldán Molina, MónicaMoreno-Pelayo, Miguel ÁngelJiménez-Mallebrera, CeciliaDistròfia muscularMutació (Biologia)Muscular dystrophyMutation (Biology)Collagen VI-related disorders are the second most common congenital muscular dystrophies for which no treatments are presently available. They are mostly caused by dominant-negative pathogenic variants in the genes encoding α chains of collagen VI, a heteromeric network forming collagen; for example, the c.877G>A; p.Gly293Arg COL6A1 variant, which alters the proper association of the tetramers to form microfibrils. We tested the potential of CRISPR/Cas9-based genome editing to silence or correct (using a donor template) a mutant allele in the dermal fibroblasts of four individuals bearing the c.877G>A pathogenic variant. Evaluation of gene-edited cells by next-generation sequencing revealed that correction of the mutant allele by homologous-directed repair occurred at a frequency lower than 1%. However, the presence of frameshift variants and others that provoked the silencing of the mutant allele were found in >40% of reads, with no effects on the wild-type allele. This was confirmed by droplet digital PCR with allele-specific probes, which revealed a reduction in the expression of the mutant allele. Finally, immunofluorescence analyses revealed a recovery in the collagen VI extracellular matrix. In summary, we demonstrate that CRISPR/Cas9 gene-edition can specifically reverse the pathogenic effects of a dominant negative variant in COL6A1.MDPI2022info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionapplication/pdfhttps://hdl.handle.net/2445/186222Articles publicats en revistes (Genètica, Microbiologia i Estadística)reponame:Dipòsit Digital de la UBinstname:Universidad de BarcelonaInglésReproducció del document publicat a: https://doi.org/10.3390/ijms23084410International Journal of Molecular Sciences, 2022, vol. 23, num. 8, p. 4410https://doi.org/10.3390/ijms23084410cc-by (c) López-Márquez, Arístides et al., 2022https://creativecommons.org/licenses/by/4.0/info:eu-repo/semantics/openAccessoai:diposit.ub.edu:2445/1862222026-05-27T06:46:51Z |
| dc.title.none.fl_str_mv |
CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts |
| title |
CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts |
| spellingShingle |
CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts López Márquez, Arístides Distròfia muscular Mutació (Biologia) Muscular dystrophy Mutation (Biology) |
| title_short |
CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts |
| title_full |
CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts |
| title_fullStr |
CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts |
| title_full_unstemmed |
CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts |
| title_sort |
CRISPR/Cas9-mediated allele-specific disruption of a dominant COL6A1 pathogenic variant improves collagen VI network in patient fibroblasts |
| dc.creator.none.fl_str_mv |
López Márquez, Arístides Morín, Matías Fernández-Peñalver, Sergio Badosa, Carmen Hernández-Delgado, Alejandro Natera-de Benito, Daniel Ortez, Carlos Nascimento, Andrés Grinberg Vaisman, Daniel Raúl Balcells Comas, Susana Roldán Molina, Mónica Moreno-Pelayo, Miguel Ángel Jiménez-Mallebrera, Cecilia |
| author |
López Márquez, Arístides |
| author_facet |
López Márquez, Arístides Morín, Matías Fernández-Peñalver, Sergio Badosa, Carmen Hernández-Delgado, Alejandro Natera-de Benito, Daniel Ortez, Carlos Nascimento, Andrés Grinberg Vaisman, Daniel Raúl Balcells Comas, Susana Roldán Molina, Mónica Moreno-Pelayo, Miguel Ángel Jiménez-Mallebrera, Cecilia |
| author_role |
author |
| author2 |
Morín, Matías Fernández-Peñalver, Sergio Badosa, Carmen Hernández-Delgado, Alejandro Natera-de Benito, Daniel Ortez, Carlos Nascimento, Andrés Grinberg Vaisman, Daniel Raúl Balcells Comas, Susana Roldán Molina, Mónica Moreno-Pelayo, Miguel Ángel Jiménez-Mallebrera, Cecilia |
| author2_role |
author author author author author author author author author author author author |
| dc.subject.none.fl_str_mv |
Distròfia muscular Mutació (Biologia) Muscular dystrophy Mutation (Biology) |
| topic |
Distròfia muscular Mutació (Biologia) Muscular dystrophy Mutation (Biology) |
| description |
Collagen VI-related disorders are the second most common congenital muscular dystrophies for which no treatments are presently available. They are mostly caused by dominant-negative pathogenic variants in the genes encoding α chains of collagen VI, a heteromeric network forming collagen; for example, the c.877G>A; p.Gly293Arg COL6A1 variant, which alters the proper association of the tetramers to form microfibrils. We tested the potential of CRISPR/Cas9-based genome editing to silence or correct (using a donor template) a mutant allele in the dermal fibroblasts of four individuals bearing the c.877G>A pathogenic variant. Evaluation of gene-edited cells by next-generation sequencing revealed that correction of the mutant allele by homologous-directed repair occurred at a frequency lower than 1%. However, the presence of frameshift variants and others that provoked the silencing of the mutant allele were found in >40% of reads, with no effects on the wild-type allele. This was confirmed by droplet digital PCR with allele-specific probes, which revealed a reduction in the expression of the mutant allele. Finally, immunofluorescence analyses revealed a recovery in the collagen VI extracellular matrix. In summary, we demonstrate that CRISPR/Cas9 gene-edition can specifically reverse the pathogenic effects of a dominant negative variant in COL6A1. |
| publishDate |
2022 |
| dc.date.none.fl_str_mv |
2022 |
| dc.type.none.fl_str_mv |
info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion |
| format |
article |
| status_str |
publishedVersion |
| dc.identifier.none.fl_str_mv |
https://hdl.handle.net/2445/186222 |
| url |
https://hdl.handle.net/2445/186222 |
| dc.language.none.fl_str_mv |
Inglés |
| language_invalid_str_mv |
Inglés |
| dc.relation.none.fl_str_mv |
Reproducció del document publicat a: https://doi.org/10.3390/ijms23084410 International Journal of Molecular Sciences, 2022, vol. 23, num. 8, p. 4410 https://doi.org/10.3390/ijms23084410 |
| dc.rights.none.fl_str_mv |
cc-by (c) López-Márquez, Arístides et al., 2022 https://creativecommons.org/licenses/by/4.0/ info:eu-repo/semantics/openAccess |
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cc-by (c) López-Márquez, Arístides et al., 2022 https://creativecommons.org/licenses/by/4.0/ |
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openAccess |
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application/pdf |
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MDPI |
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MDPI |
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Articles publicats en revistes (Genètica, Microbiologia i Estadística) reponame:Dipòsit Digital de la UB instname:Universidad de Barcelona |
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Universidad de Barcelona |
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Dipòsit Digital de la UB |
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Dipòsit Digital de la UB |
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