Deep indel mutagenesis reveals the regulatory and modulatory architecture of alternative exon splicing
While altered pre-mRNA splicing is a frequent mechanism by which genetic variants cause disease, the regulatory architecture of human exons remains poorly understood. Antisense oligonucleotides (AONs) that target pre-mRNA splicing have been approved as therapeutics for various pathologies including patient-customised treatments for rare diseases, but AON discovery is currently slow and expensive, limiting the wider adoption of the approach. Here we show that deep indel mutagenesis (DIM) -which can be made experimentally at very low cost - provides an efficient strategy to chart the regulatory landscape of human exons and rapidly identify candidate splicing-modulating oligonucleotides. DIM reveals autonomous effects of insertions, while systematic deletion scans delineate the checkerboard architecture of sequential enhancers and silencers in a model alternative exon. The results also suggest a mechanism for repression of transmembrane domain-encoding exons and for the generation of microexons. Leveraging deep learning tools, we provide a resource, DANGO, that predicts the splicing regulatory landscape of all human exons and can help to identify effective splicing-modulating antisense oligonucleotides.
Authors
- Juan Valcárcel (ORCID: https://orcid.org/0000-0001-5398-3571)
- Michael Thompson (ORCID: https://orcid.org/0000-0003-1546-0512)
- Pablo Baeza-Centurión (ORCID: https://orcid.org/0000-0002-8141-8075)
- André J. Faure (ORCID: https://orcid.org/0000-0002-4471-5994)
- Joseph Clarke (ORCID: https://orcid.org/0009-0002-2466-9932)
- Sophie Bonnal (ORCID: https://orcid.org/0000-0001-6096-3042)
- Belén Miñana (ORCID: https://orcid.org/0000-0001-7676-5788)
- Gioia Quarantani (ORCID: https://orcid.org/0009-0008-2794-2678)
Institutions
- Institució Catalana de Recerca i Estudis Avançats (ES)
- Universitat Pompeu Fabra (ES)
- German Cancer Research Center (DE)
- Heidelberg University (DE)
- University of Cambridge (GB)
- Wellcome Sanger Institute (GB)
- Centre for Genomic Regulation (ES)
Publication Details
- Journal
- Nature Communications
- Published
- 2025-08-30
- DOI
- https://doi.org/10.1038/s41467-025-62957-7
- Citations
- 1
- Primary Topic
- RNA Research and Splicing
- Type
- article
- Field-Weighted Citation Impact
- 0.81
Funders
- Centres de Recerca de Catalunya
- European Commission
- AXA Research Fund
- Generalitat de Catalunya
- Ministerio de Ciencia e Innovación
- Fondation Bettencourt Schueller
- National Institutes of Health
- Directorate for Biological Sciences
- HORIZON EUROPE Framework Programme
- Biotechnology and Biological Sciences Research Council
- Agència de Gestió d'Ajuts Universitaris i de Recerca
- Agencia Estatal de Investigación
- National Heart, Lung, and Blood Institute
- National Institute of Mental Health
- National Institute on Drug Abuse
- National Human Genome Research Institute
- NIH Office of the Director
- National Cancer Institute
- National Institute of Neurological Disorders and Stroke
- Common Fund