Abstract
Although most disease-causing variants are within coding region of genes, it is now well established that cis-acting regulatory sequences, depending on 3D-chromatin organization, are required for temporal and spatial control of gene expression. Disruptions of such regulatory elements and/or chromatin conformation are likely to play a critical role in human genetic disease. Hence, recurrent monoallelic cases, who present the most common hereditary type of nonsyndromic hearing loss (i.e., DFNB1), carry only one identified pathogenic allele. This strongly suggests the presence of uncharacterized distal cis-acting elements in the missing allele. Here within, we study the spatial organization of a large DFNB1 locus encompassing the gap junction protein beta 2 (GJB2) gene, the most frequently mutated gene in this inherited hearing loss phenotype, with the chromosome conformation capture carbon copy technology (5C). By combining this approach with functional activity reporter assays and mapping of CCCTC-binding factor (CTCF) along the DFNB1 locus, we identify a novel set of cooperating GJB2 cis-acting elements and suggest a DFNB1 three-dimensional looping regulation model.
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Data availability statement
All data generated during this study are included in this published article and the supplementary files. The 5C datasets generated during the current study are available in the GEO databases: 5C-Seq data: Gene Expression Omnibus GSE128881 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE128881).
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Acknowledgements
This work was supported by grants from the French foundation “La Fondation pour l’Audition”, the “Région Bretagne” and the association “Gaétan Salaün”. We thank Pr Laurent Misery and its team for keratinocytes. We are grateful to the members of Josée Dostie Lab for access to their Torrent 5C data transformation pipeline on Galaxy and for their excellent technical assistance. We thank Anthony Herzig for grammatical/editorial English corrections of this manuscript.
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SM: Designed and supervised the study, carried out the experiments, analyzed the data, wrote the paper. ALN: Carried out luciferase assays and ChIP experiments, analyzed the data. AQ: Carried out 5C experiments, analyzed the data. CLM: Edited the paper. CF: Supervised lab work, edited the paper. All authors read and approved the manuscript.
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Moisan, S., Le Nabec, A., Quillévéré, A. et al. Characterization of GJB2 cis-regulatory elements in the DFNB1 locus. Hum Genet 138, 1275–1286 (2019). https://doi.org/10.1007/s00439-019-02068-8
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DOI: https://doi.org/10.1007/s00439-019-02068-8