{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,27]],"date-time":"2026-05-27T11:36:01Z","timestamp":1779881761106,"version":"3.53.1"},"reference-count":17,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2025,5,30]],"date-time":"2025-05-30T00:00:00Z","timestamp":1748563200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Fundaci\u00f3n Mar\u00eda Cristina Masaveu Peterson"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Genes"],"abstract":"<jats:p>Background\/Objectives: Pathogenic recessive GJB2 variants are the main genetic cause of non-syndromic sensorineural hearing loss. However, following GJB2 testing, a significant proportion of deaf patients are only found to be heterozygous carriers of pathogenic GJB2 alleles. Five large deletions not affecting GJB2 but encompassing a minimal common 62 kb region within the neighbouring CRYL1 gene have been described to cause loss of cis GJB2 expression and, as a result, produce hearing loss when in trans with pathogenic GJB2 variants. We describe the identification and characterization of a novel deletion of this type in deaf patients from northwestern Spain. Methods: We used panel NGS sequencing to detect the deletion, MLPA to validate it, whole-genome sequencing to map its breakpoints, PCR + Sanger sequencing to finely characterize it and triple-primer PCR to screen for it. Results: We identified a novel 200 kb deletion spanning the whole CRYL1 gene in two unrelated deaf patients from Asturias (in northwestern Spain) who were heterozygous for the pathogenic GJB2 c.35delG variant. Although the large deletion was absent from gnomAD v4.1.0 and 2052 local control alleles, screening for it in 20 additional deaf carriers of monoallelic pathogenic GJB2 variants detected it in another patient from Galicia (also in northwestern Spain). The novel deletion, termed del(200 kb)insATTATA, explained hearing loss in 3\/43 (7%) deaf patients from our cohort that were otherwise heterozygous for pathogenic GJB2 variants. Conclusions: This work highlights the importance of comprehensively testing all genomic regions known to be clinically relevant for a given genetic condition, including thorough CRYL1 CNV screening for DFNB1A diagnostics.<\/jats:p>","DOI":"10.3390\/genes16060670","type":"journal-article","created":{"date-parts":[[2025,5,30]],"date-time":"2025-05-30T08:07:58Z","timestamp":1748592478000},"page":"670","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["A Novel Recurrent 200 kb CRYL1 Deletion Underlies DFNB1A Hearing Loss in Patients from Northwestern Spain"],"prefix":"10.3390","volume":"16","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-0042-0051","authenticated-orcid":false,"given":"Guadalupe A.","family":"Cifuentes","sequence":"first","affiliation":[{"name":"Fundaci\u00f3n Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"},{"name":"Instituto de Medicina Oncol\u00f3gica y Molecular de Asturias (IMOMA), 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marta","family":"Di\u00f1eiro","sequence":"additional","affiliation":[{"name":"Instituto de Medicina Oncol\u00f3gica y Molecular de Asturias (IMOMA), 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alicia R.","family":"Huete","sequence":"additional","affiliation":[{"name":"Fundaci\u00f3n Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7812-4861","authenticated-orcid":false,"given":"Raquel","family":"Cap\u00edn","sequence":"additional","affiliation":[{"name":"Instituto de Medicina Oncol\u00f3gica y Molecular de Asturias (IMOMA), 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0008-7869-1637","authenticated-orcid":false,"given":"Adri\u00e1n","family":"Santiago","sequence":"additional","affiliation":[{"name":"Fundaci\u00f3n Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"},{"name":"Instituto de Medicina Oncol\u00f3gica y Molecular de Asturias (IMOMA), 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alberto A. R.","family":"Vargas","sequence":"additional","affiliation":[{"name":"Fundaci\u00f3n Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dido","family":"Carrero","sequence":"additional","affiliation":[{"name":"Fundaci\u00f3n Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"},{"name":"Instituto de Medicina Oncol\u00f3gica y Molecular de Asturias (IMOMA), 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Esther L\u00f3pez","family":"Mart\u00ednez","sequence":"additional","affiliation":[{"name":"Instituto de Medicina Oncol\u00f3gica y Molecular de Asturias (IMOMA), 33193 Oviedo, Spain"},{"name":"Hospital Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Beatriz","family":"Aguiar","sequence":"additional","affiliation":[{"name":"Hospital Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anja","family":"Fischer","sequence":"additional","affiliation":[{"name":"Institute of Molecular Oncology and Functional Genomics, School of Medicine, Technische Universit\u00e4t M\u00fcnchen, 81675 Munich, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Roland","family":"Rad","sequence":"additional","affiliation":[{"name":"Institute of Molecular Oncology and Functional Genomics, School of Medicine, Technische Universit\u00e4t M\u00fcnchen, 81675 Munich, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mar\u00eda","family":"Costales","sequence":"additional","affiliation":[{"name":"Hospital Universitario Central de Asturias, 33011 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rub\u00e9n","family":"Cabanillas","sequence":"additional","affiliation":[{"name":"Instituto de Medicina Oncol\u00f3gica y Molecular de Asturias (IMOMA), 33193 Oviedo, Spain"},{"name":"Cabanillas Precision Consulting, 8620 Z\u00fcrich, Switzerland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7561-7759","authenticated-orcid":false,"given":"Juan","family":"Cadi\u00f1anos","sequence":"additional","affiliation":[{"name":"Fundaci\u00f3n Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"},{"name":"Instituto de Medicina Oncol\u00f3gica y Molecular de Asturias (IMOMA), 33193 Oviedo, Spain"},{"name":"Hospital Centro M\u00e9dico de Asturias, 33193 Oviedo, Spain"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,5,30]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"a033258","DOI":"10.1101\/cshperspect.a033258","article-title":"The Epidemiology of Deafness","volume":"9","author":"Sheffield","year":"2019","journal-title":"Cold Spring Harb Perspect. Med."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Butcher, E., Dezateux, C., Cortina-Borja, M., and Knowles, R.L. (2019). Prevalence of permanent childhood hearing loss detected at the universal newborn hearing screen: Systematic review and meta-analysis. PLoS ONE, 14.","DOI":"10.1371\/journal.pone.0219600"},{"key":"ref_3","unstructured":"Flint, P.W. (2015). Genetic sensorineural hearing loss. Cummings Otolaryngology Head and Neck Surgery, Elsevier."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"741","DOI":"10.1016\/j.ijporl.2007.01.010","article-title":"High prevalences of carriers of the 35delG mutation of connexin 26 in the Mediterranean area","volume":"71","author":"Lucotte","year":"2007","journal-title":"Int. J. Pediatr. Otorhinolaryngol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1325","DOI":"10.1038\/ejhg.2013.83","article-title":"EMQN Best Practice guidelines for diagnostic testing of mutations causing non-syndromic hearing impairment at the DFNB1 locus","volume":"21","author":"Hoefsloot","year":"2013","journal-title":"Eur. J. Hum. Genet."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"243","DOI":"10.1056\/NEJMoa012052","article-title":"A deletion involving the connexin 30 gene in nonsyndromic hearing impairment","volume":"346","author":"Villamar","year":"2002","journal-title":"N. Engl. J. Med."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"588","DOI":"10.1136\/jmg.2004.028324","article-title":"A novel deletion involving the connexin-30 gene, del(GJB6-d13s1854), found in trans with mutations in the GJB2 gene (connexin-26) in subjects with DFNB1 non-syndromic hearing impairment","volume":"42","author":"Alvarez","year":"2005","journal-title":"J. Med. Genet."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"267","DOI":"10.1111\/j.1399-0004.2010.01387.x","article-title":"A novel DFNB1 deletion allele supports the existence of a distant cis-regulatory region that controls GJB2 and GJB6 expression","volume":"78","author":"Wilch","year":"2010","journal-title":"Clin. Genet."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"119","DOI":"10.1002\/humu.22912","article-title":"Targeted Droplet-Digital PCR as a Tool for Novel Deletion Discovery at the DFNB1 Locus","volume":"37","author":"Tayoun","year":"2016","journal-title":"Hum. Mutat."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"9517114","DOI":"10.1155\/2024\/9517114","article-title":"Genome sequencing unveils the role of copy number variants in hearing loss and identifies novel deletions with founder effect in the DFNB1 Locus","volume":"2024","author":"Lin","year":"2024","journal-title":"Hum. Mutat."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"688","DOI":"10.1111\/j.1365-2230.2005.01878.x","article-title":"Specific loss of connexin 26 expression in ductal sweat gland epithelium associated with the deletion mutation del(GJB6-D13S1830)","volume":"30","author":"Common","year":"2005","journal-title":"Clin. Exp. Dermatol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"354","DOI":"10.1016\/j.bbrc.2009.08.152","article-title":"The digenic hypothesis unraveled: The GJB6 del(GJB6-D13S1830) mutation causes allele-specific loss of GJB2 expression in cis","volume":"389","author":"Schrijver","year":"2009","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Rodriguez-Paris, J., Tamayo, M.L., Gelvez, N., and Schrijver, I. (2011). Allele-specific impairment of GJB2 expression by GJB6 deletion del(GJB6-D13S1854). PLoS ONE, 6.","DOI":"10.1371\/journal.pone.0021665"},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Le Nabec, A., Blotas, C., Briset, A., Collobert, M., Ferec, C., and Moisan, S. (2022). 3D Chromatin Organization Involving MEIS1 Factor in the cis-Regulatory Landscape of GJB2. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23136964"},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Cabanillas, R., Dineiro, M., Cifuentes, G.A., Castillo, D., Pruneda, P.C., Alvarez, R., Sanchez-Duran, N., Capin, R., Plasencia, A., and Viejo-Diaz, M. (2018). Comprehensive genomic diagnosis of non-syndromic and syndromic hereditary hearing loss in Spanish patients. BMC Med. Genom., 11.","DOI":"10.1186\/s12920-018-0375-5"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"405","DOI":"10.1038\/gim.2015.30","article-title":"Standards and guidelines for the interpretation of sequence variants: A joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology","volume":"17","author":"Richards","year":"2015","journal-title":"Genet. Med."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1593","DOI":"10.1002\/humu.23630","article-title":"Expert specification of the ACMG\/AMP variant interpretation guidelines for genetic hearing loss","volume":"39","author":"Oza","year":"2018","journal-title":"Hum. 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