Genetic Screening Of Mutation Hot-spots For Subjects With Nonsyndromic Hearing Loss And A Novel MYO7A Gene Mutation Analysis In A Deafness Family

PartⅠGJB2, SLC26A4 and mtDNA12S rRNA hot-spots in 156 subjects with nonsyndromic hearing loss inTengzhou, ChinaObjectives Auditory dysfunction is a common sensory dysfunction, intheincidence of 1:1000, about half of them have hereditary factors. Researchers had shown that the most common causes of deafness in China were GJB2, SLC26A4 and mtDNA12S rRNA gene mutation. But so far, in most areas of our country there was no systemical analysis of genetic survey for deaf people. Inthisstudy, we performed the molecula repidemiological investigation for nonsyndromic deaf populationin Tengzhou area, so as to draw the pathogenic genes pectrum for deaf patients in this area.Methods According to the inclusion criteria and exclusion criteria, we selected severe to profound nonsyndromic deafness patients as the research objects for genetic screening in Tengzhou area. By completing the "deaf patient information registration form" and the telephone, we collected the deafness history. Improve the examination of patients with deafness （including general physical examination, physical examination specialized in ENT, audiological examinations and imaging examinations）. The peripheral blood specimens of deafnes spatients were obtained, the genomic DNA were extracted. Using multiple rapid genetic screening SNP scan TM detection technology, we detected 115 common allele mutation sites of GJB2, SLC26A4 and mtDNA12S rRNA gene. Finally, we use the SPSS20.0 software for statistical analysis. Summary the deafness alleles found in this deaf population, compare the mutation hotspots detection rates, then draw the pathogenic genes pectrum for patients with deafness in Tengzhou area.Results Among the 156 analyzed patients,74 patients were demonstrated with deafness genes, accounting for 47.44%（74/156）, including GJB2 （26.92%,42/156）, SLC26A4 （17.95%,28/156） and mtDNA12S rRNA （2.56%,4/156）.60 patients were identified with explicit gene causes, accunting for 38.46%（60/156）, including GJB2 （22.44%,35/156）, SLC26A4 （13.66%,22/156） and mtDNA12S rRNA （2.56%,4/156）. In this study, we confirmed 27 deafness-causing mutations and 27 different allelic combinations including GJB2 （11 variants,11 allelic combinations）, SLC26A4 （14 variants,16 allelic combinations） and mtDNA12S rRNA （2 variants）.Conclusion In this cohort of 156 nonsyndromic hearing-impaired subjects of Tengzhou area, the deafness-associated genes GJB2, SLC26A4 and mtDNA12S rRNA were investigated by SNPscan detection technique efficiently. The deafness mutant alleles were complex and diversified in Tengzhou Region, GJB2 and SLC26A4 gene mutations were still the major causative genes in Tengzhou region. The deafness gene mutation hot-spots in the region were c.235delC, c.299-300delAT, c.176₁91del16, c.109G> A in GJB2 gene; c.IVS7-2A> G, c.2168A> Q c.1226G> A in SLC26A4 gene; m.1555A> G in mtDNA12S rRNA gene. GJB2 c.235delC and SLC26A4 c.IVS7-2A>G were the most common mutationsites. For deaf patients and high-risk groups, the molecular genetic screening was the first step to prevent and control hereditary deafness in Tengzhou region. Together with prenatal diagnosis, medication guide and clinical interventions, genetic diagnosis can be effective to not only reduce the incidence of deafness but also provid the basis for drawing deafness gene profiles in Shandong Province.PartⅡIdentification of a novel MYO7A gene mutation in a Chinese family with autosomal recessive nonsyndromic hearing lossObjective Mutations in MYO7A gene have been reported to be associated with Usher Syndrome type 1B （USH1B） and nonsyndromic hearing loss （DFNB2, DFNA11）. Most mutations in MY07A gene caused USH1B, whereas only a few reported mutations led to DFNB2 and DFNA11. Using SNPscan detection technology, we detected 115 common gene mutations of GJB2, SLC26A4 and mtDNA12S rRNA gene for 27 deafness families in Tengzhou City. Among them,13 cases of deafness pedigrees have not been detected a common mutation. The purpose of this study is to find the gene mutation for one family with autosomal recessive Chinese hereditary nonsyndromic sensorineural deafness.Methods First of all, collect the acquisition history by questionnaire for this autosomal recessive noncomprehensive syndrome sensorineural deafness family in China. The patients and family members were detected with pure toneaudiometry, ophthalmic examination, temporal bone CT and MRI. The peripheral venous blood samples of the family members were collected, genomic DNA were extracted. Then, using target gene capture and next-generation sequencing methods, we detected the genomic DNA to find possible new deafness gene mutations. To validate the new gene mutation, we detect DNA of all the family members and 120 normal cases by Sanger sequencing. Determine the new mutation loci corresponding region structure conservation. Finally, the amino acid changes caused by the mutations were analysised.Results Patients in the Chinese family with autosomal recessive nonsyndromic sensorineural deafness showed bilateral sensorineural hearing loss after birth. Both patients of this family had no visual disorder. Through the target gene capture and next-generation sequencing, we identified a new compound heterozygous mutationin MY07A gene:c.3671C>A （p.A1224D） and c.390₃91insC （p.P81PfsX9）. The c.3671C>A （p.A1224D） is a new site, c.390₃91insC （p.P81PfsX9） has been reported. The deafness phenotype was consistent with DFNB2. Sanger sequencing confirmed: in the gene alogies, the new compound heterozygous mutations c.3671C>A （p.A1224D） and c.390₃91insC （p.P81PfsX9） in MY07A gene were co-segregated with autosomal recessive hearing loss phenotype. Sanger sequencing was employed to examine those variant mutations in the members of this family and other ethnicity-matched controls. The gene mutation analysis and protein sequence alignment further supported that the novel compound heterozygous mutations in MY07A gene were pathogenic. Although the performance is similar to the phenotype of DFNB2, the probability of developing retinopathy later can not be ruled out. We still need a long time to clear clinical observation for real DFNB2 or USH1B.Conclusion In this study, we identified the novel compound heterozygous mutations c.367C>A （p.A1224D） and c.390₃91insC （p.P81PfsX9） in MY07A gene causing autosomal recessive sensorineural hearing loss in a Chinese family.