Genetic and epigenomic determinants of hearing loss in Hispanic populations

感音神经性听力损失（SNHL）是导致残疾的主要原因，影响约14亿人 全球各地的人，包括不同年龄组和种族。尽管已经鉴定出大约150个基因 对于SNHL，西班牙裔人口仍然对SNHL研究不足，大多数西班牙裔研究集中在 单基因GJB 2。研究的几个国家--智利、墨西哥/拉美裔、尼加拉瓜、菲律宾-- GJB 2变异的患病率较低，这表明来自这些国家的SNHL队列具有新的基因 或变体以供发现。此外，西班牙裔儿童面临环境暴露于化学品的风险 这可能导致表观遗传修饰并导致SNHL。我们的首要假设是，SNHL有一个 西班牙裔人群中独特的群体特异性等位基因和表观遗传谱。我们 组建了一个国际研究小组，在耳科，遗传学， 表观基因组学和功能基因组学，与以前的合作经验，标志着这个项目将 高生产力。在我们以前的研究中，我们在西班牙裔美国人和菲律宾患者中发现了新的变异， SNHL，包括与颞骨异常相关的遗传变异， 人工耳蜗植入结果。哥伦比亚家庭提交外显子组测序，并为阴性， 这些家族可能含有非编码变体或具有SNHL的表观遗传机制。We have in 为在家族和差异甲基化区域中鉴定新的SNHL基因提供有效的途径 （DMR）的病例对照组，以及在动物模型和上皮细胞的验证方法。我们的目标是 确定西班牙裔SNHL儿童的遗传和表观遗传风险因素。对于目标1，我们将确定 使用分层方法，包括桑格测序， 筛选、纯合性作图、连锁分析和传递不平衡检验。我们将招募500名 西班牙裔家庭，包括足以进行全基因组显著连锁的大家庭，并提交DNA 样品进行测序和分析。在这些家族中发现的新SNHL基因和变异体将被跟踪 在小鼠和斑马鱼模型中进行的蛋白质定位和听力研究，以及 上皮细胞对于目标2，我们将通过分析一个基因组的甲基化组来进行表观基因组范围的关联研究。 一个由500名SNHL先证者和500名听力正常儿童组成的高效能队列，按年龄、性别和人口进行匹配， 以识别与SNHL相关的DMR。对于最常见的DMR，我们将利用CRISPR-dCas 9 在上皮细胞上的技术，以确定靶向甲基化位点是否会影响基因表达。一体化 使用甲基化数量性状基因座分析的甲基化图谱和遗传数据将有助于 了解遗传与环境对SNHL的贡献。总体而言，该项目将影响遗传筛查 协议和遗传咨询，特别是在西班牙裔人口，以及提高对 从而为SNHL的治疗提供新的靶点。

ABSTRACT Sensorineural hearing loss (SNHL) is a leading cause of disability and affects ~1.4 billion people globally, including different age groups and ethnicities. Although around 150 genes have been identified for SNHL, the Hispanic population remains understudied for SNHL, with most Hispanic studies focused on a single gene GJB2. A few countries studied – Chile, Mexico/Hispanic-American, Nicaragua, the Philippines – have a low prevalence of GJB2 variants, suggesting that SNHL cohorts from these countries have novel genes or variants for discovery. Additionally, Hispanic children are at risk for environmental exposures to chemicals that may lead to epigenetic modifications and cause SNHL. Our overarching hypothesis is that SNHL has a unique, population-specific allelic and epigenetic spectrum in Hispanic-descent populations. We assembled an international group of researchers with complementary expertise in otology, genetics, epigenomics and functional genomics, with previous collaborative experience that signals this project will be highly productive. In our previous studies, we identified novel variants in Hispanic-American and Filipino patients with SNHL, including genetic variants that were associated with temporal bone anomalies and predictive of cochlear implant outcomes. Nicaraguan families were submitted for exome sequencing and were negative for variants; these families likely harbor non-coding variants or have epigenetic mechanisms of SNHL. We have in place efficient pipelines for the identification of novel SNHL genes in families and differentially methylated regions (DMRs) in case-control cohorts, as well as validation methods in animal models and epithelial cells. Our goal is to determine genetic and epigenetic risk factors in Hispanic children with SNHL. For Aim 1, we will identify SNHL variants from next-generation sequence data using a tiered approach, which includes Sanger sequencing, filtering, homozygosity mapping, linkage analysis and transmission disequilibrium tests. We will recruit 500 Hispanic families, including large families sufficient for genome-wide significant linkage, and submit DNA samples to sequencing and analyses. Novel SNHL genes and variants identified in these families will be followed up with protein localization and hearing studies in mouse and zebrafish models and mutation constructs in epithelial cells. For Aim 2, we will perform an epigenome-wide association study by profiling the methylome of a well-powered cohort of 500 SNHL probands and 500 hearing children matched by age, sex and population, in order to identify DMRs that are associated with SNHL. For the top identified DMRs, we will utilize CRISPR-dCas9 technology on epithelial cells to determine if targeting the methylation site will affect gene expression. Integration of methylation profiles and genetic data using methylation quantitative trait locus analysis will aid in understanding genetic vs. environmental contributions to SNHL. Overall this project will impact genetic screening protocols and genetic counseling particularly in Hispanic populations, as well as improve understanding of the hearing mechanism and lead to new targets for the development of treatment of SNHL.