ABSTRACT Hearing loss (HL) is the most common sensory disorder affecting more than 28 million Americans and affects 15-26% of the world's population. Amongst the elderly, it ranks as the most common neurological disability, impacting ~50% of octogenarians. Clinically significant HL is present in at least 1 per 500 infants at birth. Recent advances in molecular technology continues to revolutionize human genetics. However, there is a pressing need to continue identifying new human HL genes and determining causative variants in known NSHL genes for completing a genomic and phenotypic database. This is critical as we and others have provided direct evidence of further genetic heterogeneity of HL with a number of genes/mutations yet to be identified. Importantly, as shown in our preliminary studies, we established the comprehensive and integrated scope - Miami Ear Institute Otogenetic Program (MEIOP) and pipelines including existence of aggregated, clinically well-defined WES/WGS/RNAseq/non-coding causative variation and multi-omics and gene panel datasets; data analysis platform, established the diverse Miami Otogenetic Repository (MOR) (including 60% US minorities) that contains biological samples and clinical data on families with NSHL, a productive functional follow-up, including model systems (cells, zebrafish); and in silico approaches of machine learning supported pathogenicity predictions, and statistical variant burden analysis for the genetic and functional analysis of variants. In this competitive renewal, our overarching goals are to further improve the clinical care of persons with HL and provide a more robust foundation for therapies that target specific types of genetic HL. We will achieve these goals by addressing current knowledge gaps as reflected in the following specific aim 1,1: To determine causative variants in known NSHL genes and expand our minority focused populations using our well established multi-tier genomic profiling approach. Aim 1.2: To identify novel NSHL genes using targeted sequence capture/ whole exome/genome analysis using a customized local pipeline platform and functional and multi-omics analysis. Aim 2: To enhance the investigation of novel disease mechanisms through cellular/tissue assays and animal models of rare and common human HL genes/variants. Our approaches will include functional analysis of wild type and mutant proteins in iPSCs, inner ear organoids, and animal models, followed by characterization of their inner ear phenotypes and function. The completion of these aims will be discoveries of novel coding and non-coding variants in genes and pathways involved in the pathophysiology of deafness, improve clinical care for persons with hearing loss, provide a better genetic foundation for precision medicine for the hearing impaired, and identify new targets for gene therapy for deafness in USA populations, especially in USA minorities.