Molecular Basis of Inherited Deafness

描述（由申请人提供）：转录因子Math 1是内耳毛细胞发育和存活所必需的。因此，毛细胞中至少有一些被Math 1激活的基因是毛细胞发育所必需的，因此是遗传性耳聋的候选基因。虽然已经发现了70多个人类非综合征性耳聋的基因位点，但只有14个基因被确定。这项研究将有助于确定其他人。此外，如果用Math 1转染，耳蜗中相邻的支持细胞可以分化为毛细胞，尽管其他细胞类型不能。因此，鉴定由Math 1激活的基因，以及Math 1的毛细胞特异性结合伴侣，可以揭示可以促进毛细胞再生的基因。与听力损失相关的听力损失影响着数千万美国人，通常是由毛细胞死亡引起的，可以通过支持细胞再生毛细胞来改善。拟议的项目旨在识别毛细胞中Math 1激活的基因，使用基因芯片筛选来自几种不同方法的RNA。它将首先确定I类基本螺旋-环-螺旋转录因子，这些转录因子在毛细胞中与Math 1形成异多聚体，使用来自正常毛细胞的基因芯片数据，RT-PCR和转染细胞中的免疫共沉淀。然后，它将使用基因芯片来识别Math 1的候选靶基因，在转染Math 1的耳蜗细胞系中，在四环素阻遏物下稳定转染Math 1的骨肉瘤细胞中，以及在缺乏Math 1的小鼠中。在第三个目标中，它将通过确定它们是否在适当的发育阶段在毛细胞中表达，以及Math 1是否可以在候选基因启动子的控制下激活报告基因来测试这些候选基因。最后，将确认的候选基因的染色体位置与人和小鼠的耳聋基因座进行比较，以产生耳聋基因的其他候选基因。由于Math 1对于毛细胞的生成是必要的，并且（在某些细胞类型中）足够，因此这项研究应该告诉我们很多关于毛细胞如何成为毛细胞的信息。

DESCRIPTION (provided by applicant): The transcription factor Math1 is necessary for the development and survival of hair cells in the inner ear. At least some of the genes that are activated by Math1 in hair cells are therefore necessary for development of hair cells, and consequently are candidate genes for inherited deafness. Although over 70 gene loci for human nonsyndromic deafness have been found, the genes for only 14 have been identified. This study will help identify others. Moreover, adjacent supporting cells in cochlea can differentiate into hair cells if transfected with Math1, although other cell types cannot. Thus identification of the genes activated by Math1, and of the hair-cell-specific binding partners of Math1, may reveal genes that could promote regeneration of hair cells. Age-related hearing loss, which affects tens of millions of Americans, often results from the death of hair cells and could be ameliorated by the regeneration of hair cells from supporting cells. The proposed project seeks to identify genes activated by Math1 in hair cells, using GeneChips to screen RNAs derived from several different approaches. It will first identify class I basic helix-loop-helix transcription factors that form heteromultimers with Math1 in hair cells, using GeneChip data from normal hair cells, RT-PCR, and co-immunoprecipitation in transfected cells. It will then use GeneChips to identify candidate target genes of Math1, in a cochlea cell line transfected with Math1, in stablytransfected osteosarcoma cells with Math1 under a tetracycline repressor, and in mice lacking Math1. In a third aim, it will test these candidates by determining whether they are expressed in hair cells at an appropriate developmental phase, and whether Math1 can activate a reporter gene under the control of the candidate-gene promoter. Finally, the chromosomal locations of confirmed candidates will be compared to deafness loci in human and mouse, to produce additional candidates for deafness genes. Since Math1 is necessary and (in some cell types) sufficient for the generation of hair cells, this study should teach us much about how a hair cell becomes a hair cell.