Molecular Basis of Inherited Deafness

描述（由申请人提供）：转录因子Math1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Math1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。尽管已经发现了70多个人类非综合征性耳聋的基因位点，但仅鉴定出14个基因位点。这项研究将有助于识别其他疾病。此外，耳蜗内邻近的支持细胞在转染Math1后可以分化为毛细胞，而其他类型的细胞则不能。因此，鉴定由Math1激活的基因，以及Math1的毛细胞特异性结合伴侣，可能会揭示促进毛细胞再生的基因。与年龄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造成的，可以通过支持细胞的毛细胞再生来改善。该项目旨在鉴定毛细胞中由Math1激活的基因，使用GeneChips筛选来自几种不同方法的rna。它将首先利用来自正常毛细胞的基因芯片数据、RT-PCR和转染细胞的共免疫沉淀，鉴定与毛细胞中Math1形成异聚体的I类基本螺旋-环-螺旋转录因子。然后，它将使用GeneChips在转染了Math1的耳蜗细胞系、稳定转染了四环素抑制因子下的Math1的骨肉瘤细胞以及缺乏Math1的小鼠中鉴定Math1的候选靶基因。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Math1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类和小鼠的耳聋基因位点进行比较，以产生额外的耳聋基因候选基因。由于Math1对于毛细胞的产生是必要的，并且（在某些细胞类型中）是充分的，这项研究应该告诉我们很多关于毛细胞如何变成毛细胞的知识。

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.