DIFFERENTIAL GENE EXPRESSION IN NORMAL, MUTANT AND AGED

描述(申请人摘要)：本研究的总体科学目标 建议是确定其表达变化的基因作为两个结果 小鼠感音神经性耳聋的类型：与年龄相关的听力损失 菌株C57BL6/J和3个突变型先天性耳聋小鼠。这些 密歇根大学的两位科学家正在研究突变体 这笔拨款的联合调查员。Shaker-2已被证明携带突变 坎珀博士的团队发现了一种非传统肌球蛋白MyoXV的基因。旋转木马 旋转者突变体已经被定位，突变的基因正在被识别 科尔曼博士的团队。我们建议确定对 单基因引起的内耳转录谱系(转录组) 导致小鼠耳聋的突变。这种方法基于两个 相关假设：(1)这些突变改变了正常发育 在细胞成熟过程中运行的过程和动态平衡机制 和(2)，这些变化将反映在 在细胞中工作的基因的稳态转录水平的变化 相关的发育和动态平衡途径。通过使用高度并行的 识别基因表达变化的方法，我们将开始表征 直接或间接受突变影响的调节电路， 而它们本身也可能在正常的内耳中发挥关键作用 发展、动态平衡和功能。AIMS 1和2目前建议使用 可用的试剂和技术，如尼龙膜上的基因阵列，以 检测增龄相关小鼠模型C57BL/6J的基因表达变化 听力损失(目标1)，以及在三种先天性耳聋和 前庭功能障碍(目标2)。最后两个目标解决了 应用新的和新兴的技术来提高泳池的“完整性” 通过数据库比对和SAGE获得内耳基因图谱(AIM 3) 分析，以及通过开发DNA进行表达谱分析的“吞吐量” 玻片上含有在小鼠内耳表达的基因的微阵列 (目标4)。这些研究应该会加深我们对分子事件的理解。 年龄相关性听力损失和先天性耳聋的研究并提供新的试剂 用于评估耳朵中基因表达的变化。

DESCRIPTION (Applicant's abstract): The overall scientific objectives of this proposal are to identify genes whose expression changes as a consequence of two types of sensorineural deafness in the mouse: age-related hearing loss in strain C57BL6/J, and congenital deafness in three deafness mouse mutants. These mutants are being studied at the University of Michigan by two of the co-investigators on this grant. Shaker-2 has been shown to carry mutations in the gene for an unconventional myosin, MyoXV, by Dr. Camper's group. Pirouette and spinner mutants have been mapped and the mutated genes are being identified by Dr. Kohrman's group. We propose to determine the broad effects on the transcriptional repertoire (transcriptome) of the inner ear due to single gene mutations that result in deafness in the mouse. This approach is based on two related hypotheses: (1) these mutations alter the normal developmental processes and homeostatic mechanisms that operate during maturation of the cochlea or in the adult organism, and (2), the alterations will be reflected in changes in the steady state transcript levels of genes that operate in the relevant developmental and homeostatic pathways. By using highly parallel methods to identify gene expression changes, we will begin to characterize the regulatory circuits that are affected directly or indirectly by the mutations, and which themselves may also play critical roles in normal inner ear development, homeostasis and function. Aims 1 and 2 propose to use currently available reagents and technique, such as gene arrays on nylon membranes, to examine gene expression changes in C57BL/6J, a mouse model of age-related hearing loss (Aim 1), and in the three mouse models of congenital deafness and vestibular dysfunction (Aim 2). The final two Aims address the feasibility of applying new and emerging techniques to increase the 'completeness' of the pool of profiled inner ear genes (Aim 3) through database comparisons and SAGE analysis, and the 'throughput' of expression profiling by developing DNA microarrays on glass slides containing genes expressed in the mouse inner ear (Aim 4). These studies should enhance our understanding of the molecular events of age-related hearing loss and congenital deafness and provide new reagents for assessing changes in gene expression in the ear.