Drosophila deafness genes: analysis of chordotonal organ functions

这项建议的长期目标是了解约翰斯顿的发展和功能 器官(JO)，这是遗传模式生物果蝇的听觉器官。这个 果蝇JO与哺乳动物的内耳是同源的，因为它们都依赖于 无机盐家族中高度保守的转录因子的活性。小鼠无调性同源基因1 (Mathl)和苍蝇无性基因在互惠转基因拯救中可以替代对方的功能 实验。此外，人类无调性同系物1(Atohl)可以调节听觉毛发的再生。 药物性耳聋哺乳动物体内的细胞。因此，苍蝇JO代表了一个强大的基因发现 听证资源。Jo在几个方面不同于其他脉络膜器官，这是它专门用于听力的。 在幼虫发育的早期阶段，当JO发育的许多关键事件发生时，包括不对称的分裂 前体细胞，感觉器官细胞谱系的规范，以及正确的细胞形状变化 在分化过程中，JO在蛹内被遮蔽，很容易被解剖。我们的总体战略是 设计更好地可视化正在开发的JO的方法，并将这些方法用作系统化的基础 用于基因发现的表达微阵列分析。首先，我们将描述几个JO基因的作用 在这些早期的蛹阶段。为了确定这些基因的医学相关性，我们将筛选 他们的人类同源基因与分离耳聋的家庭的联系。第二，我们将对文化进行解剖 触角盘能在体内动态成像细胞谱系和标记表达，远离遮蔽 蚕蛹。我们将用几个标志来确定文化发展的保真度，并定义 这些阶段的突出事件。第三，我们将利用此方法的较高吞吐量来恢复 在这些关键阶段，来自野生型和突变型触角圆盘的足够的RNA来比较基因的表达 使用微阵列分析。我们计划最初使用CUT突变体作为范例。被切割的转录因子是 CDP/CUX1是正常JO发育所必需的，而哺乳动物的同源基因CDP/CUX1在内耳表达。 因此，我们希望我们的实验结果将允许我们识别作用于 这些关键阶段。总体而言，这些研究将为未来关于发育和功能的研究提供信息 哺乳动物内耳的生物学，并加速我们对人类听觉障碍的理解。

The long term objectives of this proposal are to understand the development and function of Johnston's organ (JO), which is the auditory organ in the genetic model organism, the fruit fly Drosophila. The Drosophila JO is homologous to the mammalian inner ear because they both rely for their specification on the activity of highly conserved transcription factors of the atonal family. The mouse atonal homolog 1 (Mathl) and the fly atonal genes can substitute for each other's function in reciprocal transgenic rescue experiments. Furthermore, the human atonal homolog 1(Atohl) can mediate regeneration of auditory hair cells in pharmacologically deafened mammals. Thus, the fly JO represents a powerful gene discovery resource for hearing. JO differs from other chordotonal organs in several ways that specialize it for hearing. At early pupal stages when many critical events of JO development occur, including asymmetric divisions of precursor cells, specification of sense organ cell lineages, and cell shape changes essential for correct differentiation, the JO is obscured within the puparium and very fragile to dissect. Our general strategy is to devise methods to better visualize the developing JO, and to use these methods as the basis for systematic expression microarray analysis for gene discovery. First we will characterize the roles of several JO genes at these early pupal stages. To characterize the medical relevance of these genes, we will screen for associations of their human homologs with families segregating deafness. Second, we will culture dissected antennal disks to image cell lineages and marker expression dynamically in vivo, away from the obscuring puparium. We will determine the fidelity of development in culture with several markers, and define the salient events at these stages. Third, we will exploit the higher throughput of this approach to recover sufficient RNA from wild-type and mutant antennal disks at these critical stages to compare gene expression using microarray analysis. We plan to use cut mutants initially as a paradigm. The cut transcription factor is required for normal JO development, and the mammalian homolog, CDP/Cux1 is expressed in the inner ear. Thus, we expect that the results of our experiments will permit us to identify target genes of cut that act at these critical stages. Overall, these studies will inform future research on the developmental and functional biology of the mammalian inner ear, and accelerate our understanding of human auditory disorders.