MOLECULAR ANALYSES OF TEMPORAL BONE DEVELOPMENT

该提案的主要目标是研究POU结构域基因Brain-4（Brn 4）在内耳发育过程中的功能。Brn 4基因（POU 3F 4）的人类直系同源物中的突变产生“镫骨手术期间与外淋巴涌出相关的混合传导性和感觉神经性耳聋”（DFN 3; McKusick目录号304400）。 畸形的镫骨和颞骨发生在这些患者。最近，我们的实验室已经证明，类似的畸形发生在一个小鼠谱系的内耳中，该谱系包含Brn 4基因的靶向“敲除”突变。 在该动物模型中发育异常的个体发生分析将提供表征导致突变表型的机制的手段，以及洞察颞骨的正常发育。 该提案的具体目标是：1.充分表征Brn 4/Pou 3f 4基因靶向突变小鼠发育异常的个体发生。2.描述Brn 4基因敲除小鼠发生发育缺陷的细胞机制。 我们建议测试两种可能的机制，可以解释在突变体中观察到的颞骨质量减少：假设1）Brn 4抑制耳囊形成过程中的细胞凋亡，或假设2）Brn 4调节耳间充质细胞的细胞增殖。3.为了充分描述Brn 4突变等位基因的分子基础，性连锁坐立不安。4.表征Brn 4基因中耳特异性转录调控元件。5.对Brn 4缺失家系的听觉功能进行功能分析。 为了解决导致Brn 4无效突变体听力损失的病理机制，我们建议更详细地评估Brn 4无效谱系的听觉功能。

The broad goal of this proposal is to study the function of the POU-domain gene, Brain-4 (Brn4), during inner ear development. Mutations in the human ortholog of the Brn4 gene (POU3F4) produce "mixed conductive and sensorineural deafness associated with perilymphatic gusher during stapes surgery" (DFN3; McKusick catalog number 304400). Malformations of both the stapes and the temporal bone occur in these patients. Recently, our laboratory has demonstrated that similar malformations occur in the inner ear of a mouse pedigree containing a targeted "knockout" mutation in the Brn4 gene. Analysis of the ontogeny of developmental abnormalities in this animal model will provide a means to characterize the mechanisms that lead to the mutant phenotype, as well as, insight into the normal development of the temporal bone. The specific aims of this proposal are: 1. To fully characterize the ontogeny of developmental anomalies in mice with a targeted mutation in the Brn4/Pou3f4 gene. 2. To characterize the cellular mechanisms by which developmental defects occur in the Brn4 knockout mice. We propose to test two possible mechanisms that would explain the decreased temporal bone mass observed in the mutant: Hypothesis 1) Brn4 inhibits apoptosis during otic capsule formation, or Hypothesis 2) Brn4 regulates cell proliferation of otic mesenchyme cells. 3. To fully characterize the molecular basis of a Brn4 mutant allele, sex- linked fidget. 4. To characterize otic-specific transcriptional regulatory elements in the Brn4 gene. 5. To undertake functional analyses of auditory function of the Brn4 null pedigree. To address the pathological mechanisms leading to hearing loss of the Brn4 null mutants, we propose to assess auditory function of the Brn4 null pedigree in greater detail.