Sensory cell fate specification in the inner ear

描述：细胞外信号分子在脊椎动物内耳发育的多个阶段为其提供位置信息，以产生前庭和耳蜗所需的生长和分化模式，这两个内耳器官分别负责感觉、平衡和声音。在以前的工作中，我们确定了分泌因子Shh和Wnt1/Wnt3a在小鼠耳囊沿其背腹轴的极化中所起的作用。我们的研究确定，从脊索分泌的Shh直接向耳上皮腹侧区域发出信号，指导耳蜗管的生长。而Wnt1/Wnt3a从背侧后脑分泌，向背侧耳囊发出信号，调节前庭的形态发生。除了这些早期的作用外，Hedgehog(HH)和Wnt/2catenin信号通路也在耳朵发育后期的关键时刻活跃，此时感觉上皮祖细胞正在进行指定和分化为毛细胞和支持细胞。我们现在建议使用一种条件基因打靶策略来研究Wnt/2catenin和HH信号通路在小鼠内耳发育后期的特殊要求，该策略在耳蜗和前庭发育的特定时期灭活这些通路的基本中介。我们的条件性基因打靶方法利用了我们最近的发现，即起源于背侧耳囊的Wnt反应细胞随着时间的推移延伸到腹侧，并有助于耳蜗管的感觉上皮。使用他莫昔芬诱导形式的cre重组酶，在Wnt Response Top启动子(Topcreer)的转录控制下表达，2catenin和Smoothated的花等位基因在感觉上皮祖细胞中将被灭活。初步研究结果表明，Wnt/2catenin信号通路是半规管Corti器和棘内毛细胞和支持细胞规范和/或分化所必需的。这项提议中的实验将进一步阐述WNT/2catenin和HH信号通路调节内耳感觉细胞命运的机制。我们还开发了一种基于遗传重组的策略来不可磨灭地标记背侧耳囊中的Wnt反应细胞，以便在内耳发育过程中追踪它们的命运。这项建议中描述的其他实验将检验这一假设，即Corti器官中的毛细胞和支持细胞来自背侧耳囊中一群Wnt反应的祖细胞。这些研究的结果应该会提高我们对内耳听觉和前庭区域感觉发育的基本理解。公共卫生相关性内耳的听觉和前庭结构分别调节我们的听觉和平衡感。在确定人类遗传性耳聋和前庭疾病的病因方面继续取得进展。尽管如此，对协调内耳发育的遗传途径的详细了解仍然有限。通过阐明调控内耳细胞命运决定的遗传网络，我们的研究不仅应该提高我们对这个复杂器官的基本了解，而且还应该提高我们对先天性耳聋的发病机制的认识。

DESCRIPTION: Extracellular signaling molecules provide the vertebrate inner ear with positional information at multiple stages of its development to produce the required patterns of growth and differentiation necessary for the formation of the vestibulum and cochlea, the two inner ear organs responsible for sensing, balance and sound, respectively. In previous work, we identified roles for the secreted factors, Shh and Wnt1/Wnt3a, in the polarization of the mouse otic vesicle along its dorsal-ventral axis. Our studies determined that Shh, secreted from the notochord, signals directly to ventral regions of the otic epithelium to direct the outgrowth of the cochlear duct. Whereas, Wnt1/Wnt3a, secreted from the dorsal hindbrain, signals to the dorsal otocyst to regulate vestibular morphogenesis. In addition to these early roles, Hedgehog (Hh) and Wnt/2catenin signaling pathways are also active at critical junctures later in ear development when sensory epithelial progenitors are undergoing their specification and differentiation into hair cells and support cells. We now propose to investigate the specific requirements of Wnt/2catenin and Hh signaling pathways at later stages of inner ear development in the mouse, using a conditional gene targeting strategy that inactivates essential mediators of these pathways at defined periods of cochlear and vestibular development. Our conditional gene targeting approach takes advantage of our recent finding that Wnt responsive cells originating in the dorsal otocyst extend ventrally over time and contribute to the sensory epithelium of the cochlear duct. Using a tamoxifen inducible form of cre recombinase, expressed under the transcriptional control of a Wnt responsive Top promotor (TopcreER), floxed alleles of 2catenin and Smoothened will be inactivated in sensory epithelial progenitors. Preliminary results indicate that Wnt/2catenin signaling is required for the specification and/or differentiation of hair cells and support cells in the organ of corti and cristae of the semicircular canals. Experiments in this proposal will further elaborate on the mechanisms by which Wnt/2catenin and Hh signaling pathways function to mediate sensory cell fates in the inner ear. We have also developed a genetic recombination based strategy to indelibly mark Wnt responsive cells in the dorsal otocyst in order to trace their fates over the course of inner ear development. Additional experiments described in this proposal will test the hypothesis that hair cells and support cells in the organ of corti derive from a population of Wnt responsive progenitors in the dorsal otocyst. Results from these studies should improve our fundamental understanding of sensory development in auditory and vestibular regions of the inner ear. PUBLIC HEALTH RELEVANCE The auditory and vestibular structures of the inner ear mediate our senses of hearing and balance, respectively. Progress continues to be made in identifying the causes of hereditary forms of deafness and vestibular disease in humans. Nonetheless, a detailed understanding of the genetic pathways coordinating inner ear development remains limited. By elucidating the genetic networks regulating cell fate decisions in the inner ear, our studies should not only improve our fundamental understanding of this intricate organ, but also the pathogenesis of congenital forms of deafness.