Regulation of inner ear development by FGF signals and effectors

内耳上皮的形态发生需要几种信号通路的协调部署， 干扰导致听力和/或平衡异常。随着人工耳蜗植入术的出现， 听力损失，即使在内耳畸形的情况下，关键是要准确地了解这种畸形如何 影响听觉神经节和神经支配。此外，鉴于对内耳体外生成的强烈关注， 用于移植和体内操作发育信号分子的细胞类型， 本发明的目的在于阐明用于听力恢复的各种内耳细胞的分化，阐明这种分化的作用和调节。 控制耳分化和形态发生的信号和它们的效应物是促进治疗所必需的。 表达编码FGF 3和FGF 10的基因，它们是通过FGFR 2b和FGFR 1b发出信号的配体， 在上皮和神经节结构域的整个耳发育过程中动态变化。进行的研究 Mansour实验室对常规的Fgf 3和Fgf 10条件性敲除小鼠以及表达Fgf 10基因的小鼠进行了研究。 强力霉素诱导配体陷阱（dnFGFR 2b），快速抑制通过FGFR 1b和 FGFR 2b显示，Fgf 3和Fgf 10在耳囊形成的基板谱系中不是必需的，但在耳囊形成的基板谱系中是必需的。 所需的随后耳囊肿图案，神经母细胞的维护，上皮细胞增殖和两者 前庭和耳蜗形态发生。此外，第一个全基因组的耳囊mRNA分析显示， FGFR 2b/1b信号转导靶点，定义了参与耳形态发生和功能的新候选基因。 该提议具有两个目标，解决了以下假设：1）连续需要FGFR 2b/1b信令 对于耳神经母细胞的特化和维持，以及在后期阶段，间充质信号传导，以及 与上皮和神经节结构域中的相同，是耳蜗上皮分化和神经节 2）FGFR 2b/1b下游靶基因介导FGFR 2b/1b的部分或全部作用 在耳形态发生和神经节发生中的信号传导。为了确定FGFR 2b/1b信号通路在耳部疾病中的早期作用， 神经节形成及其在上皮分化和神经节维持中的后期作用，DOX诱导的 将使用dnFGFR 2b的普遍和CRE限制性表达，并将使用形态学和分子标记物 将评估两种组织中的耳模式、增殖和存活。确定的角色 FGFR 2b/1b信号传导的下游靶点，两种编码转录因子的基因， FGFR 2b/1b信号传导和一个编码受FGFR 2b/1b抑制的BMP信号传导调节因子的基因 将研究信号。将为每个基因产生耳条件突变体，并且它们的形态学和生物学特性将被改变。 将评估功能发展。此外，BMP调节剂在多大程度上有助于 将评估dnFGFR 2b表型和过表达BMP调节剂的作用。 这些结果将有助于新的知识，将有助于未来的努力，操纵FGF信号转导 听力恢复系统。

Morphogenesis of the inner ear epithelium requires coordinated deployment of several signaling pathways and disruptions cause abnormalities of hearing and/or balance. With the advent of cochlear implantation to treat hearing loss even in cases of inner ear malformation, it is critical to understand exactly how such malformations affect the auditory ganglia and innervation. Also, in light of the intense focus on in vitro generation of inner ear cell types for transplantation and in vivo manipulation of developmental signaling molecules to promote differentiation of various inner ear cells for hearing restoration, elucidating the roles and regulation of such signals and their effectors governing otic differentiation and morphogenesis are necessary to advance treatment. The genes encoding FGF3 and FGF10, ligands that signal through FGFR2b and FGFR1b, are expressed dynamically throughout otic development in both epithelial and ganglion domains. Studies conducted by the Mansour Lab of both conventional Fgf3 and Fgf10 conditional knockout mice and those expressing a doxycycline-inducible ligand trap (dnFGFR2b) that rapidly inhibits signaling through both FGFR1b and FGFR2b, showed that Fgf3 and Fgf10 are not required in the placode lineage for otocyst formation, but are required subsequently for otocyst patterning, neuroblast maintenance, epithelial proliferation and both vestibular and cochlear morphogenesis. Furthermore, the first genome wide analyses of otocyst mRNA revealed FGFR2b/1b signaling targets that define novel candidates for genes involved in otic morphogenesis and function. This proposal has two Aims addressing the hypotheses that 1) FGFR2b/1b signaling is required continuously for both otic neuroblast specification and maintenance, and that at later stages, mesenchymal signaling, as well as that in the epithelial and ganglion domains, is required for cochlear epithelial differentiation and ganglion maintenance and 2) FGFR2b/1b downstream target genes mediate some or all of the effects of FGFR2b/1b signaling on otic morphogenesis and gangliogenesis. To determine the early role of FGFR2b/1b signaling in otic ganglion formation and its later role in epithelial differentiation and ganglion maintenance, DOX-induced ubiquitous and CRE-limited expression of dnFGFR2b will be employed and morphology and molecular markers of otic patterning, proliferation and survival in both tissues will be assessed. To determine the roles of downstream targets of FGFR2b/1b signaling, two genes encoding transcription factors that are activated by FGFR2b/1b signaling and one gene encoding a BMP signaling regulator that is repressed by FGFR2b/1b signaling will be studied. Otic conditional mutants will be generated for each gene, and their morphologic and functional development will be assessed. In addition, the extent to which the BMP regulator contributes to the dnFGFR2b phenotypes and the effects of overexpressing the BMP regulator will be assessed. The results will contribute new knowledge that will facilitate future efforts to manipulate the FGF signaling system for hearing restoration.