Development studies of the inner ear

描述(申请人提供)：形态因子是在胚胎发育过程中作为梯度作用的分子，根据暴露在形态因子中的时间和/或浓度，赋予细胞不同的命运。鸟类和哺乳动物发育中的内耳中存在几种已知在其他系统中起形态原作用的分泌配体，包括Wnt基因家族的分子。WNT转录本在胚胎耳蜗径向轴上分布不对称。目前的研究表明，通过异位基因转移在体内破坏这种局部表达可以改变鸡胚胎发育中的耳蜗细胞的命运。这一新出现的证据表明，Wnt形态原可能在耳蜗中发挥作用，这导致了3条线索的调查。在目标1中，将对转录本进行深度测序，以比较对照耳和通过异位Wnt传递将听力器官的非神经侧转变为神经侧命运的耳朵。这有望揭示感觉器官两侧差异使用的基因，其中一些基因将有能力直接影响放射状同一性的各个方面，如细胞分裂的时间调节、细胞类型的选择以及轴突神经的分布和分类。越来越多的新证据表明，以转录抑制靶基因转录而闻名的microRNAs可以与癌细胞和胚胎中参与Wnt信号通路的分子形成相互作用的网络。在目标2中，microRNA图谱的变化将与异位Wnt传递诱导的鸡听力器官径向细胞类型的转变有关。这应该提供特定的候选microRNAs，它们与Wnt途径相互作用，形成反馈或前馈网络，从而影响听力器官放射轴上的细胞表型。目标3将转向研究发育中的小鼠耳蜗，以确定哪些Wnt信号分子以与该物种的形态生成功能一致的方式分布。然后，可用的阻断所有Wnt配体分泌的小鼠突变体将为揭示特定的Wnt配体是否与特定的microRNA子网络提供了宝贵的资源。为此，将在不同Wnt配体存在的情况下培养缺乏Wnt分泌的胚胎耳蜗组织，并比较这些处理对下游microRNAs的影响。这些研究的总体意义在于，它们可能有助于更好地理解有助于形成和形成耳蜗细胞类型的基因网络，并为人类患者听力损失生物治疗的设计和实施提供长期目标。

DESCRIPTION (provided by applicant): Morphogens are molecules that operate as gradients in developing embryos to endow cells with different fates depending upon the duration and/or concentration of morphogen exposure. Several secreted ligands known to function as morphogens in other systems are present in the developing inner ears of birds and mammals, including molecules in the Wnt gene family. Wnt transcripts are asymmetrically distributed across the radial axis of the embryonic cochlea. Current research reveals that disruption of this localized expression in vivo by ectopic gene transfer can transform cell fates in the developing cochlea of the chicken embryo. This emerging evidence that a Wnt morphogen may be active in the cochlea leads to 3 lines of inquiry. In Aim 1, deep-sequencing of transcripts will be undertaken to compare control ears to those in which the abneural-side of the hearing organ has been converted to a neural-side fate by ectopic Wnt delivery. This is expected to reveal genes that are differentially employed on the two sides of the sensory organ, some of which will have the ability to directly influence aspects of radial identity, such as the temporal regulation f cell division, the choice of cell types, and the distribution and class of axonal innervation. Accumulating new evidence suggests that microRNAs, known for their ability to transcriptionally repress target transcripts, can form interacting networks with molecules involved in the Wnt signaling pathway in cancer cells and in embryos. In Aim 2, changes in the microRNA profile will be linked to the transformation of cell types across the radial axis of the chicken hearing organ that are induced by ectopic Wnt delivery. This should provide specific candidate microRNAs that interact with the Wnt pathway to form either feedback or feedforward networks and thereby influence cellular phenotypes across the radial axis of the hearing organ. Aim 3 will switch to studying the developing mouse cochlea to identify which Wnt signaling molecules are distributed in a manner consistent with a morphogen function in this species. Then, available mouse mutants that block the secretion of all Wnt ligands will provide a valuable resource to reveal whether specific Wnt ligands form sub-networks with specific microRNAs. To accomplish this, embryonic cochleas lacking Wnt secretion will be cultured in the presence of different Wnt ligands and the downstream microRNAs affected by these treatments will be compared. The overall significance of these studies is that they may offer a better understanding of the gene networks that contribute to forming and patterning cochlear cell types, with a long-term goal of informing the design and implementation of biological-based treatments of hearing loss in human patients.