Local modulation of retinoic acid signaling in cranial placode formation

摘要 这是关于“局部调节视黄酸”的申请#1F32DE027599-01的修订提交文件 在颅基板形成中的信号传导”。颅基板是胚胎外胚层的局灶性增厚， 在发育过程中分化，以指定脊椎动物头部的关键结构，包括成对的感觉器官， 器官和感觉颅神经节。所有的颅基板都起源于一个共同的祖先领域， 基板前区（PPR）。作为对周围组织信号的反应，PPR最终分离 沿着胚胎的前后轴，以形成不同的基板域，其含有细胞类型特异性 到每个感觉基板：腺垂体，嗅觉，透镜，三叉神经，鳃上和耳 基板破坏基板特化的突变导致广泛的先天性出生缺陷， 以失明或耳聋形式的感觉丧失为特征，或由于垂体 缺陷和口面区域感觉神经支配的丧失。因为颅基板的机制 规范了解甚少，干预和临床改善结果的策略有限 这些缺陷。圣让内实验室使用非洲爪蟾研究颅面的基本机制 在所有脊椎动物中高度保守。之前的实验室研究 表明锌指转录因子Zic 1在非细胞自主的细胞内促进基板的命运。 通过激活视黄酸（RA）信号通路的方式。在上调的基因中， 分解代谢酶Cyp 26 c1和RA调节的同源结构域转录因子Pitx 2c。初步 结果表明Cyp 26 c1对PPR的特化很重要，并在预防过量RA中起作用 积累此外，发现Pitx 2c和Cyp 26 c1共享共同的表达结构域， 占据前神经板之间的区域，其中Zic 1诱导RA合成，和预期的PPR。 基于这些观察，我假设Cyp 26 c1参与RA的局部降解， 建立适当的RA水平阈值，随后可激活Pitx 2c介导的 PPR形成的基因表达。我会综合运用发展基因和高等教育 通过筛选方法确定Cyp 26 c1和Pitx 2c如何调节RA水平， PPR的适当空间定位，并确定在PPR形成中发挥关键作用的新基因。这 这项研究将解决我们目前知识中的空白，即形态发生梯度如何局部调节， 调节特定细胞中的基因表达，还将揭示与颅神经系统相关的新基因。 基板规格这项研究的结果将为基板的机制提供更深入的见解 规范，并将阐明影响感觉器官的颅面缺陷的分子基础。

ABSTRACT This is a revised submission of application #1F32DE027599-01 on the “local modulation of retinoic acid signaling in cranial placode formation”. Cranial placodes are focal thickenings of the embryonic ectoderm that differentiate during development to specify key structures in the vertebrate head, including paired sensory organs and sensory cranial ganglia. All cranial placodes arise from a common progenitor territory called the pre-placodal region (PPR). In response to cues from the surrounding tissues, the PPR eventually segregates along the anterior-posterior axis of the embryo to form distinct placodal domains that contain cell-types specific to each of the sensory placodes: the adenohypophyseal, olfactory, lens, trigeminal, epibranchial and otic placodes. Mutations that disrupt placode specification cause a wide range of congenital birth defects that are characterize by sensory loss in the form of blindess or deafness, or hormone imbalances due to pituitary defects, and loss of sensory enervation to the orofacial region. Because the mechanisms of cranial placode specification are poorly understood, there are limited strategies to intervene and clinically improve the outcome of these defects. The Saint-Jeannet Lab uses Xenopus laevis to study the basic mechanisms of craniofacial development as their blueprint highly conserved across all vertebrates. Previous studies in the lab have demonstrated that the zinc-finger transcription factor Zic1 promotes placodal fate in a non-cell autonomous manner by activating retinoic acid (RA) signaling pathway. Among the genes upregulated are the RA catabolizing enzyme Cyp26c1 and an RA-regulated homeodomain transcription factor Pitx2c. Preliminary results suggest that Cyp26c1 is important for PPR specification and plays a role in preventing excess RA accumulation. Additionally, Pitx2c and Cyp26c1 were found to share a common domain of expression that occupies the region between anterior neural plate, where Zic1 induces RA synthesis, and the prospective PPR. Based on these observations, I hypothesize that Cyp26c1 participates in local degradation of RA in order to establish the appropriate threshold of RA levels, which can subsequently activated Pitx2c-mediated gene expression for the formation of PPR. I will use a combination of developmental, genetic and high throughput screening approaches to determine how Cyp26c1 and Pitx2c modulate RA levels for the appropriate spatial positioning of PPR, and also identify novel genes that play key roles in PPR formation. This study will address current gaps in our knowledge as to how morphogen gradients are locally modulated to regulate gene expression in particular cells, and will also uncover new genes that are involved in cranial placode specification. The findings from this study will provide deeper insights into the mechanisms of placode specification and will shed light on the molecular basis for craniofacial defects affecting sensory organs.