Regulation of primary cilia formation in endoderm development and repair

项目摘要/摘要 与普遍认为的几乎所有脊椎动物细胞都有初级纤毛的观点相反，我们的实验室最近发现 胚胎外的内脏内胚层细胞缺乏初生纤毛。我们的实验室也是第一个证明 在哺乳动物中，所有通过Hedgehog通路的信号都需要初级纤毛，这与 数据表明，胚胎外细胞对刺猬配体没有反应。然而，胚胎外细胞可以 分泌Hedgehog配体，这是胚胎来源的血管系统正常发育所必需的 胎盘。我推测，对刺猬配体产生但不起反应的其他类型的细胞也可能 缺少纤毛。与这一假设一致，我发现一些成人内胚层器官的上皮细胞，其中 分泌但对刺猬配体无反应，缺乏初生纤毛。但是我也发现， 内胚层上皮细胞在发育早期有初生纤毛。这种初级纤毛早期出现在肠道中。 形态发生，但在成年前就消失了，这表明来自邻近组织的动态信号可以抑制 初生纤毛的形成。 在拟议的研究中，我将确定内胚层上皮中初级纤毛在什么时候消失， 将使我能够确定在这个时间点存在的外部信号。候选因素的表达将 被操纵来测试初级纤毛是否可以异位组装在通常缺乏 这些细胞器。我还将测试这些细胞中缺少自分泌刺猬信号的功能相关性 组织中通过表达刺猬主要转录激活因子Gli2的结构性活性突变体，在 胚胎和成人内皮细胞，并分析其对肠道发育和恢复的影响 肠子受伤。为了寻找抑制纤毛发生的其他因素，我建议进行转录组研究。 纤毛和非纤毛内皮细胞的分析。我还建议进行筛选，以识别小 可以在无纤毛的胚胎外内脏内皮干细胞(XEN)上诱导纤毛的分子，在一个 努力寻找能够诱导异位纤毛生成的治疗靶点，这可能被证明是非常有价值的 治疗各种发育障碍和癌症。

Project Summary/Abstract Contrary to the widely-held belief that nearly all vertebrate cells have primary cilia, our lab recently discovered that extra-embryonic visceral endoderm cells lack primary cilia. Our lab was also the first to demonstrate that primary cilia are required for all signaling through the Hedgehog pathway in mammals, which is consistent with data that extra-embryonic cells do not respond to Hedgehog ligands. However, extra-embryonic cells do secrete Hedgehog ligands, which are required for normal development of the embryo-derived vasculature of the placenta. I hypothesized that other cell types that make, but do not respond, to Hedgehog ligands may also lack cilia. Consistent with this hypothesis, I find that epithelia of a number of adult endodermal organs, which secrete but do not respond to Hedgehog ligands, lack primary cilia. However, I have also found that endodermal epithelia have primary cilia in early development. This primary cilia are present early in gut tube morphogenesis but are lost before adulthood, suggesting that dynamic signals from adjacent tissues can inhibit formation of primary cilia. In the proposed research, I will determine at what point primary cilia are lost in endodermal epithelium, which will allow me to identify the exogenous signals present at this time point. Expression of candidate factors will be manipulated to test whether primary cilia can be ectopically assembled in cells that would normally lack these organelles. I will also test the functional relevance the lack of autocrine hedgehog signaling in these tissue by expressing a constitutively active mutant of Gli2, the major Hedgehog transcriptional activator, in embryonic and adult endodermal epithelium, and assaying its effects on gut development and recovery from gut injury. To find additional factors responsible for inhibition of ciliogenesis, I propose to perform transcriptome analysis of ciliated and non-ciliated endodermal epithelium. I also propose to perform screens to identify small molecules that can induce cilia on non-ciliated extra embryonic visceral endodermal stem cells (XEN), in an effort to find therapeutic targets capable of inducing ectopic ciliogenesis, which may prove invaluable in treatment of a variety of developmental disorders and cancers.