Control of TGF-beta signals by Rab23 and Megf8 in mammalian left-right patterning

项目摘要 未能正确地规划人类胚胎身体计划的左右轴，导致了各种 出生缺陷，包括人类最常见的一类出生缺陷，先天性心脏病。 尽管身体规划的左右模式对人类健康很重要，但这种机制 人们对它的产生只有部分了解。哺乳动物的左右模式始于结节，在那里 对称性的破坏是通过活动纤毛的旋转而发生的，这会导致流体向左流动。上的单元格 左侧的节点感觉到这种不对称，并通过一个在很大程度上仍不清楚的过程做出反应， 通过向身体左侧的侧板中胚层(LPM)细胞发送信号。使用的信号是 转化生长因子-β家族成员Nodal和Gdf-1。在LPM中，节点诱导出自己的表达式，即 通过左侧LPM传播，以及其抑制物Lefty1和Lefty2的表达。激活 左侧LPM中的结节通路的位置驱动形态不对称 内脏器官以及器官的结构不对称(例如，心脏和肺)。 我们的数据表明，囊泡运输蛋白Rab23对于生产功能性 转化生长因子-β信号将信息传递到老鼠和斑马鱼的左侧LPM。我们的数据表明 Rab23在节点和LPM中都起作用，左侧节点的细胞可能对不对称 通过调节液体流动水平来调节Rab23的活性。在该模型下，主动Rab23流量节点和 GDF-1蛋白通过分泌途径，因此它们可以被加工并有效地释放到 左侧LPM中的信号单元。该模型的第二部分是本文提出的研究主题。 基于人类和小鼠的表型数据，我们假设这一过程还涉及 多个EGF重复蛋白Megf8。 根据目前的提议，一套三个目标将涉及这一模式的基本方面。 在第一个目标下，Rab23调节Nodal/GDF-1运输和分泌的机制将 使用显微镜和生化方法进行研究。在第二个目标下，我们将确定 Rab23和Nodal在胚胎中的定位。在最终目标下，我们将调查 假设Megf8与Rab23共同作用于控制转化生长因子-β的运输和分泌 Nodal和GDF-1信号。 总的来说，这些实验将为解决长期存在的问题提供基础 关于纤毛驱动流体流动形式的生物物理信息如何的左右图案场 翻译成不对称的基因表达，随后，不对称的形态发生 内脏。

Project Summary Failure to properly pattern the left-right axis of the human embryonic body plan results in a variety of birth defects, including the most common category of human birth defects, congenital heart defects. Despite the importance of left-right patterning of the body plan for human health, the mechanisms that generate it are only partially understood. Left-right patterning in mammals begins in the node where symmetry breaking occurs by the rotation of motile cilia, which causes a leftward fluid flow. Cells on the left side of the node sense this asymmetry and respond, through a process that remains largely unclear, by signaling to the lateral plate mesoderm (LPM) cells on the left side of the body. The signals used are the TGF-β family members Nodal and GDF-1. In the LPM, Nodal induces its own expression, which propagates through the left LPM, as well as the expression of its inhibitors, Lefty1 and Lefty2. Activation of the Nodal pathway in the left LPM drives morphological asymmetry with respect to the placement of visceral organs as well as structural asymmetries with an organ (e.g., the heart and lungs). Our data indicate the vesicle trafficking protein Rab23 is critical for the production of functional TGF-β signals that relay information to the left LPM in both mouse and zebrafish. Our data indicate that Rab23 functions in both the node and the LPM and that cells of the left node may respond to asymmetric fluid flow by regulating the level of Rab23 activity. Under this model, active Rab23 traffics Nodal and GDF-1 proteins through the secretory pathway so they may be processed and efficiently released to signal cells in the left LPM. This second part of the model is the subject of the research proposed here. Based on human and mouse phenotypic data, we hypothesize that this process also involves the multiple EGF-repeat protein Megf8. Under the current proposal, a set of three aims will address fundamental aspects of this model. Under the first aim, the mechanism by which Rab23 regulates Nodal/GDF-1 trafficking and secretion will be investigated using microscopy and biochemical approaches. Under the second aim, we will determin the localization of Rab23 and Nodal in the embryo proper. Under the final aim, we will investigate the hypothesis that Megf8 functions together with Rab23 to control trafficking and secretion of the TGF-beta signals Nodal and GDF-1. Collectively, these experiments will provide the basis for addressing the long-standing question in the left-right patterning field regarding how biophysical information in the form of cilium-driven fluid flow is translated into asymmetric gene expression and, subsequently, asymmetric morphogenesis of the internal organs.