Cell-cell communication mediated by fluid flows

项目摘要 细胞可以探测到通过其表面的外部流体流动。这样的流及其引发的流信令事件， 对器官发育、动态平衡、癌症传播和宿主-微生物相互作用至关重要，然而我们 对细胞如何感知和响应其水动力环境知之甚少。这项研究计划旨在 以揭示细胞通过流体流动进行交流的基本机制。具体地说，我们调查 细胞如何产生和感知液体流动，以及感觉细胞如何对流动衍生的信号做出反应。生成 关于流信号的新基础知识，我们使用成熟且高度易处理的左-右 斑马鱼胚胎图案化系统。在这个系统中，它容易受到遗传扰动以及 高分辨率和定量成像，流动信号抑制关键靶基因dand5的表达 感觉细胞。在我们之前的工作中，我们发现Pkd1l1，一个大的多囊藻膜蛋白，对 流动诱导的dand5抑制。在我们提议的工作的一个方面，我们将测试我们的假设，即Pkd1l1是 通过确定细胞类型以及亚细胞作用部位来确定流信号感觉组件，其中 Pkd1l1起着流信号下游的作用。我们还研究了机械敏感的功能作用。 Pkd1l1内的结构域在流调节信号中。在第二个项目中，我们解决了 了解流信号如何在感觉细胞内传递。这项工作将使用我们优化的 CRISPR发现和验证新的流量信号转导机制的方法，这将是 显著扩展我们对感官事件下游作用机制的理解 细胞内的信号。在第三个项目中，我们将使用已知的dand5转录后抑制 具体发生在流信号下游的左侧，作为生成新知识的模型 感官细胞如何对流动信号做出反应。这包括确定3‘非翻译区的作用和 RNA结合蛋白和microRNAs如何作用于流信号下游以抑制基因表达。这 这项工作将得到我们正在开发的量化流动信号通路输出的新型体内报告的帮助。 总体而言，这项研究将揭示细胞通过流信号进行通信的新机制。而当 化学信号转导级联反应(Hedgehog、Wnt等)都被广泛研究，流信号通路 代表着一个新的前沿，有许多基本原理等待着人们去发现。我们的工作使用了高度的 易于处理的斑马鱼左右图案系统将揭示新的流动信号感觉和 转导，以及细胞如何对流动信号作出反应。由于异常血流信号出现在许多疾病中 这将为改善人类健康提供新的机会。

Project Summary Cells can detect external fluid flows across their surface. Such flows, and the flow signaling events they induce, are critical for organ development, homeostasis, cancer dissemination, and host-microbe interactions, yet we know little about how cells sense and respond to their hydrodynamic environment. This research program aims to reveal fundamental mechanisms by which cells communicate through fluid flows. Specifically, we investigate how cells generate and sense fluid flows, and how sensory cells respond to flow-derived signals. To generate new fundamental knowledge about flow signaling, we use the well-established and highly tractable left-right patterning system of zebrafish embryos. In this system, which is amenable to genetic perturbation as well as high-resolution and quantitative imaging, a flow signal represses the expression of a key target gene, dand5, in sensory cells. In our prior work, we discovered that Pkd1l1, a large Polycystin membrane protein, is critical for flow-induced dand5 repression. In one aspect of our proposed work, we will test our hypothesis that Pkd1l1 is a flow signal sensory component by determining the cell types, as well as sub-cellular site of action, in which Pkd1l1 functions downstream of flow signals. We also investigate the functional role of mechanosensitive domains within Pkd1l1 in flow-regulated signaling. In a second project, we address the major gap in understanding of how flow signals are transduced within sensory cells. This work will use our optimized CRISPR approaches to discover and validate new flow signal transduction machinery, something which will markedly expand our understanding of the mechanisms acting downstream of sensory events to transduce signals intracellularly. In a third project, we will use the known post-transcriptional repression of dand5 that occurs specifically on the left side downstream of the flow signal as a model for generating new knowledge of how sensory cells respond to flow signals. This includes determining the role of the 3’untranslated region and how RNA-binding proteins and microRNAs act downstream of flow signals to repress gene expression. This work will be aided by novel in vivo reporters we are developing which quantify flow signaling pathway outputs. Overall, this research will uncover novel mechanisms by which cells communicate through flow signals. While chemical signal transduction cascades (Hedgehog, Wnt, etc.) are widely studied, flow signaling pathways represent a new frontier, with many fundamental principles waiting to be discovered. Our work using the highly tractable left-right patterning system of zebrafish will reveal new principles of flow signal sensation and transduction, as well as how cells respond to flow signals. Since aberrant flow signals occur in many disease states, this will open new opportunities to improve human health.