Transcriptional control of collective cell migration

项目总结/摘要 关键的生理和病理过程，如伤口愈合、血管形成和癌症 转移依赖于定向的集体细胞迁移，由此细胞群集体迁移和移动 有序地聚集在一起。细胞群体定向迁移的能力部分取决于细胞的生长。 组织特异性转录输入，定义表达基因的互补性，从而定义它们的能力 迁徙该项目的长期目标是了解组织特异性转录调节因子如何 控制基本的细胞过程，这些细胞过程是定向集体细胞迁移的基础。为此，简化的 一个脊索动物物种的胚胎，海鞘玻璃海鞘，将被用来研究迁移前， 心脏中胚层细胞，称为“躯干腹侧细胞”（TVC）。TVC提供了最简单的模型 在活的胚胎中指导集体细胞迁移。在胚胎的每一侧，只有两个细胞一起迁移 并显示与迁移方向一致的清晰的引导-尾随极性：引导TVC显示 宽的前缘和比拖车更明显的突起。此前已确定，梅斯普， 成纤维细胞生长因子（Fgf）信号传导和FoxF转录输入决定TVC的能力， 迁移。此外，TVC严格地在内胚层和外胚层之间迁移，这是TVC的标志。 中胚层心脏祖细胞已确定这些周围组织有助于疏通 TVC对集体极性和定向迁移的先天能动性。拟议研究的目标是 了解转录控制的内在TVC属性如何与外在信号相互作用， 决定集体极性和胚胎中的定向迁移。初步研究表明， 编码盘状结构域受体（Ddr）的蛋白在细胞中被Mesp、FGF和FoxF转录输入上调， TVC，在那里它促进与表皮的粘附。使用新开发的定量成像方法， 将分析控制Ddr表达、定位和活性的详细机制。的假设 Ddr和血管内皮生长因子受体（vascular endothelial growth factor receptor，VEGFR）之间的细胞自主拮抗作用 将测试迁移的TVC在表皮和内胚层之间的信号传导位置。初步 观察表明Ddr通过调节囊泡运输促进与表皮的粘附。的 Ddr在Rab 4/Rab 11阳性内体中起作用以促进整合素再循环至血浆的假设 膜将被测试。最后，集体迁移的受调控候选效应器的功能将是 使用TVC特异性CRISPR/Cas9介导的功能丧失测定和高含量的 表型分析控制亚细胞过程的生物分子网络的初步模型 TVC的基本行为将被建立。将特别关注Ddr的候选调节剂， Vegfr和整合素功能。该项目的完成将阐明系统的层次机制， 内在转录输入和外在信号来定义细胞特异性行为。

PROJECT SUMMARY / ABSTRACT Critical physiological and pathological processes, such as wound healing, blood vessel formation and cancer metastasis, rely on directed collective cell migrations, whereby groups of cells collectively polarize and move together in an orderly fashion. The ability of cell collectives to migrate directionally is determined in part by the tissue-specific transcriptional inputs that define the complement of expressed genes and thus their competence to migrate. The long-term goal of this project is to understand how tissue-specific transcription regulators control the basic cellular processes underlying directed collective cell migration. To this aim, the simplified embryos of a chordate species, the ascidian Ciona intestinalis, will be used to study the migration of pre- cardiac mesoderm cells, called “trunk ventral cells” (TVCs). The TVCs provide the simplest possible model of directed collective cell migration in live embryos. On each side of the embryo, only two cells migrate together and display a clear Leader-Trailer polarity aligned with the direction of migration: the leader TVC displays a broad leading edge and more conspicuous protrusions than the trailer. It was previously established that Mesp, Fibroblast growth factor (Fgf) signaling and FoxF transcriptional inputs determine the ability of TVCs to migrate. Moreover, TVCs migrate strictly between the endodermal and ectodermal germ layers, a hallmark of mesodermal cardiac progenitors. It was determine that these surrounding tissues contribute to canalizing TVCs' innate motility towards collective polarity and directed migration. The goal of the proposed research is to understand how transcriptionally-controlled intrinsic TVC properties interface with extrinsic signals to determine collective polarity and directed migration in the embryo. Preliminary studies suggested that the gene encoding the discoidin domain receptor (Ddr) is upregulated by Mesp, FGF and FoxF transcriptional inputs in the TVCs, where it promotes adhesion to the epidermis. Using newly developed quantitative imaging methods, the detailed mechanisms controlling Ddr expression, localization and activity will be analyzed. The hypothesis that a cell-autonomous antagonism between Ddr and vascular endothelial growth factor receptor (Vegfr) signaling positions the migrating TVCs between the epidermis and endoderm will be tested. Preliminary observations suggest that Ddr promotes adhesion to the epidermis by regulating vesicle trafficking. The hypothesis that Ddr acts in Rab4/Rab11-positive endosomes to promote the recycling of integrins to the plasma membrane will be tested. Finally, the functions of regulated candidate effectors of collective migration will be studied extensively using TVC-specific CRISPR/Cas9-mediated loss-of-function assays and high-content phenotypic analyses. A provisional model of the biomolecular network controlling the subcellular processes underlying TVC behavior will be built. Particular attention will be paid to the candidate modulators of Ddr, Vegfr and integrin functions. Completion of this project will illuminate the systems' level mechanisms linking intrinsic transcriptional inputs and extrinsic signals to define cell-specific behaviors.