Transcriptional control of collective cell migration

描述(由首席调查员提供)：关键的生理和病理过程，如伤口愈合、新生血管生成和癌症转移，依赖于定向的集体细胞迁移，借此细胞群变得极化并以有序的方式一起移动。细胞组集体迁移的能力部分取决于特定于组织的转录输入，这些转录输入定义了它们所表达的基因的补充性，从而决定了它们的迁移能力。该项目的长期目标是了解组织特异性转录调控因子如何控制潜在的定向集体细胞迁移的基本细胞过程。为此，一种脊索动物的简化胚胎将被用来研究心脏前中胚层细胞的迁移，这种细胞被称为“躯干腹细胞”(TVCs)。TVCs提供了活体胚胎中定向集体细胞迁移的最简单的可能模型。在胚胎的每一侧，只有两个细胞一起迁移，并显示出与迁移方向一致的明显的前导-拖尾(L-T)极性：前导TVC显示出比拖尾更宽广的前沿和更明显的突起。先前已证实，MESP、成纤维细胞生长因子信号转导和FoxF转录输入决定血管内皮细胞的迁移能力。这项拟议研究的具体目标是了解转录输入如何影响领导者TVC的规格并影响其突出活动。在初步观察的基础上，将检验以下假设：(1)腹侧来源的骨形态发生蛋白有助于下游目标基因的L-T极化表达，包括Lef/Tcf转录因子；(2)这些转录输入决定了Rho GTP酶RhoDf和CDc42的L-T极化活性，这是膜突起形成所必需的。为了解决这些可能性，我们将首先分析Lef/Tcf在TVCs中的表达、顺式调控和功能，以了解L-T极化的转录输入。其次，我们将利用定量成像方法记录野生型条件下以及转录调节因子和其他候选Rho GTP酶作用后RhoDf、Cdc42和突起活性的L-T极化，以试图表征它们对L-T极化膜突起的影响。最后，将通过TVC特异性全基因组微阵列分析确定集体细胞极化和突起活动的候选调控效应器，然后分别通过原位杂交和靶向过度表达进行系统表达和功能分析。我们的期望是通过表征转录调控因子、受调控的效应物及其对Rho GTPase信号和肌动蛋白细丝动力学的影响来确定集体细胞迁移的转录控制机制。 公共卫生相关性：在包括人类在内的动物中，各种称为集体细胞迁移的形态发生过程对于胚胎发育、伤口愈合、新生血管形成和癌症转移等生理和病理现象至关重要。我们建议使用最简单的模型来研究控制定向集体细胞迁移的分子和细胞机制，该模型由两个细胞作为极化对在海洋无脊椎动物被囊状肠衣动物的胚胎中迁移组成。由于与脊椎动物在进化上的相似性，使用这一简单模型的研究将破译推动细胞集体迁移的基本机制，并可能应用于再生医学和预防癌症转移。

DESCRIPTION (provided by Principal Investigator): Critical physiological and pathological processes, such as wound healing, de novo vessel angiogenesis and cancer metastasis, rely on directed collective cell migrations, whereby groups of cells become polarized and move together in an orderly fashion. The ability of cell groups to migrate collectively is determined in part by the tissue-specific transcriptional inputs that define the complement of genes that they express 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 (L-T) 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. The specific goal of the proposed research is to understand how transcriptional inputs contribute to the specification of the leader TVC and influence its protrusive activity. On the basis of preliminary observations, the hypotheses that (1) a ventral source of bone morphogenetic proteins (BMP) contributes to the L-T polarized expression of downstream target genes, including the Lef/Tcf transcription factor; and (2) these transcriptional inputs determine the L-T polarized activities of the Rho GTPases RhoDF and Cdc42, which are required for the formation of membrane protrusions will be tested. To address these possibilities, the expression, cis-regulation and function of Lef/Tcf in the TVCs will first be analyzed in order to understand L-T polarized transcriptional inputs. Second, quantitative imaging methods will be used to document the L-T polarization of RhoDF, Cdc42 and protrusive activities in wild-type conditions and after manipulations of transcription regulators and other candidate Rho GTPases in an attempt to characterize their effects on L-T polarized membrane protrusions. Finally, candidate regulated effectors of collective cell polarization and protrusive activity will be identified using TVC-specific whole genome microarray analysis followed by systematic expression and functional analyses by in situ hybridization and targeted over-expression, respectively. The expectation is to determine a mechanism of transcriptional control for collective cell migration by characterizing the transcription regulators, the regulated effectors and their effects on the Rho GTPase signaling and actin filament dynamics that underlie collective cell polarization and the formation of membrane protrusions. PUBLIC HEALTH RELEVANCE: In animals, including humans, a variety of morphogenetic processes known as collective cell migrations are essential for physiological and pathological phenomena such as embryonic development, wound healing, de novo vessel formation and cancer metastasis. We propose to study the molecular and cellular mechanisms that control directed collective cell migration using the simplest possible model, which consists of two cells migrating as a polarized pair in embryos of the marine invertebrate, the tunicate Ciona intestinalis. Because of the evolutionary proximity with vertebrates, studies using this simple model will decipher the basic mechanisms driving collective cell migration with potential applications for regenerative medicine and the prevention of cancer metastasis.