Transcriptional control of collective cell migration

描述（由首席研究员提供）：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，细胞群变得极化，并以有序的方式一起移动。细胞群集体迁移的能力部分是由组织特异性转录输入决定的，这些转录输入决定了它们表达的基因的补体，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节因子如何控制定向集体细胞迁移的基本细胞过程。为此，将使用脊索动物海鞘的简化胚胎来研究心脏前中胚层细胞（称为“躯干腹侧细胞”（tvc））的迁移。tvc为活胚胎中定向集体细胞迁移提供了最简单的模型。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-后端（L-T）极性：前导TVC前缘较宽，突起比后端更明显。先前已经确定Mesp、成纤维细胞生长因子（FGF）信号和FoxF转录输入决定了tvc的迁移能力。提出的研究的具体目标是了解转录输入如何有助于领导TVC的规范和影响其突出活动。在初步观察的基础上，假设：(1)骨形态发生蛋白（BMP）的腹侧来源有助于下游靶基因的L-T极化表达，包括left /Tcf转录因子；(2)这些转录输入决定了Rho gtpase （RhoDF和Cdc42）的L-T极化活性，它们是形成膜突起所必需的。为了解决这些可能性，我们将首先分析left /Tcf在tvc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，定量成像方法将用于记录RhoDF、Cdc42的L-T极化和在野生型条件下以及转录调控因子和其他候选Rho gtpase操作后的突出活性，以试图表征它们对L-T极化膜突出的影响。最后，利用tvc特异性全基因组微阵列分析鉴定细胞集体极化和突起活性的候选调控效应物，然后分别通过原位杂交和靶向过表达进行系统表达和功能分析。期望通过表征转录调控因子，调控效应因子及其对Rho GTPase信号传导和肌动蛋白丝动力学的影响来确定集体细胞迁移的转录控制机制，而Rho GTPase信号传导和肌动蛋白丝动力学是集体细胞极化和膜突起形成的基础。

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.