Elucidating the mechanisms underlying cell cycle regulation of invasive behavior

阐明侵袭行为的细胞周期调控机制

• 批准号: 10080739
• 负责人: David Matus
• 金额: $ 45.15万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-01-13 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10080739
• 关键词: Adopted; Alleles; Asthma; Basement membrane; Behavior; Biological Assay; Caenorhabditis elegans; Cardiac development; Cell Cycle; Cell Cycle Arrest; Cell Cycle Regulation; Cell division; Cells; Chromatin; Chromatin Remodeling Factor; Clustered Regularly Interspaced Short Palindromic Repeats; Cytoskeleton; Data; Defect; Development; Disease; Disease Progression; Disseminated Malignant Neoplasm; Embryonic Development; Epigenetic Process; Epithelial; Euchromatin; F-Actin; Failure; G1 Arrest; G1 Phase; Gene Expression; Genes; Genetic; Genetic Diseases; Genetic Transcription; Genome engineering; Genomic approach; Goals; Heterochromatin; Histone Deacetylase; Image; Immune System Diseases; Immunologic Surveillance; Invaded; Laboratories; Lead; Leukocytes; Lighting; Link; Location; Malignant Neoplasms; Matrix Metalloproteinases; Mediating; Methods; Mitotic; Molecular; Neoplasm Metastasis; Neural Crest; Nuclear; Nuclear Hormone Receptors; Organ; Outcome; Paper; Pathogenicity; Pathology; Phase; Phenotype; Pre-Eclampsia; Process; Publishing; RNA Interference; RNA interference screen; Research; Resolution; Rheumatoid Arthritis; Structure; Testing; Therapeutic; Tissues; Visual; Work; Zinc Fingers; base; cdc Genes; cell behavior; cell type; design; developmental disease; developmental genetics; experimental study; functional genomics; gastrulation; genetic approach; genomic locus; high throughput screening; implantation; improved; in vivo; in vivo Model; innovation; insight; migration; natural Blastocyst Implantation; neoplastic cell; prevent; public health relevance; screening; stem; therapeutically effective; tool; transcription factor; transgene expression; trophoblast

Project Summary/Abstract Cell invasion through the basement membrane is a key mechanism underlying cell dispersal and organ formation during normal development, immune surveillance and is dysregulated during cancer metastasis. Yet due to difficulties of studying these dynamic behaviors in vivo, it is the least understood aspect of the metastatic cascade. My laboratory utilizes a powerful in vivo model to examine cell invasive behavior, by combining functional genomic and genetic tools with single-cell visual analyses. We examine anchor cell (AC) invasion into the vulval epithelium during C. elegans larval development. Data from our laboratory has shown a functional link between G1 phase cell cycle arrest and the acquisition of an invasive phenotype. Through high- throughput screens, we have identified that the activity of a single conserved NR2E1 class nuclear hormone receptor, the transcription factor, nhr-67 (TLX), is required to maintain the invasive AC in G1 cell cycle arrest. Loss of nhr-67 results in mitotic ACs that fail to invade. Strikingly, AC invasion can be rescued by preventing cell division through induction of G1 cell cycle arrest. Downstream of G1 arrest, chromatin modifiers, including the histone deacetylase, hda-1, are required for expression of pro-invasive genes, including matrix metalloproteinases (MMPs) and regulators of the F-actin cytoskeleton, leading to differentiation of the invasive phenotype. For this proposal we plan to elucidate how cell cycle arrest is functionally linked to invasion. In Aim 1, using molecular epistatic interaction experiments and inducible transgene expression, we will identify the upstream network of transcription factors that mediate NHR-67 activity. In Aim 2, we will generate CRISPR/Cas9-mediated conditional alleles of cell cycle control genes to determine how the AC maintains G1 arrest. In Aim 3, we will pair tissue-specific RNAi screening of chromatin modifiers with high resolution confocal and structured illumination (SIM) imaging of sub-nuclear organization, to understand the link between invasion and differentiation. The results of these aims will provide the first mechanistic view of the transcriptional and epigenetic control of cell cycle arrest and invasive behavior. Thus, our proposed work has the potential to provide a mechanistic understanding of how cells acquire and maintain an invasive phenotype, a critical aspect of many developmental and genetic disorders.

项目摘要/摘要 细胞通过基底膜的侵袭是细胞扩散的关键机制。 和器官在正常发育过程中的形成，免疫监视和失调 在癌症转移过程中。然而，由于在活体内研究这些动态行为的文化不同，它是 转移级联反应最不为人所知的方面。我的实验室利用体内强大的 结合功能基因组和遗传工具检测细胞侵袭行为的模型 用单细胞视觉分析。我们检查锚细胞(AC)对外阴上皮的侵袭。 在线虫幼虫发育期间。我们实验室的数据显示了 在G1期细胞周期停滞和获得侵袭性表型之间。通过高中生- 吞吐量筛选，我们已经识别出fi单个保守的NR2E1类的活性 核激素受体，转录因子NHR-67(TLX)，是维持 侵袭性AC在G1期细胞周期阻滞中的作用NHR-67的缺失会导致有丝分裂的急性冠脉综合征无法侵袭。 值得注意的是，AC的侵袭可以通过诱导G1细胞来阻止细胞分裂来挽救 周期拘禁。在G1期停滞的下游，染色质Modifier，包括组蛋白脱乙酰酶， HDA-1是表达包括基质金属蛋白酶在内的促侵袭基因所必需的 (MMPs)和F-肌动蛋白细胞骨架调节因子，导致侵袭性细胞分化 表型。对于这项提议，我们计划阐明细胞周期停滞是如何在功能上与 入侵。在目标1中，利用分子上位相互作用实验和诱导转基因 表达，我们将确定介导NHR-67的上游转录因子网络 活动。在目标2中，我们将产生CRISPR/Cas9介导的细胞周期条件等位基因 控制基因，以确定AC如何维持G1期停滞。在目标3中，我们将配对组织物种fic 用高分辨率共聚焦和结构光筛选染色质Modifi的RNAi (SIM)亚核组织成像，以了解入侵和 差异化。这些目标的结果将提供fi第一个机械观点 转录和表观遗传对细胞周期停滞和侵袭行为的控制。因此，我们的 拟议的工作有可能提供对细胞如何获得 并保持侵袭性表型，这是许多发育和遗传的关键方面 精神错乱。