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: 9919976
• 负责人: David Matus
• 金额: $ 2.58万
• 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 2021-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9919976
• 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; Epithelium; 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; Treatment Efficacy; Visual; Work; Zinc Fingers; base; cdc Genes; cell behavior; cell type; design; developmental disease; developmental genetics; experimental study; functional genomics; gastrulation; genetic approach; high throughput screening; implantation; improved; in vivo; in vivo Model; innovation; insight; migration; natural Blastocyst Implantation; neoplastic cell; prevent; public health relevance; screening; stem; 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 细胞通过基底膜侵袭是细胞扩散基础的关键机制 在正常发育，免疫监视期间形成器官和器官的形成失调 在癌症转移期间。然而，由于在体内研究这些动态行为的困难，这是 转移性级联反应最少的方面。我的实验室利用一个强大的体内 通过结合功能基因组和遗传工具来检查细胞侵入性行为的模型 带有单细胞视觉分析。我们检查锚固细胞（AC）入侵到外皮上皮 在秀丽隐杆线虫幼虫发育期间。我们实验室的数据显示了功能链接 在G1相细胞周期停滞与侵入性表型的获取之间。通过高 - 吞吐量屏幕，我们已经确定了单个构成NR2E1类的活动 核骑马受体，转录因子NHR-67（TLX），需要维持 G1细胞周期停滞中的侵入性交流。 NHR-67的损失导致无法入侵的有丝分裂AC。 引人注目的是，可以通过诱导G1细胞来防止细胞分裂来挽救AC侵袭 周期逮捕。 G1停滞的下游，染色质修饰器，包括组蛋白脱乙酰基酶， HDA-1是促侵入性基因的表达所必需的，包括基质金属蛋白酶 （MMP）和F-肌动蛋白细胞骨架的调节剂，导致侵入性的分化 表型。对于此提案，我们计划阐明细胞周期停滞在功能上如何链接 入侵。在AIM 1中，使用分子上皮相互作用实验和诱导转化 表达，我们将确定介导NHR-67的转录因子的上游网络 活动。在AIM 2中，我们将生成CRISPR/CAS9介导的细胞周期条件等位基因 控制基因以确定AC如何保持G1停滞。在AIM 3中，我们将配对组织特异性 具有高分辨率和结构化照明的染色质修饰者的RNAi筛选 （SIM）亚核组织的成像，以了解入侵与 分化。这些目标的结果将提供第一个机械观点 细胞周期停滞和侵入性行为的转录和表观遗传控制。那，我们的 拟议的工作有可能提供对细胞如何获得的机械理解 并保持侵入性表型，这是许多发育和遗传的关键方面 疾病。