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

基本信息

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
项目类别：
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

项目摘要

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.

项目概要/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形成过程中的正常发育、免疫监视和失调然而，由于在体内研究这些动态行为的困难，转移级联反应中最不为人所知的方面我的实验室利用了强大的体内功能。通过结合功能基因组和遗传工具来检查细胞侵袭行为的模型通过单细胞视觉分析，我们检查了锚细胞 (AC) 侵入外阴上皮的情况。我们实验室的数据显示了线虫幼虫发育过程中的功能联系。 G1 期细胞周期停滞和侵袭性表型的获得之间通过高- 通过吞吐量筛选，我们发现单个保守 NR2E1 类的活性核激素受体，即转录因子 nhr-67 (TLX)，是维持 G1 细胞周期停滞中的侵袭性 AC 缺失 nhr-67 导致有丝分裂 AC 无法侵袭。引人注目的是，AC 入侵可以通过诱导 G1 细胞阻止细胞分裂来挽救 G1 停滞的下游，染色质修饰剂，包括组蛋白脱乙酰酶， hda-1，是促侵袭基因（包括基质金属蛋白酶）表达所必需的 (MMP) 和 F-肌动蛋白细胞骨架的调节因子，导致侵袭性细胞的分化对于这个提议，我们计划阐明细胞周期停滞如何与功能相关。在目标 1 中，使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 的转录因子的上游网络在目标 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 如何维持 G1 停滞在目标 3 中，我们将配对组织特异性。使用高分辨率共焦和结构照明对染色质修饰剂进行 RNAi 筛选 (SIM) 亚核组织成像，以了解侵袭和侵袭之间的联系这些目标的结果将提供第一个机械视图。细胞周期停滞和侵袭行为的转录和表观遗传控制。拟议的工作有可能提供对细胞如何获得的机制的理解并维持侵入性表型，这是许多发育和遗传的一个关键方面失调。