Transcriptional regulation of morphogenetic behavior and invasive cell fate specification in C. elegans

秀丽隐杆线虫形态发生行为和侵袭细胞命运规范的转录调控

基本信息

批准号：
9980182
负责人：
Taylor Kinney
金额：
$ 3.9万
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：
2019
资助国家：
美国
起止时间：
2019-07-20 至 2022-07-19
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9980182
关键词：
Address BHLH Protein Basement membrane Behavior Binding Sites Biological Models Caenorhabditis elegans Cell Cycle Cell Cycle Arrest Cell Cycle Regulation Cell Differentiation process Cell Proliferation Cells Chemicals Congenital Abnormality Connective Tissue Cytoskeleton Data Development Disease EVI1 gene Epithelial Epithelium Event Extracellular Matrix Extracellular Matrix Proteins F-Actin Fertility Fingers Genes Genetic Screening Genetic Transcription Genomic approach Goals Gonadal structure Green Fluorescent Proteins HELB gene Hermaphroditism Human Immune System Diseases Immune response Immunity Infection Injury Invaded Knowledge Leukocytes Literature Malignant - descriptor Malignant Neoplasms Matrix Metalloproteinases Modeling Morphogenesis Neoplasm Metastasis Neural Crest Nuclear Hormone Receptors Nuclear Localization Signal Orthologous Gene Outcome Pathology Pathway interactions Pattern Phenotype Pre-Eclampsia Pregnancy Process Proteins Publishing Puncture procedure RARB gene RNA Interference Regulation Regulator Genes Regulatory Element Research Resolution Role Signal Transduction Specific qualifier value System Testing Therapeutic Tissues Transcriptional Regulation Tubal Pregnancy Up-Regulation Uterus Visual Work Zinc Fingers acronyms bZIP Domain base cell behavior cell fate specification egg genetic approach human DNA improved in vivo notch protein novel programs promoter reverse genetics sensor sex therapy development tool transcription factor translational impact trophoblast

项目摘要

PROJECT SUMMARY / ABSTRACT The ability of cells to traverse basement membranes (BMs) is a key part of fertility, development, immunity, and disease. BM invasion is facilitated through expression of extracellular matrix proteins, upregulation of matrix metalloproteinases, polarization of the F-actin cytoskeleton, and cell cycle arrest. Precise coordination of these pro-invasive programs is largely achieved through transcriptional regulation; however, our understanding of the gene regulatory networks (GRNs) involved is limited due to the lack of model systems in which cell invasion can be visualized live. Here, I propose to fill this gap in knowledge by utilizing morphogenesis of the Caenorhabditis elegans uterine-vulval connection as a tractable and visually amenable model to examine cell invasion in vivo. During development of the hermaphroditic somatic gonad, a specialized uterine cell called the anchor cell (AC) invades through the underlying BM to connect the uterus to the vulval epithelium. The AC itself is specified in a cell fate decision event earlier in development, in which two initially equipotent cells diverge via stochastic Notch asymmetry, giving rise to the presumptive AC and a proliferative ventral uterine (VU) cell. Prior research by our lab and others has identified six transcription factors (TFs) that regulate AC invasion. These include the basic leucine zipper TF fos-1 (Fos), the basic helix-loop-helix TF hlh-2 (E/Daughterless), two nuclear hormone receptors, nhr-67 (NR2E1/Tailless/TLX) and sex-1 (RARB/NR1B2), as well as two zinc-finger TFs, egl-43 (EVI1/MEL1) and mep-1. These TFs appear to be functioning in at least three distinct GRN sub-circuits to regulate AC invasion, one of which involves NHR-67, which functions upstream of CKI-1 (p21/p27) to induce G1 cell cycle arrest. Remarkably, five of the six pro-invasive TFs function reiteratively during the AC/VU cell fate decision. These include the three TFs comprising the NHR-67/cell cycle-dependent pro-invasive pathway (EGL- 43S, MEP-1, and NHR-67), as well as HLH-2 and SEX-1, which have predicted binding sites within the nhr-67 promoter. Thus, based on the literature and my preliminary studies, my central hypothesis is that the AC invasive program is dependent on the function of multiple GRN sub-circuits, one of which modulates cell cycle arrest and is reiteratively used in AC fate specification. In Aim 1 of this project, I will dissect the cis- and trans-regulation of AC invasion, focusing on the cell cycle-dependent GRN sub-circuit involving the pro-invasive TF nhr-67/TLX. In Aim 2, I will examine the roles of pro-invasive TFs that reiteratively function in AC specification and investigate if cell cycle control is the common denominator underlying these two processes. Cutting-edge functional tools, including an endogenous protein depletion system and a novel cell cycle state sensor, paired with the ability to perform high-resolution subcellular visual analyses, will allow for thorough and rigorous testing of this hypothesis.

项目概要/摘要细胞穿过基底膜 (BM) 的能力是生育、发育、免疫和发育的关键部分。疾病。通过细胞外基质蛋白的表达、基质的上调促进骨髓侵袭金属蛋白酶、F-肌动蛋白细胞骨架的极化和细胞周期停滞。这些的精确协调促侵入性程序主要是通过转录调控来实现的；然而，我们对由于缺乏细胞入侵的模型系统，所涉及的基因调控网络（GRN）有限。实时可视化。在这里，我建议通过利用秀丽隐杆线虫的形态发生来填补这一知识空白线虫子宫-外阴连接作为一种易于处理且视觉上易于接受的模型来检查体内细胞侵袭。在雌雄同体体细胞性腺的发育过程中，一种称为锚细胞（AC）的特殊子宫细胞通过下面的骨髓侵入，将子宫连接到外阴上皮。 AC 本身在一个中指定发育早期的细胞命运决定事件，其中两个最初等能的细胞通过随机缺口发生分歧不对称性，产生假定的 AC 和增殖性腹侧子宫 (VU) 细胞。我们之前的研究实验室和其他人已经确定了六种调节 AC 入侵的转录因子 (TF)。这些包括基本的亮氨酸拉链 TF fos-1 (Fos)、基本螺旋-环-螺旋 TF hlh-2 (E/Daughterless)、二核激素受体 nhr-67 (NR2E1/Tailless/TLX) 和 sex-1 (RARB/NR1B2)，以及两个锌指 TF，egl-43 (EVI1/MEL1) 和 mep-1。这些 TF 似乎在至少三个不同的 GRN 子电路中发挥作用调节 AC 侵袭，其中之一涉及 NHR-67，它在 CKI-1 (p21/p27) 上游发挥作用，诱导 G1 细胞周期停滞。值得注意的是，六个促侵袭性 TF 中的五个在 AC/VU 细胞命运期间反复发挥作用决定。其中包括构成 NHR-67/细胞周期依赖性促侵袭途径 (EGL- 43S、MEP-1 和 NHR-67)，以及 HLH-2 和 SEX-1，它们预测了 nhr-67 内的结合位点发起人。因此，根据文献和我的初步研究，我的中心假设是交流侵入性该程序依赖于多个 GRN 子电路的功能，其中之一调节细胞周期停滞和在 AC 命运规范中反复使用。在这个项目的目标 1 中，我将剖析顺式和反式监管 AC 侵袭，重点关注涉及促侵袭性 TF nhr-67/TLX 的细胞周期依赖性 GRN 子回路。在目标 2，我将研究在 AC 规范中反复发挥作用的促侵入性 TF 的作用，并研究如果细胞周期控制是这两个过程的共同点。尖端的功能工具，包括内源性蛋白质消耗系统和新型细胞周期状态传感器，并具有进行高分辨率亚细胞视觉分析，将允许对这一假设进行彻底和严格的测试。