Understanding developmental patterning's influence on morphogenesis

了解发育模式对形态发生的影响

• 批准号: 10389165
• 负责人: Emily Bowie
• 金额: $ 6.76万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10389165
• 关键词: Actins; Actomyosin; Address; Affect; Apical; Area; Biochemical; Biological; Brain; Caenorhabditis elegans; Candidate Disease Gene; Cell Polarity; Cell Shape; Cells; Cellular biology; Complement; Congenital Abnormality; Contracts; Cytoskeleton; Data Set; Defect; Development; Embryo; Endoderm; Event; Failure; Future; Genes; Genetic; Genetic Transcription; Goals; Human; Imaging Techniques; Knowledge; Lead; Link; Live Birth; Mass Spectrum Analysis; Modeling; Molecular; Morphogenesis; Myosin ATPase; Nematoda; Neural Tube Closure; Neural Tube Defects; Neural tube; Nonmuscle Myosin Type IIA; Optics; Organism; Pattern; Phosphotransferases; Principal Investigator; Process; Proteins; RNA Interference; Regulation; Reproducibility; Role; Shapes; Side; Spinal Cord; Surface; Testing; Time; Tissues; Training; Transcript; Translating; United States; Up-Regulation; Work; biochemical tools; career; cell cortex; cell fate specification; constriction; gastrulation; insight; knock-down; mutant; neuroepithelium; novel; peroxisome; precursor cell; recruit; response; screening; spatiotemporal; success; transcription factor; transcriptome; transcriptome sequencing

Abstract Morphogenetic events utilize precisely timed changes in cell shape. One of the fundamental mechanisms cells use to change their shape is apical constriction. Apical constriction relies on the contraction of cortical actomyosin networks that causes the apical side of a cell to shrink, resulting in tissue morphogenesis. In humans, apical constriction aids the internalization of the future spinal cord and brain in a process known as neural tube closure. Failure of apical constriction can lead to neural tube defects, which accounts for birth defects in 1 out of every 3,000 live births. Therefore, uncovering the processes that govern apical constriction will advance our understanding of basic mechanisms underlying cell shape changes, causes, and potential treatments for neural tube defects. Despite current knowledge of developmental patterning of apical constriction, precise genetic mechanisms that govern which cells undergo apical constriction, how the apical surface is determined, and when to constrict, remain only partially understood. I plan to use Caenorhabditis elegans (C. elegans) gastrulation, a morphogenetic event driven by apical constriction, to address these issues. Gastrulation in C. elegans starts with the internalization of the two endodermal precursor cells (EPCs), which depend on the spatial and temporal precision of the expression of cell fate specification factors end-1 and end-3. However, mechanistic links between end-1,3 and the resulting apical constriction remain largely unknown. Using the genetically tractable and optically clear C. elegans, I plan to dissect the cellular mechanisms that translate developmental patterning into specific, localized, and precisely timed cell shape changes. Comparing the transcriptome of wild-type and end-3 null embryos, I identified thirty target genes whose expression depends on end-3. After screening these genes, I identified ten new genes that contribute to C. elegans gastrulation. In Aim 1, I will use a variety of cell biological approaches to identify the mechanisms by which some of these genes couple developmental patterning to changing cell shape. Aim 2 focuses on the myosin-activating kinase MRCK-1 localizes to the apical cell cortex of EPCs and is required for apical constriction. MRCK-1 is dependent on end-1,3 expression and becomes localized apically specifically in only EPCs despite MRCK-1 being present at similar levels in all cells. I will use MRCK-1 localization as a molecular foothold for understanding how a pivotal protein becomes recruited to the apical cortex in only certain cells. Aim 2 will further investigate which domains of MRCK-1 are required for this localization pattern and identify interactors with these domains that function to initiate apical constriction, to better connect cell fate regulators and intracellular localization of a key protein. Overall, I propose the use of genetic, biochemical, and imaging techniques to advance our understanding of how transcriptional networks and other developmental patterning inputs deploy localized factors that influence cell shape changes.

摘要 形态发生事件利用细胞形状的精确定时变化。其中一个基本原则 细胞用来改变形状的机制是顶端收缩。根尖收缩依赖于收缩 一种皮质肌球蛋白网络，导致细胞顶端收缩，导致组织 形态发生。在人类中，心尖收缩有助于未来脊髓和大脑的内化 这一过程被称为神经管关闭。心尖收缩失败可导致神经管缺陷， 每3,000名活产儿中就有1人患有出生缺陷。因此，揭示治理的过程 顶端收缩将促进我们对细胞形状变化的基本机制的理解， 神经管缺陷的原因和可能的治疗方法。 尽管目前对根尖狭窄的发育模式有了解，但精确的遗传学 控制哪些细胞经历根尖收缩、如何确定根尖表面的机制，以及 什么时候该收紧，只有一部分人了解。我打算用线虫(线虫) 原肠形成，一种由顶端收缩驱动的形态发生事件，以解决这些问题。原肠胚在C. 优雅始于两个内皮祖细胞(EPC)的内化，这依赖于 细胞命运指定因子END-1和END-3表达的空间和时间精度。然而， 末端1，3和由此产生的根尖收缩之间的机械联系在很大程度上仍不清楚。使用 从基因上易驯化和光学上透明的线虫，我计划剖析翻译的细胞机制 发育模式转变为特定的、局部的和精确计时的细胞形状变化。比较一下 野生型和end-3缺失胚胎的转录组，我确定了30个目标基因，它们的表达取决于 在第三端。在筛选了这些基因后，我确定了10个与线虫原肠形成有关的新基因。在……里面 目标1，我将使用各种细胞生物学方法来确定其中一些 基因将发育模式与细胞形状的变化联系在一起。 目的2重点研究肌球蛋白激活激酶MRCK-1定位于内皮祖细胞顶端细胞皮质和 是根尖收缩所必需的。MRCK-1依赖于end-1，3的表达，并在顶端定位 具体地说，尽管MRCK-1在所有细胞中的表达水平相似，但仅在EPC中存在。我要用MRCK-1 定位作为了解关键蛋白如何被招募到根尖的分子立足点 皮质仅存在于某些细胞中。目标2将进一步调查这需要MRCK-1的哪些结构域 定位模式，并确定与这些结构域的相互作用，以启动根尖收缩，以 更好地连接细胞命运调节器和细胞内一种关键蛋白的定位。总体而言，我建议使用 遗传、生化和成像技术，以促进我们对转录网络如何 而其他发育模式的输入采用了影响细胞形状变化的局部因素。