Cytokinesis staging mechanisms

细胞分裂分期机制

• 批准号: 10441374
• 负责人: ERIC Lyle WEISS
• 金额: $ 39.29万
• 依托单位: NORTHWESTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-12 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10441374
• 关键词: Affect; Affinity; Alleles; Amino Acid Motifs; Animals; Architecture; Binding; Biochemical; Biological Assay; Cell Line; Cell Proliferation; Cell division; Cells; Cellular Morphology; Complex; Crystallization; Cytokinesis; Daughter; Dependence; Docking; Elements; Enzymes; Eukaryotic Cell; Evaluation; Event; Excision; Failure; Family; Genome Stability; Goals; Growth; Human; Human Cell Line; Knowledge; Ligands; Mammalian Cell; Maps; Mechanics; Methods; Molecular Genetics; Morphogenesis; Morphology; Output; Pathway interactions; Peptides; Phage Display; Phosphorylation; Phosphotransferases; Process; Property; Protein Family; Protein Kinase; Proteins; Proteome; Regulation; Reporting; Research; Research Design; Saccharomycetales; Signal Pathway; Signal Transduction; Site; Specificity; Staging; Structure; Substrate Interaction; System; Time; Tissues; Work; Yeasts; analog; carcinogenesis; daughter cell; gain of function; genetic analysis; in vivo; interest; novel; protein aminoacid sequence; protein transport; trafficking

PROJECT SUMMARY Eukaryotic appropriately commence daughter carcinogenesis processes cytokinesis cell division requires dependency relationships in which late events start only after early ones are completed. During cytokinesis, for example, distinct large-scale morphological processes in precise and consistent order to accomplish architectural reorganization that produces two cells. Failure of mechanical division of cells can drastically affect genome stability, contributing to and other important human maladies. mechanisms that coordinate cytokinetic are incompletely understood, and a subject of intense research interest. Notably, machinery of and regulators currently known to control it are highly conserved. Working in budding yeast, we Regulatory have discovered a system that blocks late events of cytokinesis when early ones are delayed or defective, allowing time for corrective mechanisms to function. This “checkpoint” pathway, termed “Enforcement of Cytokinesis Order” (ECO), works by stabilizing a multi-motif protein that blocks a cytokinesis-specific secretion function of the Ndr/Lats protein kinase Cbk1, a key component of a highly conserved “Ndr-hippo” signaling system. Hippo signaling pathways are crucial regulators of morphogenesis, proliferation, and differentiation of eukaryotic cells; Ndr/Lats kinases are their downstream-most signaling components. Our prior research contributed crucial analysis for understanding of these systems. We were was among the first to describe specific functions of a hippo signaling system in vivo, defined the distinctive Ndr kinase phosphorylation motif, reported the first canonical substrate docking by an AGC-family protein kinase, and contributed the first and only crystal structure of an Ndr/Lats kinase bound to a Mob co-activator. This project seeks to determine how the checkpoint mechanism we discovered blocks secretion of late cytokinesis proteins when early stages of the process are defective. It will include exploration of in vivo docking interactions with cytokinesis regulators, assessment of secretion organization during cytokinesis, and evaluation of the importance of Ndr-hippo regulation of key effector proteins of cytokinesis. We will also explore the hypothesis that human Ndr-hippo pathways participate in control of late cytokinesis, using two different approaches. In one, we will determine if human Ndr kinases engage in docking interactions with in vivo phosphorylation targets, using a highly optimized phage display approach to identify possible peptide motifs that interact with Ndr kinase – Mob co-activator complexes. We will use a new ligand footprinting method we developed to map peptide motif associations. In another approach, we will determine if human Ndr-hippo signaling functions in cytokinesis by constructing cell lines expressing only analog sensitive Ndr kinases. This will allow rapid and reversible inhibition of these kinases in cells undergoing both normal and defective cytokinesis.

项目总结 真核生物 适当地 开始 女儿 致癌作用 流程 胞质分裂 细胞分裂需要依赖关系，在这种关系中，较晚的事件仅在较早的事件 完成。例如，在细胞质分裂过程中，不同的大规模形态过程 以精确和一致的顺序完成架构重组，从而产生两个 细胞。细胞机械分裂的失败会极大地影响基因组的稳定性，导致 以及其他重要的人类疾病。协调细胞运动的机制 都是不完全了解的，是一个引起强烈研究兴趣的主题。值得注意的是， 而目前已知的控制它的监管机构是高度保守的。在萌芽酵母中工作，我们 监管 已经发现了一种系统，当早期的细胞质分裂延迟或有缺陷时，它可以阻止细胞质分裂的晚期事件， 为纠正机制发挥作用留出时间。这条“检查点”路径被称为“强制执行 胞质分裂顺序“(ECO)通过稳定多基序蛋白来阻断胞质分裂特异性的分泌。 NDR/LATS蛋白激酶Cbk1的功能，它是高度保守的“NDR-河马”信号的关键组成部分 系统。 河马信号通路是河马细胞形态发生、增殖和分化的重要调节因子 真核细胞；NDR/LATS激酶是它们下游最重要的信号成分。我们之前的研究 为理解这些系统提供了至关重要的分析。我们是第一批描述 体内河马信号系统的特定功能，定义了独特的NDR激酶磷酸化基序， 报道了第一个通过AGC家族蛋白激酶对接的规范底物，并贡献了第一个和 只有与MOB共激活剂结合的NDR/LATS激酶的晶体结构。 这个项目试图确定我们最近发现的检查点机制是如何阻止分泌物的 当过程的早期阶段有缺陷时，胞质分裂蛋白。它将包括体内的探索 与胞质分裂调节剂的相互作用，胞质分裂期间分泌组织的评估，以及 评价NDR-HIPPO调节胞质分裂关键效应蛋白的重要性。我们还将探索 假设人类NDR-河马通路参与控制晚期胞质分裂，使用两种不同的 接近了。在一项研究中，我们将确定人类NDR激酶是否参与体内的对接相互作用 磷酸化靶标，使用高度优化的噬菌体展示方法识别可能的多肽基序 与NDR激酶-Mob共激活剂复合体相互作用。我们将使用一种新的配体足迹方法 为绘制多肽基序关联图而开发。在另一种方法中，我们将确定人类NDR-河马 通过构建仅表达模拟敏感的NDR激酶的细胞系，信号在胞质分裂中发挥作用。这 将允许在经历正常和缺陷的细胞中快速和可逆地抑制这些激酶 胞质分裂。