Regulation of Quiescence in Eukaryotic Cells

真核细胞静止的调节

基本信息

批准号：
8563047
负责人：
David Gresham
金额：
$ 29.43万
依托单位：
NEW YORK UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-01 至 2018-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8563047
关键词：
Antineoplastic Agents Biological Assay Candida albicans Carbon Cell Cycle Cell division Cell physiology Cells Complement Computing Methodologies Conflict (Psychology)Coupled Cryptococcus neoformans Cyclic AMP-Dependent Protein Kinases Defect Development Diabetes Mellitus Eukaryota Eukaryotic Cell Failure Fission Yeast Gene Expression Profile Gene Expression Regulation Genes Genetic Genetic Programming Goals Growth Health Human Image Infection Inflammation Institutes Label Laboratories Libraries Malignant Neoplasms Maps Measures Messenger RNA Metabolic Methods Microbe Mitosis Modeling New York Nitrogen Nutrient Organism Pathway interactions Phosphorus Physiological Play Post-Transcriptional Regulation Process Production Protein Kinase Proteins RNA Recurrence Regulation Resistance Role Saccharomyces cerevisiae Saccharomycetales Science Signal Pathway Signal Transduction Starvation Testing Transcript Translations Universities Variant Yeast Model System base cell growth chemotherapy clinically relevant drug standard experience gene interaction genetic analysis genome-wide genome-wide analysis human disease in vivo insight loss of function mutation mRNA Stability mRNA Transcript Degradation microbial mutant neoplastic cell novel therapeutics pathogen programs ras Proteins research study response tool transcriptome sequencing tumor tumor microenvironment

项目摘要

DESCRIPTION (provided by applicant): Regulated exit from cell division and initiation of a non-proliferative quiescent state is a criticl requirement in all organisms. Failure to maintain quiescence and inappropriate reinitiation of proliferative cell growth underlies many human cancers. Conversely, subpopulations of quiescent tumor cells may play critical roles in resistance to chemotherapy and tumor recurrence as cancer drugs typically target processes active during cell growth. Similarly, quiescent pathogenic microbes are frequently insensitive to standard drug treatments. We will use the single-celled eukaryotic microbes, Saccharomyces cerevisiae (budding yeast) and Schizosaccharomyces pombe (fission yeast) to identify the conserved networks that regulate cell quiescence. Microbes and some tumor cells enter quiescent states in response to nutrient depletion and are able to survive for prolonged periods of nutrient starvation. Our preliminary studies demonstrate that initiation of quiescence in response to defined nutrient starvation is actively regulated by conserved signaling pathways including the TORC1, Ras/Protein kinase A (PKA) and AMPK pathways. In Aim 1 we will define the conserved genetic program that controls cell quiescence by quantifying the defect in quiescence attributable to loss of function mutations in each gene in both budding and fission yeast in three quiescence-inducing conditions: carbon, nitrogen and phosphorous starvation. We will complement this genetic approach with studies of the phenotypic hallmarks of quiescence in wildtype and mutant cells to identify processes defective in quiescent mutants. In Aim 2 we will study how signaling pathways integrate environmental information to initiate the quiescence program by identifying targets of quiescence-regulating pathways and interactions between pathways using genome-wide genetic interaction mapping in quiescent conditions in both species. These experiments will allow us to identify conserved functional interactions that enable the cell to initiate quiescence n response to specific pro-quiescence signals while simultaneously receiving pro-growth signals that activate parallel pathways. We hypothesize that one means of coordinating signaling pathways is by dynamic subcellular localization of their components and we will test this hypothesis using mutants in which signaling components are mislocalized. In Aim 3 we will quantify variation in mRNA synthesis and degradation rates as cells enter quiescence using in vivo metabolic labeling of mRNAs coupled with RNA-Seq. We will use this method to test whether cells alter the stability of specific transcripts as cell growth slows and they enter quiescence. We will then identify conserved determinants of mRNA degradation variation using computational methods. By focusing on conserved signaling pathways and cellular processes that regulate quiescence we will enhance our understanding of quiescence in both normal and diseased human cells as well as microbial pathogens. A detailed understanding of cell quiescence will ultimately enable new therapeutic strategies that specifically target quiescent cells in a variety of pathological settings including cancer and microbial infections.

描述（由申请人提供）：调节细胞分裂的退出和非增殖静止状态的启动是所有生物体的关键要求。未能维持静止状态和不适当地重新启动增殖细胞生长是许多人类癌症的根源。相反，静止肿瘤细胞亚群可能在化疗耐药和肿瘤复发中发挥关键作用，因为癌症药物通常针对细胞生长期间活跃的过程。同样，静止的病原微生物通常对标准药物治疗不敏感。我们将使用单细胞真核微生物酿酒酵母（芽殖酵母）和粟酒裂殖酵母（裂殖酵母）来识别调节细胞静止的保守网络。微生物和一些肿瘤细胞因营养耗尽而进入静止状态，并且能够在长时间的营养饥饿中生存。我们的初步研究表明，响应特定营养饥饿而启动的静止是由保守的信号通路主动调节的，包括 TORC1、Ras/蛋白激酶 A (PKA) 和 AMPK 通路。在目标 1 中，我们将定义保守的遗传程序，通过量化在三种静止诱导条件（碳、氮和磷饥饿）下出芽和裂殖酵母中每个基因的功能突变丧失引起的静止缺陷来控制细胞静止。我们将通过研究野生型和突变细胞的静止表型特征来补充这种遗传方法，以识别静止突变体中缺陷的过程。在目标 2 中，我们将研究信号通路如何整合环境信息来启动静止程序，方法是使用两个物种静止条件下的全基因组遗传相互作用图谱来识别静止调节通路的目标和通路之间的相互作用。这些实验将使我们能够识别保守的功能相互作用，使细胞能够启动对特定促静止信号的静止和反应，同时接收激活平行通路的促生长信号。我们假设协调信号通路的一种方法是通过其组件的动态亚细胞定位，并且我们将使用信号组件错误定位的突变体来测试这一假设。在目标 3 中，我们将使用 mRNA 体内代谢标记结合 RNA-Seq 来量化当细胞进入静止状态时 mRNA 合成和降解率的变化。我们将使用这种方法来测试当细胞生长减慢并进入静止状态时，细胞是否会改变特定转录本的稳定性。然后我们将使用计算方法确定 mRNA 降解变异的保守决定因素。通过关注调节静止的保守信号通路和细胞过程，我们将增强对正常和患病人类细胞以及微生物病原体静止的理解。对细胞静止的详细了解最终将有助于制定新的治疗策略，专门针对各种病理环境中的静止细胞，包括癌症和微生物感染。