Cell cycle regulation by C. elegans CUL-2 E3 complexes

线虫 CUL-2 E3 复合物的细胞周期调节

基本信息

批准号：
7570108
负责人：
EDWARD T. KIPREOS
金额：
$ 25.08万
依托单位：
UNIVERSITY OF GEORGIA
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-02-01 至 2011-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7570108
关键词：
Affinity Chromatography Alleles Anaphase Animals Binding Bypass CUL2 gene Caenorhabditis elegans Cancer Biology Cell Cycle Cell Cycle Regulation Cell Proliferation Cell physiology Chromosome Condensation Complex Coupled Cyclin B Cyclins Defect Development Ethylnitrosourea Event Foundations Genes Genetic Genetic Screening Goals Human In Vitro Individual Kidney Link Malignant Neoplasms Malignant neoplasm of central nervous system Mass Spectrum Analysis Mediating Meiosis Metaphase Mitosis Mitotic Molecular Mutagens Pathway interactions Pattern Phase Transition Phenotype Process Protein Analysis Protein Subunits Proteins Proteolysis Protocols documentation RNA Interference Regulation Research Role Temperature Tumor Suppressor Proteins Ubiquitin Ubiquitin-mediated Proteolysis Pathway Ubiquitination Von Hippel-Lindau Tumor Suppressor Protein anti-cancer therapeutic base beta catenin genetic analysis in vivo insight loss of function mutant novel overexpression prevent protein degradation protein function research study sex determination tumor tumor progression ubiquitin ligase yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): The long-term goal of this research is to understand cell cycle regulation in the context of animal development. Regulated proteolysis is central to the control of the cell cycle, and defects in ubquitin- mediated proteolysis have been linked to cancer. Cell cycle-dependent protein degradation occurs principally through the ubiquitin proteolytic pathway. The human CUL2 ubiquitin ligase functions as part of a tumor suppressor complex to prevent cancer, but its role in regulating the cell cycle has not been directly studied. In C. elegans, CUL-2 is required for multiple aspects of cell cycle regulation, including: the G1 to S phase transition; chromosome condensation; the meiosis II metaphase to anaphase transition; mitotic progression; and the degradation of mitotic cyclin B. CUL-2 has additional roles in the regulation of polarity and sex determination. CUL-2 functions as a central component of a multisubunit ubiquitin ligase that employs distinct substrate recognition subunits (SRSs). The identity of the SRSs that recognize substrates in several of the CUL-2-dependent cell cycle pathways are unknown, as are the critical substrates, whose ubiquitination allows the cell cycle events to occur. The experiments in this proposal will clarify the functions of the CUL-2 ubiquitin ligase in three major ways. First, SRSs and proteins that physically associate with the CUL-2 complex will be identified by immunoaffinity purification and two-hybrid screens. Interacting proteins will be characterized for cellular function and expression pattern. Second, the substrates that are ubiquitinated by CUL-2 complexes to regulate the cell cycle will be identified by a modified affinity purification protocol and two-hybrid screen. These substrates will be characterized to determine the CUL-2-dependent process in which they function and their role in that process. Finally, genetic screens for suppressors of CUL-2 complex SRS genes will identify interacting genes that either modulate CUL-2 complex activity, allow a bypass of CUL-2 function, or are critical substrates of CUL-2. Understanding the molecular pathways through which CUL-2 regulates the cell cycle will provide important insights into the control of fundamental cell cycle events. Cancer derives from unregulated cell proliferation, and a complete understanding of cell cycle control will provide a foundation for understanding cancer biology and developing anti-cancer therapeutics.

描述（由申请人提供）：这项研究的长期目标是在动物发育的背景下了解细胞周期调节。调节的蛋白水解是控制细胞周期的核心，紫bqu素介导的蛋白水解中的缺陷与癌症有关。细胞周期依赖性蛋白质降解主要通过泛素蛋白水解途径发生。人CUL2泛素连接酶是预防癌症的肿瘤抑制复合物的一部分，但尚未直接研究其在调节细胞周期中的作用。在秀丽隐杆线虫中，CUL-2对于细胞周期调节的多个方面需要CUL-2，包括：G1至S相变；染色体凝结；减数分裂II中期向后期过渡；有丝分裂进展；并且有丝分裂细胞周期蛋白B的降解在极性和性别确定的调节中具有额外的作用。 CUL-2充当使用不同底物识别亚基（SRSS）的多亚基泛素连接酶的中心成分。识别几个CUL-2依赖性细胞周期途径中底物的SRS的身份以及临界底物也未知，其泛素化允许发生细胞周期事件。该提案中的实验将以三种主要方式阐明CUL-2泛素连接酶的功能。首先，通过免疫亲和力纯化和两个杂交筛网来鉴定与CUL-2复合物物理相关的SRSS和蛋白质。相互作用的蛋白质将以细胞功能和表达模式为特征。其次，通过修改后的亲和力纯化方案和两杂交屏幕，将识别由CUL-2复合物泛素化的基板以调节细胞周期。这些底物将被表征以确定它们在该过程中发挥作用的CUL-2依赖性过程。最后，用于CUL-2复合物SRS基因抑制子的遗传筛选将识别调节CUL-2复合活性的相互作用基因，允许CUL-2功能旁路，或者是CUL-2的关键底物。了解CUL-2调节细胞周期的分子途径将为控制基本细胞周期事件的控制提供重要的见解。癌症来自不受管制的细胞增殖，对细胞周期控制的完全了解将为了解癌症生物学和发展抗癌治疗剂提供基础。