Role of Inositol Polyphosphates in Kinetochore Function and Transcription

肌醇多磷酸在着丝粒功能和转录中的作用

基本信息

批准号：
7011303
负责人：
Ales Vancura
金额：
$ 24.75万
依托单位：
ST. JOHN'S UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-01-01 至 2009-12-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The long-term goal of our laboratory is to identify and characterize eukaryotic cellular processes regulated by phospholipase C (PLC), an enzyme which plays vital roles in signal transduction pathways. PLC hydrolyzes phosphatidylinositol 4,5-bisphosphate [Ptdlns(4,5)P2] to produce two important second messengers: inositol 1,4,5-trisphosphate [lns(1,4,5)P3] and diacylglycerol (DAG). Since Plc1p (encoded by PLC1 gene) is the only PLC enzyme in the budding yeast Saccharomyces cerevisiae and hydrolysis of Ptdlns(4,5)P2 is the only pathway for synthesis of lns(1,4,5)P3 and other inositol polyphosphates (InsPs), cells with deletion of PLC1 gene (plc1-delta) are completely devoid of all InsPs and S. cerevisiae thus represents an ideal system in which to study metabolism and cellular roles of InsPs. The central hypothesis of this proposal is that InsPs play important role(s) in nuclear events, such as chromosome segregation/cell cycle progression and transcriptional regulation. Our results demonstrate that plc1-delta cells display alterations in the structure of core centromeric chromatin, higher frequency of chromosome loss, and mitotic delay (Lin et al., 2000; DeLillo et al., 2003). The mechanism of Plc1p's involvement in kinetochore function, not apparent at first, was suggested by recent work that showed that InsPs regulate the activity of chromatin remodeling complexes (Shen et al., 2003; Steger et al., 2003) and that the RSC complex, one of the chromatin remodelers, associates with centromeric loci and is important for high fidelity chromosome segregation (Hsu et al., 2003). The Specific Aim 1 of this proposal will test the hypothesis that Plc1p and InsPs affect recruitment, integrity, or other aspects of RSC function at the kinetochore. Genetic experiments and genome-wide expression analysis indicate that in addition to chromatin remodeling, InsPs affect transcription also by a mechanism independent of chromatin remodeling complexes. The Specific Aim 2 will determine whether Plc1p and InsPs affect transcriptional regulation of stress-induced genes by affecting recruitment of Msn2p/Msn4p activators, Tup1p/Ssn6p represser complex, or Srb10p component of the Srb/mediator complex. The Specific Aim 3 will utilize the well-studied GAL1 promoter as a model and determine, why plc1-delta cells fail to express GAL1 gene. The focus of this proposal is to elucidate the role of Plc1p and InsPs in two very important aspects of cell physiology: chromosome segregation/cell cycle progression and transcriptional regulation, processes that are misregulated in human disease, including cancer. The results of this work will ultimately contribute to identification of novel molecular targets for cancer therapy.

描述（由申请人提供）：我们实验室的长期目标是识别和表征由磷脂酶C（PLC）调节的真核细胞过程，磷脂酶C（PLC）是一种在信号传输途径中起着至关重要的作用的酶。 PLC水解磷脂酰肌醇4,5-双磷酸[PTDLNS（4,5）P2]产生两个重要的第二个使者：肌醇1,4,5-三磷酸[LNS（1,4,5）P3]和二酰基甘油（DAG）。 Since Plc1p (encoded by PLC1 gene) is the only PLC enzyme in the budding yeast Saccharomyces cerevisiae and hydrolysis of Ptdlns(4,5)P2 is the only pathway for synthesis of lns(1,4,5)P3 and other inositol polyphosphates (InsPs), cells with deletion of PLC1 gene (plc1-delta) are completely因此，没有所有Insps和酿酒酵母，代表了一个理想的系统，在其中研究了INSP的新陈代谢和细胞作用。该提案的中心假设是INSP在核事件中起重要作用，例如染色体隔离/细胞周期进程和转录调控。我们的结果表明，PLC1-DELTA细胞在核心centromeric染色质，较高的染色体损失频率和有丝分裂延迟的结构中显示出改变（Lin等，2000； Delillo等，2003）。最近的工作表明，INSP调节染色质重塑复合物的活性（Shen等，2003; Steger et et al。，2003），RSC复合物与Chromatin Remodecters，IS IS IS IS IS IS IS INSTROMECERITER（IS IS IS INSTROMECERITER）相关，这表明INSP调节了INSP的活性（Shen等人，2003; Steger等人）（Shen等人，2003; Shege foreme and Isextipe consecte）（Shen et et al。等，2003）。该提案的具体目的1将检验以下假设：PLC1P和INSP会影响动力学上RSC功能的募集，完整性或其他方面。遗传实验和全基因组表达分析表明，除了染色质重塑外，INSP还通过独立于染色质重塑络合物的机制影响转录。具体目标2将通过影响MSN2P/MSN4P激活剂，TUP1P/SSN6P抑制剂复合物或SRB/介体介体复合物的SRB10P成分来确定PLC1P和INSP是否会影响压力诱导的基因的转录调节。特定的目标3将利用良好的GAL1启动子作为模型，并确定为什么PLC1-DELTA细胞无法表达GAL1基因。该提案的重点是阐明PLC1P和INSP在细胞生理学的两个非常重要方面的作用：染色体隔离/细胞周期进程和转录调控，这些过程在包括癌症在内的人类疾病中不正常的过程。这项工作的结果最终将有助于确定癌症治疗的新分子靶标。