CONTROL OF CELL PROLIFERATION

细胞增殖的控制

• 批准号: 3204816
• 负责人: James M Roberts
• 金额: $ 26.12万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-09-01 至 1998-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3204816
• 关键词: SDS polyacrylamide gel electrophoresis; Saccharomyces cerevisiae; biological signal transduction; cell cycle; cell cycle proteins; cell growth regulation; cinematography; cytogenetics; fibroblasts; fungal genetics; gene expression; human tissue; laboratory rabbit; lymphocyte; microinjections; mitogens; protein kinase; protein structure function; synchronous cell division

The proliferation of all eukaryotic cells is primarily controlled by a single decision which occurs in the G1 phase of the cell cycle -- either to complete the cell cycle and divide or to withdraw from the cell cycle and enter a quiescent state. In normal cells this decision is regulated by specific intracellular and extracellular signals, such as mitogenic growth factors. In transformed or tumorigenic cells, however, these controls are suppressed and cell proliferation is unconstrained. Important advances have been made recently in understanding the molecular basis for this critical regulatory process. In S. cerevisiae, this decision is called START and completion of START culminates in DNA replication and commits the cell to complete the remainder of the cell division cycle. The biochemical event that underlies this cellular decision is the assembly and activation of complex between a protein kinase, encoded by the CDC28 gene, and a positively acting regulatory subunit of the kinase, a cyclin. Efforts to understand the molecular basis for controlling the proliferation of mammalian cells, therefore, have recently focused upon this family of protein kinases, named the cyclin-dependent kinases, and their regulators, the cyclins. We have isolated a new human cyclin, cyclin E, based upon its ability to functionally substitute for the S. cerevisiae G1 cyclins. We have found, by biochemical and physiological assays, that it probably plays a major role in regulating proliferation during the G1 phase of the mammalian cell cycle --1) Cyclin E binds to and activates a cyclin-dependent kinase specifically during the G1 phase of the human cell cycle; 2) Constitutive expression of cyclin E in primary human fibroblasts accelerates G1, reduces cell size and decreases growth factor requirements for cell proliferation; 3) Two factors that inhibit the cell cycle in G1, rapamycin and TGF-beta, block activation of the cyclin B/kinase complex. Our general goal is to develop a more complete picture of the physiological, biochemical and molecular actions of cyclin E during the mammalian cell cycle, both in fibroblasts and lymphocytes, and to understand how these actions contribute to the control of cell proliferation. Our specific approaches will include analyzing the physiological effects of constitutive cyclin E expression and inhibition of cyclin E expression on regulation of the mammalian cell cycle; studying, by high resolution two dimensional PAGE, cell cycle dependent modifications of cyclin E and cell cycle dependent interactions of cyclin B with other proteins; and correlating the physiological actions of cyclin B with its molecular and biochemical properties by analysis of cyclin E mutations.

所有真核细胞的增殖主要由一种 发生在细胞周期G1期的单一决定-- 完成细胞周期并分裂或退出细胞周期 进入静止状态。在正常细胞中， 通过特定的细胞内和细胞外信号，如促有丝分裂信号， 生长因子然而，在转化或致瘤细胞中，这些 对照被抑制，细胞增殖不受限制。重要 最近在理解分子基础方面取得了进展， 这一关键的监管过程。In S.酿酒厂这个决定 称为START，START的完成在DNA复制中达到高潮， 使细胞完成细胞分裂周期的剩余部分。的 作为这种细胞决定基础的生化事件是 与活化蛋白激酶之间的复合物，由CDC 28编码 基因，以及激酶的正作用调节亚基，细胞周期蛋白。 努力了解控制的分子基础， 因此，哺乳动物细胞的增殖最近集中在 这个蛋白激酶家族，称为细胞周期蛋白依赖性激酶， 它们的调节因子细胞周期蛋白。我们分离到一种新的人类细胞周期蛋白， E，基于其在功能上替代S的能力。酿酒酵母 G1细胞周期蛋白。通过生物化学和生理学分析，我们发现， 它可能在G1期调节增殖中起主要作用， 1）细胞周期蛋白E结合并激活细胞周期中的一个阶段。 细胞周期蛋白依赖性激酶，特异性在人细胞的G1期 2）细胞周期蛋白E在原代人成纤维细胞中的组成型表达 加速G1期，缩小细胞大小，减少生长因子需求 用于细胞增殖; 3）抑制G1期细胞周期的两个因子， 雷帕霉素和TGF-β阻断细胞周期蛋白B/激酶复合物的活化。 我们的总体目标是更全面地了解 细胞周期蛋白E在细胞周期中的生理、生化和分子作用 哺乳动物细胞周期，无论是在成纤维细胞和淋巴细胞， 了解这些行动如何有助于控制细胞 增殖我们的具体方法将包括分析 组成型细胞周期蛋白E表达和抑制的生理效应 细胞周期蛋白E表达对哺乳动物细胞周期的调控; 通过高分辨率二维PAGE研究细胞周期依赖性， 细胞周期蛋白E的修饰和细胞周期依赖的细胞周期蛋白相互作用 B与其他蛋白质的关系，以及细胞周期蛋白的生理作用 B及其分子生物学特性的研究 突变。