FUNCTION OF THE RETINOBLASTOMA PROTEIN

视网膜母细胞瘤蛋白的功能

• 批准号: 6172317
• 负责人: EDWARD E HARLOW
• 金额: $ 48.41万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-08-05 至 2003-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6172317
• 关键词: 3T3 cells; DNA binding protein; cell cycle; cell differentiation; cell transformation; gene expression; gene mutation; genetic regulation; laboratory mouse; microinjections; oncogenes; phosphorylation; protein kinase; protein structure function; retinoblastoma protein; tissue /cell culture; transcription factor; transfection

The retinoblastoma gene is the prototype of a class of cancer genes known as tumor suppressor genes. Tumor suppressor genes are defined by two key characteristics: (1) mutation of these genes promotes tumorigenesis (2) the selective advantage conferred by mutation of these genes is due to the loss of gene function. We are investigating the function of the retinoblastoma gene product (pRB). The best understood function of pRB is as a regulator of transcription factors. During the last grant cycle we characterized the ability of pRB and related proteins to regulate the E2F transcription factor and to control the expression E2F-target genes. The activation of E2F promotes cell cycle progression and, in normal cells, pRB acts to repress of E2F-dependent transcription. A great deal is known about the biochemistry of protein complexes formed between E2F and pRB-family proteins but the in vivo functions of this regulatory system are poorly understood. In the last two years we have established two experimental systems to examine the functions of these proteins. First, using homologous recombination we generated mutant alleles of E2F-1 in the mouse and studied the effects of eliminating E2F-1 function. Previous work had shown that E2F-1 behaved as an oncogene when over-expressed in tissue culture. However mice lacking E2F-1 developed a broad range of tumors revealing that E2F-1 acts to suppress tumor formation in vivo. In the next grant cycle we propose to continue this investigation of E2F function. We will concentrate on the characterization of the phenotype of E2F-1 null cells, on the phenotype of cells lacking DP-1 (the heterodimeric partner of E2F-1), and on the phenotype of cells lacking multiple pRB-regulated E2F s. To test role of the pRB/E2F-1 complex in pRB and E2F-1 functions we will characterize mouse strains carrying specific mutant alleles of E2F-1 that eliminate this interaction. A second area of functional studies that we initiated in the last grant cycle has also given unexpected findings. pRB is one member of a family of proteins, including p107 and p130, which share strong structual homologies. During the characterization of 3T3 cell lines prepared from pRB, p107 or p30 nullizygous mouse embryos, we observed that pRB, and p107 or p130 have specific and opposite effects on adipocyte differentiation. Whereas pRB promotes differentiation, p107 and p130 block adipocyte differentiation. We have finished the initial characterization of these differences and in the next round of this grant will investigate the functional differences between pRB p107 and p130 in adipogenesis and in other defined differentiation systems.

视网膜母细胞瘤基因是一类癌症基因的原型 称为肿瘤抑制基因。 肿瘤抑制基因的定义如下： 两个关键特征：（1）这些基因的突变促进 （2）这些突变所赋予的选择性优势 这是由于基因功能的丧失。 我们正在调查 视网膜母细胞瘤基因产物（pRB）的功能。 最好的理解 pRB的功能是作为转录因子的调节剂。 期间 上一个赠款周期，我们的特点是能力的pRB和相关的 蛋白质来调节E2 F转录因子和控制 表达E2 F靶基因。 E2 F的激活促进细胞周期 在正常细胞中，pRB的作用是抑制E2 F依赖性的细胞凋亡。 转录。 关于蛋白质复合物形成的生物化学， E2 F和pRB家族蛋白之间的差异，但这种差异的体内功能 监管体系知之甚少。 在过去的两年里， 建立了两个实验系统来检查这些功能 proteins. 首先，使用同源重组，我们产生突变体， E2 F-1等位基因，并研究了消除E2 F-1等位基因的影响。 E2 F-1功能。以前的研究表明，E2 F-1表现为一种 癌基因在组织培养中过度表达。 然而，老鼠缺乏 E2 F-1发展了广泛的肿瘤，揭示了E2 F-1的作用， 抑制体内肿瘤形成。 在下一个赠款周期，我们建议 继续E2 F功能的研究 我们将集中于 E2 F-1缺失细胞表型的表征， 缺乏DP-1（E2 F-1的异二聚体伴侣）的细胞表型， 和缺乏多个pRB调节的E2 F的细胞的表型。 到 测试pRB/E2 F-1复合物在pRB和E2 F-1功能中的作用，我们将 表征携带E2 F-1的特定突变等位基因的小鼠品系 消除这种相互作用。 第二个功能研究领域是我们在上一次拨款中启动的 周期也有意想不到的发现。 pRB是家族中的一员 蛋白质，包括p107和p130，它们具有很强的结构特征， 同源性 在表征由以下制备的3 T3细胞系期间： pRB、p107或p30缺失的小鼠胚胎中，我们观察到pRB和 p107和p130对脂肪细胞具有特异性和相反的作用 分化而pRB促进分化，p107和p130 阻断脂肪细胞分化。 我们已经完成了最初的 这些差异的特点，并在下一轮的这一点， 格兰特将研究pRB p107和 p130在脂肪形成和其他确定的分化系统中的作用。