INTRAGENIC SUPPRESSOR MUTATIONS FOR P53 CANCER MUTATIONS

P53 癌症突变的基因内抑制突变

• 批准号: 2842129
• 负责人: RAINER K BRACHMANN
• 金额: $ 21.12万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-01 至 2004-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2842129
• 关键词: DNA binding protein; Escherichia coli; apoptosis; gene expression; gene mutation; immunoprecipitation; neoplasm /cancer genetics; nucleic acid sequence; oncoproteins; p53 gene /protein; plasmids; polymerase chain reaction; protein structure function; suppressor mutations; tissue /cell culture; western blottings; yeasts

The p53 protein exerts its tumor suppressor function mainly as a transcription factor that induces GI and G2 cell cycle arrest and/or apoptosis. Approximately 50 percent of human cancers have p53 missense mutations that result in high levels of p53 cancer mutants with a structurally altered core domain unable to bind to P53 DNA-binding sites upstream of effector genes. The same cancers (corresponding to 2.4 million cancer patients per year worldwide) carry a poor prognosis since p53 is an important mediator of cancer therapy-induced apoptosis. If one could restore function to this large pool of p53 mutant proteins, the impact on cancer therapy would be substantial. A detailed knowledge is therefore needed of the extent to which the most common p53 cancer mutations destabilize the structure of the p53 core domain. More importantly, suppressor mechanisms must be identified that can override the deleterious effects of p53 cancer mutations. This goal can be achieved through the study of intragenic suppressor mutations. A genetic approach using our functional p53 yeast assay showed in a pilot study that such suppressor mutations exist for selected p53 cancer mutations and that they can be isolated very efficiently. We now propose to identify intragenic suppressor mutations for the eight most common p53 cancer mutations and test them against the 35 most common p53 cancer mutations (accounting for 37 percent of human cancers with p53 mutations) using yeast and mammalian assays. This systematic study will define the structural motifs that are absolutely essential for the structural integrity of the p53 core domain and may identify suppressor mutations that act as global suppressors. The crystal structures for the core domains of the most promising suppressor mutations, the most common cancer mutations and combinations of both will then be determined to acquire a precise understanding of the structural dynamics of the p53 core domain. Preliminary experiments indicate that p53 proteins with a suppressor mutation alone have characteristics different from wild-type p53. Some of them may, in fact, behave like "super p53" proteins that are resistant to the dominant-negative effects of p53 cancer mutants and that may be more efficient in inducing cell cycle arrest and/or apoptosis. We will identify such p53 proteins and determine the mechanisms that make them resistant to dominant-negative p53 cancer mutants. We will further study how some of them are able to induce selectively and/or strongly G1 or G2 arrest or apoptosis. This will determine the relative importance of p53 downstream effector genes and may ultimately lead to the identification of new p53 pathways.

p53蛋白施加其肿瘤抑制剂的功能主要是 诱导GI和G2细胞周期停滞和/或的转录因子 凋亡。 大约50％的人类癌症有P53错过 导致高水平的p53癌症突变体的突变 结构上改变的核心结构域无法与p53 DNA结合位点结合 效应基因上游。 相同的癌症（对应于2.4 每年全球百万癌症患者）的预后不佳 p53是癌症治疗引起的凋亡的重要介质。 如果 一个人可以将功​​能恢复到这个庞大的p53突变蛋白， 对癌症治疗的影响将是很大的。 详细的知识 因此，需要最常见的p53癌的程度 突变破坏了p53核心结构域的结构。 更多的 重要的是，必须确定可以覆盖的抑制机制 p53癌症突变的有害作用。 这个目标可以是 通过研究基因内抑制突变实现。 一个 使用我们的功能性p53酵母测定法中显示的遗传方法 研究认为某些p53癌症存在这种抑制突变 突变，可以非常有效地隔离它们。 我们现在 提议确定八个最大的基因内抑制突变 常见的p53癌症突变并针对35个最常见的p53进行测试 癌症突变（占P53人类癌的37％ 突变）使用酵母和哺乳动物测定法。这项系统的研究将 定义对对 p53核心域的结构完整性，可以识别抑制器 充当全球抑制器的突变。 晶体结构 最有希望的抑制剂突变的核心领域，最多 然后将确定两者的常见癌症突变和组合 要精确理解p53的结构动力学 核心域。 初步实验表明p53蛋白具有抑制剂 仅突变的特征与野生型p53不同。 一些 实际上，它们的行为可能像“超级P53”蛋白 对p53癌症突变体和 这可能更有效地诱导细胞周期停滞和/或 凋亡。 我们将确定此类p53蛋白，并确定 使它们能够抵抗优势阴性p53癌的机制 突变体。 我们将进一步研究其中一些能够诱导的 有选择地和/或强烈的G1或G2停滞或凋亡。 这会 确定p53下游效应子基因的相对重要性和 最终可能导致识别新的p53途径。