Identification of mechanisms and consequences of chromos

识别染色体的机制和后果

• 批准号: 6758335
• 负责人: Thomas E. Ried
• 金额: --
• 依托单位: DIVISION OF CLINICAL SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6758335
• 关键词: adenoma; aneuploidy; animal tissue; carcinoma; cell line; cell transformation; chromosome movement; chromosome translocation; colorectal neoplasms; cytogenetics; gel electrophoresis; genetically modified animals; green fluorescent proteins; karyotype; laboratory mouse; microarray technology; molecular genetics; neoplasm /cancer genetics

The review of CGH analyses performed in our laboratory has revealed that the acquisition of whole chromosomes or chromosome arm gains and losses is a frequent event, particularly at early stages of carcinogenesis. The comparison of 25 cell lines established from solid tumors by SKY and CGH has provided compelling evidence that the vast majority of epithelial cancers have either numerical or structural chromosomal aberrations resulting in genomic imbalances. The comparison of diploid, mismatch repair deficient colorectal carcinomas with aneuploid ones indicates that numerical chromosomal aberrations are 60 times more prevalent in the aneuploid tumors. Abnormalities of the centrosome correlated with chromosomal aneuploidy both in human cancer cell lines and in animal cells deficient for cell cycle regulator genes such as p53 and BRCA1. All these results taken together support the notion that chromosomal aneuploidy is a major theme in epithelial cancers. In order to elucidate mechanisms leading to aneuploidy, to establish the functional relevance of chromosomal aneuploidy, and to identify whether aneuploidy is a cause or a consequence of genetic changes in solid tumors, we have focused on the following projects: Structural and functional analysis of abnormal centrosomes and their relationship to chromosome segregation fidelity. Sequential inhibition of p53 and Rb-function with E6 and E7-genes from human papilloma virus 16 and subsequent analyses of chromosomal instability. Microcell mediated chromosome transfer of chromosomes that are frequently gained in colorectal carcinomas into cells derived from normal colorectal epithelium and adenomas followed by cytogenetic analysis and assays for cellular immortalization and transformation. In vitro induction of chromosomal aneuploidy in primary cultures derived from normal murine cells of different organ origin with aneugens followed by cytogenetic analysis and assays for confirmation of cellular immortalization and transformation. Comparison of cells carrying chromosomal aneuploidies with their wildtype parental cells using cDNA microarrays and 2D-gel protein electrophoresis. In two model systems, we have also studied what general consequences recurrent chromosomal aneuploidies exert on gene expression levels. The results suggest that only a few candidate genes are the target for the acquisition and maintenance of genomic imbalances. This suggests that genomic amplification events are accompanied by the silencing of large stretches of chromatin.

在我们实验室进行的CGH分析的审查表明，收购整个染色体或染色体臂的增益和损失是一个常见的事件，特别是在早期阶段的致癌作用。通过SKY和CGH建立的25种实体瘤细胞系的比较提供了令人信服的证据，即绝大多数上皮癌具有导致基因组失衡的染色体数目或结构畸变。二倍体、错配修复缺陷型结直肠癌与非整倍体结直肠癌的比较表明，染色体数目畸变在非整倍体肿瘤中的发生率高60倍。在人类癌细胞系和缺乏细胞周期调节基因如p53和BRCA 1的动物细胞中，中心体的缺失与染色体非整倍性相关。所有这些结果加在一起支持了染色体非整倍性是上皮癌的一个主要主题的观点。为了阐明导致非整倍体的机制，建立染色体非整倍体的功能相关性，并确定非整倍体是实体瘤遗传变化的原因还是结果，我们专注于以下项目： 异常中心体的结构和功能分析及其与染色体分离保真度的关系。 人乳头瘤病毒16型E6和E7基因对p53和Rb功能的顺序抑制及随后的染色体不稳定性分析 微细胞介导的染色体转移，通常在结直肠癌中获得的染色体转移到来自正常结直肠上皮和腺瘤的细胞中，然后进行细胞遗传学分析和细胞永生化和转化的测定。 用非整倍体原代培养物体外诱导不同器官来源的正常鼠细胞的染色体非整倍体，然后进行细胞遗传学分析和测定，以确认细胞永生化和转化。 使用cDNA微阵列和2D凝胶蛋白电泳比较携带染色体非整倍性的细胞与它们的野生型亲本细胞。 在两个模型系统中，我们还研究了复发性染色体非整倍性对基因表达水平的一般影响。结果表明，只有少数候选基因是获得和维持基因组不平衡的目标。这表明，基因组扩增事件伴随着大的染色质延伸的沉默。