CELLULAR PROCESSING OF DAMAGED DNA--ROLE IN ONCOGENESIS

这个项目涉及癌症的分子流行病学， 首先意识到，多达六个独立的 遗传和表观遗传事件可能参与了从 从正常细胞到恶性细胞基因组DNA的变化发生在特定的 位点，并可导致原癌基因的激活或失活， 肿瘤抑制基因通过突变，重组，基因 扩增、易位或其他染色体异常。在一些 人类遗传性疾病，肿瘤的发病率增加， 与受损DNA的修复和/或复制缺陷相关。 我们的最终目标是了解受损DNA的处理过程 与致癌作用有关。 在开发敏感技术方面处于领先地位， 定量特定DNA损伤的限制性片段， 区域的基因组，我们将扩展我们的分析基因组内的罚款 DNA修复的结构，以了解控制效率的因素 在染色质和不同的功能域的过程中， 基因组，如复制起点和表达的基因转录 不同的RNA聚合酶。在发现了优先修复 转录的DNA链在表达的基因，我们将测试一个模型， 基于增强转录因子的转录偶联修复 通过RNA聚合酶II的5 '-3'外切核酸酶活性缩短。我们 将严格测试链特异性DNA修复可以 用作转录的灵敏测定。核基质相关 DNA将被表征，以确定这是否是 转录偶联修复干皮病的区域限制性修复 色素沉着，互补组C，将评估学习的基础 对于癌症易感表型，以及表达的修复缺陷， 将研究Cockayne综合征中的基因，以了解 缺陷和缺乏癌症倾向。修理中的差异 特定基因的风险可能解释了一些深刻的 在不同组织的致癌反应中看到的差异， 不同生物体中的相同组织。携带病变的质粒探针， 定义的位点将用于分析特定序列修复， 不同遗传背景的细胞。定义的嵌合质粒将 也可用于将基因导入不同的基因组结构域进行研究 损伤处理的具体特点，导致增强的 将受损的DNA整合到人体细胞中。我们亦会探讨 在非增殖细胞中局部DNA更新的可能作用 突变的精细结构，以测试我们的假设，转录- 相关的DNA更新可能导致异常的高突变频率 在某些领域。 这项研究将大大有助于我们了解 DNA损伤处理缺陷的基础，在某些癌症易感 遗传性疾病，它也应该导致新的，敏感的探针， 分析人体细胞的损伤和修复。此外，我们的研究 应该有助于解释DNA损伤和DNA周转在 生物学终点，如存活、诱变和致癌。

This project is concerned with the molecular epidemiology of cancer, beginning with the realization that as many as half a dozen independent genetic and epigenetic events may be involved in the transformation from a normal cell to malignancy. The changes in genomic DNA occur at specific sites and can lead to activation of protooncogenes or inactivation of tumor suppressor genes through mutation, recombination, gene amplification, translocation, or other chromosomal abnormalities. In some human hereditary diseases an increased incidence of neoplasia is correlated with a defect in the repair and/or replication of damaged DNA. Our ultimate objective is to understand how the processing of damaged DNA in mammalian cells relates to carcinogenesis. Having pioneered in the development of sensitive techniques for quantifying particular DNA lesions in restriction fragments from specific regions of the genome we will extend our analysis of intragenomic fine structure of DNA repair, to learn the factors that control the efficiency of the process in chromatin and in different functional domains of the genome, such as replication origins and expressed genes transcribed by different RNA polymerases. Having discovered preferential repair of the transcribed DNA strand in expressed genes, we will test a model for transcription-coupled repair based upon factors that enhance transcript shortening by the 5'- 3' exonuclease activity of RNA polymerase II. We will critically test the possibility that strand-specific DNA repair can be used as a sensitive assay for transcription. Nuclear matrix associated DNA will be characterized to determine whether that is the site of transcription-coupled repair. Domain limited repair in xeroderma pigmentosum, complementation group C, will be assessed to learn the basis for the cancer prone phenotype, and the deficiency in repair of expressed genes in Cockayne's syndrome will be studied to understand the basis for the defect and the absence of cancer proneness. Differences in the repair of particular genes at risk may account for some of the profound differences seen in the carcinogenic responses of different tissues and of the same tissue in different organisms. Plasmid probes carrying lesions at defined sites will be used in the analysis of specific sequence repair in cells of different genetic background. The defined chimeric plasmids will also be used to introduce genes into different genomic domains to study the specific features of damage processing that result in the enhanced integration of damaged DNA in human cells. We will also explore the possible role of localized DNA turnover in non-proliferating cells in the fine structure of mutagenesis to test our hypothesis that transcription- associated DNA turnover may result in anomalous high mutation frequencies in some domains. This research should contribute substantially to our understanding of the basis for DNA damage processing deficiencies in certain cancer-prone hereditary diseases and it should also result in new, sensitive probes for the analysis of damage and repair in human cells. In addition, our studies should help to interpret the role of DNA damage and DNA turnover in biological end points such as survival, mutagenesis, and carcinogenesis.