RNA Polymerase Transcription Past DNA Adducts

RNA 聚合酶转录 DNA 加合物

• 批准号: 6916582
• 负责人: David A Scicchitano
• 金额: $ 29.5万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-30 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6916582
• 关键词: DNA damage; DNA directed RNA polymerase; DNA repair; RNA biosynthesis; active sites; adduct; carbopolycyclic compound; cell line; computer simulation; enzyme activity; genetic transcription; nucleic acid sequence; polymerase chain reaction; protein structure function; ribonucleotides; stereochemistry; tissue /cell culture; transposon /insertion element

DESCRIPTION (provided by applicant): The long range goal of this research is to gain a detailed understanding of how covalently modified bases in DNA affect RNA polymerase behavior during the elongation phase of transcription, with the ultimate objective of producing detailed structural models that portray RNA polymerase interactions at bulky and small adducts found in DNA. The following four Specific Aims will be targeted toward this: (1) to assess the effects of transcription past site-specific lesions on elongation past the damaged or modified site; (2) to determine the base sequence of full-length transcripts, and to characterize the base composition at the 3'- ends of truncated transcripts; (3) to determine Km, and Vrnax values for base addition at DNA adducts during transcription; and (4) to employ computer-modeling techniques to provide characteristics of transcription complexes stalled at DNA adducts. It is now quite clear that certain DNA lesions can cause RNA polymerase to stall at the modified site, resulting in a truncated transcript, or progress past the altered base, producing full-length RNA. Stalled transcription complexes signal transcription-coupled DNA repair. The importance of transcription-coupled DNA repair is evidenced by the clinical phenotype observed when it is aberrant, as is seen in patients with Cockayne's syndrome, a disease characterized by severe growth and developmental defects. There are broad and significant implications for preferential clearance of DNA damage from discrete, active genetic loci: Biases in mutagenesis can exist; correlations of DNA damage and repair with tumorigenesis might be stronger when preferential clearance rates for a particular adduct are used for making the comparisons, rather than total genomic repair; and the actual removal of adducts from the transcribed strand of an expressed gene might be very dependent on their ability to impede RNA synthesis.

描述（由申请人提供）：本研究的长期目标是详细了解DNA中共价修饰的碱基在转录延伸阶段如何影响RNA聚合酶行为，最终目标是产生详细的结构模型，描述DNA中发现的大体积和小加合物处的RNA聚合酶相互作用。本研究的主要目的是：（1）评估转录经过位点特异性损伤后对延伸经过损伤或修饰位点的影响;（2）确定全长转录本的碱基序列，并表征截短转录本3 '端的碱基组成;（3）确定转录过程中DNA加合物处碱基加成的Km和Vmax值;（4）采用计算机建模技术提供停滞在DNA加合物处的转录复合物的特征。现在已经很清楚，某些DNA损伤可以导致RNA聚合酶在修饰位点停滞，导致截短的转录物，或者通过改变的碱基，产生全长RNA。停滞的转录复合物信号转录偶联DNA修复。转录偶联DNA修复的重要性通过其异常时观察到的临床表型来证明，如在Cockayne综合征患者中所见，Cockayne综合征是一种以严重生长和发育缺陷为特征的疾病。从离散的、活跃的遗传位点优先清除DNA损伤有广泛而重要的意义：诱变中可能存在偏倚;当使用特定加合物的优先清除率而不是总基因组修复率进行比较时，DNA损伤和修复与肿瘤发生的相关性可能更强;加合物从表达基因的转录链上的实际去除可能非常依赖于它们阻碍RNA合成的能力。