SEQUENCE CONTEXT EFFECTS ON DAMAGE RECOGNITION BY NER

核苷酸切除修复（NER）缺陷与许多人类疾病直接相关，包括色素性干皮病、柯凯因综合征和癌症。在所有细胞模型中，DNA损伤识别的机制仍然知之甚少。损伤识别难题的一个核心特征是，DNA双工的局部扭曲（可能是由损伤位点附近的碱基对分离引起的）是否是使DNA损伤位点被识别并随后被NER机制去除的关键步骤。如果损伤位点附近的碱基对不配对确实是识别损伤的关键特征，那么直接遵循的是，任何形式的损伤周围的序列都会对损伤的可识别性产生影响，即更容易不配对的序列应该更容易被识别。这项拟议的研究有两个目标。主要目标将是确定在酿酒酵母中是否存在DNA序列上下文偏好，以识别已知仅由NER修复的两种类型的DNA损伤：2-乙酰氨基芴-DNA加合物和苯并[a]芘二醇环氧化物-DNA加合物。两种损伤形式的可识别性将在损伤位点序列背景不同的一组DNA底物中进行比较。该项目的第二个目标是通过明确确定NER机制中哪些蛋白质成分会产生任何观察到的序列上下文偏好，从而与NER蛋白本身建立相关性。

Defects in Nucleotide Excision Repair (NER) have been linked directly to numerous human diseases including xeroderma pigmentosum, Cockayne syndrome and cancer. The mechanism of DNA damage recognition remains poorly understood in all cellular models. A central feature to the conundrum of damage recognition is whether localized distortion of the DNA duplex, possibly by the unpairing of base pairs near the damage site, is the crucial step enabling a site of DNA damage to be identified and subsequently removed by the NER machinery. If the unpairing of base pairs near the damage site is truly a crucial feature in identifying damage then it follows directly that sequences immediately surrounding any form of damage will bear on the damage recognizability, i.e. sequences that unpair more easily should be more easily recognized. The proposed study has two goals. The primary goal will be to determine in Saccharomyces cerevesiae if here is a DNA sequence context preference for recognition of two types of DNA damage known to be repaired exclusively by NER:2-acetylaminofluorene-DNA adducts and benzo[a]pyrene diol epoxide-DNA adducts. The recognizability of both forms of damage will be compared within a panel of DNA substrates that differ in the sequence context of the damage site. The secondary goal of this project is to make a correlation with the NER proteins themselves by identifying specifically which protein components within the NER machinery give rise to any observed sequence context preference.