Cellular Responses to DNA-Protein Crosslinks

环境和内源性暴露于产生dna -蛋白质和dna -肽交联的化学物质与几种癌症、哮喘和其他疾病的风险增加有关。目前，美国人口中有成千上万的人暴露在最常见的dna -蛋白质交联诱导剂甲醛中。这些暴露既发生在职业环境中，也发生在家庭环境中，受影响的个人经常经历多年的慢性暴露，远远超过典型的室内空气质量标准。尽管目前对这类DNA损伤的细胞修复和反应机制知之甚少，但我们之前的研究已经严格建立了合成化学程序，以创建和利用含有位点特异性修饰的DNA-蛋白质交联的DNA，并确定了剂量依赖性的全基因组测定，以鉴定其产物功能限制DNA-蛋白质交联诱导的细胞毒性的基因。这些研究产生了一系列假设，假设真核细胞暴露于慢性低水平的DNA-蛋白交联剂，通过同源重组使细胞毒性和诱变最小化，而在急性高剂量暴露后，细胞将转向核蛋白酶体依赖性共价连接蛋白降解引发的途径，然后可以通过核苷酸切除修复或翻译DNA聚合酶进行处理。为了实现我们确定慢性和急性暴露下dna -蛋白交联修复和耐受的基本途径的目标，基因特异性缺失分析将确定在限制细胞毒性和诱变方面至关重要的基因群和相关途径。单个基因产物在调节细胞对dna -蛋白质交联反应中的作用将包括修复、重组、翻译合成、细胞周期检查点、染色质重塑和蛋白质水解途径。翻译合成聚合酶的活性将通过含有位点特异性dna -肽交联和随机加合的DPCs的dna来确定。总的来说，这些调查将产生对这类病变的修复和耐受性的综合分析

Environmental and endogenous exposure to chemicals that produce DNA-protein and DNA-peptide crosslinks are correlated with an increased risk of several cancers, asthma, and other diseases. Currently many thousands of individuals in the US population are exposed to one of the most common DNA-protein crosslink-inducing agents, formaldehyde. These exposures take place in both occupational and in-home settings and affected individuals often experience multi-year chronic exposures that are far in excess of typical indoor air quality standards. Although little is currently known concerning the cellular repair and response mechanisms for this class of DNA lesions, our prior investigations have both rigorously established synthetic chemical procedures to create and utilize DNAs containing site-specifically modified DNA-protein crosslinks and determined dose-dependent, genome-wide assays that identify genes whose products function to limit DNA-protein crosslink-induced cytotoxicity. These investigations have generated a series of hypotheses which postulate that eukaryotic cells exposed to chronic, low levels of DNA-protein crosslinking agents minimize cytotoxicity and mutagenesis through homologous recombination, while following acute high dose exposure, cells will shift to pathways initiated by nuclear proteasome-dependent degradation of covalently linked proteins that can then be processed via either nucleotide excision repair or translesion DNA polymerases. To accomplish our objective of determining the fundamental pathways for the repair and tolerance of DNA-protein crosslinks under chronic and acute exposures, gene-specific deletion analyses will identify the constellation of genes and interrelated pathways that are critical in limiting cellular toxicity and mutagenesis. The roles of individual gene products in modulating cellular response to DNA-protein crosslinks will include repair, recombination, translesion synthesis, cell cycle check points, chromatin remodeling and proteolytic pathways. The activities of translesion synthesis polymerases will be established using DNAs containing site-specific DNA-peptide crosslinks and randomly adducted DPCs. Collectively, these investigations will yield comprehensive analyses of repair and tolerance of this class of lesions