Interplay between benzo(a)pyrene-induced senescence and transcriptonal repression of DNA repair

苯并芘诱导的衰老与 DNA 修复转录抑制之间的相互作用

• 批准号: 388225306
• 负责人: Professor Dr. Markus Christmann
• 金额: --
• 依托单位: Institut für Toxikologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2017
• 资助国家: 德国
• 起止时间: 2016-12-31 至 2018-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/388225306?language=en
• 关键词: Interplay; between; benzo; pyrene; induced

A coordinated and faithful DNA repair is of central importance for maintaining genomic integrity and survival. A significant influence on the outcome of DNA damage processing is expected if the relative abundance of key repair factors is altered. This could disrupt the balance between individual repair pathways and the effectiveness of the DNA repair. Transcriptional activation of DNA repair genes is an important regulatory mechanism contributing to the adaptation of cells to genotoxic stress conditions. However, it appears that the inverse strategy, i.e. downregulation of gene expression in response to DNA damage, also plays a role in the fine-tuned regulation of complex DNA repair pathways. In our previous work, we analysed the regulation of DNA repair in response to benzo(a)pyrene 9,10-diol-7,8-epoxide (BPDE), the active metabolite of benzo(a)pyrene (B[a]P), which is the most important carcinogen formed by incomplete combustion during food preparation and smoking. We showed that low (nontoxic) BPDE concentrations cause AP-1 and p53 dependent upregulation of several NER genes, leading to enhanced NER activity and consequently reducing the effectiveness of a challenge dose (adaptive response). To further elucidate alterations in the expression of DNA repair genes following BPDE exposure, qPCR microarrays were performed, revealing a strong repression of the mismatch repair factors MSH2, MSH6 and EXO1, as well as of Rad51, the central component of the homologous recombination. The repression of these genes was mediated by abrogation of the E2F1 pathway. As part of the present application, we will investigate the molecular mechanisms leading to the repression of MSH2, MSH6, EXO1 and RAD51, focussing on mechanisms underlying abrogation of the E2F1 pathway and on the impact of histone modification.Since we could show that repression of these DNA repair mechanisms is a specific trait of senescent cells, the repression may represent a great danger for the organism. In absence of these important DNA repair mechanisms, smoking induced DNA lesions, may lead to accumulation of mutations and chromosomal aberrations in senescent cells. Furthermore, since the nontoxic BPDE concentrations only induce a transient DNA damage response, the cells may exit senescence with unrepaired genomic alterations, which could contribute to the carcinogenic potential of smoking. To test this hypothesis, the mechanisms responsible for induction and maintenance of senescence will be analysed using senescent and non-senescent cells separated by FACS upon BPDE exposure. In detail, we will focus on the impact of the DNA damage response and the SASP phenotype. In addition open approaches using kinome and transcriptome profiling will be performed. Finally we will analyse whether cells can escape from B[a]P/BPDE-induced senescence and whether these cells harbour increased genomic alterations.

协调和忠实的DNA修复对于维持基因组完整性和生存至关重要。如果关键修复因子的相对丰度改变，则预期对DNA损伤处理的结果有重大影响。这可能会破坏单个修复途径和DNA修复有效性之间的平衡。 DNA修复基因的转录激活是有助于细胞对遗传毒性应激条件适应的重要调节机制。然而，看来的反向策略，即基因表达对DNA损伤的下调，也在复杂DNA修复途径的微调调节中起作用。在以前的工作中，我们分析了对苯并（A）pyrene 9,10-二醇-7,8-环氧（BPDE）的调节，这是苯并（a）pyrene（a）pyrene（b [a] p）的活性代谢物，这是由不含食物制备过程中的燃烧和烟雾制成和烟雾制成和烟熏剂形成的最重要的癌。我们表明，低（无毒的）BPDE浓度导致AP-1和p53依赖于几个NER基因的上调，从而增强了NER活性，因此降低了挑战剂量的有效性（自适应反应）。为了进一步阐明BPDE暴露后DNA修复基因表达的改变，进行了QPCR微阵列，揭示了对不匹配修复因子MSH2，MSH6和EXO1以及RAD51的强烈抑制，RAD51是同源重组的中心成分。这些基因的抑制是通过废除E2F1途径介导的。作为本应用的一部分，我们将研究导致MSH2，MSH6，EXO1和RAD51抑制的分子机制，重点关注E2F1途径废除的机制，并且在组蛋白修饰的影响上，我们可以表明，这些DNA修复机制的抑制作用可能是一种危险的抑制，这可能是一种危险的特定性状，这可能是众所周知的特定性状。在没有这些重要的DNA修复机制的情况下，吸烟引起的DNA病变可能导致突变和染色体畸变的积累。此外，由于无毒的BPDE浓度仅诱导瞬时DNA损伤反应，因此细胞可能会以未修复的基因组改变而退出衰老，这可能有助于吸烟的致癌潜力。为了检验这一假设，将使用BPDE暴露后FACS分离的衰老和非元素细胞来分析负责衰老诱导和维持的机制。详细说明，我们将重点关注DNA损伤反应和SASP表型的影响。此外，将执行使用Kinome和转录组分析的开放方法。最后，我们将分析细胞是否可以从B [A] P/BPDE诱导的衰老以及这些细胞携带的基因组改变增加。