Repair of Genome Destabilizing DNA Structures

描述（由申请人提供）：DNA损伤和修复对人类健康和疾病至关重要。该申请的长期目标是：扩大我们对DNA螺旋扭曲在DNA损伤识别和处理中的作用的理解;确定影响DNA结构诱导的遗传不稳定性的因素;阐明与某些癌症相关的易位中涉及的潜在机制;以及进一步开发新的方法来减少人类细胞中的遗传不稳定性。在短期内，我们将继续研究我们最近的发现，即天然存在的Z-DNA和H-DNA结构诱导的螺旋扭曲具有高度致突变性，并可诱导哺乳动物细胞中的DNA双链断裂（DSB）。我们建议研究DNA螺旋扭曲对基于质粒的系统中基因组不稳定性的影响，以及对人类细胞和转基因突变报告小鼠中染色体的影响。我们将集中在位于人类c-MYC启动子易位断裂点附近的H-DNA形成序列和位于人类BCL-2基因染色体断裂点的Z-DNA序列，这些序列在淋巴瘤和白血病中发现。我们将确定DNA修复，复制和转录在结构诱导的遗传不稳定性中的作用。我们将利用我们在引入位点特异性DNA螺旋扭曲中的专业知识，以明确的分子间三链体结构的形式来测试我们的假设，即某些类型的DNA螺旋扭曲（在存在或不存在DNA损伤本身的情况下）被人类细胞中的DNA修复机制所识别。从这些研究中获得的新信息将深入了解非B DNA诱导的遗传不稳定性的机制;人类DNA修复中的限速步骤（即扭曲/损伤识别）;以及处理DNA螺旋扭曲中核苷酸切除修复和错配修复之间的重叠。它还将鉴定参与DSB生成的蛋白质，DSB由在人类癌症中映射到易位断点的序列处形成的非规范DNA结构诱导。这些发现将使我们更好地了解由DNA损伤和自然发生的螺旋扭曲引起的癌症和其他疾病的发病机制，并最终开发新的治疗和预防方法。

DESCRIPTION (provided by applicant): DNA damage and repair are fundamental to human health and disease. The long-term objectives of this application are to: expand our understanding of the role of DNA helical distortions in DNA damage recognition and processing; determine the factors influencing DNA structure-induced genetic instability; elucidate potential mechanisms involved in translocations associated with certain cancers; and further the development of novel approaches to reduce genetic instability in human cells. In the short term, we will pursue our recent discovery that helical distortions induced by naturally occurring Z-DNA and H-DNA structures are highly mutagenic and can induce DNA double-strand breaks (DSBs) in mammalian cells. We propose to study the effect of DNA helical distortions on genomic instability in plasmid-based systems as well as on chromosomes in human cells and in transgenic mutation-reporter mice. We will focus on the H- DNA-forming sequence located near the translocation breakpoint in the human c-MYC promoter and the Z- DNA sequence located at a chromosomal breakpoint in the human BCL-2 gene, found in lymphomas and leukemias. We will determine the role(s) of DNA repair, replication and transcription in the structure-induced genetic instability. We will use our expertise in the introduction of site-specific DNA helical distortions in the form of well-defined intermolecular triplex structures to test our hypothesis that certain types of DNA helical distortions (in the presence or absence of DNA damage per se) are recognized by the DNA repair machinery in human cells. The new information obtained from these studies will provide insight into the mechanisms of non-B DNA-induced genetic instability; the rate-limiting step in human DNA repair (i.e. distortion/damage recognition); and the overlap between nucleotide excision repair and mismatch repair in processing DNA helical distortions. It will also identify the proteins involved in the generation of DSBs induced by non- canonical DNA structures formed at sequences that map to translocation breakpoints in human cancers. These discoveries should lead to a better understanding of the pathogenesis of cancers and other diseases that are caused by DNA damage and naturally occurring helical distortions, and ultimately to the development of new approaches to treatment and prevention.