G-Quadruplex forming sequence motifs and genome instability

G-四链体形成序列基序和基因组不稳定性

• 批准号: 9247232
• 负责人: Nayun Kim
• 金额: $ 30.8万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9247232
• 关键词: Affect; Affinity Chromatography; Antibodies; Antineoplastic Agents; Binding; Binding Proteins; Biological Assay; Cancer Biology; Cancer Etiology; Cells; Characteristics; Chemicals; Chromatin; Chromosome Arm; Chromosomes; Complex; DNA; DNA Damage; DNA Restriction Enzymes; DNA Sequence; DNA Structure; DNA replication fork; Data; Development; Drug resistance; Elements; Enzymes; Etoposide; Eukaryota; Event; Frequencies; G-Quartets; Genetic; Genetic Recombination; Genetic Transcription; Genome; Genomic Instability; Genomics; Goals; Guanine; Human; In Vitro; Knowledge; Lead; Left; Ligands; Link; Maintenance; Malignant Neoplasms; Maps; Mass Spectrum Analysis; Measurement; Measures; Methods; Mission; Modeling; Molecular; Mutagenesis; Mutation; Neoplasm Metastasis; Pathway interactions; Proteins; Public Health; Repetitive Sequence; Reporter; Research; Resistance; Role; Running; Saccharomyces cerevisiae; Saccharomycetales; Second Primary Cancers; Site; Stress; Structure; Testing; Topoisomerase; Trans-Activators; Transact; Transcription Coactivator; Tumorigenicity; Type I DNA Topoisomerases; United States National Institutes of Health; Work; Yeasts; Zeocin; base; cancer cell; cancer genome; cancer therapy; chemotherapy; chromosome loss; design; experimental study; genetic approach; grasp; in vivo; insight; mutant; novel; prevent; public health relevance; repaired; small molecule; tumor; tumorigenesis; tumorigenic; yeast genome

 DESCRIPTION (provided by applicant): Mutations and chromosome rearrangements are ubiquitous feature of cancer genomes and linked to the anomalous cellular activities associated with oncogenesis. Recombination hotspots, where genome rearrangements initiate at incongruently higher frequencies, are significantly enriched with repetitive sequence elements with the potential to fold into non-helical DNA secondary structures. When left unresolved, such DNA structures can impede transcription and replication leading to genome instability. G-quadruplex or G4 DNA is a tetra-helical structure consisting of runs of guanines and one of the DNA structures that presents a challenge to normal DNA transactions. Comprehensive understanding of how a subset of the guanine run- containing sequences interspersed in the genome becomes genetically unstable, however, is still lacking. Also not understood is how such potential hotspots of genome instability are kept under control. The long-term research goal of the applicant is to better understand how spontaneous mutagenesis and rearrangements occur at disparate rates throughout the genome. The goal of the proposed research is to characterize what potentiates the marked elevation in instability associated with certain guanine-run containing sequences in the model eukaryote, S. cerevisiae. The central hypothesis of this proposal is that genome instability at a G4 motif is contingent on both genomic context and trans-acting factors that modulate the structural transformation of a guanine-run containing sequence into G4 DNA structure. This hypothesis is founded on the preliminary data generated by the applicant that identified two critical factors in modulating instability at a model G4 DNA- forming sequence - the level of transcription and the activity of topoisomerase I. Our specific aims are designed to (1) define the role of topoisomerase I in the G4 DNA-associated genome instability (2) test whether DNA damage cooperatively aggravates the instability of G4 forming sequences and (3) Identify proteins binding specifically to co-transcriptionally formed G4 DNA in vivo. In combination with molecular approaches to detect G4 DNA in vivo, genetic assays developed for quantitative measurement of recombination occurring at a G4 motif in transcription-dependent manner will be employed in AIM 1 and 2 to determine the effect of disruption in Top1 function and accumulation of DNA damage on the stability of co- transcriptionally formed G4 DNA. The genetic assays have proved effective and productive in generating the relevant preliminary data. Under AIM3, novel trans-activating factors with possible roles in suppressing instability at the G4 DNA will be identified by chromatin affinity purification approach. The stability of G4 motifs is particularly relevant to th biology of cancer since there is a significant overlap between genomic loci containing G4 sequences and the sites of translocations observed in various human cancers. The result of the proposed study can ultimately enhance our understanding of the molecular events instigating the tumorigenic rearrangements as well as therapy-related secondary malignancy or drug-resistant tumor development.

 描述（由应用提供）：突变和染色体重排是癌症基因组的普遍特征，与与肿瘤发生有关的异常细胞活性有关。重组热点，基因组重排以不一致的频率启动，它具有重复性序列元件的显着富集，并且可能会折叠成非螺旋DNA二级结构。当尚未解决时，这种DNA结构会阻碍转录和复制导致基因组不稳定性。 G-四链体或G4 DNA是一种四螺旋结构，由鸟嘌呤的运行和DNA结构之一组成，对正常DNA交易构成了挑战。然而，人们对基因组中杂种序列的子集的全面了解仍然缺乏一般不稳定。同样不明白的是，这种潜在的基因组不稳定性的热点如何保持控制。适用的长期研究目标是更好地了解整个基因组中的赞助诱变和重排如何以不同的速率发生。拟议的研究的目的是表征与模型真核生物S. cerevisiae中包含某些鸟嘌呤运行的序列相关的不稳定性显着升高的潜力。该提议的中心假设是，G4基序的基因组不稳定性均取决于基因组环境和反式作用因子，这些因素调节了鸟嘌呤运行的结构转化，其中包含鸟嘌呤的序列到G4 DNA结构中。该假设建立在适用的初步数据基础上，该数据确定了在模型G4 DNA形成序列调节不稳定性时的两个关键因素 - 转录水平和拓扑异构酶I的活性I。我们的具体目的I我们的具体目的旨在（1）定义TOPOISomerase I在G4 DNA基因组中的作用（2）是否损害了dna issi nistiention（2）的作用（2） G4形成序列和（3）鉴定在体内特异性结合的蛋白质与共转录形成的G4 DNA。结合在体内检测G4 DNA的分子方法，将在AIM 1和2中开发出用于以转录依赖性方式进行重新组合的定量测量重组的遗传测定，以确定TOP1功能中的破坏和DNA损害对共同传输型G4 DNA的稳定性的损害的影响的影响。遗传测定已被证明有效，并产生了相关的初步数据。在AIM3下，将通过染色质亲和力纯化方法鉴定出具有可能作用在G4 DNA处抑制不稳定性的新型反激活因子。 G4基序的稳定性与癌症的TH生物学特别相关，因为含有G4序列的基因组基因座与在各种人类癌症中观察到的易位部位存在显着重叠。拟议的研究的结果最终可以增强我们对煽动肿瘤重排的分子事件以及与治疗相关的继发性恶性肿瘤或耐药性肿瘤发育的理解。