Suppression of translocations by RecQ-like DNA helicases

描述（由申请人提供）：某些具有极端癌症风险的人类遗传疾病，如布鲁姆综合征和沃纳综合征，与RecQ家族DNA解旋酶突变有关。缺乏Sgs1（该酵母中唯一与RecQ相关的DNA解旋酶）的酵母突变体已被证明是Bloom's综合征和Werner综合征的一些细胞表型的优秀模型系统，特别是关于它们的超重组表型。尽管在sgs1突变体中，累积的总染色体重排率仅适度升高，但最近的研究表明，缺乏sgs1的细胞在高度分化的基因（60- 65%同一性）中，特别容易发生复杂的、反复发生的易位。虽然Sgs1和错配修复蛋白是相似但不相同（同源）序列之间重组的抑制剂，但只有Sgs1才能抑制这些复合物和反复易位。本提案的目的是研究在没有Sgs1的情况下引起同源性驱动易位的染色体内和染色体间重组机制。本研究的具体目的是：(1)确定缺乏DNA损伤检查点传感器或染色质组装因子的sgs1突变体中基因结构和染色体环境在多大程度上控制同源性驱动易位的速率和结构。这将通过修改易位靶点的位置、取向和拷贝数以及同源块距和DNA序列一致性来实现。(2)阐明在sgs1突变体中抑制和形成反复易位的dna损伤检查点组分的不同需求。(3)研究Sgs1的功能域和已知的物理相互作用在抑制同源性驱动易位中的作用。(4)确定5种人类recq样DNA解旋酶替代Sgs1抑制同源性驱动易位的能力。SGS1将被人类RecQ同源的cdna取代，其中一些已经在酵母中成功表达并抑制了SGS1突变表型的某些方面。这些研究将提供关于RecQ解旋酶在维持基因组完整性中的作用的机制见解，并将阐明导致染色体重排的一般机制，特别是基因易位，这通常与人类癌症有关。公共卫生相关性：基因组不稳定是大多数癌症的标志，其原因尚不清楚。拟议的研究将为DNA解绕酶在维持基因组完整性中的作用提供机制见解，并将阐明导致染色体重排的一般机制，特别是基因易位，这通常与人类癌症和染色体断裂综合征有关。

DESCRIPTION (provided by applicant): Certain human genetic disorders with extreme cancer risk, such as Bloom's syndrome and Werner syndrome, are associated with mutations in DNA helicases of the RecQ family. Mutants of the yeast Saccharomyces cerevisiae that lack Sgs1, the only RecQ- related DNA helicase in this yeast, have proven to be excellent model systems for some cellular phenotypes of the Bloom's and Werner syndromes, especially with respect to their hyperrecombination phenotype. Although rates of accumulating gross- chromosomal rearrangements are only moderately elevated in sgs1 mutants, it was recently shown that cells lacking Sgs1 are uniquely susceptible to undergoing complex, recurring translocations driven by small regions of homology in highly diverged genes (60-65 % identity). Although Sgs1 and mismatch repair proteins are inhibitors of recombination between similar but nonidentical (homeologous) sequences, only Sgs1 is required for the suppression of these complexes and recurring translocations. The objective of this proposal is to study the intra- and interchromosomal recombination mechanisms that cause homeology-driven translocations in the absence of Sgs1. The specific aims of this study are to: (1) Determine the extent to which gene structure and chromosome environment control the rate and structure of homeology-driven translocations in sgs1 mutants lacking DNA damage checkpoint sensors or chromatin assembly factors. This will be achieved by modifying location, orientation and copy number of translocation targets as well as homology block distance and DNA sequence identity. (2) Elucidate the differential requirement of DNA-damage checkpoint components for the suppression and formation of recurring translocations in sgs1 mutants. (3) Examine the role of functional domains and known physical interactions of Sgs1 in the inhibition of homeology-driven translocations. (4) Determine the ability of the five human RecQ-like DNA helicases to substitute for Sgs1 in the suppression of homeology-driven translocations. SGS1 will be replaced with cDNAs of human RecQ homologs, some of which have been successfully expressed in yeast and suppress some aspects of the sgs1 mutant phenotype. These studies will provide mechanistic insights into the role of RecQ helicases in the maintenance of genome integrity and will shed light on the general mechanisms leading to gross-chromosomal rearrangements, especially gene translocations, which are often associated with human cancers. PUBLIC HEALTH RELEVANCE: The causes of genome instability, which is a hallmark of most cancers, are still unclear. The proposed studies will provide mechanistic insights into the role of DNA unwinding enzymes in the maintenance of genome integrity and will shed light on the general mechanisms leading to chromosomal rearrangements, especially gene translocations, which are often associated with human cancers and chromosome breakage syndromes.