TRI-NUCLEOTIDE REPEAT AND FRAGILE SITES IN YEAST

描述(改编自调查人员摘要)：一个不断增长的 人类遗传性疾病的数量被归因于 三核苷酸重复序列(TNRs)。例如脆性X综合征，第二个 智力低下的主要原因，是由于CGG束扩张和 强直性肌营养不良是由于CTG束扩张所致。此外， CGG束的扩张导致人类五条已知染色体的断裂 其中一些与人类疾病相关。染色体断裂发生在 脆性部位也与染色体重排有关 具有人类肿瘤特征的。与人类疾病相关的TNRs是 1~2成熟。TNR和脆弱部位的研究将得到基因的极大帮助 束扩张和/或染色体脆性的检测。这样做的目的是 格兰特将开发酿酒酵母作为TNR扩张的模型 和三联体介导的染色体脆性。 这位研究人员已经在酵母染色体上插入了130个重复的CTG。 这一区域得到了较大的扩张，第一次较大的扩张 在人类之外报告的。CTG束也可能是依赖于长度的脆弱的 酵母菌中的站点。这一观察结果为一种简单的、遗传学的 束扩张和染色体脆性的检测。将使用这些化验方法 鉴定突变或过度表达的酵母和人类基因 增加CTG扩展和/或脆弱性。拟议工作的一个前提是 大多数影响TNR稳定性的基因将编码参与TNR稳定性的蛋白质 复制、修复或染色质结构，其中许多是保守的 从酵母到人类。还将研究CGG区域的行为。 核酸酶和DNA甲基酶将用于分析染色质结构 CTG和CGG束，因为这两个DNA都显示异常的核小体形成 体外性质。最后，脆弱的部位被认为是由 TNR DNA的晚期复制。该模型将使用密度进行测试 转移和双向凝胶电泳法确定是否复制 时间或复制分叉进程受TNR的存在的影响。

DESCRIPTION (adapted from investigator's abstract): An ever-increasing number of human genetic diseases are attributed to expansion of trinucleotide repeats (TNRs). For example fragile X syndrome, the second leading cause of mental retardation, is due to expansion of a CGG tract and myotonic muscular dystrophy, is due to expansion of a CTG tract. Moreover, expansion of CGG tracts induce breakage at five known human chromosomal loci, some of which correlate with human disease. Chromosome breakage at fragile site is also implicated in the chromosomal rearrangements characteristic of human tumors. TNRs associated with human disease are rare. TNR and fragile site research would be greatly aided by genetic assays for tract expansion and/or chromosome fragility. The goal of this grant is to develop Saccharomyces cerevisiae as a model for TNR expansion and triplet-mediate chromosome fragility. The investigator has inserted 130 repeats of CTG onto a yeast chromosome. Large expansion of this tract were obtained, the first large expansion reported outside of humans. CTG tracts may also be length-dependent fragile sites in yeast. This observation provides the basis for a simple, genetic assay for tract expansion and chromosome fragility. The assays will be used to identify yeast and human genes whose mutation or over-expression increases CTG expansion and/or fragility. A premise of the proposed work is that most genes that affect TNR stability will encode proteins involved in replication, repair, or chromatin structure, many of which are conserved from yeast to humans. The behavior of CGG tracts will also be studied. Nucleases and DNA methylases will be used to analyze the chromatin structure of CTG and CGG tracts as both DNAs display unusual nucleosome forming properties in vitro. Finally, fragile sites are thought to be caused by late replication of TNR DNA. This model will be tested using density transfer and two-dimensional gel electrophoresis to determine if replication timing or replication fork progression is affected by the presence of TNRs.