STRUCTURE AND BEHAVIOR OF YEAST TELOMERES

端粒是真核生物染色体的物理末端， 线性染色体的稳定维持和完全复制。 的 端粒的特殊结构和归因于端粒的各种功能， 他们认为，多个基因产物参与了它们的复制， 和功能 描述了遗传和生物化学方法来鉴定和 表征与酵母端粒相互作用的蛋白质 啤酒。 Yalpha26和RAP 1，两个端粒编码基因的候选者 结合交叉反应与已知结合的蛋白质产生的抗血清 特别是纤毛原生动物新尖毛藻的端粒。 此外，Yalpha 26的过量拷贝影响酵母中的端粒长度。 DNA 测序将揭示Yalpha26是否真的与尖毛藻基因有关。 无效和过表达等位基因的表型对生活力、端粒 长度和染色体稳定性也将被确定。 RAP 1，一个 必需基因，在体外对端粒序列也具有高亲和力 对于各种基因的转录控制区。 到 确定RAP 1蛋白是否在体内与端粒相互作用， 表型（端粒长度，染色体稳定性，转录水平） 将确定RAP 1过表达或缺陷的菌株。 生物化学方法也将被用来鉴定相互作用的蛋白质 有端粒。 染色体端粒的染色质结构将是 在正常细胞和在所示条件下生长的细胞中测定 影响端粒结构。 亲和纯化方法 端粒染色质的结构将得以发展， 如果Yalpha26（或RAP 1）在端粒上被发现， 鉴定其他端粒相关蛋白。 最后，六个lps（线性 质粒稳定性）突变体，所述突变体在 频率升高，但在野生型率失去环状染色体将是 通过分子生物学和遗传学方法进行分析，以确定 他们的产品如何与酵母端粒相互作用。

Telomeres, the physical ends of eukaryotic chromosomes, are required for the stable maintenance and complete duplication of linear chromosomes. The special structure of telomeres and the variety of functions ascribed to them suggest that multiple gene products are involved in their replication and function. Genetic and biochemical approaches are described to identify and characterize proteins that interact with the telomeres of Saccharomyces cerevisiae. Yalpha26 and RAP1, two candidates for genes encoding telomere binding cross reacts with antiserum raised to a protein known to bind specifically to telomeres in the ciliated protozoan Oxytricha nova. Moreover, excess copies of Yalpha 26 affect telomere length in yeast. DNA sequencing will reveal if Yalpha26 is indeed related to the Oxytricha gene. The phenotype of null and overexpressing alleles on viability, telomere lengths, and chromosome stability will also be determined. RAP1, an essential gene, has high affinity in vitro for telomeric sequences as well as for the transcription control regions of a variety of genes. To determine if the RAP1 protein interacts with telomeres in vivo, the phenotype (telomere lengths, chromosome stability, transcription levels) of strains overexpressing or deficient in RAP1 will be determined. Biochemical methods will also be used to identify proteins that interact with telomeres. The chromatin structure of a chromosomal telomere will be determined in normal cells and in cell grown under conditions shown previously to affect telomere structure. Methods for affinity purification of telomeric chromatin will be developed that will enable us to ask directly if Yalpha26 (or RAP1) is found on telomeres, and, ultimately, to identify other telomere associated proteins. Finally, six lps (linear plasmid stability) mutants that lose linear artificial chromosomes at elevated frequency but lose circular chromosomes at wild-type rates will be analyzed by molecular biological and genetic methods to determine if and how their products interact with yeast telomeres.