DNA REPAIR IN CANCER AND SENESCENCE

癌症和衰老中的 DNA 修复

• 批准号: 6097867
• 负责人: VILHELM A. BOHR
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6097867
• 关键词: DNA damage; DNA directed RNA polymerase; DNA repair; Werner's syndrome; aging; biomarker; cell line; genetic transcription; helicase; human genetic material tag; phenotype; premature aging; protein purification; transfection

Summary of work: Werner's Syndrome (WS) is a homozygous recessive disease characterized by early onset of many characteristics of normal aging, such as wrinkling of the skin, graying of the hair, cataracts, diabetes, and osteoporosis. The symptoms of WS begin to appear around the age of puberty, and most patients die before age 50. Because of the acceleration of aging in WS, the study of this disease will hopefully shed light on the degenerative processes that occur in normal aging. Cells from WS patients grow more slowly and senescence at an earlier population doubling than age-matched normal cells, possibly because these cells appear to lose the telomeric ends of their chromosomes at an accelerated rate. In general, WS cells have a high level of genomic instability, with increased amounts of DNA deletions, insertions, and rearrangements. These effects could potentially be the result of defects in DNA repair, replication, and/or recombination, although the actual biochemical defect remains unknown. The gene that is defective in WS, the WRN gene, has recently been identified and characterized as a helicase. Thus, the genetic evidence also points to a role for the WRN protein in some aspect of DNA metabolism. We are using several avenues to identify and characterize the biochemical defect in WS cells. WS cells appear to have a subtle defect in transcription-coupled repair, the highly efficient removal of lesions from the transcribed strand of active genes. We have taken various approaches to analysis of transcriptional capacity in Werners cells and extracts. We observe a defect which can be complemented by addition of purified werners protein. We are making purified WRN protein for use in a number of basic and complex biochemical assays. The protein has helicase activity and will unwind small and large DNa duplex constructs. The helicase activity is sensitive to a compound, distamycin, which is a DNA minor groove binder, and the helicase will work on blunt duplexes. We are currently characterizing the helicase activity and searching for proteins that will enhance it and thus may be interacting with the Werner helicase. Our ongoing and future studies will be directed towards elucidation of the causes of the accelerated aging phenotype in WS, with hope that this knowledge can also be applied to our current understanding of both the aging of cells and organisms in general.

Werner综合征（WS）是一种 一种以多个基因的早期发病为特征的纯合子隐性疾病 正常衰老的特征，如皮肤的皱纹， 头发变白、白内障、糖尿病和骨质疏松症。的 WS的症状开始出现在青春期左右， 大多数患者在50岁之前死亡。由于加速度 在WS中的衰老，对这种疾病的研究将有望阐明 在正常衰老过程中发生的退化过程。细胞 WS患者生长较慢，衰老较早， 与年龄匹配的正常细胞相比， 因为这些细胞似乎失去了端粒末端， 染色体加速繁殖一般来说，WS细胞具有 基因组高度不稳定，DNA含量增加 缺失、插入和重排。这些影响可以 可能是DNA修复，复制， 和/或重组，尽管实际的生化缺陷 仍然未知。WS中有缺陷的基因WRN 基因，最近已被确定和表征为解旋酶。 因此，遗传证据也指出了WRN的作用 蛋白质在DNA代谢的某些方面。我们正在使用几个 识别和表征WS生化缺陷的途径 细胞WS细胞似乎有一个微妙的缺陷， 转录偶联修复， 从活性基因的转录链中分离出来我们采取了各种 分析Werners细胞转录能力的方法 和提取物。我们观察到一个缺陷，可以补充， 加入纯化的Werners蛋白。我们正在制作纯化的WRN 蛋白质用于许多基本和复杂的生物化学 测定。该蛋白质具有解旋酶活性， 大DNA双链体构建体。解旋酶活性对a 化合物偏端霉素，其为DNA小沟结合剂，和 解旋酶将作用于平的双链体。我们目前正在 表征解旋酶活性并寻找 会增强它，因此可能会与沃纳 解旋酶我们正在进行的和未来的研究将针对 阐明WS中加速老化表型的原因， 希望这些知识也能应用到我们目前的 了解细胞和生物体的衰老。