CONTROL MECHANISMS IN HUMAN TUMOR CELLS--SMALL RNAS

我们正在研究主要环境致癌物质的一些影响， 紫外线(UV)辐射，对人类和其他真核细胞。三 在这个实验室里进行了史无前例的观察，观察了 紫外光照射对新合成的小核RNA积累的影响 (短链RNA)U1、U2、U3、U4和U5种：(1)尽管U短链RNA很小 (例如，人类U1 RNA长165个核苷酸)，它们的生物合成是 对紫外线非常敏感；(2)它们的失活曲线 综合是单相的，而不是通常的单斜率；以及(3) 在最初的两小时内，对U1-U5 SnRNA合成的抑制作用增加 在紫外线照射后(尽管抑制的转录通常会恢复到 当嘧啶二聚体从DNA中移除时是正常的)。 我们现在希望调查这些令人费解的背后的机制 观察。这包括确定U SnRNA内的步骤 受紫外线辐射影响的生物合成途径，例如，启动， 延长，还是终止转录？)。另一个目标是找出 在这种抑制中涉及的关键改变是什么；例如， 嘧啶二聚体的形成与辐射后修复的超敏 DNA的缺口，或随之而来的DNA扭转应力的变化。一项额外的 我们的目标是定义获得 紫外线辐射对U1-U5单链RNA合成的抑制作用。这个 可使用的方法是：整个细胞的放射性标记实验 包括杂交选择试验；在分离的核和在 一种可溶的体外系统；胸苷激酶缺陷型转录 将克隆的U1基因导入细胞。(H)

We are studying some effects of the major environmental carcinogen, ultraviolet (UV) radiation, on human and other eukaryotic cells. Three unprecedented observations were made in this laboratory about the effect of UV radiation on the accumulation of newly synthesized small nuclear RNA (snRNA) species U1, U2, U3, U4, and U5: (1) although U snRNAs are small (for example, human U1 RNA is 165 nucleotides long), their biosynthesis is very sensitive to UV radiation; (2) the inactivation curve of their synthesis is monophasic, instead of the usual single slope; and (3) the inhibition of U1-U5 snRNA synthesis increases within the first two hours after UV irradiation (although inhibited transcription usually returns to normal as pyrimidine dimers are removed from DNA). We now wish to investigate the mechanisms behind these puzzling observations. This includes determining the step within the U snRNA biosynthesis pathway which is affected by UV radiation, e.g., initiation, elongation, or termination of transcription?). Another goal is to find out what the critical alteration involved in this inhibition is; e.g., formation of pyrimidine dimers, hypersensitivity to postirradiation repair nicks in DNA, or ensuing changes in DNA torsional stress. An additional objective is to define the portion of genomic DNA required to obtain the inhibitory effects of UV radiation on U1-U5 snRNA synthesis. The approaches to be used are: radiolabeling experiments with whole cells including hybrid selection assays; transcription in isolated nuclei and in a soluble in vitro system; transcription in thymidine kinase-deficient cells that have been transfected with cloned U1 genes. (H)