CONTROL MECHANISMS IN HUMAN TUMOR CELLS--SMALL RNAS

我们正在研究主要环境致癌物的一些影响， 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前所未有的观察， 紫外辐射对新合成的小核RNA积累的影响 （snRNA）种U1，U2，U3，U4和U 5：（1）尽管U snRNA很小， (for例如，人U1 RNA是165个核苷酸长），它们的生物合成是 对紫外线非常敏感;（2）它们的灭活曲线 合成是双相的，而不是通常的单斜率;和（3） U1-U 5 snRNA合成的抑制在前两小时内增加 紫外线照射后（尽管抑制的转录通常恢复到 正常的，因为嘧啶二聚体从DNA中除去）。 我们现在希望调查这些令人困惑的背后的机制 意见。 这包括确定U snRNA内的步骤 受UV辐射影响的生物合成途径，例如，启动， 延伸或转录终止？）。 另一个目标是找出 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，对辐射后修复的超敏反应 DNA中的缺口，或随之而来的DNA扭转应力的变化。 额外 目的是确定获得所需的基因组DNA的部分， UV辐射对U1-U 5 snRNA合成的抑制作用。 的 使用的方法是：用全细胞进行放射性标记实验 包括杂交选择测定;在分离的细胞核中的转录和在细胞中的转录。 一种可溶性体外系统;胸苷激酶缺陷的转录 用克隆的U1基因转染的细胞。 （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)