EFFECTS OF RADIATION ON MAMALIAN CELLS

这项研究计划的长期目标是获得知识 关于应对气候变化所涉机制的报告 哺乳动物细胞的辐射，特别是其 致癌和致突变作用。 该方法是一个多- 多方面的，调查的终点包括 诱变、恶性转化、细胞存活和 诱发染色体异常。 该计划有一个双重 目的：第一，更好地界定低水平辐射的风险 通过检查总剂量等因素的作用， 剂量率、LET和细胞内剂量分布 诱变和转化;第二，了解更多关于 这些效应的细胞和分子机制。 的 具体的目标是：1）识别癌基因激活过程中， 辐射引起的恶性转化过程。 转染 定期将DNA导入NIH 3 T3细胞 照射10 T1/2细胞培养物后， 将通过Southern印迹法检查次级转染子， 对特异性单克隆抗体的反应性。 2)审查 各种类型的电离辐射的直接致突变效应 通过转染照射的细胞， 质粒载体中的基因。 3)为了研究影响 人的形态转化和永生化 二倍体成纤维细胞 一个特别的重点将是研究一个 来自c-myc第二内含子的DNA片段， 诱导部分菌株的特定形态学改变， 转化细胞 我们建议确定这一假定 通过缺失分析和测序的调节元件，和 进一步研究其在转化中的作用。 4)分析 自发和诱导突变的分子结构 TK 6人细胞系中的常染色体tk基因座。 适当 将获得探针，并将限制性片段或序列 寻找多态性标记。 分子生物学技术将 以确定是否采用大规模 我们观察到的变化包括基因转换，有丝分裂， 重组或多基因座缺失事件，并估计 他们的程度。 5)对突变株进行细胞遗传学分析 克隆，以确定是否大规模的DNA结构 这些变化与可见的染色体异常有关。 结果将与剂量印迹结果相关。 6)到 分析由以下物质引起的分子结构变化的光谱 不同类型的辐射，包括快中子，重离子 和俄歇放射性核素。 7)继续研究 转化的相对生物有效性， 低剂量、低剂量率中子辐照诱变。 8)到 研究高LET剂量微分布的作用 用俄歇仪研究细胞内辐射对生物学效应的影响 发射电子的放射性核素。 重点是诱变 在人类TK 6细胞中。

The long term goal of this research program is to gain knowledge concerning the mechanisms involved in the response of mammalian cells to radiation, with particular reference to its carcinogenic and mutagenic effects. The approach is a multi- faceted one, and the endpoints under investigation include mutagenesis, malignant transformation, cell survival and the induction of chromosomal abnormalities. The program has a dual purpose: first, to better define the risks of low-level radiation exposure by examining the role of factors such as total dose, dose-rate, LET and the intracellular distribution of dose on mutagenesis and transformation; and second, to learn more about the cellular and molecular mechanisms for these effects. The specific aims are: 1) To identify oncogenes activated during the process of malignant transformation by radiation. Transfection of DNA into NIH 3T3 cells will be carried out at regular intervals after irradiation of 10T1/2 cell cultures; DNA from primary and secondary transfectants will be examined by Southern blotting and reactivity to specific monoclonal antibodies. 2) To examine the direct mutagenic effects of various types of ionizing radiation on the c-Ha-ras proto-oncogene by transfection of the irradiated gene in a plasmid vector. 3) To examine factors involved in the morphological transformation and immortalization of human diploid fibroblasts. A particular focus will be on the study of a DNA fragment derived from the second intron of c-myc which induces specific morphologic alterations in a strain of partially transformed cells. We propose to identify this presumptive regulatory element by deletion analysis and sequencing, and furthur study its role in transformation. 4) To analyze the molecular structure of spontaneous and induced mutations at the autosomal tk locus in the TK6 human cell line. Appropriate probes will be obtained and restriction fragment or sequence polymorphic markers sought. Molecular biological techniques will be employed in order to determine whether the large-scale changes we have observed involve gene conversion, mitotic recombination or multi-locus deletion events and to estimate their extent. 5) To carry out cytogenetic analysis of mutant clones in order to determine whether large scale DNA structural changes are associated with visible chromosomal abnormalities. Results will be correlated with those of dosage blots. 6) To analyse the spectrum of molecular structural changes induced by different types of radiation including fast neutrons, heavy ions and Auger emitting radionuclides. 7) To continue studies of the relative biological effectiveness for transformation and mutagenesis of low dose, low dose-rate neutron exposure. 8) To investigate the role of microdistribution of dose of high LET radiation within the cell on its biologic effects by use of Auger electron emitting radionuclides. Emphasis will be on mutagenesis in human TK6 cells.