TWO-DIMENSIONAL GEL ANALYSIS OF ONCOGENE-MEDIATED TRANSFORMATION

本项目的目的是研究致癌机制。 采用定量双向凝胶电泳法。这 技术可以让我们检查质量和数量上的变化 细胞经历过程中数千种细胞多肽的合成 肿瘤性转化。研究的重点是(1)继续 凝胶分析计算机系统(简称ELSSIE)的开发和(2) 用ELSIE软件分析需要计算机分析的实验 2DG。埃尔西系统正在实验室中用于研究 不同转化癌基因对蛋白质合成的影响。 大鼠肝上皮(RLE)细胞的分离和单细胞培养 克隆的。这些细胞具有正常的二倍体核型，代表一种 均质、克隆系统用于转化研究。不同 含有转化癌基因的逆转录病毒，如v-H-ras，v-N-ras， V-raf或v-myc/v-raf嵌合体已被用于转化RLE细胞或 人二倍体成纤维细胞系(LG-1)。大黄鱼的多肽图谱 转化的细胞与未转化的细胞显著不同 祖先。虽然质量差异很少，但超过30%的 斑点可能在数量上有所不同。非常相似的癌基因，如v-H-ras 和v-N-ras，导致基本上无法区分的2DG模式。这些 模式不同于v-raf和v-myc/v-raf介导的模式 转型。V-raf和v-myc/v-raf模式也很容易 可区分的，但彼此更相似，而不是 未转化或转化为ras的细胞。

The objective of this project is to study the mechanism of carcinogenesis using quantitative two-dimensional gel electrophoresis (2DG). This technique lets us examine both qualitative and quantitative changes in the synthesis of thousands of cellular polypeptides as the cell undergoes neoplastic transformation. Research is focused on (1) the continued development of the computer system (dubbed ELSIE) to analyze gels and (2) the use of ELSIE to analyze experiments requiring computerized analysis of 2DGs. The ELSIE system is being used in the laboratory to study the effects of different transforming oncogenes on the synthesis of proteins. Rat liver epithelial (RLE) cells have been isolated and single-cell cloned. These cells have a normal diploid karyotype and represent a homogeneous, clonal system for the study of transformation. Different retrovirus containing transforming oncogenes, such as v-H-ras, v-N-ras, v-raf, or a v-myc/v-raf chimera have been used to transform RLE cells or a human diploid fibroblast line (LG-1). The polypeptide patterns of the transformed cells differ significantly from their non-transformed progenitors. While qualitative differences are rare, over 30% of the spots may vary quantitatively. Very similar oncogenes, such as v-H-ras and v-N-ras, cause essentially indistinguishable 2DG patterns. These patterns are distinct from those caused by v-raf and v-myc/v-raf mediated transformation. The v-raf and v-myc/v-raf patterns are also readily distinguishable, but are more similar to each other than to non-transformed or to ras transformed cells.