ACTIVATION OF PROTO-ONCOGENES ENCODING GROWTH FACTOR RECEPTOR PROTEINS

与v-erbB采用宽松严格的杂交条件 作为探测器，我们之前已经发现了一个新的成员 人基因组中ErbB/EGF受体基因家族的扩增 一种乳腺癌。在一系列人乳腺肿瘤细胞中 线条，转录分析显示表达水平升高 ErbB-2的表达水平是正常水平的8到128倍 对照组8例。一个异常大小的erbB-2 在这些细胞系中未检测到转录本。免疫印迹 分析表明，185kd的erbB-2产物水平升高 由这些细胞表达。在四个品系中，erbB-2基因扩增 在没有明显基因重排的情况下 演示了。代表性细胞中扩增的基因拷贝 SK-BR-3系定位于异常染色体上 原位杂交定位。在另外四个细胞系中，4-到 在缺失的情况下，可以观察到8倍的erbB-2mRNA的过度表达 基因扩增。在具有代表性的细胞系7R-75-1中， 确定了erbB-2的正常染色体定位。 此外，在10%的患者中观察到了erbB-2基因的扩增。 对人乳腺肿瘤组织进行分析。这些发现与 一种表面正常的erbB-2基因产物的过表达 人类乳腺肿瘤。为了评估转型 这种生长因子受体样基因的潜力，我们介绍了 用DNA方法测定NIH/3T3细胞中erbB-2的正常编码序列 表达不同表达的基因产物的转染法 级别。在SV40启动子的作用下，该基因缺乏转化 尽管表达了erbB-2蛋白水平，但仍具有活性。A五比一 在LTR影响下，其表达增加了10倍 与作为有效癌基因的erbB-2的激活有关。这个 乳腺癌组织中erbB-2蛋白表达水平较高 扩增了erbB-2基因的细胞系。

Employing relaxed stringency hybridization conditions with v-erbB as probe, we previously had identified a novel member of the erbB/EGF receptor gene family in the human genome amplified in a mammary carcinoma. In a series of human mammary tumor cell lines, transcript analysis demonstrated elevated expression levels of erbB-2 ranging from 8- to 128-fold above those of normal controls in 8 out of 16 cases. An aberrantly sized erbB-2 transcript was not detected in these cell lines. Immunoblot analysis indicated elevated levels of the 185-kd product of erbB-2 expressed by these cells. In four lines, erbB-2 gene amplification in the absence of an apparent gene rearrangement was demonstrated. Amplified gene copies in a representative cell line, SK-BR-3, were localized in an aberrant chromosomal location by in situ hybridization. In four additional cell lines, 4-to 8-fold erbB-2 mRNA overexpression was observed in the absence of gene amplification. In a representative cell line, 7R-75-1, normal chromosomal location of erbB-2 was determined. Moreover, gene amplification of erbB-2 was observed in 10% of human mammary tumor tissues analyzed. These findings linked overexpression of an apparently normal erbB-2 gene product with human mammary neoplasia. In order to assess the transforming potential of this growth factor receptor-like gene, we introduced the normal coding sequence of erbB-2 in NIH/3T3 cells by DNA transfection expressing the gene product at different expression levels. Under SV40 promoter, the gene lacked transforming activity despite expression of erbB-2 protein levels. A five to tenfold increase in its expression under LTR influence was associated with activation of erbB-2 as a potent oncogene. The higher levels of erbB-2 protein were observed in mammary tumor cell lines with erbB-2 gene amplification.