Osei-Frimpong J, Sepulveda J, Rangdaeng S, Lebovitz R M
Department of Pathology, Baylor College of Medicine, Houston, Texas 77030.
Mol Carcinog. 1994 Jun;10(2):72-81. doi: 10.1002/mc.2940100204.
Prolonged expression of activated ras mutants resulted in both neoplastic transformation and suppression of serum-induced c-fos expression in Rat1 fibroblasts. Expression of other serum-inducible genes, including c-jun and beta-actin, was not suppressed in ras-transformed Rat1 cells, indicating that these effects are specific for c-fos and that growth-factor signal transduction pathways remain essentially intact. Run-on transcription studies indicated that c-fos transcription was blocked at the level of initiation in these cells. Transient transfection studies using 360 bp from the wild-type c-fos promoter as well as a series of mutated c-fos promoter fragments linked to the chloramphenicol acetyltransferase gene indicated that repression of c-fos was mediated by approximately 49 bp immediately upstream of the dyad symmetry element (DSE). Deletion of this region, referred to as the upstream repressor region (URR), restored serum inducibility to the c-fos promoter in ras-transformed cells. In contrast, suppression of c-fos transcription was not affected by either deletion of 240 bp between the DSE and the TATA element or by base-substitution mutations that inactive the ternary complex factor and fos-AP-1-like binding sites. In addition, in vitro competition studies indicated that ras-transformed cells express one or more repressor factors that interact with as-yet-unidentified elements within the c-fos promoter (possibly the URR) and block serum induction of c-fos. These findings suggest that prolonged expression of activated ras results in the activation of one or more as-yet-unidentified proteins that suppress transcription of the c-fos gene by interacting with the URR.
活化的ras突变体的长期表达导致Rat1成纤维细胞发生肿瘤转化并抑制血清诱导的c-fos表达。在ras转化的Rat1细胞中，包括c-jun和β-肌动蛋白在内的其他血清诱导基因的表达未受抑制，这表明这些效应是c-fos特有的，且生长因子信号转导途径基本保持完整。连续转录研究表明，在这些细胞中，c-fos转录在起始水平被阻断。使用来自野生型c-fos启动子的360 bp以及与氯霉素乙酰转移酶基因相连的一系列突变的c-fos启动子片段进行的瞬时转染研究表明，c-fos的抑制是由紧邻二元对称元件（DSE）上游约49 bp介导的。删除该区域（称为上游抑制区域，URR）可恢复ras转化细胞中c-fos启动子的血清诱导性。相比之下，DSE和TATA元件之间240 bp的缺失或使三元复合因子和fos-AP-1样结合位点失活的碱基取代突变均不影响c-fos转录的抑制。此外，体外竞争研究表明，ras转化细胞表达一种或多种抑制因子，这些因子与c-fos启动子内尚未鉴定的元件（可能是URR）相互作用，并阻断c-fos的血清诱导。这些发现表明，活化的ras的长期表达导致一种或多种尚未鉴定的蛋白质活化，这些蛋白质通过与URR相互作用抑制c-fos基因的转录。
