Albanese C, Johnson J, Watanabe G, Eklund N, Vu D, Arnold A, Pestell R G
Division of Endocrinology, Metabolism, and Molecular Medicine, Northwestern University Medical School, Chicago, Illinois 60611, USA.
J Biol Chem. 1995 Oct 6;270(40):23589-97. doi: 10.1074/jbc.270.40.23589.
Several different oncogenes and growth factors promote G1 phase progression. Cyclin D1, the regulatory subunit of several cyclin-dependent kinases, is required for, and capable of shortening, the G1 phase of the cell cycle. The present study demonstrates that transforming mutants of p21ras (Ras Val-12, Ras Leu-61) induce the cyclin D1 promoter in human trophoblasts (JEG-3), mink lung epithelial (Mv1.Lu), and in Chinese hamster ovary fibroblast cell lines. Site-directed mutagenesis of AP-1-like sequences at -954 abolished p21ras-dependent activation of cyclin D1 expression. The AP-1-like sequences were also required for activation of the cyclin D1 promoter by c-Jun. In electrophoretic mobility shift assays using nuclear extracts from cultured cells and primary tissues, several AP-1 proteins (c-Jun, JunB, JunD, and c-Fos) bound the cyclin D1 -954 region. Cyclin D1 promoter activity was stimulated by overexpression of mitogen-activated protein kinase (p41MAPK) or c-Ets-2 through the proximal 22 base pairs. Expression of plasmids encoding either dominant negative MAPK (p41MAPKi) or dominant negatives of ETS activation (Ets-LacZ), antagonized MAPK-dependent induction of cyclin D1 promoter activity. Epidermal growth factor induction of cyclin D1 transcription, through the proximal promoter region, was antagonized by either p41MAPKi or Ets-LacZ, suggesting that ETS functions downstream of epidermal growth factor and MAPK in the context of the cyclin D1 promoter. The activation of cyclin D1 transcription by p21ras provides evidence for cross-talk between the p21ras and cell cycle regulatory pathways.
几种不同的癌基因和生长因子可促进G1期进程。细胞周期蛋白D1是几种细胞周期蛋白依赖性激酶的调节亚基，是细胞周期G1期所必需的，并且能够缩短该时期。本研究表明，p21ras的转化突变体（Ras Val-12、Ras Leu-61）可在人滋养层细胞（JEG-3）、貂肺上皮细胞（Mv1.Lu）和中国仓鼠卵巢成纤维细胞系中诱导细胞周期蛋白D1启动子。-954处AP-1样序列的定点诱变消除了p21ras依赖性的细胞周期蛋白D1表达激活。c-Jun激活细胞周期蛋白D1启动子也需要AP-1样序列。在使用培养细胞和原代组织的核提取物进行的电泳迁移率变动分析中，几种AP-1蛋白（c-Jun、JunB、JunD和c-Fos）与细胞周期蛋白D1 -954区域结合。有丝分裂原激活蛋白激酶（p41MAPK）或c-Ets-2的过表达通过近端22个碱基对刺激细胞周期蛋白D1启动子活性。编码显性负性MAPK（p41MAPKi）或ETS激活显性负性蛋白（Ets-LacZ）的质粒表达可拮抗MAPK依赖性的细胞周期蛋白D1启动子活性诱导。p41MAPKi或Ets-LacZ拮抗表皮生长因子通过近端启动子区域对细胞周期蛋白D1转录的诱导，这表明在细胞周期蛋白D1启动子的背景下，ETS在表皮生长因子和MAPK的下游发挥作用。p21ras对细胞周期蛋白D1转录的激活为p21ras与细胞周期调节途径之间的相互作用提供了证据。
