Goya L, Maiyar A C, Ge Y, Firestone G L
Department of Molecular and Cell Biology, University of California, Berkeley 94720.
Mol Endocrinol. 1993 Sep;7(9):1121-32. doi: 10.1210/mend.7.9.8247014.
Con8 mammary tumor cells are an epithelial cell line derived from the 7,12-dimethylbenz(alpha)anthracene-induced 13762NF rat mammary adenocarcinoma. The synthetic glucocorticoid dexamethasone suppresses the growth of Con8 cells, and after 5 days of treatment with this steroid, Con8 cells undergo less than 0.5 population doublings. This growth arrest is accompanied by a 30-fold elevation in c-jun transcript levels, no change in c-fos expression, and a moderate increase in total AP-1 transcriptional activity. Dexamethasone inhibited DNA synthesis within one cell cycle, and flow cytometry of propidium iodide-stained nuclei demonstrated that dexamethasone growth-suppressed cells had a DNA content indicative of a specific cell cycle block in either G1 or G0. Consistent with a G1/G0 arrest of the cell cycle, dexamethasone did not prevent Con8 cells from entering the S phase after release from synchronization at the G1/S boundary by a double thymidine block. Analysis of [3H]thymidine incorporation and autoradiography of [3H]thymidine-labeled nuclei revealed that after either dexamethasone withdrawal or the addition of transforming growth factor-alpha (TGF alpha), Con8 cells synchronously reinitiate cell cycle progression. Northern blot analysis demonstrated that an induction of transcripts for the G1 marker genes c-myc and cyclin D1 occurs before cells enter the S-phase. After dexamethasone withdrawal, c-myc and cyclin D1 expression transiently peak at 2 and 4 h, respectively. In contrast, c-myc expression peaked at 0.5-1 h, whereas cyclin D1 expression was induced at 2 h and maintained at a high level after the addition of TGF alpha. Our results demonstrate that glucocorticoids induce a specific block of the cell cycle progression of a rat mammary tumor cell, and that after synchronous progression through the cell cycle, the temporal expression pattern for c-myc and cyclin D1 is distinct for dexamethasone release vs. the addition of TGF alpha to glucocorticoid-suppressed cells.
Con8乳腺肿瘤细胞是一种上皮细胞系，源自7,12-二甲基苯并(α)蒽诱导的13762NF大鼠乳腺腺癌。合成糖皮质激素地塞米松可抑制Con8细胞的生长，用这种类固醇处理5天后，Con8细胞的群体倍增数少于0.5。这种生长停滞伴随着c-jun转录水平升高30倍，c-fos表达无变化，以及总AP-1转录活性适度增加。地塞米松在一个细胞周期内抑制DNA合成，碘化丙啶染色细胞核的流式细胞术表明，地塞米松生长抑制的细胞具有指示G1或G0期特定细胞周期阻滞的DNA含量。与细胞周期的G1/G0期停滞一致，地塞米松不会阻止Con8细胞在通过双胸腺嘧啶阻断在G1/S边界同步化释放后进入S期。对[3H]胸腺嘧啶掺入的分析以及[3H]胸腺嘧啶标记细胞核的放射自显影显示，在地塞米松撤药或添加转化生长因子-α(TGFα)后，Con8细胞同步重新启动细胞周期进程。Northern印迹分析表明，在细胞进入S期之前，G1标记基因c-myc和细胞周期蛋白D1的转录本会被诱导。地塞米松撤药后，c-myc和细胞周期蛋白D1的表达分别在第2小时和第4小时短暂达到峰值。相比之下，添加TGFα后，c-myc表达在0.5-1小时达到峰值，而细胞周期蛋白D1表达在第2小时被诱导并维持在高水平。我们的结果表明，糖皮质激素可诱导大鼠乳腺肿瘤细胞的细胞周期进程出现特定阻滞，并且在通过细胞周期同步进展后，地塞米松释放与向糖皮质激素抑制的细胞中添加TGFα相比，c-myc和细胞周期蛋白D1的时间表达模式是不同的。
