Choo C K, Rorke E A, Eckert R L
Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
Exp Cell Res. 1993 Sep;208(1):161-9. doi: 10.1006/excr.1993.1234.
Calcium is an important regulator of normal human cervical cell differentiation; however, it is not known whether the calcium responsiveness of the cells would be altered by immortalization with human papillomavirus. In the present study we examine the effects of extracellular calcium on morphology, expression of biochemical markers of cervical cell differentiation, and HPV16 oncogene transcription in an HPV16-immortalized human cervical cell line, ECE16-1. ECE16-1 cells respond to increased calcium with increased levels of mRNA encoding cytokeratin K13, transglutaminase, and involucrin. Dose-response studies indicate that these markers are coordinately regulated and that maximal levels are present at calcium concentrations of > or = 0.4 mM. RNA levels begin to increase within 24 h after cells are shifted to medium containing high calcium and are maximal by 3 days. Protein levels change directly with the change in mRNA, suggesting that mRNA level determines protein concentration; however, transcription runoff experiments indicate that the increased mRNA does not result from new synthesis. Parallel studies indicate that expression of the HPV16 oncogenes, E6 and E7, are not regulated by extracellular calcium. These results are consistent with in vivo experiments showing that in advanced cervical intraepithelial neoplasia, where HPV16 DNA is integrated in host cell DNA, E6/E7 transcript production is not regulated in a differentiation-dependent manner.
钙是正常人类宫颈细胞分化的重要调节因子；然而，尚不清楚人乳头瘤病毒永生化是否会改变细胞的钙反应性。在本研究中，我们检测了细胞外钙对人乳头瘤病毒16型（HPV16）永生化的人宫颈细胞系ECE16 - 1的形态、宫颈细胞分化生化标志物表达以及HPV16癌基因转录的影响。ECE16 - 1细胞对钙增加的反应是，编码细胞角蛋白K13、转谷氨酰胺酶和内披蛋白的mRNA水平升高。剂量反应研究表明，这些标志物受到协同调节，在钙浓度≥0.4 mM时达到最高水平。细胞转移至高钙培养基后24小时内，RNA水平开始升高，3天时达到最高。蛋白质水平随mRNA的变化而直接变化，表明mRNA水平决定蛋白质浓度；然而，转录延伸实验表明，mRNA增加并非新合成所致。平行研究表明，HPV16癌基因E6和E7的表达不受细胞外钙的调节。这些结果与体内实验一致，体内实验表明，在HPV16 DNA整合到宿主细胞DNA中的晚期宫颈上皮内瘤变中，E6/E7转录产物的产生不受分化依赖性调节。
