Regulation of the Gl-S-Phase Transition in Non-Transformed and Transformed Human Breast Epithelial Cells

摘要

The goal of this work was to study the regulation of the thymidine kinase (TK) gene in non-tumorigenic and tumorigenic human breast epithelial cells. Previous studies in our laboratory had shown that this gene is regulated during the cell cycle by both transcriptional and post-transcriptional mechanisms. The approach used in the work reported here was to examine the expression of 2 hybrid genes. TK-Luc, which contains the human TK promoter linked to a luciferase reporter gene, was used to study transcriptional regulation. CMV-TK(*), which contains an epitope-tagged human TK cDNA expressed from a constitutive CMV promoter, was used to study post-transcriptional regulation. These constructs were stably transfected into non-tumorigenic (184B5) and tumorigenic (MCF-7) human breast epithelial cells. No transfectants were obtained in the 184B5 cells, despite using several transfection protocols. The TK-Luc construct was expressed at low, constitutive levels in MCF-7 cells, indicating that the TK promoter fragment used was not sufficient to confer high, regulated levels of expression to linked genes in these cells. The CMV-TK(*) construct was regulated by serum, but not by estrogen/antiestrogen treatment, indicating that post-transcriptional regulation of this gene occurs in response to serum growth factors, and not as a function of cell cycle position.