Expression of cyclin D1 mRNA is not upregulated by Myc in rat fibroblasts

Conflicting results have been published regarding the regulation of cyclin D1 mRNA in rat fibroblasts expressing a hormone-regulated Myc protein, MycER. We confirm that activation of MycER with oestrogen rapidly induces cyclin D1 mRNA, even in the presence of cycloheximide. However, we show that this is an artefact resulting from an oestrogen-activated transcriptional activation domain in the oestrogen receptor part of the MycER chimaera. First, addition of 4-hydroxy-tamoxifen (4OHT), which does not activate this domain, allows association of MycER with Max and induces cell proliferation in serum-starved Rat-1-MycER cells without affecting cyclin D1 mRNA levels or the activity of D1 promoter-luciferase constructs. Second, Rat-1 cells expressing a mutant MycER with a hormone-binding domain that still binds 4OHT but no longer binds oestrogen, are driven into the cell cycle in response to 4OHT but fail to up-regulate cyclin D1 mRNA. Finally, Rat-1 cells in which wild-type human c-Myc expression can be induced, also progress into the cell cycle without increased D1 mRNA expression.