Abstract
Metallothionein (MT) gene expression is increased in cadmium resistant Chinese hamster ovary cells (CHO Cd R ) upon medium (regular or serum-free) change during culturing. Among the major components of the medium, NaHCO 3 was found to be able to induce MT gene expression in a dose- and time-dependent manner. The same effect was observed with other alkaline solutions, such as HEPES and NaOH. Using MT promoter-luciferase reporter gene constructs, we found that the presence of metal response elements (MREs) in the promoter region is necessary for NaHCO 3 -induced MT gene transcription. This finding is further supported by the observation that the binding activity between the metal-responsive transcription factor 1 (MTF-1) and the MRE were increased after NaHCO 3 treatment. Following NaHCO 3 treatment, an increase in cell proliferation was observed in Cd R cells but not in the parental CHO K1 cells that do not express MT transcripts due to MT gene methylation. Using synchronized cells, an increase in cell proliferation was observed 9 h after NaHCO 3 addition. Notably, proliferation of CHO K1 cells was increased when transfected with an MT gene. The effect of MT on cell growth was affirmed by treating CHO K1 cells with 5-azacytidine (Aza) to demethylate the MT gene. Proliferation increased in Aza-treated CHO K1 cells after NaHCO 3 treatment. These results demonstrate that NaHCO 3 stimulates MT gene expression and causes an enhancement of cell proliferation in CHO cells. © 2005 Wiley-Liss, Inc.