BASAL LEVELS OF METALLOTHIONEIN I AND II EXPRESSION IN MOUSE EMBRYO FIBROBLASTS ENHANCE GROWTH IN LOW FOLATE THROUGH A CELL CYCLE MEDIATED PATHWAY

The impact of basal (non‐induced) expression levels of metallothionein I and II on the growth of mouse embryo fibroblasts in standard DMEM/F‐12 containing 8.8μm folic acid, and in DMEM/F12 without hypoxanthine, thymidine or folic acid, containing 15n m or 15p m[6S]‐folinic acid, was assessed by comparing wild‐type MT (+/+) and homozygous null MT (−/−) cell lines. No difference in growth rate was observed between the two in DMEM/F12, although MT (−/−) cells displayed a 6‐fold decrease in p27 Kip1 , a two fold increase in p53 and a slight increase in p21 Waf1 . After 6 days in culture, the growth rate for MT (−/−) cells in 15n m or 15p m[6S]‐folinic acid was half that of MT (+/+). After an additional 6 days in 15n m folate, both MT (+/+) and (−/−) cells maintained their respective growth rates, while those in 15p m had ceased to grow. During the initial 6 days in 15n m folate, neither cell population displayed an increase in apoptosis or a change in cell cycle distribution, even though MT (−/−) cells sustained an additional 4‐fold increase in p21 Waf1 and a 6‐fold decrease in cyclin E expression. At day 12, however, the MT (−/−) population, but not MT (+/+), underwent a 7‐fold increase in apoptosis coupled with a 3 fold increase in S phase cells. Hence, the basal level of MT I and II constitutively expressed in MT (+/+) cells enhances growth in 15nM [6S]‐folinic acid by preventing S phase arrest and apoptosis.