Cellular adaptation to down-regulated iron transport into lymphoid leukaemic cells: effects on the expression of the gene for ribonucleotide reductase

Ribonucleotide reductase is an iron-containing enzyme that is essential for DNA synthesis. Whereas previous studies have used various iron chelators t o examine the relationship between cellular iron metabolism and ribonucleot ide reductase activity in cells, they have not elucidated the relationship between iron transport into cells and the expression of the gene for ribonu cleotide reductase. To investigate this, we examined ribonucleotide reducta se mRNA, protein and enzyme activity in a novel line of CCRF-CEM cells (DFe -T cells) that display an approx. 60 %, decrease in their uptake of iron co mpared with the parental wild-type cell line. We found that DFe-T cells dis played an approx. 40 % decrease in ribonucleotide reductase specific enzyme activity relative to wild-type cells without a change in their proliferati on. Kinetic analysis of CDP reductase activity revealed an approx. 60 %, de crease in V-max in DFe-T cells without a change in K-m. Despite the decreas e in enzyme activity, the mRNA and protein for the R1 and R2 subunits of ri bonucleotide reductase in DFe-T cells were similar to those of wild-type ce lls. ESR spectroscopy studies revealed that DFe-T cells had a 22 %, decreas e in the tyrosyl free radical of the R2 subunit, suggesting that a larger a mount of R2 protein was present as functionally inactive apo-R2 in these ce lls. Our studies indicate that ribonucleotide reductase activity in CCRF-CE M cells can be downregulated by more than 50 % in response to down-regulate d iron transport without an adverse effect on cell proliferation. Furthermo re, our studies suggest a regulatory link between ribonucleotide reductase activity and iron transport into these cells.