Translational Regulation of Ribonucleotide Reductase by Eukaryotic Initiation Factor 4E Links Protein Synthesis to the Control of DNA Replication

Ribonucleotide reductase synthesizes dNDPs, a specific and limiting step in DNA synthesis, and can participate in neoplastic transformation when overexpressed. The small subunit (ribonucleotide reductase 2 (RNR2)) was cloned as a major product in a subtraction library from eukaryotic initiation factor 4E (eIF4E)-transformed cells (Chinese hamster ovary-4E (CHO-4E)). CHO-4E cells have 20–40-fold elevated RNR2 protein, reflecting an increased distribution of RNR2 mRNA to the heavy polysomes. CHO-4E cells display an altered cell cycle with shortened S phase, similar to cells selected for RNR2 overexpression with hydroxyurea. The function of ribonucleotide reductase as a checkpoint component of S progression was studied in yeast in which elevated eIF4E rescued S-arrested rnr2–68 ts cells, by increasing recruitment of its mRNA to polysomes. Crosses between rnr2–68 ts and mutant eIF4E ( cdc33–1 ts ) engendered conditional synthetic lethality, with extreme sensitivity to hydroxyurea and the microtubule depolymerizing agent, benomyl. The double mutant ( cdc33–1 rnr2–68 ) also identified a unique terminal phenotype, arrested with small bud and a randomly distributed single nucleus, which is distinct from those of both parental single mutants. This phenotype defines eIF4E and RNR2 as determinants in an important cell cycle checkpoint, in early/mid-S phase. These results also provide a link between protein and DNA synthesis and provide an explanation for cell cycle alterations induced by elevated eIF4E.