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 overe xpressed, The small subunit (ribonucleotide reductase 2 (RNR2)) was cloned as a major product in a subtraction library from eukaryotic initiation fact or 4E (eIF4E)-transformed cells (Chinese hamster ovary-4E (CHO-4E)), CHO-4E cells have 20-40-fold elevated RNR2 protein, reflecting an increased distr ibution of RNR2 mRNA to the heavy polysomes, CHO-4E cells display an altere d cell cycle with shortened S phase, similar to cells selected for RNR2 ove rexpression with hydroxyurea, The function of ribonucleotide reductase as a checkpoint component of S progression was studied in yeast in which elevat ed 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 sensitivit y to hydroxyurea and the microtubule depolymerizing agent, benomyl. The dou ble mutant (cdc33-1 rnr2-68) also identified a unique terminal phenotype, a rrested 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 e arly/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.