In Saccharomyces cerevisiae, many genes encoding enzymes involved in deoxyribonucleotide synthesis are expressed preferentially near the G(1)/S boundary of the! cell cycle. The relationship between the induction of deoxyribonucleotide-synthesizing genes, deoxyribonucleoside triphosphate levels, and replication initiation was investigated using a factor-synchronized wildtype yeast or dbf4 yeast that are temperature-sensitive for replication initiation. Neither the timing nor extent of gene induction was inhibited when a factor-arrested dbf4 cells were released into medium containing the ribonucleotide reductase inhibitor hydroxyurea, which blocks replication fork progression, or were released at 37 degreesC, which blocks replication origin firing. Thus, the induction of deoxyribonucleotide-synthesizing genes at G(1)/S was fully independent of DNA chain elongation or initiation. Deoxyribonucleoside triphosphate levels increased severalfold at G(1)/S in wild-type cells and in,dbf4 mutants incubated at the non-permissive temperature. Thus, deoxyribonucleoside triphosphate accumulation, like the induction of deoxyribonucleotide-synthesizing genes, was not dependent on replication initiation. Deoxyribonucleoside triphosphate accumulation at G(1)/S was suppressed in cells lacking Swi6, a transcription factor required for normal cell cycle regulation of deoxyribonucleotide-synthesizing genes. The results suggest that cells use gene induction at G(1)/S as a mechanism to pre-emptively, rather than reflexively, increase the synthesis of DNA precursors to meet the demand of the replication forks for deoxyribonucleotides.