Fibroblasts produce a variety of cytokines including granulocyte/macrophage colony-stimulating factor (GM-CSF). GM-CSF is pivotal for proliferation and function of myeloid cells. In this report, we describe the regulation of GM-CSF gene by irradiation in human fibroblasts. We found that fibroblasts constitutively produced GM-CSF; irradiation markedly increased the production of GM-CSF. The increase in GM-CSF transcripts by irradiation was both time- and dose-dependent. Moreover, irradiation increased GM-CSF mRNA in cells with prolonged exposure to 12-O-tetradecanoyl-phorbol-13-acetate (TPA). WI38 fibroblasts constitutively produce low levels of IL-1. Induction of GM-CSF mRNA by irradiation was partially blocked by anti-IL-1 antibodies. On the other hand, inhibition of prostaglandin synthesis did not affect induction of GM-CSF RNA. Transcriptional run-on analysis showed that irradiation increased the rate of GM-CSF transcription. Stability studies of GM-CSF mRNA in these cells showed that half-life (t1/2) increased from < 20 min in unirradiated cells to > 100 min in irradiated cells. These findings suggest that the increase in GM-CSF mRNA observed after irradiation is regulated by transcriptional and post-transcriptional mechanisms. Our results indicate that induction of GM-CSF gene by irradiation requires de novo protein synthesis and increased levels of GM-CSF transcripts also occur through a pathway distinct from protein kinase C activation.