Regulation of CSF-1 gene expression was investigated in human monocytes. CSF-1 transcripts were at low or undetectable levels in resting monocytes. However, in monocytes treated with 12-O-tetradecanoylphorbol-13-acetate (TPA), CSF-1 mRNA was increased by 3 h and reached maximal levels by 12 h of drug exposure. When nuclear run-on assays were used, CSF-1 gene transcription was also at low or undetectable levels in resting monocytes but was activated after TPA exposure. TPA-treated monocytes exposed to actinomycin D further demonstrated that the half-life of the CSF-1 mRNA is 0.9 h. The results also demonstrated that the protein synthesis inhibitor, cycloheximide (CHX), increases CSF-1 mRNA levels in both resting and TPA-treated monocytes. These effects of CHX occurred in the absence of detectable increases in CSF-1 gene transcription. Moreover, treatment of monocytes with CHX and actinomycin D demonstrated that inhibition of protein synthesis is associated with stabilization of the CSF-1 transcript. Taken together, these findings indicated that CSF-1 gene expression is controlled at both transcriptional and posttranscriptional levels in human monocytes.