Since many lines of evidence suggest that expression of the transforming growth factor-β1 (TGF-β1) gene may be regulated post-transcriptionally, we examined the effect of the 5′-untranslated region (UTR) of this gene on TGF-β1 expression. For this purpose, fragments of the 840-nucleotide highly GC-rich TGF-β1 5′-UTR were inserted into the 5′-UTR of the structural gene for human growth hormone driven by the simian virus 40 early promoter. A portion of the 5′-UTR of TGF-β1 mRNA spanning the sequences from +11 to + 147 was shown to inhibit growth hormone expression by as much as 22-fold. This effect was cell-specific; growth hormone production was inhibited in PC-3 human prostate adenocarcinoma and A-549 human lung adenocarcinoma cells, while no effect was seen in rat pheochromocytoma PC12 cells, which show efficient translation of endogenous TGF-β1 mRNA. Computer analysis showed that this region of the 5′-UTR contained a stable secondary stem-loop structure spanning sequences +49 to + 76. This stem-loop region alone is sufficient to inhibit expression of the growth hormone gene, suggesting that it plays an important role in post-transcriptional regulation of TGF-β1 gene expression.