Inhibition of spermidine synthase gene expression by transforming growth factor-β₁ in hepatoma cells

We screened genes responsive to transforming growth factor-β (TGF-β₁) protein in a human hepatoma cell line (Hep3B) using a PCR-mediated differential display, technique, in order to investigate the mechanisms involved in TGF-β-induced growth suppression. We found a gene that was down-regulated by TGF-β₁ to be completely identical in an approx. 620 bp segment to the gene for the enzyme spermidine synthase, which mediates the conversion of putrescine into spermidine. Both spermidine synthase mRNA expression and its enzyme activity were decreased after TGF-β₁ treatment of Hep3B cells. The inhibition of spermidine synthase gene expression by TGF-β₁, protein was also observed in other hepatoma cell lines. The expression of genes for other biosynthetic enzymes in polyamine metabolism (ornithine decarboxylase and S-adenosylmethionine decarboxylase) was also inhibited to the same extent as for spermidine synthase, while the gene expression of spermidine/spermine N¹-acetyltransferase, a catabolic enzyme, was relatively resistant to TGF-β₁. Spermine levels in Hep3B cells were decreased by TGF-β₁ treatment, although the levels of spermidine and putrescine were unchanged, probably due to compensation by remaining spermidine/spermine N¹-acetyltransferase activity. Exogenously added spermidine or spermine, but not putrescine, partially antagonized the growth-inhibitory effects of TGF-β₁, on Hep3B cells. Our data suggest that down-regulation of gene expression of the enzymes involved in polyamine metabolism, including spermidine synthase, may be associated with the mechanism of TGF-β-induced growth suppression.