delta-Aminolaevulinate synthase in human HepG2 hepatoma cells. Repression by haemin and induction by chemicals

Abstract

delta-Aminolaevulinate (ALA) synthase, the rate-limiting enzyme in haem biosynthesis in the normal liver, was examined in human HepG2 hepatoma cells. Haemin, up to 100 microM, had no effect on ALA synthase activity in vitro; it did, however, exhibit a dose-dependent inhibitory action when added to cells growing in culture (half-maximal inhibition at 1 microM). The half-life of ALA synthase activity after haemin treatment was 2 h, which was similar to that found after treatment with cycloheximide. Cells treated with actinomycin D showed a longer half-life of the enzyme activity, i.e. 4 h, compared with haemin or cycloheximide treatment. Treatment of cells with succinylacetone markedly inhibited the activity of ALA dehydratase and 59Fe incorporation into haem, but in increased ALA synthase activity. Both the haemin-induced repression and the succinylacetone-mediated de-repression of ALA synthase activity were reversible within 4 h after replacing the medium with fresh medium without the chemical. In addition to succinylacetone, dimethyl sulphoxide and 3-methylcholanthrene induced the enzyme. Induction of ALA synthase by these chemicals was also suppressed by treatment of cells with haemin. These findings indicate that the level of ALA synthase in HepG2 cells is maintained by both synthesis and degradation of the enzyme, and that the synthesis of the enzyme is regulated by the concentration of regulatory free haem in the cell.