Regulation of the expression of human ferrochelatase by intracellular ironlevels

Mammalian ferrochelatase, the terminal enzyme of the heme biosynthetic path way, catalyzes the insertion of a ferrous ion into protoporphyrin and conta ins a labile [2Fe-2S] cluster center at the C-terminus. To clarify the role s of the iron-sulfur cluster in the expression of mammalian ferrochelatase, enzyme activity in human erythroleukemia K562 cells under iron-depleted co nditions was examined. Treatment of cells with an iron chelator, desferriox amine, resulted in a decrease in enzyme activity, in a dose- and time-depen dent manner. Heme content decreased during desferrioxamine treatment of the cells. Addition of ferric ion-nitrilotriacetate [Fe(III)NTA] to desferriox amine-containing cultures led to restoration of the reduction in the enzyme activity. While RNA blots showed that the amount of ferrochelatase mRNA re mained unchanged during these treatments, the amount of ferrochelatase decr eased with a concomitant decrease in enzyme activity. When full-length huma n ferrochelatase was expressed in Cos7 cells, the activity was found mainly in the mitochondria and was decreased markedly by treatment with desferrio xamine. The activity in Cos7 cells expressing human ferrochelatase in cytop lasm decreased with desferrioxamine, but to a lesser extent. When Escherich ia coli ferrochelatase, which lacks the iron-sulfur cluster, was expressed in Cos7 cells, the activity did not change following any treatment. Convers ely, the addition of Fe(III)NTA to the culture of K562 and Cos7 cells led t o an increase in ferrochelatase activity. These results indicate that the e xpression of mammalian ferrochelatase is regulated by intracellular iron le vels, via the iron-sulfur cluster center at the C-terminus, and this contri butes to the regulation of the biosynthesis of heme at the terminal step.