The effect of protoporphyrin IX (hemin without iron) on the expression of transferrin receptor and ferritin was investigated in Friend leukemia cells. Cells treated with protoporphyrin IX exhibit enhanced transferrin-receptor expression and markedly reduced ferritin synthesis. Stimulation of transferrin-receptor expression is observed at both the mRNA and protein level. The effect on ferritin synthesis is mediated by translational inhibition of the mRNA, which, in contrast, is transcriptionally stimulated by protoporphyrin IX treatment. The regulation of transferrin receptor and ferritin in response to iron perturbations has been studied extensively and is mediated by the binding of iron-regulatory proteins (IRP) to the iron-responsive elements (IRE) present in the 3' and 5' untranslated regions of the transferrin-receptor and ferritin mRNA, respectively. To elucidate the molecular mechanisms underlying the effects of protoporphyrin IX on ferritin and transferrin-receptor expression, the role of the IRE sequence was investigated both in vivo by transfection experiments, with a construct containing the coding region for the chloramphenicol acetyltransferase (CAT) reporter gene under the translational control of the ferritin IRE, and in vitro by RNA band-shift assays. Whereas, examination of IRP binding to the IRE by in vitro assays suggests an apparent inactivation of IRP by protoporphyrin IX treatment, CAT assays indicate that protoporphyrin IX is able to induce in vivo a translational inhibition similar to that obtained by treatment with the iron chelator Desferal. This observation raises the possibility of different effects on the IRP activity exerted by porphyrin treatment in intact tissue-culture cells and in vitro. We conclude that translation of ferritin mRNA and degradation of transferrin-receptor mRNA are inhibited in intact tissue-culture cells by protoporphyrin IX through a mechanism similar to that exerted by iron chelation, thus involving depletion of the intracellular iron pool. These results can improve the understanding of the regulation of ferritin gene expression in some pathological conditions associated with disturbed heme synthesis.