Erythropoietin induction in Hep3B cells is not affected by inhibition of heme biosynthesis

Erythropoietin (Epo) is one of the physiologically important genes whose tr anscription is up-regulated by hypoxia. Our laboratory previously proposed that the sensor of this event is a heme protein which turns over rapidly. W e have investigated the effects of four inhibitors of heme synthesis (4,6-d ioxoheptanoic acid (DHA), isoniazid (INH), N-methyl protoporphyrin IX (MPP) , and deferoxamine mesylate (DSF)) on hypoxia-, cobalt-, and DSF-induced er ythropoietin (Epo) mRNA expression, heme biosynthesis, and cell viability i n Hep3B cells. DHA (0.1-1.0 mM) inhibited heme biosynthesis more than 85%, but did not suppress Epo mRNA expression. Epo mRNA expression was inhibited only at higher concentrations of DHA (2, 4 mM) which also inhibited cell v iability. No suppression of Epo mRNA expression by INH was observed at dose s known to inhibit heme biosynthesis. MPP did not suppress Epo mRNA express ion although it showed an inhibitory effect on heme biosynthesis without an y decreased cell viability. 130 mu M DSF, a dose which inhibited heme biosy nthesis without cell toxicity, suppressed hypoxia-induced Epo mRNA expressi on, but enhanced cobalt-induced Epo mRNA expression. These results show tha t although the oxygen sensor is probably a heme protein it does not turn ov er rapidly. Therefore, cobalt is unlikely to act by substituting for heme i ron. (C) 2000 Elsevier Science B.V. All rights reserved.