Erythropoietin induction in Hep3B cells is not affected by inhibition of heme biosynthesis.
Erythropoietin (Epo) is one of the physiologically important genes whose transcription is up-regulated by hypoxia. Our laboratory previously proposed that the sensor of this event is a heme protein which turns over rapidly. We have investigated the effects of four inhibitors of heme synthesis (4,6-dioxoheptanoic acid (DHA), isoniazid (INH), N-methyl protoporphyrin IX (MPP), and deferoxamine mesylate (DSF)) on hypoxia-, cobalt-, and DSF-induced erythropoietin (Epo) mRNA expression, heme biosynthesis, and cell viability in 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 viability. No suppression of Epo mRNA expression by INH was observed at doses known to inhibit heme biosynthesis. MPP did not suppress Epo mRNA expression although it showed an inhibitory effect on heme biosynthesis without any decreased cell viability. 130 microM DSF, a dose which inhibited heme biosynthesis without cell toxicity, suppressed hypoxia-induced Epo mRNA expression, but enhanced cobalt-induced Epo mRNA expression. These results show that although the oxygen sensor is probably a heme protein it does not turn over rapidly. Therefore, cobalt is unlikely to act by substituting for heme iron.
Horiguchi
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Hematology Division, Brigham and Women's Hospital, Harvard Medical School, Room 223 LMRC, 221 Longwood Avenue, Boston, MA 02115, USA.
Franklin Bunn
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D002470:Cell Survival / Q000187:drug effects; D003676:Deferoxamine / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004921:Erythropoietin / Q000096:biosynthesis*; D006418:Heme / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D006537:Heptanoates / Q000494:pharmacology*; D006801:Humans; D007538:Isoniazid / Q000494:pharmacology; D011524:Protoporphyrins / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D004791:Enzyme Inhibitors; D006537:Heptanoates; D011524:Protoporphyrins; D004921:Erythropoietin; D006418:Heme; C020804:succinylacetone; C028842:N-methylprotoporphyrin IX; D003676:Deferoxamine; D007538:Isoniazid