Expression of coproporphyrinogen oxidase and synthesis of hemoglobin in human erythroleukemia K562 cells

Coproporphyrinogen oxidase (CPOX), the sixth enzyme in the heme-biosyntheti c pathway, catalyzes oxidative decarboxylation of coproporphyrinogen to pro toporphyrinogen and is located in the intermembrane space of mitochondria. To clarify the importance of CPOX in the regulation of heme biosynthesis in erythroid cells, we established human erythroleukemia K562 cells stably ex pressing mouse CPOX. The CPOX cDNA-transfected cells had sevenfold higher C POX activity than cells transfected with vector only. Expression of ferroch elatase and heme content in the transfected cells increased slightly compar ed with the control. When K562 cells overexpressing CPOX were treated with delta -aminolevulinic acid (ALA), most became benzidine-positive without in duction of the expression of CPOX or ferrochelatase, and the heme content w as about twofold higher than that in ALA-treated control cells. Increases i n cellular heme concomitant with a marked induction of the expression of he me-biosynthetic enzymes, including CPOX, ferrochelatase and erythroid-speci fic delta -aminolevulinic acid synthase, as well as of alpha -globin synthe sis, were observed when cells were treated with transforming growth factor (TGF)beta1. These increases in the transfected cells were twice those in co ntrol cells, indicating that overexpression of CPOX enhanced induction of t he differentiation of K562 cells mediated by TGF beta1 or ALA. Conversely, the transfection of antisense oligonucleotide to human CPOX mRNA into untre ated and TGF beta1-treated K562 cells led to a decrease in heme production compared with sense oligonucleotide-transfected cells. These results sugges t that CPOX plays an important role in the regulation of heme biosynthesis during erythroid differentiation.