MITOCHONDRIAL ANTISENSE RNA FOR CYTOCHROME-C-OXIDASE (MARCO) CAN INDUCE MORPHOLOGIC CHANGES AND CELL-DEATH IN HUMAN HEMATOPOIETIC-CELL LINES

To identify essential molecules capable of inducing terminal morpholog ic maturation and cell death of myeloid progenitor cells, we isolated cDNA clones by functional expression cloning using a library construct ed from all-trans retinoic acid (ATRA)-treated human promyelocytic HL- 60 cells. Clones which induced morphologic changes in HL-60 cells from blastic cells to mature neutrophilic granulocytes were selected. The isolated positive cDNA clone was demonstrated to encode an antisense R NA for cytochrome c oxidase/serine tRNA derived from a mitochondrial g ene (MARCO). When MARCO was expressed in HL-60 cells with the lac swit ch system, blastic cell morphology became neutrophilic after 48-hour i ncubation with IPTG, and cell death was observed after 3 days. Also, h igh molecular weight DNA fragmentation was observed after 36 hours in culture. Similar results were observed using transformants from human K562 cells and CMK cells, RT-PCR analysis revealed that MARCO was tran scribed in both ATRA and TNF-alpha systems, and also in human blood ne utrophilic granulocytes. Following transfection with cytochrome c oxid ase expression plasmids, TNF-alpha-induced high molecular weight DNA f ragmentation in U937 cells and HL-60 cells was inhibited in these tran sformants. These results indicate that maturational changes in hematop oietic cells and the process of cell death may be induced by mitochond rial respiratory insufficiency, and also that the mitochondrial gene M ARCO may be used as one of the candidates for gene supplementation the rapy for the acute leukemias. (C) 1997 by The American Society of Hema tology.