FORCED GATA-1 EXPRESSION IN THE MURINE MYELOID CELL-LINE M1 - INDUCTION OF C-MPL EXPRESSION AND MEGAKARYOCYTIC ERYTHROID DIFFERENTIATION/

The ''zinc-finger'' transcription factor GATA-1 was first shown in cel ls of erythroid lineage. It is also expressed in cells of other hemato poietic lineages including megakaryocytes, mast cells, and eosinophils . GATA-1 is now considered to be one of the central regulators in hema topoietic cell differentiation, To further analyze the role of GATA-1 in controlling differentiation from hematopoietic stem cells, we inves tigated the phenotypic changes induced by the overexpression of murine GATA-1 in the murine myeloid leukemic cell line, M1. Forced expressio n of GATA-1 induced the appearance of erythroid cells and megakaryocyt es as assessed by cellular morphology, acetylcholinesterase activity, and expression of platelet factor 4 and beta-globin mRNA synthesis. Be cause the c-mpl ligand, thrombopoietin, plays an important role in meg akaryopoiesis, the expression of c-mpl and c-mpl ligand (thrombopoieti n) mRNA was analyzed by Northern blot and reverse transcription-polyme rase chain reaction (RT-PCR) in M1 cells overexpressing GATA-1. The c- mpl ligand mRNA was equally expressed both in parental M1 cells and in those transfected with the GATA-1 expression vector. In contrast, the mRNA expression of c-mpl was increased only in GATA-1 expressing M1 c ells differentiated towards erythroid and megakaryocyte lineages. The increased expression of c-mpl mRNA induced by GATA-1 raised the questi on as to whether or not GATA-1 transactivated the c-mpl promoter. The activity of the c-mpl promoter in the presence of cotransfected GATA-1 was significantly increased compared with that of the control. A plas mid with the mutated GATA-binding site did not show transactivation ab ility in the cotransfection with a GATA expression vector. These findi ngs suggest that the upregulation of c-mpl induced by GATA-1 expressio n in M1 cells is closely associated with erythroid and megakaryocytic differentiation. (C) 1998 by The American Society of Hematology.