Biologic significance of GATA-1 activities in Ras-mediated megakaryocytic differentiation of hematopoietic cell lines

Lineage-specific transcription factors play crucial roles in the developmen t of hematopoietic cells. In a previous study, it was demonstrated that Has activation was involved in thrombopoietin-induced megakaryocytic different iation. In this study, constitutive Has activation by H-ras(G12V) evoked me gakaryocytic maturation of erythroleukemia cell lines F-36P and K562, but n ot of myeloid cell line 32D c13 that lacks GATA-1. However, the introductio n of GATA-1 led to reprogramming of 32D c13 toward erythrocytic/megakaryocy tic lineage and enabled it to undergo megakaryocytic differentiation in res ponse to H-ras(G12V). In contrast, the overexpression of PU.1 and c-Myb cha nged the phenotype of K562 from erythroid to myeloid/monocytic lineage and rendered K562 to differentiate into granulocytes and macrophages in respons e to H-ras(G12V), respectively. In GATA-1-transfected 32D c13, the endogeno us expression of PU.1 and c-Myb was easily detectable, but their activities were reduced severely. Endogenous GATA-1 activities were markedly suppress ed in PU.1-transfected and c-myb-transfected K562. As for the mechanisms of these reciprocal inhibitions, GATA-1 and PU.1 were found to associate thro ugh their DNA-binding domains and to inhibit the respective DNA-binding act ivities of each other. In addition, c-Myb bound to GATA-1 and inhibited its DNA-binding activities. Mutant GATA-1 and PU.1 that retained their own tra nscriptional activities but could not inhibit the reciprocal partner were l ess effective in changing the lineage phenotype of 32D c13 and K562. These results suggested that GATA-1 activities may be crucial for Has-mediated me gakaryocytic differentiation and that its activities may be regulated by th e direct interaction with other lineage-specific transcription factors such as PU.1 and c-Myb. (Blood. 2000;96:2440-2450) (C) 2000 by The American Soc iety of Hematology.