Transcriptional regulation in megakaryocytes: the thrombopoietin receptor gene as a model.

The MPL gene codes for the thrombopoietin receptor, whose ligand specifically controls megakaryocytic differentiation. In order to understand the molecular basis for the megakaryocyte-specific expression of MPL, we analyzed the regulatory elements of this gene. Two regions are hypersensitive to DNase I in nuclei of cells that express MPL: the promoter and a portion of intron 6. The latter behaves as a chromatin-dependent enhancer. A 200 bp fragment of the promoter is sufficient for high-level specific expression. This fragment can bind several transacting factors in vitro, including GATA-1 and members of the Ets family. GATA-1 binds with low affinity to a unique GATA motif at -70 in the MPL promoter, and destruction of this site yields only a modest decrease in expression level in human erythroleukemia (HEL) cells. Ets proteins also bind with low affinity to two sites. One is located at position -15 and its destruction reduces expression to 50%; the other is located immediately downstream of the GATA motif and plays a crucial role in expression of the promoter in HEL cells, as its inactivation reduces expression to 15%. This study indicates a molecular basis for the coregulation of markers of megakaryocyte differentiation. Finally, we describe other nuclear factor binding sites that may be involved in the cell-type-specific expression of MPL.