P45 NF-E2 REGULATES EXPRESSION OF THROMBOXANE SYNTHASE IN MEGAKARYOCYTES

Transcription factor p45 NF-E2 is highly expressed in the erythroid an d megakaryocytic lineages, Although p45 recognizes regulatory regions of several erythroid genes, mice deficient for this protein display on ly mild dyserythropoiesis but have abnormal megakaryocytes and lack ci rculating platelets, A number of megakaryocytic marker genes have been extensively studied, but none of them is regulated by NF-E2, To find target genes for p45 NF-E2 in megakaryopoiesis, we used an in vivo imm unoselection assay: genomic fragments bound to p45 NF-E2 in the chroma tin of a megakaryocytic cell line were immunoprecipitated with an anti -p45 antiserum and cloned, One of these fragments belongs to the secon d intron of the thromboxane synthase gene (TXS), We demonstrate that t he TXS gene, which is mainly expressed in megakaryocytes, is indeed di rectly regulated by p45 NF-E2, First, its promoter contains a function al NF-E2 binding site; second, the intronic NF-E2 binding site is loca ted within a chromatin-dependent enhancer element; third, p45-null mur ine megakaryocytes do not express detectable TXS mRNA, although TXS ex pression can be detected in other cells. These data, and the structure of the TXS promoter and enhancer, suggest that TXS belongs to a disti nct subgroup of genes involved in platelet formation and function.