Differential GATA-1 Sensitivities of Target Loci.

Abstract

The transcription factor GATA-1 is a key regulator of red cell differentiation, activating numerous erythroid-specific genes while downregulating genes that permit proliferation of erythroid precursors. We have demonstrated that GATA-1 only occupies a limited number of the high affinity WGATAR motifs present within target gene loci, FOG-1 is required for GATA-1 to occupy a subset of chromatin sites, and GATA-1 occupancy often coincides with GATA-2 displacement or a “GATA switch”. Once bound, GATA-1 elicits changes in the histone modification patterns and the three dimensional structure of target gene loci via recruitment of cofactors such as FOG-1 and CBP. As detailed mechanistic studies have not been conducted with most GATA-1 target genes, it is unclear whether these genes are equally sensitive to GATA-1 or if they respond differently to the rising GATA-1 levels during erythropoiesis. Using GATA-1 fusions to the estrogen receptor ligand binding domain (ER-GATA-1) in the GATA-1-null erythroid precursor cell line, G1E, we analyzed the responses of endogenous GATA-1 target genes to varied levels of GATA-1 activity. We found that transcriptional activation of Tac-2 required higher concentrations of ER-GATA-1 than is required for other GATA-1 target genes. Previously, we showed that Tac-2, which encodes the neurokinin-B precursor protein preprotachykinin B, is regulated by GATA-1 in erythroid cell lines and is induced upon ex vivo differentiation of human CD34+ peripheral blood cells. Importantly, whereas regulation of many GATA-1 target genes is only partially disrupted by removal of the N-terminal 193 amino acids from ER-GATA-1 (ER-GATA-1 ΔN), Tac-2 expression was very sensitive to this truncation. Whereas NK-B signals through G-protein-coupled receptors to modulate neuronal function, its functions beyond the nervous system are poorly understood. Although erythroid cells do not express NK-B receptors, the receptors, but not NK-B, are expressed in certain endothelial cell subtypes, and elevated levels of NK-B are implicated in the pregnancy-associated disorder pre-eclampsia. Tac-2 represents the first GATA-1 target gene that critically requires the N-terminus. Studies are underway to elucidate mechanisms underlying the exquisite sensitivity of Tac-2 to deletion of the GATA-1 N-terminus, the relationship between Tac-2 deregulation and GATA-1 N-terminal deletions in megakaryoblastic leukemia, and the function of erythroid cell-derived NK-B.