Neurokinin-B Transcription in Erythroid Cells

Abstract

The GATA family of transcription factors establishes genetic networks that control developmental processes including hematopoiesis, vasculogenesis, and cardiogenesis. We found that GATA-1 strongly activates transcription of the Tac-2 gene, which encodes proneurokinin-B, a precursor of neurokinin-B (NK-B). Neurokinins function through G protein-coupled transmembrane receptors to mediate diverse physiological responses including pain perception and the control of vascular tone. Whereas an elevated level of NK-B was implicated in pregnancy-associated pre-eclampsia (Page, N. M., Woods, R. J., Gardiner, S. M., Lomthaisong, K., Gladwell, R. T., Butlin, D. J., Manyonda, I. T., and Lowry, P. J. (2000) Nature 405, 797-800), the regulation of NK-B synthesis and function are poorly understood. Tac-2 was expressed in normal murine erythroid cells and was induced upon ex vivo erythropoiesis. An estrogen receptor fusion to GATA-1 (ER-GATA-1) and endogenous GATA-1 both occupied a region of Tac-2 intron-7, which contains two conserved GATA motifs. Genetic complementation analysis in GATA-1-null G1E cells revealed that endogenous GATA-2 occupied the same region of intron-7, and expression of ER-GATA-1 displaced GATA-2 and activated Tac-2 transcription. Erythroid cells did not express neurokinin receptors, whereas aortic and yolk sac endothelial cells differentially expressed neurokinin receptor subtypes. Since NK-B induced cAMP accumulation in yolk sac endothelial cells, these results suggest a new mode of vascular regulation in which GATA-1 controls NK-B synthesis in erythroid cells.The development of blood cells from hematopoietic stem cells is critically dependent upon integration of the activities of diverse transcription factors. Of particular importance are three members of the GATA family of transcription factors that regulate hematopoiesis via both unique and overlapping activities (1). GATA-1 is required for definitive or adult erythropoiesis, for megakaryocyte maturation, and for eosinophil production (2-7), whereas GATA-2 is important for the survival and function of multipotent hematopoietic precursor cells (8 -10). GATA-1 and GATA-2 have redundant activities to control primitive or embryonic erythropoiesis (11). GATA-3 uniquely regulates aspects of lymphopoiesis and central nervous system development (12-16). Understanding how GATA factors regulate development requires the elucidation of genetic circuits that control expression of GATA factor target genes and the GATA factors themselves.The C-terminal zinc finger of dual zinc finger GATA factors recognizes a simple DNA motif (WGATAR) (17-20) that is distributed abundantly throughout genomes. In contrast, the N-terminal zinc finger mediates interactions with the friend of GATA-1 (FOG-1) 1 coregulator (21, 22) and stabilizes DNA binding at certain GATA motifs (17,23). Despite the high frequency of WGATAR in genomes, analyses of GATA-1 and GATA-2 binding to chromatin by quantitative ChIP analysis have revealed an exquisite specificity of chromatin occupancy. For exampl...

Differential sensitivities of transcription factor target genes underlie cell type-specific gene expression profiles

“…2C), Tac-2 activation requires a considerably higher ER-GATA-1 level͞activity than other genes. Tac-2 is highly induced upon differentiation of human peripheral blood erythroid precursors and is also expressed in mouse Ter119ϩ bone marrow erythroid cells (45).…”

“…To establish whether differential chromatin access underlies the distinct sensitivities, the ER-GATA-1 level͞activity was titrated with 5, 10, or 40 nM ␤-estradiol for 16 h, and chromatin occupancy was measured at sites defined previously (13,14,20,22,28,(43)(44)(45). ER-GATA-1 occupied ␤-globin HSs1-4 maximally and the ␤major promoter nearly maximally in 10 nM ␤-estradiol-treated cells ( Fig.…”

Dynamic GATA Factor Interplay at a Multicomponent Regulatory Region of the GATA-2 Locus

“…Real-time PCR reactions (15 l) contained 2 l of cDNA, 7. (24,25). For analysis of CBP and FOG-1 occupancy, cells were cross-linked with 1% formaldehyde.…”

“…Although ER-GATA-1 and GATA-1 are seemingly indistinguishable in binding DNA and activating transcription (18,21,25,28,29), the DNA binding activity of ER-GATA-1 has not been analyzed in detail. Alternatively, the distinct patterns might reflect intrinsic differences in the competence of GATA-1 and GATA-2 to occupy GATA motifs within chromatin.…”

Context-dependent GATA Factor Function

“…In principle, context-dependent functions can arise at the level of chromatin occupancy or postchromatin occupancy. Although GATA motifs are abundant throughout the genome, only a small subset are occupied in cells (29,(37)(38)(39). Parameters that govern GATA motif occupancy include intrinsic features of the motifs, their relationship to nearest neighbor cis-elements, and the surrounding chromatin environment (39).…”