Abstract
Recent evidence supports a role for GATA transcription factors as important signal intermediates in differentiated endothelial cells. The goal of this study was to identify proteins that interact with endothelial-derived GATA transcription factors. Using yeast two-hybrid screening, we identified hematopoietically expressed homeobox (Hex) as a GATA-binding partner in endothelial cells. The physical association between Hex and GATA was confirmed with immunoprecipitation in cultured cells. Hex overexpression resulted in decreased flk-1/KDR expression, both at the level of the promoter and the endogenous gene, and attenuated vascular endothelial growth factor-mediated tube formation in primary endothelial cell cultures. In electrophoretic mobility shift assays, Hex inhibited the binding of GATA-2 to the flk-1/KDR 5'-untranslated region GATA motif. Finally, in RNase protection assays, transforming growth factor beta1, which has been previously shown to decrease flk-1 expression by interfering with GATA binding activity, was shown to increase Hex expression in endothelial cells. Taken together, the present study provides evidence for a novel association between Hex and GATA and suggests that transforming growth factor beta-mediated repression of flk-1/KDR and vascular endothelial growth factor signaling involves the inducible formation of inhibitory Hex-GATA complexes.
Highlights
The present study provides evidence for a novel association between Hex and GATA and suggests that transforming growth factor -mediated repression of flk-1/KDR and vascular endothelial growth factor signaling involves the inducible formation of inhibitory Hex-GATA complexes
Several members of the GATA family have been identified in endothelial cells, including GATA-2, -3, and -6
To confirm the specificity of interaction between Hex and GATA-6, constructs containing the Gal4-activating domain fused with full-length Hex and either the Gal4-binding domain fused with GATA-6 or the Gal4-binding domain alone were co-expressed in yeast AH109
Summary
The present study provides evidence for a novel association between Hex and GATA and suggests that transforming growth factor -mediated repression of flk-1/KDR and vascular endothelial growth factor signaling involves the inducible formation of inhibitory Hex-GATA complexes. GATA factors may be associated with cofactors, such as basal transcription factors, FOG-1, FOG-2, CBP/p300, and histone deacetylase-3 [23,24,25,26,27,28,29] Taken together, these data suggest that GATA transcription factors are involved in highly complex regulatory pathways and that the dissection of these networks may provide valuable insight into the transcriptional control of endothelial phenotypes. These transacting factors were believed to play a singular role in the differentiation of endothelial cells during development
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