GATA Transcription Factors Regulate the Expression of the Human Eosinophil-derived Neurotoxin (RNase 2) Gene

Zhijun Qiu,Kimberly D Dyer,Zhihui Xie,Madeleine Rådinger,Helene F Rosenberg

doi:10.1074/jbc.m807307200

Abstract

The transcription factors GATA-1 and GATA-2 have been implicated in promoting differentiation of eosinophilic leukocytes. In this study, we examined the roles of GATA-1 and GATA-2 in activating transcription of the secretory ribonuclease, the eosinophil-derived neurotoxin (EDN/RNase 2). Augmented expression of both GATA-1 and GATA-2 was detected in eosinophil promyelocyte HL-60 clone 15 cells in response to biochemical differentiation with butyric acid. Deletion or mutation of one or both of the two consensus GATA-binding sites in the extended 1000-bp 5' promoter of the EDN gene resulted in profound reduction in reporter gene activity. Antibody-augmented electrophoretic mobility shift and chromatin immunoprecipitation analyses indicate that GATA-1 and GATA-2 proteins bind to both functional GATA consensus sequences in the EDN promoter. Interestingly, RNA silencing of GATA-1 alone had no impact on EDN expression; silencing of GATA-2 resulted in diminished expression of EDN, and also diminished expression of GATA-1 in both butyric acid-induced HL-60 clone 15 cells and in differentiating human eosinophils derived from CD34(+) hematopoietic progenitors. Likewise, overexpression of GATA-2 in uninduced HL-60 clone 15 cells resulted in augmented transcription of both EDN and GATA-1. Taken together, our data suggest that GATA-2 functions directly via interactions with the EDN promoter and also indirectly, via its ability to regulate the expression of GATA-1 in differentiating eosinophils and eosinophil cell lines.

Highlights

Eosinophils remain among the most enigmatic of the mammalian leukocytes, consensus opinion is that they contribute in some fashion to the pathophysiology of allergic asthma and gastrointestinal dysfunction [1]
Functional GATA-1 enhancer sites direct the expression of the eosinophil granule major basic protein [13], and GATA-1 directs the eosinophil-specific expression of gp91phox, with GATA-2 functioning as a site-specific repressor [14]
BA-induced differentiation of HL-60 clone 15 cells results in augmented transcription of GATA-1 (Fig. 1B) and GATA-2 (Fig. 1C), reaching levels

Summary

Introduction

Eosinophils remain among the most enigmatic of the mammalian leukocytes, consensus opinion is that they contribute in some fashion to the pathophysiology of allergic asthma and gastrointestinal dysfunction [1]. Cells were transfected with the full-length or truncated EDN promoter reporter constructs, with or without mutations in the GATA consensus binding sites.

Results

Conclusion