Differential Contribution of the Gata1 Gene Hematopoietic Enhancer to Erythroid Differentiation

Abstract

GATA1 is a key regulator of erythroid cell differentiation. To examine how Gata1 gene expression is regulated in a stage-specific manner, transgenic mouse lines expressing green fluorescent protein (GFP) reporter from the Gata1 locus in a bacterial artificial chromosome (G1BAC-GFP) were prepared. We found that the GFP reporter expression faithfully recapitulated Gata1 gene expression. Using GFP fluorescence in combination with hematopoietic surface markers, we established a purification protocol for two erythroid progenitor fractions, referred to as burst-forming units-erythroid cell-related erythroid progenitor (BREP) and CFU-erythroid cell-related erythroid progenitor (CREP) fractions. We examined the functions of the Gata1 gene hematopoietic enhancer (G1HE) and the highly conserved GATA box in the enhancer core. Both deletion of the G1HE and substitution mutation of the GATA box caused almost complete loss of GFP expression in the BREP fraction, but the CREP stage expression was suppressed only partially, indicating the critical contribution of the GATA box to the BREP stage expression of Gata1. Consistently, targeted deletion of G1HE from the chromosomal Gata1 locus provoked suppressed expression of the Gata1 gene in the BREP fraction, which led to aberrant accumulation of BREP stage hematopoietic progenitor cells. These results demonstrate the physiological significance of the dynamic regulation of Gata1 gene expression in a differentiation stage-specific manner.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://mcb.asm.org/ .

ACKNOWLEDGMENTS

We thank Norio Suzuki, Naoshi Obara, and Harumi Yamamoto-Mukai for help and discussion and Yuko Kikuchi, Reiko Kawai, Naomi Kaneko, and Mitsuru Okano for technical assistance. We also thank Tania O'Connor for critical reading of the manuscript.

This work was supported in part by grants from ERATO-JST (M.Y.), Takeda Science Foundation (T.M.), and Naito foundation (M.Y.) and Grants-in-Aid for Creative Scientific Research (M.Y.), for Scientific Research on Priority Areas (M.Y.), for Scientific Research (T.M. and M.Y.), and for Exploratory Research (M.Y.) from the Ministry of Education, Science, Sports and Culture.