Tohoku J. Exp. Med., 2018 January, 244(1)

Establishment of a Screening System to Identify Novel GATA-2 Transcriptional Regulators

KEIICHI OHASHI,¹ TOHRU FUJIWARA,¹ KOICHI ONODERA,¹ YO SAITO,¹ SATOSHI ICHIKAWA,¹ MASAHIRO KOBAYASHI,¹ YOKO OKITSU,¹ NORIKO FUKUHARA,¹ YASUSHI ONISHI¹ and HIDEO HARIGAEUHA¹

¹Department of Hematology and Rheumatology, Tohoku University Graduate School of Medicine, Sendai, Miyagi, Japan

Hematopoietic stem cells can self-renew and differentiate into all blood cell types. The transcription factor GATA-2 is expressed in hematopoietic stem and progenitor cells and is essential for cell proliferation and differentiation. Heterozygous germline GATA2 mutations induce GATA-2 deficiency syndrome, characterized by monocytopenia, a predisposition to myelodysplasia and acute myeloid leukemia, and a profoundly reduced dendritic cell (DC) population, which is associated with increased susceptibility to viral infections. Because patients with GATA-2 deficiency syndrome could retain a wild-type copy of GATA-2, boosting residual wild-type GATA-2 activity may represent a novel therapeutic strategy for the disease. Here, we sought to establish a screening system to identify GATA-2 activators using human U937 monocytic cells as a potential model of the DC progenitor. Enforced GATA-2 expression in U937 cells induces CD205 expression, a marker of DC differentiation, indicating U937 cells as a surrogate of human primary DC progenitors. Transient luciferase reporter assays in U937 cells reveals a high promoter activity of the −0.5 kb GATA-2 hematopoietic-specific promoter (1S promoter) fused with two tandemly connected GATA-2 +9.9 kb intronic enhancers. We thus established U937-derived cell lines stably expressing tandem +9.9 kb/−0.5 kb 1S-luciferase. Importantly, forced GATA-1 expression, a repressor for GATA-2 expression, in the stable clones caused significant decreases in the luciferase activities. In conclusion, our system represents a potential tool for identifying novel regulators of GATA-2, thereby contributing to the development of novel therapeutic approaches.

keywords —— dendritic cells; GATA-1; GATA-2; GATA-2 deficiency syndrome; U937

© 2018 Tohoku University Medical Press

===============================

Tohoku J. Exp. Med., 2018, 244, 41-52

Correspondence: Hideo Harigae, M.D., Ph.D., Department of Hematology and Rheumatology, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8575, Japan.

e-mail: harigae@med.tohoku.ac.jp