TY - JOUR
T1 - High-throughput siRNA screening to reveal GATA-2 upstream transcriptional mechanisms in hematopoietic cells
AU - Saito, Yo
AU - Fujiwara, Tohru
AU - Ohashi, Keiichi
AU - Okitsu, Yoko
AU - Fukuhara, Noriko
AU - Onishi, Yasushi
AU - Ishizawa, Kenichi
AU - Harigae, Hideo
N1 - Funding Information:
This study was supported by KANAE Foundation for the Promotion of Medical Science (TF) ( http://www.kanae-zaidan.com ) which receives financial support from Sanofi KK, and by a research grant from Chugai Pharmaceutical Co., Ltd. There are no patents, products in development or marketed products to declare. This does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials.
Publisher Copyright:
© 2015 Saito et al.
PY - 2015/9/1
Y1 - 2015/9/1
N2 - Hematopoietic stem cells can self-renew and differentiate into all blood cell types. The transcription factor GATA-2 is expressed in both hematopoietic stem and progenitor cells and is essential for cell proliferation, survival, and differentiation. Recently, evidence from studies of aplastic anemia, MonoMAC syndrome, and lung cancer has demonstrated a mechanistic link between GATA-2 and human pathophysiology. GATA-2-dependent disease processes have been extensively analyzed; however, the transcriptional mechanisms upstream of GATA-2 remain less understood. Here, we conducted high-throughput small-interfering-RNA (siRNA) library screening and showed that YN-1, a human erythroleukemia cell line, expressed high levels of GATA-2 following the activation of the hematopoietic-specific 1S promoter. As transient luciferase reporter assay in YN-1 cells revealed the highest promoter activity in the 1S promoter fused with GATA-2 intronic enhancer (+9.9 kb/1S); therefore, we established a cell line capable of stably expressing +9.9 kb/1S-Luciferase. Subsequently, we screened 995 transcription factor genes and revealed that CITED2 acts as a GATA-2 activator in human hematopoietic cells. These results provide novel insights into and further identify the regulatory mechanism of GATA-2.
AB - Hematopoietic stem cells can self-renew and differentiate into all blood cell types. The transcription factor GATA-2 is expressed in both hematopoietic stem and progenitor cells and is essential for cell proliferation, survival, and differentiation. Recently, evidence from studies of aplastic anemia, MonoMAC syndrome, and lung cancer has demonstrated a mechanistic link between GATA-2 and human pathophysiology. GATA-2-dependent disease processes have been extensively analyzed; however, the transcriptional mechanisms upstream of GATA-2 remain less understood. Here, we conducted high-throughput small-interfering-RNA (siRNA) library screening and showed that YN-1, a human erythroleukemia cell line, expressed high levels of GATA-2 following the activation of the hematopoietic-specific 1S promoter. As transient luciferase reporter assay in YN-1 cells revealed the highest promoter activity in the 1S promoter fused with GATA-2 intronic enhancer (+9.9 kb/1S); therefore, we established a cell line capable of stably expressing +9.9 kb/1S-Luciferase. Subsequently, we screened 995 transcription factor genes and revealed that CITED2 acts as a GATA-2 activator in human hematopoietic cells. These results provide novel insights into and further identify the regulatory mechanism of GATA-2.
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U2 - 10.1371/journal.pone.0137079
DO - 10.1371/journal.pone.0137079
M3 - Article
C2 - 26325290
AN - SCOPUS:84943224652
SN - 1932-6203
VL - 10
JO - PLoS ONE
JF - PLoS ONE
IS - 9
M1 - e0137079
ER -
