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Enforced Expression of the GATA-2 Transcription Factor Blocks Normal Hematopoiesis

Persons, Derek A., Allay, James A., Allay, Esther R., Ashmun, Richard A., Orlic, Donald, Jane, Stephen M., Cunningham, John M., Nienhuis, Arthur W.·1999·Blood
DOI 10.1182/blood.v93.2.488
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Abstract

The zinc finger transcription factor GATA-2 is highly expressed in immature hematopoietic cells and declines with blood cell maturation. To investigate its role in normal adult hematopoiesis, a bicistronic retroviral vector encoding GATA-2 and the green fluorescent protein (GFP) was used to maintain the high levels of GATA-2 that are normally present in primitive hematopoietic cells. Coexpression of the GFP marker facilitated identification and quantitation of vector-expressing cells. Bone marrow cells transduced with the GATA-2 vector expressed GFP as judged by flow cytometry and GATA-2 as assessed by immunoblot analysis. A 50% to 80% reduction in hematopoietic progenitor-derived colony formation was observed with GATA-2/GFP-transduced marrow, compared with marrow transduced with a GFP-containing vector lacking the GATA-2 cDNA. Culture of purified populations of GATA-2/GFP-expressing and nonexpressing cells confirmed a specific ablation of the colony-forming ability of GATA-2/GFP-expressing progenitor cells. Similarly, loss of spleen colony-forming ability was observed for GATA-2/GFP-expressing bone marrow cells. Despite enforced GATA-2 expression, marrow cells remained viable and were negative in assays to evaluate apoptosis. Although efficient transduction of primitive Sca-1+Lin- cells was observed with the GATA-2/GFP vector, GATA-2/GFP-expressing stem cells failed to substantially contribute to the multilineage hematopoietic reconstitution of transplanted mice. Additionally, mice transplanted with purified, GATA-2/GFP-expressing cells showed post-transplant cytopenias and decreased numbers of total and gene-modified bone marrow Sca-1+ Lin−cells. Although Sca-1+ Lin− bone marrow cells expressing the GATA-2/GFP vector were detected after transplantation, no appreciable expansion in their numbers occurred. In contrast, control GFP-expressing Sca-1+Lin− cells expanded at least 40-fold after transplantation. Thus, enforced expression of GATA-2 in pluripotent hematopoietic cells blocked both their amplification and differentiation. There appears to be a critical dose-dependent effect of GATA-2 on blood cell differentiation in that downregulation of GATA-2 expression is necessary for stem cells to contribute to hematopoiesis in vivo.

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Metadata

Authors
Persons, D. A., Allay, J. A., Allay, E. R., Ashmun, R. A., Orlic, D., Jane, S. M., Cunningham, J. M., & Nienhuis, A. W.
Affiliations
From the Division of Experimental Hematology, Department of Hematology/Oncology and the Department of Experimental Oncology, St Jude Children’s Research Hospital, Memphis, TN; and the Hematopoiesis Section, Genetic and Molecular Biology Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD.
Venue
Blood
Publisher
American Society of Hematology
Published
1999-01-15
Volume
93
Issue
2
Pages
488-499
ISSN
  • 1528-0020
  • 0006-4971
DOI
10.1182/blood.v93.2.488
Cited by
181
References
2
Language
en

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