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 Blood, 15 August 2006, Vol. 108, No. 4, pp. 1223-1229.
Prepublished online as a Blood First Edition Paper on April 27, 2006; DOI 10.1182/blood-2005-12-008763.

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HEMATOPOIESIS

C/EBP{alpha} directs monocytic commitment of primary myeloid progenitors

Dehua Wang, Jenice D'Costa, Curt I. Civin, and Alan D. Friedman

From the Divisions of Pediatric Oncology and Immunology and Hematopoiesis, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University, Baltimore, MD.

C/EBP{alpha} is required for generation of granulocyte-monocyte progenitors, but the subsequent role of C/EBP{alpha} in myeloid lineage commitment remains uncertain. We transduced murine marrow cells with C/EBP{alpha}-estradiol receptor (ER) or empty vector and subjected these to lineage depletion just prior to culture in estradiol with myeloid cytokines. This protocol limits biases due to lineage-specific effects on developmental kinetics, proliferation, and apoptosis. Also, lowering the dose of estradiol reduced activated C/EBP{alpha}-ER to near the physiologic range. C/EBP{alpha}-ER increased Mac1+/Gr1–/MPO–/low monocytes 1.9-fold while reducing Mac1+/Gr1+/MPOhi granulocytes 2.5-fold at 48 hours, even in 0.01 µM estradiol. This pattern was confirmed morphologically and by quantitative polymerase chain reaction (PCR) assay of lineage markers. To directly assess effects on immature progenitors, transduced cells were cultured for 1 day with and then in methylcellulose without estradiol. A 2-fold increase in monocytic compared with granulocytic colonies was observed in IL-3/IL-6/SCF or GM-CSF, but not G-CSF, even in 0.01 µM estradiol. C/EBP{alpha}-ER induced PU.1 mRNA, and PU.1-ER stimulated monocytic development, suggesting that transcriptional induction of PU.1 by C/EBP{alpha} contributes to monopoiesis. A C/EBP{alpha} variant incapable of zippering with c-Jun did not induce monopoiesis, and a variant unable to bind NF-{kappa}B p50 stimulated granulopoiesis, suggesting their cooperation with C/EBP{alpha} during monocytic commitment.


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