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Molecular and Cellular Biology, December 2008, p. 7394-7401, Vol. 28, No. 24
0270-7306/08/$08.00+0     doi:10.1128/MCB.01087-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Failure of Terminal Erythroid Differentiation in EKLF-Deficient Mice Is Associated with Cell Cycle Perturbation and Reduced Expression of E2F2 ^,

Andre M. Pilon,^1,2 Murat O. Arcasoy,³ Holly K. Dressman,⁴ Serena E. Vayda,¹ Yelena D. Maksimova,⁵ Jose I. Sangerman,⁵ Patrick G. Gallagher,⁵ and David M. Bodine^1*

Hematopoiesis Section, Genetics and Molecular Biology Branch, NHGRI, NIH, 49 Convent Drive, Bethesda, Maryland 20892-4442,¹ Department of Biochemistry and Molecular Biology, The George Washington University Medical Center, 2300 Eye Street NW, Washington, DC 20037,² Duke University Medical Center, DUMC Box 3912, Durham, North Carolina 27710,³ Duke Institute for Genome Sciences and Policy 2177B CIEMAS, 101 Science Drive, DUMC Box 3382, Durham, North Carolina 27708,⁴ Department of Pediatrics, Yale University School of Medicine, 333 Cedar Street, Box 208064, New Haven, Connecticut 06520-8064⁵

Received 10 July 2008/ Returned for modification 3 September 2008/ Accepted 1 October 2008

Erythroid Krüppel-like factor (EKLF) is a Krüppel-like transcription factor identified as a transcriptional activator and chromatin modifier in erythroid cells. EKLF-deficient (Eklf^–/–) mice die at day 14.5 of gestation from severe anemia. In this study, we demonstrate that early progenitor cells fail to undergo terminal erythroid differentiation in Eklf^–/– embryos. To discover potential EKLF target genes responsible for the failure of erythropoiesis, transcriptional profiling was performed with RNA from wild-type and Eklf^–/– early erythroid progenitor cells. These analyses identified significant perturbation of a network of genes involved in cell cycle regulation, with the critical regulator of the cell cycle, E2f2, at a hub. E2f2 mRNA and protein levels were markedly decreased in Eklf^–/– early erythroid progenitor cells, which showed a delay in the G₁-to-S-phase transition. Chromatin immunoprecipitation analysis demonstrated EKLF occupancy at the proximal E2f2 promoter in vivo. Consistent with the role of EKLF as a chromatin modifier, EKLF binding sites in the E2f2 promoter were located in a region of EKLF-dependent DNase I sensitivity in early erythroid progenitor cells. We propose a model in which EKLF-dependent activation and modification of the E2f2 locus is required for cell cycle progression preceding terminal erythroid differentiation.

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* Corresponding author. Mailing address: National Human Genome Research Institute, Hematopoiesis Section, 49 Convent Drive, MSC-4442, Bldg. 49, Room 4A04, Bethesda, MD 20892. Phone: (301) 402-0902. Fax: (301) 402-4929. E-mail: tedyaz{at}mail.nih.gov

Published ahead of print on 13 October 2008.

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

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Molecular and Cellular Biology, December 2008, p. 7394-7401, Vol. 28, No. 24
0270-7306/08/$08.00+0     doi:10.1128/MCB.01087-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.