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Prepublished online as a Blood First Edition Paper on August 14, 2003; DOI 10.1182/blood-2002-12-3766.
Submitted December 12, 2002
Accepted August 6, 2003
Induction of an embryonic globin gene promoter by short chain fatty acids
Nancy J Dempsey, Laureen S Ojalvo, Davina W Wu, and Jane A Little*
Hematology, Oncology, and Transplantation, Department of Internal Medicine, University of Minnesota, Minneapolis, MN, USA
Laboratory of Cellular and Developmental Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
* Corresponding author; email: littl002{at}umn.edu.
Short-chain fatty acids (SCFAs) and dimethyl sulfoxide (DMSO) induce adult erythroid differentiation in murine erythroleukemia (MEL) cells, but only SCFAs concurrently up-regulate expression from the endogenous embryonic globin gene  y. The y promoter, linked to a reporter gene and stably transfected into MEL cells, was tested during adult erythroid differentiation. Both the y-CACCC site at -114 bps and enhancer sequences (HS2) from the  -globin locus control region (LCR) were essential to maximal SCFA-mediated induction of expression from these constructs in MEL cells. Gel-shift analyses of binding activity from SCFA-induced MEL cell nuclear extracts showed in vitro binding by SP1, SP3, BKLF, and EKLF at the y-CACCC site. Transient cotransfections in non-erythroid NIH/3T3 cells of SP1, SP3, BKLF, or EKLF and HS2 y promoter luciferase constructs, with or without co-activators (p300, CBP, or PCAF) and SCFAs, were performed. SP1, SP3, and EKLF further increased expression from HS2 y promoter constructs following exposure to SCFAs. This effect was variably augmented by coactivators and was diminished in EKLF mutants that were unable to undergo H/FAT-mediated acetylation. In addition, acetylation of SP1 was detectable in NIH/3T3 cells following exposure to SCFAs. In sum, LCR sequence and an embryonic globin gene promoter CACCC site were essential to that promoter's up-regulation during SCFA-mediated induction of adult erythroid differentiation in vitro. Of factors that interact at the CACCC site, SCFA-mediated acetylation is implicated for SP1 and EKLF, and may be a mechanism through which SCFAs induce embryonic/fetal globin gene promoters during adult erythroid differentiation.
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