Induction of an embryonic globin gene promoter by short-chain fatty acids

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 epsilony. The epsilony promoter, linked to a reporter gene and stably transfected into MEL cells, was tested during adult erythroid differentiation. Both the epsilony-CACCC site at -114 bp and enhancer sequences (hypersensitive site 2 [HS2]) from the beta-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 specificity proteins 1 and 3 (SP1, SP3) and basic or erythroid Krüppel-like factors (BKLF, EKLF) at the epsilony-CACCC site. In a functional analysis, transient cotransfections in nonerythroid NIH/3T3 cells of SP1, SP3, BKLF, or EKLF and HS2 epsilony promoter-luciferase constructs, with or without coactivators (p300, CREB-binding protein [CBP], or p300/CBP-associated factor [PCAF]) and SCFAs, were performed. SP1, SP3, and EKLF further increased expression from HS2 epsilony promoter constructs following exposure to SCFAs. This effect was variably augmented by coactivators and was diminished in EKLF mutants that were unable to undergo histone/factor-acetyl transferase (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 in SP1 and EKLF, and may be a mechanism through which SCFAs induce embryonic/fetal globin gene promoters during adult erythroid differentiation.