Chromatin immunoprecipitation (ChIP) studies indicate a role for CCAAT enhancer binding proteins alpha and epsilon (C/EBPα and C/EBPε) and CDP/cut in myeloid maturation-induced lactoferrin gene expression

Abstract

IntroductionAlthough a number of transcription factors have been implicated in myelopoiesis, 1-4 several members of the CCAAT enhancer binding protein (C/EBP) family of transcription factors have been shown to be indispensable for normal development of the myeloid lineage. The C/EBP family of transcription factors is a class of basic region/leucine zipper (bZIP) factors that recognize the consensus DNA-binding sequence, 5Ј-ATTGCGCAAT-3Ј, as obligate dimers. Currently, 6 members of this family of transcription factors have been described, including C/EBP ␣, ␤, ␥, ␦, ⑀, and CHOP-10/ GADD 153, all of which contain highly homologous dimerization (leucine-zipper) domains and DNA-binding (basic-region) motifs. These proteins have been implicated in regulating gene expression in a variety of cell types, including adipocytes (reviewed in Darlington et al 5 ), constitutive and acute phase response genes in the liver (reviewed in Diehl 6 ), and myelomonocytic cells (reviewed in Lekstrom-Himes and Xanthopoulos 7 ).Severe hematopoietic abnormalities have been reported for mice nullizygous for C/EBP␣ and ⑀. Although C/EBP␣ Ϫ/Ϫ mice die perinatally due to defects in gluconeogenesis, 8,9 they also demonstrate an early block in the differentiation of granulocytes. C/EBP␣ has been postulated to be a master regulator of the granulopoietic developmental program. Although C/EBP␣ is expressed in several tissues, its expression in the hematopoietic compartment is restricted to the granulocytic lineage, where it is expressed in the most primitive precursors. Recently, mutations within the C/EBP␣ gene have been demonstrated in patients with acute myeloid leukemia (AML) (French-American-British [FAB] classification M1 and M2). 10 Additionally, the AML1/ETO fusion protein that results from the t(8;21) translocation in patients with M2 AML has been shown to down-regulate C/EBP␣ expression, 11 suggesting that loss of C/EBP␣ function in AML patients may contribute to the observed early block in the granulocytic maturation pathway.C/EBP⑀ is expressed relatively late in the granulocytic maturation pathway and has been shown to play a role in late myeloid gene expression. [12][13][14] Absence of C/EBP⑀ in mutant mice causes a late block in the differentiation of granulocytes. C/EBP⑀ Ϫ/Ϫ mice produce hyposegmented granulocytes that are functionally defective. Mutant mice usually survive 2 to 5 months but eventually succumb to opportunistic bacterial infections. 15 It has been demonstrated that C/EBP⑀ Ϫ/Ϫ mice express absent or low levels of RNA for several genes, including the secondary granule protein (SGP) genes (murine lactoferrin [mLF], murine neutrophil gelatinase [mNG], and murine neutrophil collagenase [mNC]). Studies from our laboratory have demonstrated that expression of mLF and mNC in the developing neutrophil is dependent on intact C/EBP binding sites within their gene promoters (Khanna-Gupta et al 16 ; and A.K.-G. and N.B., manuscript in preparation, 2003). Mutations within the C/EBP⑀ gene have been described in patients with spec...

Vitamin D3 induces expression of human cathelicidin antimicrobial peptide 18 in newborns

“…As shown in prior studies [30,49], myeloid-specific transcription factor C/EBPe is closely related to the induction of hCAP18 and NGAL expression in human neutrophils. Moreover, BPI and lactoferrin, the expressions of which are regulated by C/EBPe, are also reduced in neonates [34,50]. On the other hand, primary granule protein MPO, which is regulated independently of C/EBPe, was not reduced in neonates.…”

Activation of Protein Kinase Cδ by All-trans-retinoic Acid

“…Chromatin Immunoprecipitation (ChIP) Assays-ChIP assays were performed essentially as described previously (44,45). ChIP DNA was used as a template for PCR using DR5 RARE forward (5Ј-CAC TGC AGA AAC AGC CAG-3Ј) and reverse (5Ј-CAT GGG CAG GCT GAT AAG-3Ј) primers.…”

RARα-PLZF oncogene inhibits C/EBPα function in myeloid cells

“…C/EBPα is important for optimal Csf3r expression in vivo (22), although the induction of granulocytic differentiation by C/EBPα could be Csf3r-independent (23). Therefore, we analyzed the expression levels of additional C/EBPα target genes that are involved in myeloid differentiation, i.e., Ltf, Mpo, Ela2 as well as miR-223 (24)(25)(26)(27), all produced at the promyelocyte stage of differentiation. Interestingly, these C/EBPα target genes were strongly reduced in RP7, RP11, and RP-TAP19 clones compared with parental cells or control cells expressing the empty TAP vector ( Fig.…”