Skip to main content
SpringerLink
Log in

C/EBPε participates in all-trans retinoic acid induction of PI3Kγ in U937 cells via an intronic matrix attachment region sequence

  • Published:
Molecular Biology Reports Aims and scope Submit manuscript

Abstract

ATRA (all-trans retinoic acid) regulates gene expression by binding as a ligand to its specific receptors like C/EBPε which is directly induced. In the U937 cell line, PI3Kγ is selectively induced over other PI3Ks by ATRA, although the mechanism is still unclear. Here, we show that C/EBPε and PI3Kγ are induced in U937 cells by ATRA both in levels of mRNA and protein. Reporter gene assay revealed that C/EBPε is able to interact with a previously identified 2 kb MAR (matrix attachment region) sequence in the last intron of PI3Kγ gene, and increases its linked heterogenous reporter gene expression. ChIP assay showed that induction of endogenous PI3Kγ is at least partially caused by enhanced, direct C/EBPε binding to a 15 bp sequence at nucleotides 1428–1442 within this MAR sequence, and EMSA analysis confirmed this binding in vitro. The results above collectively show that C/EBPε participates in ATRA induction of PI3Kγ.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Balmer JE, Blomhoff R (2002) Gene expression regulation by retinoic acid. J Lipid Res 43:1773–1808

    Article  CAS  PubMed  Google Scholar 

  2. Wang ZY (2002) Mechanism of action of all-trans retinoic acid and arsenic trioxide in the treatment of acute promyelocytic leukemia. Gan To Kagaku Ryoho 29:214–218

    CAS  PubMed  Google Scholar 

  3. Lennartsson A, Vidovic K, Pass MB, Cowland JB, Gullberg U (2006) All-trans retinoic acid-induced expression of bactericidal/permeability-increasing protein (BPI) in human myeloid cells correlates to binding of C/EBPbeta and C/EBPepsilon to the BPI promoter. J Leukoc Biol 80:196–203

    Article  CAS  PubMed  Google Scholar 

  4. Wang L, Bai J, Hu Y (2008) Identification of the RA response element and transcriptional silencer in human αCaMKII promoter. Mol Biol Rep 35:37–44

    Article  PubMed  Google Scholar 

  5. Park DJ, Chumakov AM, Vuong PT, Chih DY, Gombart AF, Miller WH, Koeffler HP (1999) CCAAT/enhancer binding protein ε is a potential retinoid target gene in acute promyelocytic leukemia treatment. J Clin Invest 103:1399–1408

    Article  CAS  PubMed  Google Scholar 

  6. Chih DY, Chumakov AM, Park DJ, Silla AG, Koeffler HP (1997) Modulation of mRNA expression of a novel human myeloid-selective CCAAT/enhancer binding protein gene (C/EBPε). Blood 90:2987–2994

    CAS  PubMed  Google Scholar 

  7. Kubota T, Hirama T, Verbeek W, Kawano S, Chih DY, Chumakov AM, Taguchi H, Koeffler HP, DNase I (2001) Hypersensitivity analysis of the human CCAAT enhancer binding protein ε gene. Leu Res 25:981–995

    Article  CAS  Google Scholar 

  8. Baier R, Bondeva T, Klinger R, Bondev A, Wetzker R (1999) Retinoic acid induces selective expression of phosphoinositide 3-kinase gamma in myelomonocytic U937 cells. Cell Growth Differ 10:447–456

    CAS  PubMed  Google Scholar 

  9. Hirsch E, Wymann MP, Patrucco E, Tolosano E, Bulgarelli-Leva G, Marengo S, Rocchi M, Altruda F (2000) Analysis of the murine phosphoinositide 3-kinase γ gene. Gene 256:69–81

    Article  CAS  PubMed  Google Scholar 

  10. Dai BB, Ying L, Cai R, Li Y, Zhang X, Lu J, Qian GX (2006) Identification of a nuclear matrix attachment region like sequence in the last intronic of PI3Kγ. Biochem Biophys Res Commun 341:583–590

    Article  CAS  PubMed  Google Scholar 

  11. Cai R, Xu WR, Dai BB, Cai X, Xu RT, Lu J (2010) SATB1 binds an intronic MAR sequence in human PI3kγ in vitro. Mol Biol Rep 37:1461–1465

    Article  CAS  PubMed  Google Scholar 

  12. Li K, Cai R, Dai BB, Zhang XQ, Wang HJ, Ge SF, Xu WR, Lu J (2007) SATB1 regulates SPARC expression in K562 cell line through binding to a specific sequence in the third intron. Biochem Biophys Res Commun 356:6–12

    Article  CAS  PubMed  Google Scholar 

  13. Chumakov AM, Grillier I, Chumakova E, Chih D, Slater J, Koeffler HP (1997) Cloning of the novel human myeloid-specific C/EBP-ε transcriptional factor. Mol Cell Biol 17:1375–1386

    CAS  PubMed  Google Scholar 

  14. Lenny N, Westendorf JJ, Hiebert SW (1997) Transcriptional regulation during myelopoiesis. Mol Biol Rep 24:157–168

    Article  CAS  PubMed  Google Scholar 

  15. Hawkins PT, Stephens LR (2007) PI3Kgamma is a key regulator of inflammatory responses and cardiovascular homeostasis. Science 318:64–66

    Article  CAS  PubMed  Google Scholar 

  16. Davie JR (1997) Nuclear matrix, dynamic histone acetylation and transcriptionally active chromatin. Mol Biol Rep 24:197–207

    Article  CAS  PubMed  Google Scholar 

  17. Cockerill PN, Yuen MH, Garrard WT (1987) The enhancer of the immunoglobulin heavy chain locus is flanked by presumptive chromosomal loop anchorage elements. J Biol Chem 262:5394–5397

    CAS  PubMed  Google Scholar 

  18. Cockerill PN, Garrard WT (1986) Chromosomal loop anchorage of the kappa immunoglobulin gene occurs next to the enhancer in a region containing topoisomerase II sites. Cell 44:273–282

    Article  CAS  PubMed  Google Scholar 

  19. Chattopadhyay S, Whitehurst CE, Chen J (1998) A nuclear matrix attachment region upstream of the T cell receptor beta gene enhancer binds Cux/CDP and SATB1 and modulates enhancer-dependent reporter gene expression but not endogenous gene expression. J Biol Chem 273:29838–29846

    Article  CAS  PubMed  Google Scholar 

  20. Zhong XP, Carabana J, Kranagel MS (1999) Flanking nuclear matrix attachment region synergize with the T cell receptor δ enhancer to promote V(D)J recombination. Proc Natl Acad Sci USA 96:11970–11975

    Article  CAS  PubMed  Google Scholar 

  21. Forrester WC, van Genderen C, Jenuwein T, Grosschedl R (1994) Dependence of enhancer-mediated transcription of the immunoglobulin mu gene on nuclear matrix attachment regions. Science 26:1221–1225

    Article  Google Scholar 

  22. Blackwood EM, Kadonaga JT (1998) Going the distance: a current view of enhancer action. Science 281:61–63

    Article  Google Scholar 

  23. Williamson EA, Williamson IK, Chumako AM, Friedman AD, Koeffler HP (2005) CCAAT/enhancer binding protein epsilon: changes in function upon phosphorylation by p38 MAP kinase. Blood 105:3841–3847

    Article  CAS  PubMed  Google Scholar 

  24. Kim J, Cantwell CA, Johnson PF, Pfarr CM, Williams SC (2002) Transcriptional activity of CCAAT/enhancer-binding Proteins is controlled by a conserved inhibitory domain that is a target for sumoylation. J Biol Chem 277:38037–38044

    Article  CAS  PubMed  Google Scholar 

  25. Kim J, Sharma S, Li Y, Cobos E, Palvimo JJ, Williams SC (2005) Repression and coactivation of CCAAT/enhancer binding protein epsilon by sumoylation and protein inhibitor of activated STAT x proteins. J Biol Chem 280:12246–12254

    Article  CAS  PubMed  Google Scholar 

  26. Chumakov AM, Silla A, Williamson EA, Koeffler HP (2007) Modulation of DNA binding properties of CCAAT/enhancer binding protein epsilon by heterodimer formation and interactions with NFkappaB pathway. Blood 109:4209–4219

    Article  CAS  PubMed  Google Scholar 

  27. Gombart AF, Kwok SH, Anderson KL, Yamaguchi Y, Torbett BE, Koeffler HP (2003) Regulation of the neutrophil and eosinophil secondary granule gene expression by transcription factors C/EBPε and PU.1. Blood 101:3265–3273

    Article  CAS  PubMed  Google Scholar 

  28. Verbeek W, Gombart AF, Chumakov AM, Müller C, Friedman AD, Koeffler HP (1999) C/EBPε directly interacts with the DNA binding domain of c-myb and cooperatively activates transcription of myeloid promoters. Blood 93:3327–3337

    CAS  PubMed  Google Scholar 

  29. Gombart AF, Grewal J, Koeffler HP (2007) ATF4 differentially regulates transcriptional activation of myeloid-specific genes by C/EBPepsilon and C/EBPalpha. J Leukoc Biol 81:1535–1547

    Article  CAS  PubMed  Google Scholar 

  30. Gery S, Gombart AF, Fung YK, Koeffler HP (2004) C/EBPε interacts with retinoblastoma and E2F1 during granulopoiesis. Blood 103:828–834

    Article  CAS  PubMed  Google Scholar 

  31. Chih DY, Park DJ, Gross M, Idos G, Vuong PT, Hirama T, Chumakov AM, Said J, Koeffler HP (2004) Protein partners of C/EBPε. Exp Hem 32:1173–1181

    Article  CAS  Google Scholar 

  32. Tan JA, Hall SH, Hamil KG, Grossman G, Petrusz P, French FS (2002) Protein inhibitors of activated STAT resemble scaffold attachment factors and function as interacting nuclear receptor coregulators. J Biol Chem 277:16993–17001

    Article  CAS  PubMed  Google Scholar 

  33. Liu B, Yang Y, Chernishof V, Loo RR, Jang H, Tahk S, Yang R, Mink S, Shultz D, Bellone CJ, Loo JA, Shuai K (2007) Proinflammatory stimuli induce IKKalpha-mediated phosphorylation of PIAS1 to restrict inflammation and immunity. Cell 129:903–914

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by Grant No.30800636 from the China National Nature Science Foundation.

Author information

Authors and Affiliations

  1. Department of Molecular and Cellular Biology, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, 280# Chongqing Road(S), Shanghai, 200025, People’s Republic of China

    Rong Cai, Xia Cai, Weirong Xu & Jian Lu

  2. Department of Oncology, Shanghai Pulmonary Hospital, Tongji Univeristy School of Medicine, 507# Zhengmin Road, Shanghai, 200025, People’s Republic of China

    Bin Chen

Authors
  1. Rong Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xia Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Bin Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Weirong Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Jian Lu
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jian Lu.

Additional information

Rong Cai and Xia Cai contributed equally to this article.

Electronic supplementary material

Below is the link to the electronic supplementary material.

11033_2010_34_MOESM1_ESM.tif

ATRA-enhanced PIMAR-linked Luc activity in U937 cells. Both the COS-7 and U-937 cells were transiently transfected with the plasmids indicated, then 24 h later, treated or left untreated with 5 mM ATRA for another 24 h and lysed for relative luciferase activity measurement. In comparison with in COS-7, ATRA treatment alone increased the activity of the PIMAR by more than 3-time in the U937 cell line. This observation indicated that ATRA might control the transcriptional regulatory activity of PIMAR by affecting the expression or binding activity of regulatory transcription factors (TF) or cofactors able to interact with PIMAR. Moreover, it was observed that C/EBPε enhanced by ATRA, increased PIMAR activity by about 2-fold in COS-7, and 4-fold in the U937 cell line. These data indicate the binding activity of C/EBPε protein from the plasmid co-transfected with the PIMAR sequence was reinforced by ATRA in both of the two cell lines above. However, since there are endogenous C/EBPε and other haematopoietic tissue-specific TFs expressed in U937 cells, the enhancement of ATRA on PIMAR activity was stronger. (TIFF 74 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Cai, R., Cai, X., Chen, B. et al. C/EBPε participates in all-trans retinoic acid induction of PI3Kγ in U937 cells via an intronic matrix attachment region sequence. Mol Biol Rep 37, 3795–3800 (2010). https://doi.org/10.1007/s11033-010-0034-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11033-010-0034-1

Keywords

Search

Navigation