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. 1995 Feb;63(2):601–608. doi: 10.1128/iai.63.2.601-608.1995

Differential expression of interferon regulatory factor 1 (IRF-1), IRF-2, and interferon consensus sequence binding protein genes in lipopolysaccharide (LPS)-responsive and LPS-hyporesponsive macrophages.

S A Barber 1, M J Fultz 1, C A Salkowski 1, S N Vogel 1
PMCID: PMC173039  PMID: 7822029

Abstract

Macrophages secrete interferon (IFN), as well as other cytokines, following lipopolysaccharide (LPS) stimulation. The interferon regulatory factors (IRFs) comprise a family of DNA-binding proteins that have been implicated in the transcriptional regulation of IFN and certain IFN-inducible genes. We therefore characterized basal and LPS-inducible levels of IRF-1, IRF-2, and interferon consensus sequence binding protein (ICSBP) mRNA in LPS-responsive macrophages and compared the expression of these genes in macrophages that typify two murine models of LPS hyporesponsiveness. In the first model, the LPS-hyporesponsive phenotype of the C3H/HeJ mouse is genetically determined and maps to the Lps locus on mouse chromosome 4. In the second model, normally LPS-responsive macrophages acquire a transient LPS-hyporesponsive phenotype following a prior exposure to LPS, a phenomenon referred to as "endotoxin tolerance." Using reverse transcription PCR, we detected basal levels of IRF-1 mRNA in LPS-responsive (Lpsn) macrophages that were approximately 15 times higher than those found in LPS-hyporesponsive (Lpsd) macrophages. Conversely, Lpsd macrophages expressed basal levels of IRF-2 mRNA that were approximately 18 times higher than those expressed in Lpsn macrophages. LPS stimulation resulted in a dose- and time-dependent accumulation of IRF-1, IRF-2, and ICSBP mRNA only in Lpsn macrophages. Cycloheximide inhibited the accumulation of LPS-stimulated IRF-2 and ICSBP mRNA, but not IRF-1 mRNA, thus designating IRF-1 an immediate-early, LPS-inducible gene. Finally, macrophages rendered tolerant to endotoxin expressed elevated but nonmaximal mRNA levels for all three transcription factors that are not reinduced upon secondary challenge with LPS. Thus, the IRFs may represent yet an additional molecular pathway in the complex response to LPS.

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Selected References

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  1. Abdollahi A., Lord K. A., Hoffman-Liebermann B., Liebermann D. A. Interferon regulatory factor 1 is a myeloid differentiation primary response gene induced by interleukin 6 and leukemia inhibitory factor: role in growth inhibition. Cell Growth Differ. 1991 Aug;2(8):401–407. [PubMed] [Google Scholar]
  2. Belardelli F., Gessani S., Proietti E., Locardi C., Borghi P., Watanabe Y., Kawade Y., Gresser I. Studies on the expression of spontaneous and induced interferons in mouse peritoneal macrophages by means of monoclonal antibodies to mouse interferons. J Gen Virol. 1987 Aug;68(Pt 8):2203–2212. doi: 10.1099/0022-1317-68-8-2203. [DOI] [PubMed] [Google Scholar]
  3. Bovolenta C., Driggers P. H., Marks M. S., Medin J. A., Politis A. D., Vogel S. N., Levy D. E., Sakaguchi K., Appella E., Coligan J. E. Molecular interactions between interferon consensus sequence binding protein and members of the interferon regulatory factor family. Proc Natl Acad Sci U S A. 1994 May 24;91(11):5046–5050. doi: 10.1073/pnas.91.11.5046. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Dong Z., Qi X., Fidler I. J. Tyrosine phosphorylation of mitogen-activated protein kinases is necessary for activation of murine macrophages by natural and synthetic bacterial products. J Exp Med. 1993 Apr 1;177(4):1071–1077. doi: 10.1084/jem.177.4.1071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Driggers P. H., Ennist D. L., Gleason S. L., Mak W. H., Marks M. S., Levi B. Z., Flanagan J. R., Appella E., Ozato K. An interferon gamma-regulated protein that binds the interferon-inducible enhancer element of major histocompatibility complex class I genes. Proc Natl Acad Sci U S A. 1990 May;87(10):3743–3747. doi: 10.1073/pnas.87.10.3743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Fertsch D., Schoenberg D. R., Germain R. N., Tou J. Y., Vogel S. N. Induction of macrophage Ia antigen expression by rIFN-gamma and down-regulation by IFN-alpha/beta and dexamethasone are mediated by changes in steady-state levels of Ia mRNA. J Immunol. 1987 Jul 1;139(1):244–249. [PubMed] [Google Scholar]
  7. Fertsch D., Vogel S. N. Recombinant interferons increase macrophage Fc receptor capacity. J Immunol. 1984 May;132(5):2436–2439. [PubMed] [Google Scholar]
  8. Fujita T., Reis L. F., Watanabe N., Kimura Y., Taniguchi T., Vilcek J. Induction of the transcription factor IRF-1 and interferon-beta mRNAs by cytokines and activators of second-messenger pathways. Proc Natl Acad Sci U S A. 1989 Dec;86(24):9936–9940. doi: 10.1073/pnas.86.24.9936. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fultz M. J., Barber S. A., Dieffenbach C. W., Vogel S. N. Induction of IFN-gamma in macrophages by lipopolysaccharide. Int Immunol. 1993 Nov;5(11):1383–1392. doi: 10.1093/intimm/5.11.1383. [DOI] [PubMed] [Google Scholar]
  10. Gessani S., Belardelli F., Borghi P., Boraschi D., Gresser I. Correlation between the lipopolysaccharide response of mice and the capacity of mouse peritoneal cells to transfer an antiviral state. Role of endogenous interferon. J Immunol. 1987 Sep 15;139(6):1991–1998. [PubMed] [Google Scholar]
  11. Gessani S., Di Marzio P., Rizza P., Belardelli F., Baglioni C. Posttranscriptional regulation of interferon mRNA levels in peritoneal macrophages. J Virol. 1991 Feb;65(2):989–991. doi: 10.1128/jvi.65.2.989-991.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Harada H., Fujita T., Miyamoto M., Kimura Y., Maruyama M., Furia A., Miyata T., Taniguchi T. Structurally similar but functionally distinct factors, IRF-1 and IRF-2, bind to the same regulatory elements of IFN and IFN-inducible genes. Cell. 1989 Aug 25;58(4):729–739. doi: 10.1016/0092-8674(89)90107-4. [DOI] [PubMed] [Google Scholar]
  13. Harada H., Willison K., Sakakibara J., Miyamoto M., Fujita T., Taniguchi T. Absence of the type I IFN system in EC cells: transcriptional activator (IRF-1) and repressor (IRF-2) genes are developmentally regulated. Cell. 1990 Oct 19;63(2):303–312. doi: 10.1016/0092-8674(90)90163-9. [DOI] [PubMed] [Google Scholar]
  14. Havell E. A., Spitalny G. L. Endotoxin-induced interferon synthesis in macrophage cultures. J Reticuloendothel Soc. 1983 May;33(5):369–380. [PubMed] [Google Scholar]
  15. Hayes M. P., Zoon K. C. Priming of human monocytes for enhanced lipopolysaccharide responses: expression of alpha interferon, interferon regulatory factors, and tumor necrosis factor. Infect Immun. 1993 Aug;61(8):3222–3227. doi: 10.1128/iai.61.8.3222-3227.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Henricson B. E., Benjamin W. R., Vogel S. N. Differential cytokine induction by doses of lipopolysaccharide and monophosphoryl lipid A that result in equivalent early endotoxin tolerance. Infect Immun. 1990 Aug;58(8):2429–2437. doi: 10.1128/iai.58.8.2429-2437.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Henricson B. E., Manthey C. L., Perera P. Y., Hamilton T. A., Vogel S. N. Dissociation of lipopolysaccharide (LPS)-inducible gene expression in murine macrophages pretreated with smooth LPS versus monophosphoryl lipid A. Infect Immun. 1993 Jun;61(6):2325–2333. doi: 10.1128/iai.61.6.2325-2333.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kamijo R., Harada H., Matsuyama T., Bosland M., Gerecitano J., Shapiro D., Le J., Koh S. I., Kimura T., Green S. J. Requirement for transcription factor IRF-1 in NO synthase induction in macrophages. Science. 1994 Mar 18;263(5153):1612–1615. doi: 10.1126/science.7510419. [DOI] [PubMed] [Google Scholar]
  19. Kessler D. S., Levy D. E., Darnell J. E., Jr Two interferon-induced nuclear factors bind a single promoter element in interferon-stimulated genes. Proc Natl Acad Sci U S A. 1988 Nov;85(22):8521–8525. doi: 10.1073/pnas.85.22.8521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kessler D. S., Veals S. A., Fu X. Y., Levy D. E. Interferon-alpha regulates nuclear translocation and DNA-binding affinity of ISGF3, a multimeric transcriptional activator. Genes Dev. 1990 Oct;4(10):1753–1765. doi: 10.1101/gad.4.10.1753. [DOI] [PubMed] [Google Scholar]
  21. Leu R. W., Rummage J. A., Horn M. J. Characterization of murine macrophage Fc receptor-dependent phagocytosis and antibody-dependent cellular cytotoxicity during in vitro culture with interferons-gamma, alpha/beta and/or fetal bovine serum. Immunobiology. 1989 Feb;178(4-5):340–350. doi: 10.1016/S0171-2985(89)80057-9. [DOI] [PubMed] [Google Scholar]
  22. Manthey C. L., Brandes M. E., Perera P. Y., Vogel S. N. Taxol increases steady-state levels of lipopolysaccharide-inducible genes and protein-tyrosine phosphorylation in murine macrophages. J Immunol. 1992 Oct 1;149(7):2459–2465. [PubMed] [Google Scholar]
  23. Matsuyama T., Kimura T., Kitagawa M., Pfeffer K., Kawakami T., Watanabe N., Kündig T. M., Amakawa R., Kishihara K., Wakeham A. Targeted disruption of IRF-1 or IRF-2 results in abnormal type I IFN gene induction and aberrant lymphocyte development. Cell. 1993 Oct 8;75(1):83–97. [PubMed] [Google Scholar]
  24. McIntire F. C., Sievert H. W., Barlow G. H., Finley R. A., Lee A. Y. Chemical, physical, biological properties of a lipopolysaccharide from Escherichia coli K-235. Biochemistry. 1967 Aug;6(8):2363–2372. doi: 10.1021/bi00860a011. [DOI] [PubMed] [Google Scholar]
  25. Miyamoto M., Fujita T., Kimura Y., Maruyama M., Harada H., Sudo Y., Miyata T., Taniguchi T. Regulated expression of a gene encoding a nuclear factor, IRF-1, that specifically binds to IFN-beta gene regulatory elements. Cell. 1988 Sep 9;54(6):903–913. doi: 10.1016/s0092-8674(88)91307-4. [DOI] [PubMed] [Google Scholar]
  26. Nelson N., Marks M. S., Driggers P. H., Ozato K. Interferon consensus sequence-binding protein, a member of the interferon regulatory factor family, suppresses interferon-induced gene transcription. Mol Cell Biol. 1993 Jan;13(1):588–599. doi: 10.1128/mcb.13.1.588. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Perera P. Y., Barber S. A., Ching L. M., Vogel S. N. Activation of LPS-inducible genes by the antitumor agent 5,6-dimethylxanthenone-4-acetic acid in primary murine macrophages. Dissection of signaling pathways leading to gene induction and tyrosine phosphorylation. J Immunol. 1994 Nov 15;153(10):4684–4693. [PubMed] [Google Scholar]
  28. Pine R., Canova A., Schindler C. Tyrosine phosphorylated p91 binds to a single element in the ISGF2/IRF-1 promoter to mediate induction by IFN alpha and IFN gamma, and is likely to autoregulate the p91 gene. EMBO J. 1994 Jan 1;13(1):158–167. doi: 10.1002/j.1460-2075.1994.tb06245.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Pine R., Decker T., Kessler D. S., Levy D. E., Darnell J. E., Jr Purification and cloning of interferon-stimulated gene factor 2 (ISGF2): ISGF2 (IRF-1) can bind to the promoters of both beta interferon- and interferon-stimulated genes but is not a primary transcriptional activator of either. Mol Cell Biol. 1990 Jun;10(6):2448–2457. doi: 10.1128/mcb.10.6.2448. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Politis A. D., Ozato K., Coligan J. E., Vogel S. N. Regulation of IFN-gamma-induced nuclear expression of IFN consensus sequence binding protein in murine peritoneal macrophages. J Immunol. 1994 Mar 1;152(5):2270–2278. [PubMed] [Google Scholar]
  31. Salkowski C. A., Vogel S. N. Lipopolysaccharide increases glucocorticoid receptor expression in murine macrophages. A possible mechanism for glucocorticoid-mediated suppression of endotoxicity. J Immunol. 1992 Dec 15;149(12):4041–4047. [PubMed] [Google Scholar]
  32. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  33. Tanaka N., Kawakami T., Taniguchi T. Recognition DNA sequences of interferon regulatory factor 1 (IRF-1) and IRF-2, regulators of cell growth and the interferon system. Mol Cell Biol. 1993 Aug;13(8):4531–4538. doi: 10.1128/mcb.13.8.4531. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Tanaka N., Taniguchi T. Cytokine gene regulation: regulatory cis-elements and DNA binding factors involved in the interferon system. Adv Immunol. 1992;52:263–281. doi: 10.1016/s0065-2776(08)60877-9. [DOI] [PubMed] [Google Scholar]
  35. Tebo J. M., Hamilton T. A. Lipopolysaccharide induces DNA binding activity specific for the IFN-stimulated response element in murine peritoneal macrophages. J Immunol. 1992 Oct 1;149(7):2352–2357. [PubMed] [Google Scholar]
  36. Tokunaga K., Taniguchi H., Yoda K., Shimizu M., Sakiyama S. Nucleotide sequence of a full-length cDNA for mouse cytoskeletal beta-actin mRNA. Nucleic Acids Res. 1986 Mar 25;14(6):2829–2829. doi: 10.1093/nar/14.6.2829. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Veals S. A., Kessler D. S., Josiah S., Leonard D. G., Levy D. E. Signal transduction pathway activating interferon-alpha-stimulated gene expression. Br J Haematol. 1991 Oct;79 (Suppl 1):9–13. doi: 10.1111/j.1365-2141.1991.tb08110.x. [DOI] [PubMed] [Google Scholar]
  38. Veals S. A., Santa Maria T., Levy D. E. Two domains of ISGF3 gamma that mediate protein-DNA and protein-protein interactions during transcription factor assembly contribute to DNA-binding specificity. Mol Cell Biol. 1993 Jan;13(1):196–206. doi: 10.1128/mcb.13.1.196. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Veals S. A., Schindler C., Leonard D., Fu X. Y., Aebersold R., Darnell J. E., Jr, Levy D. E. Subunit of an alpha-interferon-responsive transcription factor is related to interferon regulatory factor and Myb families of DNA-binding proteins. Mol Cell Biol. 1992 Aug;12(8):3315–3324. doi: 10.1128/mcb.12.8.3315. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Virca G. D., Kim S. Y., Glaser K. B., Ulevitch R. J. Lipopolysaccharide induces hyporesponsiveness to its own action in RAW 264.7 cells. J Biol Chem. 1989 Dec 25;264(36):21951–21956. [PubMed] [Google Scholar]
  41. Vogel S. N., Fertsch D. Macrophages from endotoxin-hyporesponsive (Lpsd) C3H/HeJ mice are permissive for vesicular stomatitis virus because of reduced levels of endogenous interferon: possible mechanism for natural resistance to virus infection. J Virol. 1987 Mar;61(3):812–818. doi: 10.1128/jvi.61.3.812-818.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Vogel S. N., Havell E. A., Spitalny G. L. Monoclonal antibody-mediated inhibition of interferon-gamma-induced macrophage antiviral resistance and surface antigen expression. J Immunol. 1986 Apr 15;136(8):2917–2923. [PubMed] [Google Scholar]
  43. Watanabe N., Sakakibara J., Hovanessian A. G., Taniguchi T., Fujita T. Activation of IFN-beta element by IRF-1 requires a posttranslational event in addition to IRF-1 synthesis. Nucleic Acids Res. 1991 Aug 25;19(16):4421–4428. doi: 10.1093/nar/19.16.4421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Watson J., Kelly K., Largen M., Taylor B. A. The genetic mapping of a defective LPS response gene in C3H/HeJ mice. J Immunol. 1978 Feb;120(2):422–424. [PubMed] [Google Scholar]

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