title: Involvement of Notch signaling pathway in regulating IL-12 expression via c-Rel in activated macrophages.
abstract: Macrophages play an important role both in innate and adaptive immune responses. Treatment with interferon (IFN) γ together with lipopolysaccharide (LPS) activates pro-inflammatory macrophages which secrete various pro-inflammatory cytokines including IL-12. IL-12 promotes a Th1 type immune response by directly controlling the differentiation of CD4(+) T helper 1 cells. Activation of Notch signaling pathway was reported in activated macrophages but the involvement of this signaling pathway in IL-12 expression has not been documented. In this study, we investigated the role of Notch signaling in regulating expression of the IL-12/IL-23 subunit, IL-12p40. Using a gamma-secretase inhibitor (GSI) to inhibit Notch signaling, we observed a profound decrease in il12p40 mRNA levels and IL-12p70 secretion upon IFNγ/LPS stimulation. On the other hand, overexpression of activated form of Notch1 in activated RAW264.7 macrophage-like cell lines significantly increased the level of il12p40 mRNA. GSI treatment did not affect the expression of irf5, a master regulator of il12p40 transcription in macrophages. Detailed analysis of the signaling cascades that were affected by this inhibition showed that c-Rel nuclear translocation was inhibited and Erk1/2 activation was compromised by GSI treatment. Addition of exogenous tumor necrosis factor (TNF) α only partially rescued the expression of il12p40 in the presence of GSI. Unexpectedly, inhibition of Notch signaling using a dominant negative (DN) Mastermind-like (MAML) transcription co-activator, did not affect c-Rel nuclear localization upon activation or il12p40 mRNA levels, suggesting that the transcriptional activity of Notch signaling is dispensable for the activation of c-Rel. These results strongly suggest that Notch signaling in activated macrophages is involved in regulating the expression of il12p40 directly via c-Rel and indirectly via TNFα production.
authors.affiliation: Graduate Program in Biotechnology, Faculty of Science, Chulalongkorn University, Phayathai Road, Pathumwan, Bangkok, Thailand.
authors.forename: Natt
authors.initials: N
authors.lastname: Boonyatecha
authors.forename: Naunpun
authors.initials: N
authors.lastname: Sangphech
authors.forename: Wipawee
authors.initials: W
authors.lastname: Wongchana
authors.forename: Pathipark
authors.initials: P
authors.lastname: Kueanjinda
authors.forename: Tanapat
authors.initials: T
authors.lastname: Palaga
mesh_terms: D053829:Amyloid Precursor Protein Secretases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D002460:Cell Line; D005260:Female; D018398:Homeodomain Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D050835:Interferon Regulatory Factors / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D007371:Interferon-gamma / Q000276:immunology; D053762:Interleukin-12 Subunit p40 / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology; D020935:MAP Kinase Signaling System; D008262:Macrophage Activation / Q000187:drug effects / Q000235:genetics* / Q000276:immunology*; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D020569:Proto-Oncogene Proteins c-rel / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D051881:Receptor, Notch1 / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D051880:Receptors, Notch / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015398:Signal Transduction; D000072056:Transcription Factor HES-1; D015533:Transcriptional Activation / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000276:immunology / Q000378:metabolism
chemical_list: D053829:Amyloid Precursor Protein Secretases; D051792:Basic Helix-Loop-Helix Proteins; D018398:Homeodomain Proteins; D050835:Interferon Regulatory Factors; D007371:Interferon-gamma; D053762:Interleukin-12 Subunit p40; D008070:Lipopolysaccharides; D020569:Proto-Oncogene Proteins c-rel; D012333:RNA, Messenger; D051881:Receptor, Notch1; D051880:Receptors, Notch; D000072056:Transcription Factor HES-1; D014409:Tumor Necrosis Factor-alpha; C489734:Hes1 protein, mouse; C498178:Irf5 protein, mouse; C497762:Notch1 protein, mouse