Research Article| Volume 3, ISSUE 1, P94-106, January 2014

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Molecular network of ChIP-Seq-based NF-κB p65 target genes involves diverse immune functions relevant to the immunopathogenesis of multiple sclerosis


      • We identified 918 ChIP-Seq NF-κB p65 binding sites on protein-coding genes.
      • The binding sites were accumulated in promoter and intronic regions.
      • Their molecular network was associated with immune functions relevant to MS.
      • They included 10 MS risk genes and 49 MS brain lesion-specific proteins.



      The transcription factor nuclear factor-kappa B (NF-κB) acts as a central regulator of immune response, stress response, cell proliferation, and apoptosis. Aberrant regulation of NF-κB function triggers development of cancers, metabolic diseases, and autoimmune diseases. We attempted to characterize a global picture of the NF-κB target gene network relevant to the immunopathogenesis of multiple sclerosis (MS).


      We identified the comprehensive set of 918 NF-κB p65 binding sites on protein-coding genes from chromatin immunoprecipitation followed by deep sequencing (ChIP-Seq) dataset of TNFα-stimulated human B lymphoblastoid cells. The molecular network was studied by a battery of pathway analysis tools of bioinformatics.


      The GenomeJack genome viewer showed that NF-κB p65 binding sites were accumulated in promoter (35.5%) and intronic (54.9%) regions with an existence of the NF-κB consensus sequence motif. A set of 52 genes (5.7%) corresponded to known NF-κB targets by database search. KEGG, PANTHER, and Ingenuity Pathways Analysis (IPA) revealed that the NF-κB p65 target gene network is linked to regulation of immune functions and oncogenesis, including B cell receptor signaling, T cell activation pathway, Toll-like receptor signaling, and apoptosis signaling, and molecular mechanisms of cancers. KeyMolnet indicated an involvement of the complex crosstalk among core transcription factors in the NF-κB p65 target gene network. Furthermore, the set of NF-κB p65 target genes included 10 genes among 98 MS risk alleles and 49 molecules among 709 MS brain lesion-specific proteins.


      These results suggest that aberrant regulation of NF-κB-mediated gene expression, by inducing dysfunction of diverse immune functions, is closely associated with development and progression of MS.


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        • Achiron A.
        • Feldman A.
        • Mandel M.
        • Gurevich M.
        Impaired expression of peripheral blood apoptotic-related gene transcripts in acute multiple sclerosis relapse.
        Annals of the New York Academy of Sciences. 2007; 1107: 155-167
        • Barnes P.J.
        • Karin M.
        Nuclear factor-κB. A pivotal transcription factor in chronic inflammatory diseases.
        New England Journal of Medicine. 1997; 336: 1066-1071
        • Bonetti B.
        • Stegagno C.
        • Cannella B.
        • Rizzuto N.
        • Moretto G.
        • Raine C.S.
        Activation of NF-κB and c-jun transcription factors in multiple sclerosis lesions. Implications for oligodendrocyte pathology.
        American Journal of Pathology. 1999; 155: 1433-1438
        • Brüstle A.
        • Brenner D.
        • Knobbe C.B.
        • et al.
        The NF-κB regulator MALT1 determines the encephalitogenic potential of Th17 cells.
        Journal of Clinical Investigation. 2012; 122: 4698-4709
        • Collins B.E.
        • Smith B.A.
        • Bengtson P.
        • Paulson J.C.
        Ablation of CD22 in ligand-deficient mice restores B cell receptor signaling.
        Nature Immunology. 2006; 7: 199-206
        • Comabella M.
        • Khoury S.J.
        Immunopathogenesis of multiple sclerosis.
        Clinical Immunology. 2012; 142: 2-8
        • Eggert M.
        • Goertsches R.
        • Seeck U.
        • Dilk S.
        • Neeck G.
        • Zettl U.K.
        Changes in the activation level of NF-kappa B in lymphocytes of MS patients during glucocorticoid pulse therapy.
        Journal of the Neurological Sciences. 2008; 264: 145-150
        • Gerstein M.B.
        • Kundaje A.
        • Hariharan M.
        • et al.
        Architecture of the human regulatory network derived from ENCODE data.
        Nature. 2012; 489: 91-100
        • Gilmore T.D.
        Introduction to NF-κB: players, pathways, perspectives.
        Oncogene. 2006; 25: 6680-6684
        • Gregersen P.K.
        • Amos C.I.
        • Lee A.T.
        • et al.
        REL, encoding a member of the NF-κB family of transcription factors, is a newly defined risk locus for rheumatoid arthritis.
        Nature Genetics. 2009; 41: 820-823
        • Gveric D.
        • Kaltschmidt C.
        • Cuzner M.L.
        • Newcombe J.
        Transcription factor NF-κB and inhibitor IκBα are localized in macrophages in active multiple sclerosis lesions.
        Journal of Neuropathology and Experimental Neurology. 1998; 57: 168-178
        • Han M.H.
        • Hwang S.I.
        • Roy D.B.
        • et al.
        Proteomic analysis of active multiple sclerosis lesions reveals therapeutic targets.
        Nature. 2008; 451: 1076-1081
        • Hayden M.S.
        • West A.P.
        • Ghosh S.
        NF-κB and the immune response.
        Oncogene. 2006; 25: 6758-6780
        • Hilliard B.
        • Samoilova E.B.
        • Liu T.S.
        • Rostami A.
        • Chen Y.
        Experimental autoimmune encephalomyelitis in NF-κB-deficient mice: roles of NF-κB in the activation and differentiation of autoreactive T cells.
        Journal of Immunology. 1999; 163: 2937-2943
        • Huang da W.
        • Sherman B.T.
        • Lempicki R.A.
        Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources.
        Nature Protocols. 2009; 4: 44-57
        • International Multiple Sclerosis Genetics Consortium
        • Wellcome Trust Case Control Consortium 2
        • Sawcer S.
        • et al.
        Genetic risk and a primary role for cell-mediated immune mechanisms in multiple sclerosis.
        Nature. 2011; 476: 214-219
        • Kasowski M.
        • Grubert F.
        • Heffelfinger C.
        • et al.
        Variation in transcription factor binding among humans.
        Science. 2010; 328: 232-235
        • Krumbholz M.
        • Derfuss T.
        • Hohlfeld R.
        • Meinl E.
        B cells and antibodies in multiple sclerosis pathogenesis and therapy.
        Nature Reviews Neurology. 2012; 8: 613-623
        • Landt S.G.
        • Marinov G.K.
        • Kundaje A.
        • et al.
        ChIP-Seq guidelines and practices of the ENCODE and modENCODE consortia.
        Genome Research. 2012; 22: 1813-1831
        • Levin L.I.
        • Munger K.L.
        • O'Reilly E.J.
        • Falk K.I.
        • Ascherio A.
        Primary infection with the Epstein-Barr virus and risk of multiple sclerosis.
        Annals of Neurology. 2010; 67: 824-830
        • Lindsey J.W.
        • Agarwal S.K.
        • Tan F.K.
        Gene expression changes in multiple sclerosis relapse suggest activation of T and non-T cells.
        Molecular Medicine. 2011; 17: 95-102
        • Martone R.
        • Euskirchen G.
        • Bertone P.
        • et al.
        Distribution of NF-κB-binding sites across human chromosome 22.
        Proceedings of the National Academy of Sciences of the United States of America. 2003; 100: 12247-12252
        • Martín-Saavedra F.M.
        • Flores N.
        • Dorado B.
        • et al.
        Beta-interferon unbalances the peripheral T cell proinflammatory response in experimental autoimmune encephalomyelitis.
        Molecular Immunology. 2007; 44: 3597-3607
        • Miterski B.
        • Böhringer S.
        • Klein W.
        • et al.
        Inhibitors in the NFκB cascade comprise prime candidate genes predisposing to multiple sclerosis, especially in selected combinations.
        Genes and Immunity. 2002; 3: 211-219
        • Myouzen K.
        • Kochi Y.
        • Okada Y.
        • et al.
        Functional variants in NFKBIE and RTKN2 involved in activation of the NF-κB pathway are associated with rheumatoid arthritis in Japanese.
        PLoS Genetics. 2012; 8: e1002949
        • Oeckinghaus A.
        • Hayden M.S.
        • Ghosh S.
        Crosstalk in NF-κB signaling pathways.
        Nature Immunology. 2011; 12: 695-708
        • Pahan K.
        • Schmid M.
        Activation of nuclear factor-κB in the spinal cord of experimental allergic encephalomyelitis.
        Neuroscience Letters. 2000; 287: 17-20
        • Pahl H.L.
        Activators and target genes of Rel/NF-κB transcription factors.
        Oncogene. 1999; 18: 6853-6866
        • Park J.M.
        • Greten F.R.
        • Wong A.
        • et al.
        Signaling pathways and genes that inhibit pathogen-induced macrophage apoptosis. CREB and NF-κB as key regulators.
        Immunity. 2005; 23: 319-329
        • Park P.J.
        ChIP-Seq: advantages and challenges of a maturing technology.
        Nature Reviews Genetics. 2009; 10: 669-680
        • Rothwarf D.M.
        • Karin M.
        The NF-κB activation pathway: a paradigm in information transfer from membrane to nucleus.
        Science's STKE. 1999; 1999 (RE1)
        • Satoh J.
        • Misawa T.
        • Tabunoki H.
        • Yamamura T.
        Molecular network analysis of T-cell transcriptome suggests aberrant regulation of gene expression by NF-κB as a biomarker for relapse of multiple sclerosis.
        Disease Markers. 2008; 25: 27-35
      1. Satoh J, Tabunoki H. Molecular network of ChIP-Seq-based vitamin D receptor target genes. Multiple Sclerosis; 2013 [in press]. PMID: 23401126. DOI: 10.1177/1352458512471873.

        • Satoh J.
        Bioinformatics approach to identifying molecular biomarkers and networks in multiple sclerosis.
        Clinical and Experimental Neuroimmunology. 2010; 1: 127-140
        • Satoh J.I.
        • Tabunoki H.
        • Yamamura T.
        Molecular network of the comprehensive multiple sclerosis brain-lesion proteome.
        Multiple Sclerosis. 2009; 15: 531-541
        • Schneider G.
        • Henrich A.
        • Greiner G.
        • et al.
        Cross talk between stimulated NF-κB and the tumor suppressor p53.
        Oncogene. 2010; 29: 2795-2806
        • Sica A.
        • Dorman L.
        • Viggiano V.
        • et al.
        Interaction of NF-κB and NFAT with the interferon-γ promoter.
        Journal of Biological Chemistry. 1997; 272: 30412-30420
        • Viatour P.
        • Merville M.P.
        • Bours V.
        Chariot A. Phosphorylation of NF-κB and IκB proteins: implications in cancer and inflammation.
        Trends in Biochemical Sciences. 2005; 30: 43-52
        • Yan J.
        • Greer J.M.
        NF-κBa potential therapeutic target for the treatment of multiple sclerosis.
        CNS & Neurological Disorders: Drug Targets. 2008; 7: 536-557
        • van Loo G.
        • De Lorenzi R.
        • Schmidt H.
        • et al.
        Inhibition of transcription factor NF-κB in the central nervous system ameliorates autoimmune encephalomyelitis in mice.
        Nature Immunology. 2006; 7: 954-961
        • van Zelm M.C.
        • Smet J.
        • Adams B.
        • et al.
        CD81 gene defect in humans disrupts CD19 complex formation and leads to antibody deficiency.
        Journal of Clinical Investigation. 2010; 120: 1265-1274
        • Zepp J.A.
        • Liu C.
        • Qian W.
        • et al.
        Cutting edge: TNF receptor-associated factor 4 restricts IL-17-mediated pathology and signaling processes.
        Journal of Immunology. 2012; 189: 33-37
        • Zhang W.
        • Shi Q.
        • Xu X.
        • et al.
        Aberrant CD40-induced NF-κB activation in human lupus B lymphocytes.
        PLoS One. 2012; 7: e41644
      2. Ziesché E, Kettner-Buhrow D, Weber A, et al. The coactivator role of histone deacetylase 3 in IL-1-signaling involves deacetylation of p65 NF-κB. Nucleic Acids Research; 2013;41:90–109. 10.1093/nar/gks916.