title: Nfkb1 inhibits LPS-induced IFN-β and IL-12 p40 production in macrophages by distinct mechanisms.
abstract: BACKGROUND
  Nfkb1-deficient murine macrophages express higher levels of IFN-β and IL-12 p40 following LPS stimulation than control macrophages, but the molecular basis for this phenomenon has not been completely defined. Nfkb1 encodes several gene products including the NF-κB subunit p50 and its precursor p105. p50 is derived from the N-terminal of 105, and p50 homodimers can exhibit suppressive activity when overexpressed. The C-terminal region of p105 is necessary for LPS-induced ERK activation and it has been suggested that ERK activity inhibits both IFN-β and IL-12 p40 following LPS stimulation. However, the contributions of p50 and the C-terminal domain of p105 in regulating endogenous IFN-β(Ifnb) and IL-12 p40 (Il12b) gene expression in macrophages following LPS stimulation have not been directly compared.
  
  
  RESULTS
  We have used recombinant retroviruses to express p105, p50, and the C-terminal domain of p105 (p105ΔN) in Nfkb1-deficient murine bone marrow-derived macrophages at near endogenous levels. We found that both p50 and p105ΔN inhibited expression of Ifnb, and that inhibition of Ifnb by p105ΔN depended on ERK activation, because a mutant of p105ΔN (p105ΔNS930A) that lacks a key serine necessary to support ERK activation failed to inhibit. In contrast, only p105ΔN but not p50 inhibited Il12b expression. Surprisingly, p105ΔNS930A retained inhibitory activity for Il12b, indicating that ERK activation was not necessary for inhibition. The differential effects of p105ΔNS930A on Ifnb and Il12b expression inversely correlated with the function of one of its binding partners, c-Rel. This raised the possibility that p105ΔNS930A influences gene expression by interfering with the function of c-Rel.
  
  
  CONCLUSIONS
  These results demonstrate that Nfkb1 exhibits multiple gene-specific inhibitory functions following TLR stimulation of murine macrophages.
authors.affiliation: Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts, United States of America.
authors.forename: Xixing
authors.initials: X
authors.lastname: Zhao
authors.forename: Erik J
authors.initials: EJ
authors.lastname: Ross
authors.forename: Yanyan
authors.initials: Y
authors.lastname: Wang
authors.forename: Bruce H
authors.initials: BH
authors.lastname: Horwitz
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D004268:DNA-Binding Proteins / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D005786:Gene Expression Regulation / Q000502:physiology*; D016899:Interferon-beta / Q000378:metabolism*; D016753:Interleukin-10 / Q000235:genetics; D053762:Interleukin-12 Subunit p40 / Q000378:metabolism*; D008070:Lipopolysaccharides; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D052002:NF-kappa B p50 Subunit / Q000235:genetics / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D053762:Interleukin-12 Subunit p40; D008070:Lipopolysaccharides; D052002:NF-kappa B p50 Subunit; C499308:Rag2 protein, mouse; D016753:Interleukin-10; C073701:Nfkb1 protein, mouse; D016899:Interferon-beta