IκBζ augments IL-12– and IL-18–mediated IFN-γ production in human NK cells

Abstract

Interferon-γ (IFN-γ) production by natural killer (NK) cells and cytotoxic lymphocytes is a key component of innate and adaptive immune responses. Because inhibitor of κB-ζ (IκBζ), a Toll-like receptor (TLR)/interleukin-1 receptor (IL-1R) inducible transcription factor, regulates IFN-γ production in KG-1 cells, we tested IκBζ's role in the classic lymphocyte pathway of IL-12/IL-18–induced IFN-γ. Upon stimulation with IL-12/IL-18, monocyte-depleted human peripheral blood lymphocytes expressed the 79-kDa form of IκBζ and released IFN-γ. CD56+ NK cells were shown to be the IκBζ-producing lymphocyte subpopulation, which also released abundant IFN-γ in response to IL-12/IL-18. Importantly, IκBζ was undetectable in CD56− lymphocytes where IFN-γ release was 10-fold lower. In addition, small interfering RNA knockdown of IκBζ suppressed IFN-γ expression in CD56+ cells. The association of IκBζ with the IFN-γ promoter was documented by chromatin immunoprecipitation. IFN-γ promoter activity from IκBζ overexpression was confirmed by luciferase reporter assay. Finally, IκBζ coprecipitated with p65 and p50 NF-κB in NK cells in response to IL-12/IL-18, suggesting that IκBζ's effects on IFN-γ promoter activity are coregulated by NF-κB. These results suggest that IκBζ functions as an important regulator of IFN-γ in human NK cells, further expanding the class of IκBζ-modulated genes.

House Dust Mite Allergens and the Induction of Monocyte Interleukin 1β Production That Triggers an IκBζ-Dependent Granulocyte Macrophage Colony-Stimulating Factor Release from Human Lung Epithelial Cells

“…Notably, at the protein level, IkBz-L is the predominantly expressed isoform in lung epithelial cells. This preferential expression of IkBz-L over IkBz-S is consistent with previous publications from our group and others (21,26,34). To our knowledge, we are the first to demonstrate a role for IkBz in mediating inflammation associated with house dust mite-induced asthma.…”

“…Unlike typical IkB homologs that are constitutively present in the cytosol where they complex to NF-kB Rel domains to inhibit nuclear localization, IkBz is an inducible protein that binds to NF-kB inside the nucleus (20). IkBz forms a complex with either p50 homodimers or p50-p65 heterodimers of NF-kB that bind to the promoter sequences of several secondary response genes, including IL6, IL12, LCN2, IFNG, and DEFB4A (23)(24)(25)(26), and causes regulation of their transcription. Although there is a documented inhibitory role of IkBz (22), its function as a transcriptional activator predominates.…”

Fe-Blf Nanoformulation Targets Survivin to Kill Colon Cancer Stem Cells and Maintains Absorption of Iron, Calcium and Zinc

“…ACSC: Alginate-enclosed chitosan-coated; bLf: Bovine lactoferrin; NC:Nanocarrier/nanocapsule; RBC: Red blood cell. immunity as it activates NK cells to release IFN-γ [41], which may account for the high expression of IL-18 in both wild-type and nude mice. IL-12 is a proinflammatory cytokine responsible for both cell-mediated and humoral immune response, also known to stimulate TNF-α, IFN-γ and NK cells, which may account for its increased expression in both wild-type mice and nude mice [41].…”

Oxidatively Modified Low Density Lipoprotein (LDL) Inhibits TLR2 and TLR4 Cytokine Responses in Human Monocytes but Not in Macrophages

“…Preparation of Cell Lysates and Immunoblotting-Cells were lysed, and cell lysates were prepared as described previously (14). Total protein was estimated using Lowry's assay (BioRad), and equal protein (10 -20 g) was loaded per lane of NuPAGE 4 -12% Bis-Tris gel (Invitrogen).…”

“…Quantitative PCR-Monocytes were stimulated for the times indicated, and total RNA was extracted and converted to cDNA as described previously (14). Primers specific for IB, IL-1␤, IL-6, IL-8, TNF␣, peroxisome proliferator-activated receptor ␥ (PPAR␥), IL-1RII, IL-10, IL-1RN, and mannose receptor C type 1 (MRC-1) were used for analyzing mRNA expression.…”