Fig 2: Expressions of c-Rel and its inflammatory targets are increased in the cornea of mice with corneal injury. Mouse corneal epithelium was either untreated (n = 6) or scraped (n = 6) as described in the “Materials and methods” section. Corneas were isolated 24 h after epithelial scraping. A Total proteins were prepared from the corneas and c-Rel expression was determined by Western blot. β-actin was used as internal control. The blot was cropped to improve the clarity of the image and full-length blot was provided in the supplementary materials. B Total RNA was isolated from the cornea and mRNA expression of all five NF-κB family members was determined by quantitative RT-PCR. C Corneal tissue extracts were prepared and the production of inflammatory cytokines as indicated in the figure was determined by ELISA. Epi on: epithelium not scraped. Epi off: epithelium scraped off. Results shown are representative of two independent experiments. **P<0.01, ***P<0.001

Fig 3: Impact of DSG concentration on RelA and c-Rel ChIP-seq results. BMDMs stimulated with lipid A for 1 hr were crosslinked with 0, 1, 2, or 4 mM DSG and 1% formaldehyde and examined by ChIP-seq. The results shown are from a single replicate of each DSG concentration. The findings are consistent with those of other experiments performed with variable crosslinker concentrations (data not shown). (A) The total number of peaks (p-adj <0.01; peak score > 19) for each condition is plotted. For each DSG concentration, peaks are further grouped into two categories: peaks called only at the specified DSG concentration (darker color, new peaks) or peaks called with both the specified concentration and any lower concentration of DSG (lighter color, previous peaks). Analyses for RelA and c-Rel peaks are colored in orange and blue, respectively. (B) All peaks are ranked based on peak score and separated into bins with 1,000 peaks per bin. The percentages of new peaks (darker color) and previous peaks (lighter color) are plotted. (C) The scatter plots show the peak scores for each called peak at each concentration of DSG. The plots show all peaks that are bound in either condition with a peak score > 0. (D) The Integrative Genomics Viewer (IGV) tracks spanning the Il12b locus are shown for c-Rel and RelA ChIP-seq with 0, 1, 2, or 4 mM DSG. A representative peak that is not called at the 0 mM concentration for c-Rel and RelA but is called at higher concentrations is highlighted in grey.

Fig 4: WDR6/NF‐κB feedback loop upregulates TNFα expression by impairing autophagy‐dependent degradation of p65 in HCC AThe effects of NF‐κB inhibitor IMD0354 or IκBα‐SR on TNFα expression in MHCC‐97L cells with or without WDR6 overexpression.BThe effects of WDR6 knockdown on the protein levels of p65, RelB, c‐Rel, p52, and p50 in HCC cells.CiCo‐immunoprecipitation identified p65 as an interactor of LC3B in MHCC‐97L cells.CiiSchematic representation of the location of evolutionarily conserved LIR motifs in the sequence of p65 protein.Di, DiiThe effects of autophagic deficiency using 3‐MA on the levels of p65 protein and TNFα secretion in HCC cells with WDR6 knockdown.Ei WDR6 mRNA levels in Hep3B and MHCC‐97L cells treated with or without TNFα in culture media.EiiSchematic diagram showing three candidate NF‐κB‐binding sites in WDR6 promoter.EiiiThe impact of IMD0354 on the luciferase reporter of WDR6 proximal promoter in HCC cells.Eiv, EvChIP assays using HCC cells or HCC tissues from patients (P#36 and P#54). Data information: All results are representative data generated from three independent experiments. Data are presented as mean ± SD. Unpaired two‐tailed Student's t‐test (A, Dii, Ei, Eiii) was used for statistical analysis. Source data are available online for this figure.

Fig 5: Impact of formaldehyde concentration on RelA and c-Rel ChIP-seq results. BMDMs stimulated with lipid A for 1 hr were crosslinked with 0, 0.5, 1.0, or 2.0% formaldehyde and 1 mM DSG and examined by ChIP-seq. The results shown are from a single replicate of each formaldehyde concentration. The findings are consistent with those of other experiments performed with variable crosslinker concentrations (data not shown). (A) The total number of peaks (p-adj <0.01; peak score > 19) for each condition is plotted. For each formaldehyde concentration, peaks are further grouped into two categories: peaks called only at the specified formaldehyde concentration (darker color, new peaks) or peaks called with both the specified concentration and any lower concentration of formaldehyde (lighter color, previous peaks). Analyses for RelA and c-Rel peaks are colored in orange and blue, respectively. (B) All peaks are ranked based on peak score and separated into bins with 1,000 peaks per bin. The percentages of new peaks (darker color) and previous peaks (lighter color) are plotted. (C) The scatter plots show the peak scores for each called peak at each concentration of formaldehyde. The plots show all peaks that are bound in either condition with a peak score > 0. (D) The IGV tracks spanning the Ccl5 locus are shown for c-Rel and RelA ChIP-seq with each formaldehyde concentration. A representative peak that is not called at the 0% formaldehyde concentration for c-Rel and RelA but is called at higher concentrations is highlighted in grey.