Fig. 5. ACSL6 promotes NF-κB signaling to drive NF-κB–dependent gene expression.

(A and B) Kyoto Encyclopedia of Genes and Genomes (A) and gene set enrichment analysis (GSEA) analyses (B) of RNA-seq data from shNT and shACSL6 MHCC97H cells. MAPK, mitogen-activated protein kinase; FDR, false discovery rate; NES, normalized enrichment score. (C to E) ACSL6-depleted MHCC97H and Huh7 cells were infected with rACSL6 WT or S674A and then treated with or without IL-18 (20 ng ml⁻¹) for 12 hours. The mRNA (C) and protein expression (D) levels of the indicated genes and concentrations of CXCL1 and CXCL5 (E) in the medium were detected. (F) ACSL6-depleted MHCC97H cells were infected with rACSL6 WT or S674A and then treated with or without IL-18 (20 ng ml⁻¹) for 1 hour. Immunoblotting analyses were performed with indicated antibodies. (G) ChIP-qPCR analyses were performed with the indicated antibodies, and DNA was amplified with primers targeting positive sites in the GADD45B or CXCL1 gene in ACSL6-depleted MHCC97H cells with forced expression of rACSL6 WT and S674A. (H) Transwell assays of the migration ability of ACSL6-depleted MHCC97H or Huh7 cells with forced expression of rACSL6 WT, S674A, or S674A and MMP3. (I) CCK8 analyses of ACSL6-depleted MHCC97H or Huh7 cells with forced expression of rACSL6 WT, S674A, or S674A with GADD45B. *P < 0.05, **P < 0.01, and ***P < 0.001. One-way ANOVA [(C), (E), (G), and (H)] and two-way ANOVA (I).