American Society of Hematology

Figure 3.

$IL-1R pathway mediates NF-κB activation in ALK− ALCL with higher IL-1α. (A) The workflow of using RNA-seq analysis to profile gene expression changes upon sgRNAs depletion of IL-1α in ALCL lines (upper). IL-1α–upregulated genes grouped according to gene expression signatures, in both MAC1 and MAC2A cell lines (lower). (B) Gene set enrichment analysis (GSEA) of gene expression signatures that were enriched among IL-1α upregulated genes in both MAC1 (upper) and MAC2A (lower) lines. Only the overlapped and shared signatures among all sgRNA transduced samples in both cell lines were shown. (C) NF-κB-driven luciferase reporter–engineered MAC1 (upper) or Karpas299 (lower) lines were transduced with indicated sgRNAs. Relative NF-κB reporter activities were measured after 4 days of induction. P was calculated comparing sgCTRL and the indicated sgRNA groups; ∗P < .05; ∗∗P < .01. (D) Indicated ALCL lines were transduced with MYD88, IL-1α, or Ctrl sgRNAs, selected, and expression induced. Lysates were analyzed by immunoblotting for the indicated proteins. (E) Indicated ALCL lines were transduced with MYD88 or Ctrl shRNAs, selected, and expression induced. Lysates were analyzed by immunoblotting for the indicated proteins. (F) MAC1 line was stable engineered with IKKβ WT, IKKβ S176/180E, or empty control, then transduced with MYD88 or Ctrl shRNAs along with GFP. The fraction of viable, GFP+/shRNA+ cells relative to the live cell fraction is plotted at the indicated times after shRNA induction, normalized to day 0 values. Error bars denote SD of triplicates. P was calculated by comparing the IKKβ S176/180E and IKKβ WT engineered lines with MYD88 shRNA transduction. ∗∗P < .01.$

IL-1R pathway mediates NF-κB activation in ALK⁻ ALCL with higher IL-1α. (A) The workflow of using RNA-seq analysis to profile gene expression changes upon sgRNAs depletion of IL-1α in ALCL lines (upper). IL-1α–upregulated genes grouped according to gene expression signatures, in both MAC1 and MAC2A cell lines (lower). (B) Gene set enrichment analysis (GSEA) of gene expression signatures that were enriched among IL-1α upregulated genes in both MAC1 (upper) and MAC2A (lower) lines. Only the overlapped and shared signatures among all sgRNA transduced samples in both cell lines were shown. (C) NF-κB-driven luciferase reporter–engineered MAC1 (upper) or Karpas299 (lower) lines were transduced with indicated sgRNAs. Relative NF-κB reporter activities were measured after 4 days of induction. P was calculated comparing sgCTRL and the indicated sgRNA groups; ∗P < .05; ∗∗P < .01. (D) Indicated ALCL lines were transduced with MYD88, IL-1α, or Ctrl sgRNAs, selected, and expression induced. Lysates were analyzed by immunoblotting for the indicated proteins. (E) Indicated ALCL lines were transduced with MYD88 or Ctrl shRNAs, selected, and expression induced. Lysates were analyzed by immunoblotting for the indicated proteins. (F) MAC1 line was stable engineered with IKKβ WT, IKKβ S176/180E, or empty control, then transduced with MYD88 or Ctrl shRNAs along with GFP. The fraction of viable, GFP+/shRNA+ cells relative to the live cell fraction is plotted at the indicated times after shRNA induction, normalized to day 0 values. Error bars denote SD of triplicates. P was calculated by comparing the IKKβ S176/180E and IKKβ WT engineered lines with MYD88 shRNA transduction. ∗∗P < .01.

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