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. 2020 Oct 13;8(2):e001365. doi: 10.1136/jitc-2020-001365

Figure 4.

Figure 4

p50 binds to a κB sequence element in the Gzmb promoter to repress Gzmb expression in T cells. (A) The structure of the mouse Gzmb promoter region showing the putative NF-κB-binding sequences (P1–8) and locations. (B) Nuclear extracts were prepared from EL4 T cells and analyzed by EMSA using putative Gzmb promoter DNA probes 7 and 8 as shown in (A). The NF-κB consensus sequence probe (control probe) was used as positive control. The black arrow indicates the NF-κB–DNA complex and the gray arrow indicates p50 mAb-induced supershift. Shown are p50 binding to probe 8. Probe 7 is shown here as a negative control. (C) The left panel shows the mouse Gzmb promoter structure. The putative NF-κB-binding sequence (P8) and the ChIP PCR primer sequence locations are shown. The numbers under the bar and above the P8 probe sequence indicate the nucleotide locations relative to Gzmb transcription start site. EL4 T cells were analyzed by ChIP using IgG and p50-specific antibody, respectively. The immunoprecipitated DNA were quantified by qPCR using primers that amplify the Gzmb promoter DNA as shown at the left panel.