Fig 4. Promoter assay experiments of PD-L1.

A, Diagrammatic representation of the PD-L1 regulatory elements, including PD-L1 promoter fragment and the predicted enhancer containing an AP-1 binding site. Fragments cloned into luciferase constructs are annotated below, with positions relative to the PD-L1 TSS. B, Diagram of luciferase vectors. The empty pGL4.17 vector, the GL4.17 vector with the promoter cloned upstream of the luciferase gene alone (PDL-P), the promoter with the enhancer cloned downstream of the luciferase gene (PDL-P+E). C, PD-L1 promoter- and enhancer-driven luciferase activity in NCI-H1373 with or without MEK inhibition. Firefly luciferase activity normalized by Renilla luciferase activity significantly increased with PDL-P+E compared with that with PDL-P after DMSO treatment (left), while no significant difference was found between PDL-P and PDL-P+E after U0126 treatment (right). Data are representative of three independent experiments with the same results and shown as the mean of quadruplicate experiments. Error bars indicate SD. E, Binding of cJUN to the candidate PD-L1 enhancer region following ChIP-coupled qPCR. Data are representative of three independent experiments with similar results.