Abstract
BackgroundWe have previously reported significant change of epithelial to mesenchymal transition (EMT) phenotype of Eca-109 cells upon PD-L1 operation, and the cytoplasmic domain of PD-L1 played an essential role in promoting EMT of esophageal cancer cells. However, the underlying mechanism of how PD-L1 regulated EMT in esophageal cancer remained unclear.MethodsThe overexpression and knockdown expression models of PD-L1 and IFIT2 were established by using lenti-virus transfection and RNAi method. Western blotting, qRT-PCR, CCK8 assay, transwell assay and wound healing assay were chosen to investigate their impact on the cells. The expression levels of IFIT2 and EMT markers in esophageal cancer tissues were examined by immunohistochemical staining. The rescue experiments were further applied to investigate the role of STAT1/IFIT2 signal pathway in the PD-L1-mediated EMT. Luciferase reporter assays were performed to examine the IFIT2 promoter activities upon knockdown expression of PD-L1 to identify the putative targeted region of IFIT2 promoter.ResultsThe STAT1/IFIT2 signal pathway was activated when PD-L1 was knockdown in human esophageal cancer cells. Decreased IFIT2 expression significantly increased the cellular abilities of viability, invasion and migration by using RNAi method in human esophageal cancer cells. Decreased IFIT2 expression in esophageal cancer tissues significantly correlated with EMT status, and could be used as an independent prognostic predictor for the patients. Rescue experiments in PD-L1 knockdown cells further confirmed that STAT1/IFIT2 pathway was involved in the PD-L1 mediated EMT of esophageal cancer cells. Moreover, the luciferase reporter assay also confirmed that in esophageal cancer cells, the promoter region of IFIT2 (-3K~-1K) remains more active in PD-L1 knockdown expression cells compared with controls.ConclusionOur present work reveals a novel mechanism of how PD-L1 regulates EMT of cancer cells, namely STAT1/IFIT2 signal pathway is required in PD-L1 mediated EMT in human esophageal cancer.
Highlights
We have previously reported significant change of epithelial to mesenchymal transition (EMT) phenotype of Eca-109 cells upon PD-L1 operation, and the cytoplasmic domain of PD-L1 played an essential role in promoting EMT of esophageal cancer cells
Azuma et al have first established the theory that PD-L1 can serve as a bidirectional regulator, the extra-cellular domain of PD-L1 can interact with its receptor PD-1 on T cells, resulting in dampening T-cell mediated anti-tumor response, and the cytoplasmic domain of PD-L1 can trigger the cellular signaling pathways involved in the anti-apoptosis via ligation with PD-1 fusion protein [4]
We have previously reported that PD-L1 can be used as an important prognostic predictor in human esophageal cancer, and confirmed that PD-L1 can potentially contribute to the EMT of esophageal cancer cells by successfully establishing cellular models including PD-L1 knockdown expression and PD-L1 over-expression in Eca-109 cell line [6]
Summary
We have previously reported significant change of epithelial to mesenchymal transition (EMT) phenotype of Eca-109 cells upon PD-L1 operation, and the cytoplasmic domain of PD-L1 played an essential role in promoting EMT of esophageal cancer cells. Results The STAT1/IFIT2 signal pathway was activated when PD-L1 was knockdown in human esophageal cancer cells. Rescue experiments in PD-L1 knockdown cells further confirmed that STAT1/IFIT2 pathway was involved in the PD-L1 mediated EMT of esophageal cancer cells. The luciferase reporter assay confirmed that in esophageal cancer cells, the promoter region of IFIT2 (-3K~-1K) remains more active in PD-L1 knockdown expression cells compared with controls. Conclusion Our present work reveals a novel mechanism of how PD-L1 regulates EMT of cancer cells, namely STAT1/ IFIT2 signal pathway is required in PD-L1 mediated EMT in human esophageal cancer. Xu et al have demonstrated that, PD-L1 can induce EMT and enhance stemness through up-regulation of SREBP-1c in human renal cell carcinoma cells [8]
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