PLUNC Downregulates ATF3 And PD-L1 Expression By Inhibiting DDX17/β-catenin Interaction In Nasopharyngeal Carcinoma

Objective:To explore how PLUNC regulates ATF3 and PD-L1 by regulating DDX17/β-catenin interaction in nasopharyngeal carcinoma.Methods:5-8F,HNE2 cells were used to construct stably transfected cell lines overexpressing PLUNC.Stable transfection cell lines overexpressing β-catenin or DDX17 were constructed with HNE2 and S18 cells.5-8F,HNE2 cells were transfected with PLUNC interfering plasmid or DDX17 interfering plasmid.5-8F and HONE1 were transfected withβ-catenin interfering plasmids.The expression levels of β-catenin pathway-related proteins(β-catenin,p-GSK3β,TCF4),ATF3 and PD-L1 in nasopharyngeal carcinoma cells were detected by WB after overexpressing or interfering with the expression of PLUNC,DDX17 andβ-catenin.Cell immunofluorescence assay was used to detect the expression and localization of ATF3 and PD-L1 in nasopharyngeal carcinoma cells after overexpression or interference of PLUNC,overexpression or interference of β-catenin in nasopharyngeal carcinoma cells.The interaction between PLUNC,DDX17 and β-catenin was studied by Co-IP and cell immunofluorescence experiments.Results:WB results showed that in 5-8F and HNE2 cells,PLUNC inhibited the expression of β-catenin pathway-related proteins(β-catenin,p-GSK3β),ATF3 and PD-L1.The results of cell immunofluorescence experiments showed that after overexpression of PLUNC,the expression of ATF3 in the nucleus was reduced,and the expression of PD-L1 in the nucleus,cell membrane and cytoplasm was reduced.The expression ofβ-catenin pathway-related proteins(β-catenin,p-GSK3β,TCF4),ATF3 and PD-L1 were up-regulated after transfection of PLUNC-interfering plasmids into 5-8F and HNE2 cells.Co-IP and cell immunofluorescence experiments were performed on 5-8F and HNE2 overexpressing PLUNC stable transfected cell lines.The results showed that PLUNC interacted with DDX17 and co-localized in the cytoplasm and nucleus.WB showed that PLUNC inhibited the expression of DDX17 in 5-8F and HNE2 cells and the expression of ATF3 and PD-L1 was down-regulated after interfering with DDX17.Co-IP and immunofluorescence experiments were performed with HNE2 and S18 stably transfected cell lines overexpressing β-catenin or DDX17.The results showed that DDX17 interacted with β-catenin and co-localized in the nucleus.WB and cell immunofluorescence experiments were performed with HNE2 and S18 cells overexpressing β-catenin.After interfering with the β-catenin expression of 5-8F and HONE 1,WB and cell immunofluorescence experiments were performed.The results showed that β-catenin up-regulated ATF3 and PD-Expression of L1.Conclusion:PLUNC could down-regulate the expression of ATF3 and PD-L1 in nasopharyngeal carcinoma by inhibiting DDX17/β-catenin interaction.