UBE2S Activates NF-κB Signaling By Binding With IκBα and Promotes Metastasis of Lung Adenocarcinoma Cell

BackgroundNuclear factor (NF)-κB signaling in cancer cells was reported to be involved in tumorigenesis. Phosphorylation and degradation of inhibitor of NF-κBα (IκBα) is a canonical pathway of NF-κB signaling. Herein, we report non-canonical activation of NF-κB signaling without phosphorylation of IκB but by directly binding by ubiquitin-conjugating enzyme E2S (UBE2S) for degradation in adenocarcinoma cells.MethodsTCGA and the Human Atlas Protein Database were used to analyze the survival rate and expression of UBE2S. PC9, H460, H441 and A549 cells were used in this study. PC9 and H460 cells were used for further analysis because of different protein levels of UBE2S. Specific IKK inhibitors, PS1145 and SC514, were used to analyze the phosphorylation of IκBα. Western blotting experiment was used to analyze the protein levels PC9 and H460 cells. Wound-healing experiment was used to analyze the migrative ability of PC9 and H460 cells. Overexpression and knockdown of UBE2S in H460 and PC9 cells were used to analyze the downstream proteins levels. Immunoprecipitation, immunofluorescent staining, a glutathione S transferase (GST) pull-down assay, and an in vitro binding assay were used to analyze the interaction of UBE2S and IκBα. Luciferase assay was used to analyze the activation of NF-κB signaling regulated by UBE2S. Zebrafish xenograft model was used analyzed the metastasis of PC9 cells regulated by UBE2S.Results UBE2S in lung adenocarcinoma patients was negatively related to the survival rate. The protein levels of UBE2S and IκBα were shown opposite change in PC9 and H460 cells. PC9 cells showed higher UBE2S expression and migrative ability than H460 cells. Phosphorylation of IκBα was not changed by treatment with IKK specific inhibitors, PS1145 and SC514, in PC9 and H460 cells. Overexpression and knockdown of UBE2S in H460 and PC9 cells showed the protein levels of IκBα were regulated. Immunoprecipitation, immunofluorescent staining, a glutathione S transferase (GST) pull-down assay, and an in vitro binding assay showed the direct binding of UBE2S with IκBα. Protein levels of nuclear p65 and luciferase assay showed the NF-κB signaling was regulated UBE2S. EMT markers and migrative ability of cancer cells were also regulated by UBE2S. Zebrafish xenograft tumor model showed the reduction of migrative PC9 cells by knockdown of UBE2S.ConclusionHigher UBE2S expressed in lung adenocarcinomas could bind with IκBα for activation of NF-κB signaling to promote metastasis of cancer cells. UBE2S might be a potential therapeutic target for lung adenocarcinomas.