[Regulating effect of ORF128 on NF-κB signaling pathway of HEK293T cells and its mechanism].

Objective To elucidate the regulating effect of orf virus (ORFV) encoded ORF128 on NF-κB signaling pathway during the infection of HEK293T cells with ORFV and the underlying mechanism. Methods The ORF128 DNA sequences from ORFV/QH02/2010 strain were constructed into eukaryotic expression vectors pCMV-tag2B and pEGFP-N1. During viral infection of cells, the level of ORF128 mRNA was detected by reverse transcription PCR and the subcellular localization of ORF128 protein by laser confocal microscopy. A dual luciferase reporter assay system was used to analyze the regulating effect of ORF128 on NF-κB signaling pathway, and Western blot analysis to detect the nuclear translocation of NF-κBp65 and the phosphorylation of IκBα protein (p-IκBα). Results ORF128 protein was expressed at the early stage and localized in the cell nuclear during ORFV infection, and it inhibited the expression of NF-κB reporter luciferase activity. The expression of the protein blocked the nuclear translocation of NF-κBp65 and the degradation of p-IκBα. Conclusion ORF128 inhibits the activation of NF-κB signaling pathway by blocking the nuclear translocation of NF-κBp65 and the degradation of p-IκBα.