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Mylipb negatively regulates cellular antiviral response mainly dependent of its ubiquitin ligase activity. (A) ISRE reporter activity and EPC-IFN-Luc activity in Myc-empty vector or Myc-mylipb-WT and Myc-mylpib-C381S-transfected EPC cells for 24 h, followed by transfection with or without poly I:C (1μg/mL) for 24 h. Western blotting tests to detect the overexpression of mylipb-WT and mylipb-C381S. (B) ISRE reporter activity and EPC-IFN-Luc activity in Myc-empty vector or Myc-mylipb-WT and Myc-mylipb-C381S-transfected EPC cells with or without SVCV (MOI of 1) infection for 24 h. Western blotting tests to detect the overexpression of mylipb-WT and mylipb-C381S. (C) qRT-PCR of ifn, isg15, and viperin mRNA in EPC cells transfected with Myc-empty or or Myc-mylipb-WT and Myc-mylipb-C381S-transfected EPC cells for 24 h, followed by transfection with or without poly I:C (1μg/mL) for 24 h. Western blotting tests to detect the overexpression of mylipb-WT and mylipb-C381S. (D) qRT-PCR of ifn, isg15, and viperin mRNA in EPC cells transfected with Myc-empty or or Myc-mylipb-WT and Myc-mylipb-C381S-transfected EPC cells for 24 h, followed by infection with or without SVCV (MOI of 1) for 24 h. Western blotting tests to detect the overexpression of mylipb-WT and mylipb-C381S. (E) The plaque assay of EPC cells transfected with Myc-empty or or Myc-mylipb-WT and Myc-mylipb-C381S-transfected EPC cells for 24 h, followed by infection with or without SVCV(MOI:0.1–100) for 48 h. (F) Determination of SVCV titre in culture medium of EPC cells transfected with Myc-empty or or Myc-mylipb-WT and Myc-mylipb-C381S-transfected EPC cells for 24 h, followed by infection with or without SVCV(MOI:1) for 48 h. (G) qRT-PCR of N, P, and G mRNA in EPC cells transfected with Myc-empty or or Myc-mylipb-WT and Myc-mylipb-C381S-transfected EPC cells for 24 h, followed by infection with or without SVCV for 24 h. Western blotting tests to detect the overexpression of mylipb-WT and mylipb-C381S. qRT-PCR data are presented as mean values based on three repeated experiments, and error bars indicate the ± SD. *,P < 0.05, **, P<0.01; ***, P < 0.001; ****, P< 0.0001.
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