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Figure 4 | Veterinary Research

Figure 4

From: Antiviral activity of ISG15 against classical swine fever virus replication in porcine alveolar macrophages via inhibition of autophagy by ISGylating BECN1

Figure 4

ISGylation is involved in the antiviral activity of ISG15 against CSFV.A Expression of ISGylated protein was detected by Western blot with an anti-ISG15 antibody. B, C Effects of ISGylation on CSFV replication. PAMs B and shISG15 cells C were transfected with 3.1, 3.1-ISG15AA, and 3.1-ISG15GG plasmid followed by CSFV infection for another 24 h (MOI = 1). CSFV replication was quantified by RT-qPCR and IFA, respectively. D, E PAMs with stable knockdown of USP18 were constructed by lentiviral transduction. D Expression of USP18 protein was detected by Western blot. E Cell viability with stable knockdown of USP18. ShN and shUSP18 cells were pretreated with IFN-α and then infected with CSFV (MOI = 1). F CSFV replication was quantified by RT-qPCR and IFA, respectively. G Expression of ISGylated protein was detected by Western blot with an anti-ISG15 antibody. HMx1 expression was analyzed by RT-qPCR. Data (B, C, E, F, and H) represent the mean ± SD of three independent experiments and were measured in technical duplicate. Comparisons between groups were calculated using Student’s t-test. **P < 0.01; ***P < 0.001

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