Figure 1

CSFV replication is restricted by ISG15 in PAMs. PAMs stably overexpressing ISG15 were constructed by lentiviral transduction. A Expression of ISG15 protein in PAMs and, CMV and CMV-ISG15 cells was determined by Western blot using an anti-Flag and an anti-ISG15 antibody, respectively. β-actin served as an internal control. B Cell viability of cell lines stably overexpressing ISG15. C GFP reporter expression was detected on mock-infected cells (a), CMV-infected cells (b) and l CMV-ISG15-infected cells (c). Scale bars, 100 μm. CSFV genomic RNA and viral titers D were analyzed in CMV and CMV-ISG15 cells at 12 and 24 hpi (MOI = 1) by RT-qPCR and IFA, respectively. E Expression of ISG15 protein was probed by Western blot. F Cell viability of cell lines with stable knockdown of ISG15. G GFP reporter expression was detected in mock-infected PAMs (a), shN-infected cells (b) and shISG15-1-infected cells (c). Scale bars, 100 μm. ShN and shISG15 cells were incubated with CSFV (MOI = 1). G CSFV genomic RNA was quantified in shN and shISG15 cells at 12 and 24 hpi. H The extracellular viral titers were quantified and expressed as TCID₅₀/mL. Data (B, D, 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 < 0.05; **P < 0.01; ***P < 0.001