Journal: Journal of Virology
Article Title: COP I vesicles facilitate classical swine fever virus proliferation by transporting fatty acid synthase from the Golgi apparatus to the endoplasmic reticulum
doi: 10.1128/jvi.00305-25
Figure Lengend Snippet: COP I is required for CSFV RNA replication. ( A ) NTsiRNA-, siCOPA-3-, and siCOPD-1-transfected cells were infected with 10 MOI of CSFV in FBS-free medium for 1 h at 4°C. Unbound virions were washed away with pre-cooled citrate buffer (pH = 3). Total cells were collected for CSFV RNA copy number measurement by RT-qPCR. ( B ) NTsiRNA-, siCOPA-3-, and siCOPD-1-transfected cells were infected with 10 MOI CSFV (MOI = 10) in FBS-free medium for 1 h at 4°C to allow virion binding. Cells were then washed with pre-cooled citrate buffer (pH = 3) to remove unbound virions and cultured for another 2 h at 37°C. The cells were washed and collected for CSFV RNA copy number determination by RT-qPCR. (C and D) PK-15 cells were infected with 1 MOI of CSFV for 2 h, then the medium was discarded, and the fresh medium containing GCA or Exo1 was added. Samples were collected after 8 h incubation for CSFV RNA copy number determination by RT-qPCR. ( E ) The pEGFP-NS5B-transfected cells were infected with 1 MOI of CSFV for 48 h. Cells were stained for the COP I vesicle marker, COPβ, the ER marker, Sec61, and nucleocapsids. Scale bars: 5 µm.
Article Snippet: PK-15 cells were seeded into 96-well plates and incubated with H89 (Beyotime, S1643), Exo1 (TargetMol, T4609), GCA (Selleck, S7266), BFA (Selleck, S7046), tunicamycin (TargetMol, T13229 ), or control at the indicated concentrations at 37°C in a CO 2 incubator.
Techniques: Transfection, Infection, Quantitative RT-PCR, Binding Assay, Cell Culture, Incubation, Staining, Marker
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