Figure 8: Inhibition of autophagy with specific siRNA targeting endogenous ATG5 enhances TGEV replication.

(a) ST cells were transfected with siRNA duplexes against porcine endogenous ATG5 or nontargeting control siRNA for 24 h. Knockdown efficiency of specific siRNA was indicated by relative ATG5 mRNA to β-actin mRNA when siRNA duplexes at the concentration of 100 nM. (b) Detergent lysate from equivalent numbers of siRNA transfected-ST cells were subjected to immunoblotting with antibodies as indicated. (c) ST cells were transfected with either ATG5-specific siRNA duplexes 1# or control siRNA. At 24 h after transfection, cells were infected with TGEV at MOI of 1. Cells were fixed at 24 hpi and examined the TGEV infection by IFA as described in Fig. 6b. The percentage of TGEV-infected cells per view from three independent experiments is expressed as mean ± SD. (d) The extracellular virus yields were determined at 24 hpi and expressed as TCID₅₀ per 0.1 ml. Representative data from three separate experiments are shown as the mean ± SD of three separate experiments. *p < 0.05. The p value is calculated using Student’s t-test. Gels were run under the same experimental conditions. For better clarity and conciseness of the presentation, cropped blots are shown. The raw uncropped images can be found in the Supplementary Fig. S6.