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. 2022 Nov 29;10(12):2042. doi: 10.3390/vaccines10122042

Figure 2.

Figure 2

The N protein interferes with the RIG-I-like receptor signaling pathway in HEK293T cells. (A) The pcDNA3.1 vector and pcDNA3.1-E, M, N, RDRP, ORF3, ORF6, ORF-7, ORF-8, and ORF-10 were transfected into cells. Twenty-four hours after transfection, the transfected cells were stimulated with poly (I:C). Total RNA was extracted from 0- and 6-h cell samples for real-time PCR. Data represent the relative expression of IFN--β. (paired t test, n = 3, biological replicates per group, ns > 0.05, * 0.01 < p < 0.05, ** 0.001 < p < 0.01, *** p < 0.001). (B) The pcDNA3.1 vector and pcDNA3.1-N were transfected into cells. Twenty-four hours after transfection, the transfected cells were stimulated with poly (I:C). Total RNA was extracted from 0- and 6-h cell samples for real-time PCR. Data represent relative expression of IFN-β. Data are presented as means ± SD (paired t test, n = 3 biological replicates per group, ns > 0.05, * 0.01 < p < 0.05, ** 0.001 < p < 0.01, *** p < 0.001). (C) The pcDNA3.1 vector and pcDNA3.1-N-Flag were transfected into cells. Twenty-four hours after transfection, the transfected cells were stimulated with poly (I:C). Phosphorylation was detected by western blot analysis of IRF3, NF-κB and TBK1 proteins from 0- and 6-h cell samples. Grayscale results were analyzed using a Gel-Pro analyzer. Data are presented as means ± SD (paired t test, n = 3 biological replicates per group, ns > 0.05, * 0.01 < p < 0.05, ** 0.001 < p < 0.01, *** p < 0.001).