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. 2023 Nov 22;97(12):e01183-23. doi: 10.1128/jvi.01183-23

Fig 3.

Fig 3

JINR1 inhibition attenuates JEV replication, neuronal apoptosis, and the expression of genes involved in ER stress and neuroinflammation. (A) JINR1 promotes JEV replication in SH-SY5Y cells. Cells were transfected with either ASO-NS, ASO-JINR1-1, ASO-JINR1-2, empty vector (Con-Vec), or pcDNA3.1 with full-length JINR1 (JINR1-OE), and viral replication was determined by quantifying the intracellular levels of JEV RNA using qRT-PCR at 48 hpi. (B) JINR1 silencing reduces JEV titer. Quantification of viral titer upon JINR1 depletion during JEV infection is shown in Fig. S2C (upper panel). (C) JINR1 overexpression increases JEV titer. Quantification of viral titer upon JINR1 overexpression during JEV infection is shown in Fig. S2C (lower panel). (D) JINR1 knockdown attenuates JEV-mediated reduction in cell proliferation. The proliferation of SH-SY5Y cells was assessed by WST1 assay upon JINR1 depletion at 60 hpi. The graph represents the percentage of viable cells. (E) JINR1 promotes apoptosis in SH-SY5Y cells. SH-SY5Y cells were transfected with either ASO-NS, ASO-JINR1-1, or ASO-JINR1-2, and the caspase-3/7 activity was determined at 60 hpi. (F) JINR1 depletion reduces JEV-induced cleaved PARP protein expression. SH-SY5Y cells were transfected with ASO-NS or ASO-JINR1-1, and protein lysates were collected at 60 hpi. Cleaved PARP protein levels were analyzed by western blotting. A representative blot is shown from three independent experiments with similar results. Blots were reprobed for β-actin to establish equal loading. (G) JINR1 depletion attenuates JEV-induced ER stress genes. qRT-PCR analysis of indicated ER stress genes upon JINR1 depletion in SH-SY5Y cells at 48 hpi. (H) JINR1 depletion reduces JEV-induced neuroinflammatory genes. qRT-PCR analysis of indicated neuroinflammatory genes upon JINR1 depletion in SH-SY5Y cells at 48 hpi. Error bars represent the mean ± SEM from three independent experiments. (A, G, and H) RNA samples were analyzed by qRT-PCR. (A and B) *Significant change compared to JEV-ASO-NS/JEV-Con-Vec. (C) *Significant change compared to JEV-Con-Vec. (D, E, G, and H) *Significant change compared to MI-ASO-NS. #Significant change compared to JEV-ASO-NS. Statistical comparison was made using the Student’s t-test.