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Fig. 5 | Cell Communication and Signaling

Fig. 5

From: Rabies virus phosphoprotein P5 binding to BECN1 regulates self-replication by BECN1-mediated autophagy signaling pathway

Fig. 5

A ring-like structure containing P5 was beneficial to virus replication. a, b N2a cells were transfected with Flag-P5 (a) or cotransfected with Flag-P5 and two siRNAs targeting Becn1 (b) for 12 h, and infected with RABV strain HEP-Flury or CVS-11 at an MOI = 2 for 36 h, and then harvested for western blotting analysis with mouse anti-NP, anti-BECN1, anti-Flag mAbs, and rabbit anti-GAPDH antibodies. c The ratio of viral protein N to GAPDH was normalized to that of the control conditions. d N2a cells were treated as described for panels A and B. qRT-PCR analysis of cellular viral N mRNA, and the anti-genomic RNA level. All qRT-PCR data were normalized to the expression of Gapdh and the control group was used as a reference. e Cellular supernatant in panels (a and b) were harvested for virus titer detection. Virus titers in N2a cells were determined using a TCID50 assay. f N2a cells were transfected with Flag-P5 for 12 h and were pretreated with 5 mM 3-MA or 1 mM wortmannin for 2 h, and then infected with RABV HEP-Flury strain at an MOI = 2 and incubated in the absence or presence of 5 mM 3-MA or 1 mM wortmannin for 24 h, and the cells were harvested for western blotting analysis using the indicated antibodies. g The ratio of viral protein N to GAPDH was normalized to that of the control conditions. Means and SD (error bars) of three independent experiments are indicated (*, P < 0.05; **, P < 0.01; ***, P < 0.001)

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