Figure 1: Caveolin-1/caveolae/caveolar endocytotic vesicle colocalized with TFV virions at the late phase of infection. | Scientific Reports

Figure 1: Caveolin-1/caveolae/caveolar endocytotic vesicle colocalized with TFV virions at the late phase of infection.

From: Caveolae Restrict Tiger Frog Virus Release in HepG2 cells and Caveolae-Associated Proteins Incorporated into Virus Particles

Figure 1

(A) HepG2 cells grown on coverslips were either uninfected (Con.) or infected (Inf.) by TFV at an MOI of 10. The cells were fixed with paraformaldehyde at 72 h postinfection. Caveolin-1 and TFV MCP were stained as described in the Materials and Methods. Bound IgG was detected with host-specific secondary antibodies conjugated to either Alexa Fluor 488 or 555. The yellow overlay represents colocalization of TFV MCP and caveolin-1 (white arrows). (B) HepG2 cells infected with TFV after 72 h postinfection. Cells were extracted with 1% Triton X-100 at 4 °C. The lysate was loaded at the bottom of a flotation sucrose density gradient and subjected to equilibrium centrifugation. The gradient was fractionated from the top, and polypeptides were analyzed by SDS-PAGE and immunoblotting. (C) Extracted DNA from each fractions were amplified TFV gene orf031L and orf096R by PCR, then detected by nucleic acid electrophoresis experiment (+ indicates positive control,- indicates negative control). (D) Extracted DNA from each fractions and then detected TFV gene orf096R by absolute quantitative real-time PCR. Y-axe represent absolute quantitative values of TFV gene orf096R. Asterisks indicate values that are statistically significant (p < 0.05) compared with values for NC. (E) Caveolae were isolated on sucrose gradients at different time points from 1 to 72 h post-infection, and equal volume fractions were separated. The fractions 5 and 12 of each time points were detected with anti-TFV MCP antibody by SDS-PAGE and immunoblotting.

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