Figure 4.

DENV‐2 entry into Vero cells is independent of caveolae/lipid‐rafts but is dependent on dynamin.
A. DENV‐1 or DENV‐2 suspensions were incubated at 37°C with various concentrations of methyl‐β‐cyclodextrin. After 1 h remaining infectivity was determined.
B. Vero cells were pretreated with nystatin or methyl‐β‐cyclodextrin. Then monolayers were washed with PBS and infected with DENV‐1 or DENV‐2 in culture medium without serum. After 1 h internalization, cultures were treated with proteinase K and the cell pellets were plated onto Vero cells to determine internalized virus by an infectious centre assay. Results are expressed as percentage of internalized virus with respect to a control without drug treatment.
C. Cells were untreated (control) or treated with 100 μM nystatin or 5 mM methyl‐β‐cyclodextrin. Then cultures were incubated with FITC‐labelled cholera toxin.
D. Cells transiently transfected with the constructs GFP‐cav‐1 wt, GFP cav‐1 DN or GFP‐cav‐1 Y14F were infected with DENV‐2. After 24 h infection cultures were fixed and immunofluorescence staining was performed.
E. For quantification of samples shown in (D), 250 transfected cells with similar levels of GFP expression were screened and cells positive for viral antigen were scored.
F. Vero cells were treated with dynasore, infected with DENV‐1 or DENV‐2 in the presence of the drug and then processed as in (B).
G. Cells transiently transfected with the constructs GFP‐dyn II wt or GFP‐dyn II K44A were infected with DENV‐2. After 24 h infection cultures were fixed and immunofluorescence staining was performed.
H. For quantification of samples shown in (G), 250 transfected cells with similar levels of GFP expression were screened and cells positive for viral antigen were scored.
In (A), (B), (E), (F) and (H) values represent the mean ± SD of two independent experiments.