FIG 1.
A recent N1 TMD impairs the replication of an old H1N1 virus at 37°C. (A) The three domains of an NA tetramer are shown with the 7-face TMD model for its amphipathic assembly via the polar faces a and e (21). (B) The timeline depicts the different human IAV lineages by their hemagglutinin (H) and neuraminidase (N) subtypes with the predicted hydrophobicity values (ΔGapp values for membrane insertion) of the consensus N1 TMD sequences for the H1N1 lineages. (C) Split-transfection method for comparative viral production by reverse genetics. Trypsinized 293T cells are transfected with WSN plasmids encoding its old N1 TMD or the recent pN1 TMD, divided into flasks containing MDCK cells at 33°C and 37°C, and viral titers are assayed by TCID50 values. (D) The recent pN1 TMD significantly decreases the replication of the 1933 H1N1 virus WSN at 37°C compared to at 33°C. The viral titers are displayed as means from three independent experiments ± standard errors of the means (SEM). (E) WSN and WSNpN1-TMD viral particles created by reverse genetics were isolated from the culture medium 3 days after recovery by ultracentrifugation. The M1 and NP contents were assayed by immunoblotting and the NA content by activity, which is displayed as a ratio to the WSN sedimented particles from 33°C. Cells that did not receive the NA plasmid were used as a control.