Journal: Nature Communications

Article Title: A single mutation in nonstructural protein 1 is critical for the adaptive evolution of influenza B virus

doi: 10.1038/s41467-026-70211-x

Figure Lengend Snippet: a TOM20 and TIM23 expression in A549 cells infected with BLee-1940, BYam-1988, or BYam-2018 (MOI = 0.1) was detected by immunoblotting at 24 h post-infection (left). Band intensities were determined by ImageJ (right). b TOM20 and TIM23 expression in HEK293T cells transfected with Flag-BLee-1940-NS1, Flag-BYam-1988-NS1, Flag-BYam-2018-NS1, or Flag-Vector was detected by immunoblotting at 24 h post-transfection (left). Band intensities were determined by ImageJ (right). c Autophagosomes (yellow arrows) and damaged mitochondria (red arrows) were visualized by transmission electron microscopy in IBV-infected A549 cells (MOI = 0.1) at 24 h post-infection. d Immunofluorescence assay (IFA) showing the colocalization of NS1 and mitochondria in A549 cells transfected with mCherry-Mito and infected with BYam-1988 or BYam-2018 (MOI = 0.1) for 24 h, fixed with 4% paraformaldehyde, and stained with anti-NS1 (green). Nuclei were stained with DAPI in blue. Scale bar: 3 µm. e IFA showing the colocalization of NS1 and mitochondria in A549 cells transfected with Flag-BYam-1988-NS1 or Flag-BYam-2018-NS1 and mCherry-Mito for 24 h, fixed with 4% paraformaldehyde, and stained with anti-NS1 (red). Nuclei were stained with DAPI in blue. Scale bar: 3 µm. f The localization of IBV NS1 was detected by immunoelectron microscopy in Flag-BYam-2018-NS1-transfected HEK293T cells for 24 h, where gold particles against Flag (10 nm, red arrows) were observed in the mitochondria. Scale bars: 60 nm. g The colocalization of IBV NS1 and LC3 was detected by immunoelectron microscopy with 15 nm gold particles against LC3B (red arrows) and 5 nm gold particles against Flag (blue arrows) in HEK293T cells transfected with Flag-BYam-2018-NS1 and Myc-LC3. Scale bars: 150 nm. Data are shown as mean ± SD. One-way ANOVA followed by Tukey’s test. n = 3 biological replicates ( a , b ). Data are representative of 3 biological replicates ( c –g). Source data are provided as a Source Data file.

Article Snippet: Plasmid transfection was performed using a transfection reagent (Cat#40802, YEASEN, China) according to the manufacturer’s instructions.

Techniques: Expressing, Infection, Western Blot, Transfection, Plasmid Preparation, Transmission Assay, Electron Microscopy, Immunofluorescence, Staining, Immuno-Electron Microscopy