Fig. 2: The non-structural protein 1 (NS1) of ZIKV is unique and necessary for inducing TNT formation in multiple cell types.

A TNT induction by individual mCherry-tagged ZIKV proteins in A549 cells as indicated by the % number of cells with TNTs (n = 3). B–O TNT formation by expression of mCherry-tagged NS1 proteins of orthoflaviviruses Zika virus (ZIKV) MR-766 and PRVABC-59, dengue virus (DENV) 1-4, deer tick virus (DTV), West Nile virus (WNV), and Japanese encephalitis virus (JEV). B Number of TNTs per cell in A549 and C the corresponding normalized fluorescence (NF) intensity of NS1 expression, n = 5. D Number of TNTs per cell in HTR-8 and E the corresponding NF intensity of NS1 expression, n = 5. Representative confocal images of independent experiments showing control F untransfected HTR-8 cells and G, H cells transfected with NS1 proteins from ZIKV MR-766 and PRVABC-59, n = 3. I–O Representative image of independent experiments (n = 5) showing TNT induction of NS1 orthoflaviviruses. Merged figures show nuclei (blue), actin (gray), and orthoflavivirus NS1 proteins (red). P Number of TNTs per ZIKV-MR766 NS1-expressing cells (n = 10) and Q the average length of individual TNTs (A549, n = 50; U-87 MG, n = 57; SH-SY5Y, n = 36; HTR-8, n = 50; JEG-3, n = 32, BeWo, n = 20; Vero E6, n = 3). Images were acquired using Nikon A1R (60X oil objectives lens at 1.4 normal aperture (NA) at 48 hours post transfection. The confocal images shown are representative of experiments independently repeated with similar results. Data is presented as mean ± SEM. Statistical significance was determined by ANOVA followed by Dunnett’s (P) or Tukey’s (Q) multiple comparison tests, *= P ≤ 0.05, **= P ≤ 0.01, and ****= P ≤ 0.0001. Bar = 25 μm. Source data are provided as a Source Data file.