Fig. 4: ZIKV NS1 colocalization with cytoskeletal components and regulation of TNT-related genes. | Nature Communications

Fig. 4: ZIKV NS1 colocalization with cytoskeletal components and regulation of TNT-related genes.

From: Zika virus NS1 drives tunneling nanotube formation for mitochondrial transfer and stealth transmission in trophoblasts

Fig. 4

A A549 cells expressing pGFP-actin show typical cytoskeletal actin distribution, including stress fibers. B Expression of pNS1-ZIKV and pGFP-actin show remodeling of the actin cytoskeleton and the formation of TNT-like structures connecting neighboring cells. C Cells expressing pGFP-actin and pNS1ΔTNT do not induce actin remodeling and TNT formation. D Cells co-expressing pGFP-actin and pNS1-ZIKV treated with cytochalasin-D (100 nM, 30 mins) lack TNT formation. Merged figures show actin (green), pNS1 (red) and nuclei (blue). E pNS1-ZIKV-induced TNTs in HTR-8 cells associates with the cell membrane and is composed by both tubulin and F-actin, while pNS1ΔTNT is unable to form TNTs 24 hours post transfection (hpt). Merged figures show actin (gray), tubulin (green), and ZIKV-NS1 (red). F HTR-8 gene expression levels, at 24hpt, of DNML1 [untransfected=5, pNS1-ZIKV (n = 6), pNS1ΔTNT (n = 6)], MYO10 [untransfected=5, pNS1-ZIKV (n = 5), pNS1ΔTNT (n = 6)], MYO5A [untransfected=5, pNS1-ZIKV (n = 6), pNS1ΔTNT (n = 5)], RAC1 [untransfected=4, pNS1-ZIKV (n = 4), pNS1ΔTNT (n = 6)], CDC42 [untransfected=5, pNS1-ZIKV (n = 6), pNS1ΔTNT (n = 6)], and TNFAIP2 [untransfected=5, pNS1-ZIKV (n = 6), pNS1ΔTNT (n = 6)]. Statistically significant outliers identified by Grubb’s tests were excluded in MYO5A (untransfected, n = 1; pNS1ΔTNT, n = 1) and RAC1 (untransfected, n = 1; pNS1-ZIKV, n = 1) comparisons. Values are represented as mean ± SEM or median and interquartile range (MYO5A). Statistical significance was determined by one-way ANOVA (DNML1, MYO10, RAC1, CDC42, and TNFAIP2) and Kruskal-Wallis (MYO5A) followed by Tukey’s and Dunn’s multiple comparisons tests. The confocal images shown are representative of experiments (n = 3) independently repeated with similar results. Images were acquired by confocal microscopy at 60X (A–D) oil objectives lens at 1.4 normal aperture (NA) and at 40X (E) using a Nikon A1R. Images were processed using the NIS Elements software (Nikon). Bar = 10 µm (A–D) and 25 μm (E). **= P ≤ 0.01. Source data are provided as a Source Data file.

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