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. 2021 Jan 7;17(1):e1009033. doi: 10.1371/journal.ppat.1009033

Fig 3. ZIKV NS2A binds to p53.

Fig 3

(A, B) U2OS cells were transfected with ZIKV NS2A-Flag. At 18h p.t., the cells were left untreated (A) or stimulated with 10 μM etoposide for 1.5 hrs (B), fixed with methanol and stained with indicated Abs. The images were taken as Z-stack sections and subjected to a digital deconvolution. The scale bar is 50 μm. Arrows point at the cell expressing ZIKV NS2A. (C) Endogenous p53 pull-down. U2OS cells were transfected with pDEST47-ZIKV NS2A-Flag expressing ZIKV NS2A tagged with the C-terminal 3xFlag epitope or left untransfected (UN). At 18 hrs p.t., the cells were stimulated with 10 μM etoposide for 6 hrs. NS2A-Flag was immunoprecipitated with mouse α-Flag Ab. Lysates and imunoprecipitates was tested with mouse mab α-p53 DO7 and mouse α-Flag Abs. (D) ZIKV NS2A pull-down using p53-binding proteins. U2OS cells were transfected with pDEST47-ZIKV NS2A-Flag or left untransfected (UN). At 18 hrs p.t., the cells were stimulated with 10 μM etoposide for 6h. P53 was immunopreciptated with proteins binding to p53 N- or C-terminus. Presence of ZIKV NS2A-Flag and p53 in the lysates and imunoprecipitates was tested with mouse anti-Flag and mouse α-p53 DO7 Abs.