Activation of the Mitochondrial Apoptotic Signaling Platform during Rubella Virus Infection

Claus, Claudia; Manssen, Lena; Hübner, Denise; Roßmark, Sarah; Bothe, Viktoria; Petzold, Alice; Große, Claudia; Reins, Mareen; Mankertz, Annette; Frey, Teryl K.; Liebert, Uwe G.

Immunofluorescence analysis of RV-and mock-infected cells was applied to validate CytC localization during RV infection. MitoTracker Red CMXRos (Thermo Fisher Scientific, Braunschweig, Gemany) was used as a mitochondrial counterstain. Figure 4A highlights the strict mitochondrial localization of CytC in mock-infected Vero cells. (A) Immunofluorescence analysis of RV-and mock-infected Vero cells at 3 dpi with anti-CytC antibody (shown in green). As a counterstain, mitochondria were labelled with MitoTracker Red CMXRos (shown in red) before fixation. Nuclear DNA is shown in blue, yellow colour indicates overlap areas. As a positive control, overnight incubation with 0.003% H2O2 was applied; (B) Selective permeabilization of the plasma membrane with digitonin was tested for alpha-tubulin (shown in red) as a representative cytoplasmic protein; (C) Selective permeabilization with digitonin at 25 μg/mL was applied to mock-and RV-infected Vero cells. Immunofluorescence was performed with anti-CytC antibody (shown in red). Release of CytC was induced through overnight incubation with 0.003% H2O2. (B), (C) As a control, immunofluorescence analysis of mock-infected Vero cells was performed after methanol (Met-OH) permeabilization. Asterisks highlight representative cells with localization of CytC outside of mitochondria. Scale bar, 10 μm.