Sustained Autophagy Contributes to Measles Virus Infectivity
Figure 3
Syncytia formation induces autophagy in MeV-infected cells.
(A) GFP-LC3-HeLa cells were infected with the attenuated strain of MeV Schwarz wild-type (Sch-MeV) or deficient for MeV-C expression (Sch-MeVΔC). 24 h post infection, the number of GFP+ vesicles within monocucleated or multinucleated cells was assessed by confocal microscopy in infected cells detected by staining for the viral nucleoprotein N (MeV-N). (B) GFP-LC3 HeLa cells were infected or not with Ed-MeV (MOI 3) or treated with rapamycin (Rapa) (125 nM), and treated or not with the FIP peptide (10 µg/mL). 24 h post infection, the number of GFP+ vesicles per cell was assessed by confocal microscopy. (C) HeLa cells were co-transfected with a vector encoding for the H protein of Ed-MeV and one encoding for the F protein. 24 h post transfection H/F-co-transfected HeLa cells (H+F+ HeLa) were co-cultured with GFP-LC3-HeLa cells to induce cell-cell fusion. The number of GFP+ vesicles within monocucleated or multinucleated cells was assessed by confocal microscopy after 18 h of co-culture. (D) GFP-LC3 HeLa cells and HeLa cells were treated with the indicated siRNA for 24 h. HeLa were then co-transfected with MeV-H/F as in (C) and co-cultured with siRNA treated GFP-LC3 HeLa cells. The number of GFP+ vesicles within monocucleated or multinucleated cells was assessed by confocal microscopy. (E) GFP-LC3 HeLa cells were treated with Polyethylene Glycol (PEG) and treated or not with 250 nM rapamycin (Rapa) for 2 h. The number of GFP+ vesicles within monocucleated or multinucleated cells was assessed by confocal microscopy 6 h after PEG treatment. For each experiment, representative profiles are shown and are accompanied by a graph representing the number of GFP+ vesicles per cell ( = GFP+ vesicles per one nucleus). For syncytia, the number of dots was normalized to the number of nuclei. Error bars, mean ± SD of three independent experiments. Student's t test; ***p<0.005; **p<0.01; *p<0.05; #p>0.05.