Figure 5.

Expression of IBV nsp6 leads to formation of omegasomes that are sensitive to wortmannin. (A) IBVnsp6 tagged with mCherry was expressed in HEK cells, which were immunostained for Atg5 to identify Atg5 puncta indicative of autophagosomes. (B) IBVnsp6 tagged with V5 was expressed in Vero cells were incubated with MitoTracker (to stain mitochondria) before being fixed and immunostained for V5 and Atg5. The merge of all three is shown, and the individual stains of the mitochondria (blue) and Atg5 (green). The bottom right image shows a high resolution merge of the mitochondrial stain and the Atg5 stain. (C) (i) IBVnsp6 tagged with mCherry was expressed in HEK cells constitutively expressing DFCP1-GFP, omegasomes were identified as punctate DFCP1-GFP signals. (ii) The cells expressing IBVnsp6 mCherry and constitutively expressing DFCP1-GFP were analyzed by live cell microscopy. DFCP1-GFP puncta were monitored using time lapse at the times indicated, wortmannin (+wortm) was added at 4 min (4′).