Figure 5.

Clustering of ATG12 and ATG3 with viral DNA. (A) Wild-type MEF cells were untreated, amino acid and serum deprived for 2 h with Hank’s media or infected with vaccinia virus at a MOI of 1 for 8 h. Cells were fixed, immunostained for endogenous ATG12 and co-stained with DAPI. Arrows denote areas of viral activity. (B) Quantification of clustering between viral DNA and ATG12. A minimum of 50 cells was counted and scored for the localization of viral activity with ATG12. (C) Wild-type MEF cells were untreated, amino acid and serum deprived for 2 h with Hank’s media or infected with vaccinia virus at a MOI of 1 for 8 h. Cells were fixed, immunostained for endogenous ATG3 and stained with DAPI. Arrows denote areas of viral activity. (D) Quantification of clustering between viral DNA and ATG3. A minimum of 50 cells was counted and scored for the localization of viral activity with ATG3. Complete clustering was defined as all cellular ATG3/ATG12 localizing with viral DNA (cytoplasmic DAPI staining), and vice versa. Cells with partial clustering pattern were defined as those in which all cellular ATG3/ATG12 is localized with viral DNA, with a fraction of viral DNA not localized with cellular ATG3/ATG12.