Fig 4. Atg11 and Atg17 dissociate from APs in cells depleted for Ypt7 and Vps39, but remain on APs in cells depleted for the Vps21 module components.

Endogenous Atg11 (A), or Atg17 (B) was tagged with GFP at their C-terminus in indicated strains; from left-to-right: WT, vps21Δ, vps21Δ ypt7Δ, ypt7Δ, vps9Δ, vps9Δ ypt7Δ, vps8Δ, pep12Δ, and vps39Δ. The cells, which express mCherry-Atg8 (from a plasmid) as an AP marker, were grown in YPD and shifted to SD-N (as in Fig 1). The co-localization of the GFP-tagged AtgX with Atg8 was determined using live-cell fluorescence microscopy. Shown from top to bottom: DIC, GFP, mCherry, merge, insert, and the number of red puncta used for the quantification. Arrows indicate co-localizing puncta, arrowheads (in ypt7Δ and vps39Δ mutant cells) point to mCherry-Atg8 puncta that do not co-localize with the other Atg; bar, 2 μm. C. Quantification of percent AtgX-Atg8 co-localization from panels A-B in the indicated strains: Atg11 (left), and Atg17 (right), strain color legend is shown on the right. In WT cells, a single dot per cell that contains both Atgs, represents PAS or AP. Whereas Atg11 and Atg17 are removed from most APs that accumulate in ypt7Δ and vps39Δ mutant cells, they remain on most APs that accumulate in mutant cells defective in the Vps21 module as well as in the double mutant cells. Columns represent mean, error bars represent STD, and P values, **, p <0.01. Results in this figure represent at least three independent experiments.