Figure 7.
ER collapse is required for ERphagy.(A) Western blot with samples taken from cells of the indicated genotypes expressing GFP-Scs2 at the indicated times after transfer to SPO and probed for GFP and hexokinase. (B) Average and SD quantifying free GFP as a proportion of total GFP signal 8 h after transfer to SPO, using n = 3 replicates of the experiment from A. P values calculated by Student’s t test. **, P < 0.01; ****, P < 0.0001. (C) Microscopy images of cells expressing Vph1-mCherry and the indicated GFP-tagged ER protein and either an untagged (WT) or an anti-GFP nanobody–tagged allele of Pil1 (PIL1-antiGFP). Images were taken at 0 and 7 h following transfer to SPO. Scale bar = 2 µm. Arrowheads indicate pockets of vacuole. (D) Western blot with samples taken from cells expressing Rtn1-GFP and either WT Pil1 or Pil1-antiGFP. Samples were taken at the indicated times following transfer to SPO and probed for GFP and hexokinase. (E) Schematic outlining the experimental concept in F, in which a lumenal GFP-tagged protein is not accessible to Pil1-antiGFP binding and therefore does not affect tethering, but expression of an ER membrane protein with a cytosolically accessible GFP tag results in elevated tethering. (F) Western blot with samples taken from cells expressing Pil1-antiGFP and the indicated GFP-tagged proteins. Samples harvested at the indicated times following transfer to SPO and probed for GFP and hexokinase. (G) Average and SD quantifying free GFP as a proportion of total GFP signal 8 h after transfer to SPO, using n = 3 replicates of the experiment from F. P values calculated by Student’s t test. *, P < 0.05. (H) Model for ER remodeling during the meiotic program in budding yeast. ER is represented in green, PSM in purple, retained tethers in yellow, and Scs2/22 in pink. Arrows indicate progression through meiosis. Source data are available for this figure: SourceData F7.

ER collapse is required for ERphagy. (A) Western blot with samples taken from cells of the indicated genotypes expressing GFP-Scs2 at the indicated times after transfer to SPO and probed for GFP and hexokinase. (B) Average and SD quantifying free GFP as a proportion of total GFP signal 8 h after transfer to SPO, using n = 3 replicates of the experiment from A. P values calculated by Student’s t test. **, P < 0.01; ****, P < 0.0001. (C) Microscopy images of cells expressing Vph1-mCherry and the indicated GFP-tagged ER protein and either an untagged (WT) or an anti-GFP nanobody–tagged allele of Pil1 (PIL1-antiGFP). Images were taken at 0 and 7 h following transfer to SPO. Scale bar = 2 µm. Arrowheads indicate pockets of vacuole. (D) Western blot with samples taken from cells expressing Rtn1-GFP and either WT Pil1 or Pil1-antiGFP. Samples were taken at the indicated times following transfer to SPO and probed for GFP and hexokinase. (E) Schematic outlining the experimental concept in F, in which a lumenal GFP-tagged protein is not accessible to Pil1-antiGFP binding and therefore does not affect tethering, but expression of an ER membrane protein with a cytosolically accessible GFP tag results in elevated tethering. (F) Western blot with samples taken from cells expressing Pil1-antiGFP and the indicated GFP-tagged proteins. Samples harvested at the indicated times following transfer to SPO and probed for GFP and hexokinase. (G) Average and SD quantifying free GFP as a proportion of total GFP signal 8 h after transfer to SPO, using n = 3 replicates of the experiment from F. P values calculated by Student’s t test. *, P < 0.05. (H) Model for ER remodeling during the meiotic program in budding yeast. ER is represented in green, PSM in purple, retained tethers in yellow, and Scs2/22 in pink. Arrows indicate progression through meiosis. Source data are available for this figure: SourceData F7.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal