TY - DATA T1 - GFP-Snc1-PEM accumulates in APs of vps21∆ mutant cells and macro-ER-phagy is independent of UPR induction. PY - 2015/07/16 AU - Zhanna Lipatova AU - Nava Segev UR - https://plos.figshare.com/articles/figure/_GFP_Snc1_PEM_accumulates_in_APs_of_vps21_8710_mutant_cells_and_macro_ER_phagy_is_independent_of_UPR_induction_/1485807 DO - 10.1371/journal.pgen.1005390.g004 L4 - https://ndownloader.figshare.com/files/2179014 KW - erad KW - ER stress KW - degradation KW - ERQC KW - sequential steps KW - core Atgs KW - autophagy KW - use autophagic organelles KW - Ypt GTPases KW - ER membrane KW - upr KW - ap KW - system shuttles misfolded proteins KW - vacuole KW - role KW - pathway KW - ER quality control N2 - A. The ypt1-1 mutation is epistatic to vps21∆ in ER-phagy. yDsRed-Snc1-PEM was overexpressed in WT, vps21∆, ypt1-1 and ypt1-1 vps21∆ double mutant cells that also expressed the autophagosomal marker yEGFP-Atg8. Cells were analyzed by live-cell microscopy. Shown from left to right: DIC, DsRed, GFP, merge, % cells Atg8 dots, number of Atg8 dots per cell, and % cells in which the Atg8 dots co-localize with Snc1-PEM. About 50% of WT and 85% of vps21∆ mutant cells contain ~1 dot of Atg8 representing the AP. Importantly, in ~70% of the vps21∆ mutant cells Snc1-PEM co-localizes with the APs, as compared to ~4% in WT cells. In contrast, ~90% ypt1-1 and ypt1-1 vps21∆ mutant cells contain three APs per cell, and Snc1-PEM does not co-localize with them. Arrows point to co-localization; arrowheads point to either Atg8 dots or GFP-Snc1-PEM that do not co-localize. B-D. UPR induction is not required for macro-ER-phagy. The UPR regulators Ire1 or Hac1 were deleted in the WT and ypt1-1 mutant cells. The following effects of overexpression of GFP-Snc1-PEM in WT and ypt1-1 mutant cells, without and with ire1∆ or hac1∆, were analyzed as described for Fig 1A–1C, respectively: protein level (B), accumulation of GFP-Snc1-PEM in aberrant structures (C), and UPR induction (D). B. Deletion of either Ire1 or Hac1 does not affect the level of Snc1-PEM accumulation in WT or ypt1-1 mutant cells. C. Deletion of either Ire1 or Hac1 does not affect the percent of WT and ypt1-1 mutant cells that accumulate aberrant intra-cellular Snc1-PEM. Shown from top to bottom: DIC, GFP, and % cells with intracellular Snc1-PEM structures. D. Deletion of either Ire1 or Hac1 obliterate UPR in both WT and ypt1-1 mutant cells. +/- and error bars represent STDEV. Results in this figure represent at least two independent experiments. ER -