Figure 7.
Two PxL motifs in Atg13 are required for Cdc14 docking. (A) Top: Schematic of Atg13 (S. cerevisiae) with six Cdc14 preferred target sites (SP1−6: S129-P, S348-P, S454-P, S535-P, S541-P, and S646-P, in red) and three predicted docking motifs (PxL1−3: P76-PL, P207-IL, and P476-DL, in blue). HORMA, Hop1p, Rev7p, and MAD2. Bottom: Sequence alignment of the PxL3 (P476-DL) region among Atg13 homologs from indicated species. Symbols mark identical (yellow shade) and similar residues (*). (B) IB of recombinant FLAG-tagged Cdc14/Cdc14m binding to bead-immobilized 3HA-Atg13 variant proteins that were purified by IP (α-HA) from vegetative cell lysates. Top: Schematic of experimental design. (C and D) FM analysis of GFP-tagged Cdc14(IN) recruitment to the puncta of mScarlet-tagged Atg13 variants, during synchronized meiotic divisions. (C) Representative FM images. Scale bar, 5 µm. White arrows mark recruitment of Cdc14-GFP to mScarlet-Atg13 variants. (D) Quantitation (n ≥ 300 cells, t test; ***, P ≤ 0.001). (E) Sporulation was triggered in nonsynchronized meiotic cells expressing indicated ATG13 variants, which were carried by pRS303 vector in atg13Δ strain. Shown is the percentage of cells sporulated after 48 h in SPM (n ≥ 300 cells; t test; ***, P ≤ 0.001). Source data are available for this figure: SourceData F7.

Two PxL motifs in Atg13 are required for Cdc14 docking. (A) Top: Schematic of Atg13 (S. cerevisiae) with six Cdc14 preferred target sites (SP1−6: S129-P, S348-P, S454-P, S535-P, S541-P, and S646-P, in red) and three predicted docking motifs (PxL1−3: P76-PL, P207-IL, and P476-DL, in blue). HORMA, Hop1p, Rev7p, and MAD2. Bottom: Sequence alignment of the PxL3 (P476-DL) region among Atg13 homologs from indicated species. Symbols mark identical (yellow shade) and similar residues (*). (B) IB of recombinant FLAG-tagged Cdc14/Cdc14m binding to bead-immobilized 3HA-Atg13 variant proteins that were purified by IP (α-HA) from vegetative cell lysates. Top: Schematic of experimental design. (C and D) FM analysis of GFP-tagged Cdc14(IN) recruitment to the puncta of mScarlet-tagged Atg13 variants, during synchronized meiotic divisions. (C) Representative FM images. Scale bar, 5 µm. White arrows mark recruitment of Cdc14-GFP to mScarlet-Atg13 variants. (D) Quantitation (n ≥ 300 cells, t test; ***, P ≤ 0.001). (E) Sporulation was triggered in nonsynchronized meiotic cells expressing indicated ATG13 variants, which were carried by pRS303 vector in atg13Δ strain. Shown is the percentage of cells sporulated after 48 h in SPM (n ≥ 300 cells; t test; ***, P ≤ 0.001). Source data are available for this figure: SourceData F7.

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