Figure 3. Sorting of GPI-APs into the ERES is impaired in pgap1 and pgap5 mutant cells. (A–C) VFG-GPI were expressed and accumulated in the ER by incubating FF8 (WT), FPRC2 (pgap1), and C19 (pgap5) cells stably expressing myc-tagged Sec13 (Sec13-myc) with 1 µg/ml doxycycline at 40°C for 24 h. After incubation at 10°C for 0 min (A) or 30 min (B and C, high magnification of B), cells were fixed with 4% paraformaldehyde and stained with anti-myc antibody to label the ERES. Bars: (A and B) 10 µm; (C) 5 µm. (D) Relative intensity of VFG-GPI in the ERES. The ratios of average intensities of VFG-GPI in the ERES (AI(eres)) to those in the ERES surroundings (AI(surro)) were measured. The ratios between wild-type (WT) and pgap1 mutant cells and between WT and pgap5 mutant cells differed significantly at each time point. (n = 715–1,906; Student’s t test, P < 0.0001). Error bars indicate the standard error of the mean.
Figure 3.

Sorting of GPI-APs into the ERES is impaired in pgap1 and pgap5 mutant cells. (A–C) VFG-GPI were expressed and accumulated in the ER by incubating FF8 (WT), FPRC2 (pgap1), and C19 (pgap5) cells stably expressing myc-tagged Sec13 (Sec13-myc) with 1 µg/ml doxycycline at 40°C for 24 h. After incubation at 10°C for 0 min (A) or 30 min (B and C, high magnification of B), cells were fixed with 4% paraformaldehyde and stained with anti-myc antibody to label the ERES. Bars: (A and B) 10 µm; (C) 5 µm. (D) Relative intensity of VFG-GPI in the ERES. The ratios of average intensities of VFG-GPI in the ERES (AI(eres)) to those in the ERES surroundings (AI(surro)) were measured. The ratios between wild-type (WT) and pgap1 mutant cells and between WT and pgap5 mutant cells differed significantly at each time point. (n = 715–1,906; Student’s t test, P < 0.0001). Error bars indicate the standard error of the mean.

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