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Figure 1.
Pex30 and PA accumulate at ER-LD contact sites in sei1Δ. (A) Airyscan images (AS) of WT and sei1Δ cells endogenously expressing Pex30-2xmCherry and Sec63-GFP, an ER marker, on a plasmid. Cells were stained for LDs with MDH and imaged in stationary phase. White arrowheads denote Pex30-2xmCherry and LD puncta that colocalize in sei1Δ. Bar = 2 μm. (B) Quantification of Pex30-2xmCherry accumulation in WT versus sei1Δ. Bars show mean from three independent experiments and standard SEM. 100 cells per genotype from each replicate were analyzed and compared using an unpaired t test (***P < 0.0001). (C) Widefield images (WF) of sei1Δ cells endogenously expressing Pex30-2xmCherry and Opi1-GFP, a sensor for PA, in logarithmic phase. White arrowheads show Pex30-2xmCherry and Opi1-GFP puncta that colocalize; graph to the right of C shows signal intensity on the white line. Bar = 4 μm. (D) WF images of the indicated strains expressing Opi1-GFP on a plasmid in logarithmic phase. White arrowheads show large Opi1-GFP puncta in sei1Δ, and yellow arrowheads show small Opi1-GFP puncta in sei1pex30Δ. Bar = 4 μm. (E) Quantification of the percentage of cells showing Opi1-GFP puncta shown in D. Bars show mean from three independent experiments and SEM. 100 cells per genotype from each replicate were analyzed and compared using one-way ANOVA and Tukey’s multiple comparison test (****P < 0.0001). (F) Quantification of the number of Opi1-GFP puncta in sei1Δ and sei1pex30Δ from cells shown in D. Bars show mean from three independent experiments and SEM. 100 cells per genotype from each replicate were analyzed and compared using an unpaired t test (*P < 0.05). (G) Phospholipid measurements of indicated strains by liquid-chromatography high-resolution mass spectrometry (LC-HRMS) of cell pellets (n = 3). Amount of total quantitated PA relative to total quantitated phospholipids measured in indicated strains (**P < 0.01).

Pex30 and PA accumulate at ER-LD contact sites in sei1Δ. (A) Airyscan images (AS) of WT and sei1Δ cells endogenously expressing Pex30-2xmCherry and Sec63-GFP, an ER marker, on a plasmid. Cells were stained for LDs with MDH and imaged in stationary phase. White arrowheads denote Pex30-2xmCherry and LD puncta that colocalize in sei1Δ. Bar = 2 μm. (B) Quantification of Pex30-2xmCherry accumulation in WT versus sei1Δ. Bars show mean from three independent experiments and standard SEM. 100 cells per genotype from each replicate were analyzed and compared using an unpaired t test (***P < 0.0001). (C) Widefield images (WF) of sei1Δ cells endogenously expressing Pex30-2xmCherry and Opi1-GFP, a sensor for PA, in logarithmic phase. White arrowheads show Pex30-2xmCherry and Opi1-GFP puncta that colocalize; graph to the right of C shows signal intensity on the white line. Bar = 4 μm. (D) WF images of the indicated strains expressing Opi1-GFP on a plasmid in logarithmic phase. White arrowheads show large Opi1-GFP puncta in sei1Δ, and yellow arrowheads show small Opi1-GFP puncta in sei1pex30Δ. Bar = 4 μm. (E) Quantification of the percentage of cells showing Opi1-GFP puncta shown in D. Bars show mean from three independent experiments and SEM. 100 cells per genotype from each replicate were analyzed and compared using one-way ANOVA and Tukey’s multiple comparison test (****P < 0.0001). (F) Quantification of the number of Opi1-GFP puncta in sei1Δ and sei1pex30Δ from cells shown in D. Bars show mean from three independent experiments and SEM. 100 cells per genotype from each replicate were analyzed and compared using an unpaired t test (*P < 0.05). (G) Phospholipid measurements of indicated strains by liquid-chromatography high-resolution mass spectrometry (LC-HRMS) of cell pellets (n = 3). Amount of total quantitated PA relative to total quantitated phospholipids measured in indicated strains (**P < 0.01).

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