Limits...
Increase in cellular triacylglycerol content and emergence of large ER-associated lipid droplets in the absence of CDP-DG synthase function.

He Y, Yam C, Pomraning K, Chin JS, Yew JY, Freitag M, Oliferenko S - Mol. Biol. Cell (2014)

Bottom Line: Understanding what determines the cellular amount of neutral lipids and their packaging into lipid droplets is of fundamental and applied interest.Using two species of fission yeast, we show that cycling cells deficient in the function of the ER-resident CDP-DG synthase Cds1 exhibit markedly increased triacylglycerol content and assemble large lipid droplets closely associated with the ER membranes.Our results suggest that interfering with the CDP-DG route of phosphatidic acid utilization rewires cellular metabolism to adopt a triacylglycerol-rich lifestyle reliant on the Kennedy pathway.

View Article: PubMed Central - PubMed

Affiliation: Temasek Life Sciences Laboratory, 117604 Singapore Department of Biological Sciences, National University of Singapore, 117543 Singapore.

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Large ER-associated neutral lipid deposits in cds1-9 S. pombe cells are lipid droplets. (A) The BODIPY 493/503–stained neutral lipid deposits (green) in S. pombe cds1-9 mutant cells are associated with ER membranes marked by the artificial luminal marker mCherry-ADEL (red). (B) Histogram demonstrating LD size distribution in WT (blue) and cds1-9 (red) mutant cells. LDs were binned according to volume (μm3) as follows: ≤0.1, between 0.1 and 0.2, between 0.2 and 0.5, between 0.5 and 1, between 1 and 1.5, between 1.5 and 2, and ≥2. n = 309 LDs for WT and 303 LDs for cds1-9. (C, D) Time-lapse sequences showing that the Tgl4-GFP-marked lipid droplets are metabolized in both WT and cds1-9 mutant cells grown in the presence of the fatty acid synthesis inhibitor cerulenin. (A–D) Cells grown in rich yeast extract–based medium. Time is in hours and minutes. Scale bar, 5 μm.
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Figure 6: Large ER-associated neutral lipid deposits in cds1-9 S. pombe cells are lipid droplets. (A) The BODIPY 493/503–stained neutral lipid deposits (green) in S. pombe cds1-9 mutant cells are associated with ER membranes marked by the artificial luminal marker mCherry-ADEL (red). (B) Histogram demonstrating LD size distribution in WT (blue) and cds1-9 (red) mutant cells. LDs were binned according to volume (μm3) as follows: ≤0.1, between 0.1 and 0.2, between 0.2 and 0.5, between 0.5 and 1, between 1 and 1.5, between 1.5 and 2, and ≥2. n = 309 LDs for WT and 303 LDs for cds1-9. (C, D) Time-lapse sequences showing that the Tgl4-GFP-marked lipid droplets are metabolized in both WT and cds1-9 mutant cells grown in the presence of the fatty acid synthesis inhibitor cerulenin. (A–D) Cells grown in rich yeast extract–based medium. Time is in hours and minutes. Scale bar, 5 μm.

Mentions: Similar to the cds1bbl1 mutant in S. japonicus, cds1-9 S. pombe cells formed large LDs associated with the ER membranes upon shift to the restrictive temperature of 36°C (Figure 6A). In the rich yeast extract–based medium, the average droplet volume dramatically increased, whereas the number of droplets decreased (Figure 6B; number of LDs per cell at 36°C was 23 ± 4 and 6 ± 2 for the wild type and cds1-9 mutant, respectively). Of note, we observed higher number of LDs when cells were grown in minimal medium (number of LDs per cell at 36°C was 34 ± 5 and 29 ± 7 for the wild type and cds1-9 mutant, respectively). Of interest, the overall abundance of TG in both wild-type and cds1-9 cells also strongly depended on growth conditions, with cells grown in the rich yeast extract medium showing considerably higher TG content (Supplemental Figure S6A).


Increase in cellular triacylglycerol content and emergence of large ER-associated lipid droplets in the absence of CDP-DG synthase function.

He Y, Yam C, Pomraning K, Chin JS, Yew JY, Freitag M, Oliferenko S - Mol. Biol. Cell (2014)

Large ER-associated neutral lipid deposits in cds1-9 S. pombe cells are lipid droplets. (A) The BODIPY 493/503–stained neutral lipid deposits (green) in S. pombe cds1-9 mutant cells are associated with ER membranes marked by the artificial luminal marker mCherry-ADEL (red). (B) Histogram demonstrating LD size distribution in WT (blue) and cds1-9 (red) mutant cells. LDs were binned according to volume (μm3) as follows: ≤0.1, between 0.1 and 0.2, between 0.2 and 0.5, between 0.5 and 1, between 1 and 1.5, between 1.5 and 2, and ≥2. n = 309 LDs for WT and 303 LDs for cds1-9. (C, D) Time-lapse sequences showing that the Tgl4-GFP-marked lipid droplets are metabolized in both WT and cds1-9 mutant cells grown in the presence of the fatty acid synthesis inhibitor cerulenin. (A–D) Cells grown in rich yeast extract–based medium. Time is in hours and minutes. Scale bar, 5 μm.
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Related In: Results  -  Collection

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Figure 6: Large ER-associated neutral lipid deposits in cds1-9 S. pombe cells are lipid droplets. (A) The BODIPY 493/503–stained neutral lipid deposits (green) in S. pombe cds1-9 mutant cells are associated with ER membranes marked by the artificial luminal marker mCherry-ADEL (red). (B) Histogram demonstrating LD size distribution in WT (blue) and cds1-9 (red) mutant cells. LDs were binned according to volume (μm3) as follows: ≤0.1, between 0.1 and 0.2, between 0.2 and 0.5, between 0.5 and 1, between 1 and 1.5, between 1.5 and 2, and ≥2. n = 309 LDs for WT and 303 LDs for cds1-9. (C, D) Time-lapse sequences showing that the Tgl4-GFP-marked lipid droplets are metabolized in both WT and cds1-9 mutant cells grown in the presence of the fatty acid synthesis inhibitor cerulenin. (A–D) Cells grown in rich yeast extract–based medium. Time is in hours and minutes. Scale bar, 5 μm.
Mentions: Similar to the cds1bbl1 mutant in S. japonicus, cds1-9 S. pombe cells formed large LDs associated with the ER membranes upon shift to the restrictive temperature of 36°C (Figure 6A). In the rich yeast extract–based medium, the average droplet volume dramatically increased, whereas the number of droplets decreased (Figure 6B; number of LDs per cell at 36°C was 23 ± 4 and 6 ± 2 for the wild type and cds1-9 mutant, respectively). Of note, we observed higher number of LDs when cells were grown in minimal medium (number of LDs per cell at 36°C was 34 ± 5 and 29 ± 7 for the wild type and cds1-9 mutant, respectively). Of interest, the overall abundance of TG in both wild-type and cds1-9 cells also strongly depended on growth conditions, with cells grown in the rich yeast extract medium showing considerably higher TG content (Supplemental Figure S6A).

Bottom Line: Understanding what determines the cellular amount of neutral lipids and their packaging into lipid droplets is of fundamental and applied interest.Using two species of fission yeast, we show that cycling cells deficient in the function of the ER-resident CDP-DG synthase Cds1 exhibit markedly increased triacylglycerol content and assemble large lipid droplets closely associated with the ER membranes.Our results suggest that interfering with the CDP-DG route of phosphatidic acid utilization rewires cellular metabolism to adopt a triacylglycerol-rich lifestyle reliant on the Kennedy pathway.

View Article: PubMed Central - PubMed

Affiliation: Temasek Life Sciences Laboratory, 117604 Singapore Department of Biological Sciences, National University of Singapore, 117543 Singapore.

Show MeSH