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Impaired coenzyme A synthesis in fission yeast causes defective mitosis, quiescence-exit failure, histone hypoacetylation and fragile DNA.

Nakamura T, Pluskal T, Nakaseko Y, Yanagida M - Open Biol (2012)

Bottom Line: The mutant becomes auxotrophic to pantothenate at permissive temperature, displaying greatly decreased levels of CoA, acetyl-CoA and histone acetylation.Moreover, ppc1-537 mutant cells failed to restore proliferation from quiescence.Additionally, double-strand break repair is defective in the ppc1-537 mutant, producing fragile broken DNA, probably owing to diminished histone acetylation.

View Article: PubMed Central - PubMed

Affiliation: Okinawa Institute of Science and Technology Graduate University, Tancha 1919-1, Onna, Okinawa 904-0495, Japan.

ABSTRACT
Biosynthesis of coenzyme A (CoA) requires a five-step process using pantothenate and cysteine in the fission yeast Schizosaccharomyces pombe. CoA contains a thiol (SH) group, which reacts with carboxylic acid to form thioesters, giving rise to acyl-activated CoAs such as acetyl-CoA. Acetyl-CoA is essential for energy metabolism and protein acetylation, and, in higher eukaryotes, for the production of neurotransmitters. We isolated a novel S. pombe temperature-sensitive strain ppc1-537 mutated in the catalytic region of phosphopantothenoylcysteine synthetase (designated Ppc1), which is essential for CoA synthesis. The mutant becomes auxotrophic to pantothenate at permissive temperature, displaying greatly decreased levels of CoA, acetyl-CoA and histone acetylation. Moreover, ppc1-537 mutant cells failed to restore proliferation from quiescence. Ppc1 is thus the product of a super-housekeeping gene. The ppc1-537 mutant showed combined synthetic lethal defects with five of six histone deacetylase mutants, whereas sir2 deletion exceptionally rescued the ppc1-537 phenotype. In synchronous cultures, ppc1-537 cells can proceed to the S phase, but lose viability during mitosis failing in sister centromere/kinetochore segregation and nuclear division. Additionally, double-strand break repair is defective in the ppc1-537 mutant, producing fragile broken DNA, probably owing to diminished histone acetylation. The CoA-supported metabolism thus controls the state of chromosome DNA.

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Lipid homeostasis is impaired in ppc1-537 mutant. (a) WT and ppc1-537 cells grown at 26°C and at 36°C for 4 h were stained by nile red [53] to visualize lipid droplets. The number indicates the relative abundance of lipid droplets in WT and ppc1-537 cells. Scale bar, 10 μm. (b) Recovery of lipid droplets stained by nile red upon the transformation of ppc1+ gene. Scale bar, 10 μm. (c)(i) ppc1-537 mutant was sensitive to TSA at 26°C. The sensitivity was similar to that of clr6-1 mutant at 26°C. (ii) ppc1-537 was sensitive to nicotinamide (NA) at 26–30°C, while the WT cells normally grew at the same temperature.
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RSOB120117F7: Lipid homeostasis is impaired in ppc1-537 mutant. (a) WT and ppc1-537 cells grown at 26°C and at 36°C for 4 h were stained by nile red [53] to visualize lipid droplets. The number indicates the relative abundance of lipid droplets in WT and ppc1-537 cells. Scale bar, 10 μm. (b) Recovery of lipid droplets stained by nile red upon the transformation of ppc1+ gene. Scale bar, 10 μm. (c)(i) ppc1-537 mutant was sensitive to TSA at 26°C. The sensitivity was similar to that of clr6-1 mutant at 26°C. (ii) ppc1-537 was sensitive to nicotinamide (NA) at 26–30°C, while the WT cells normally grew at the same temperature.

Mentions: The neutral lipids (triacylglycerol and cholesteryl esters) serve as an energy reserve, and these molecules are stored in lipid droplets. Acetyl-CoA is also required for the production of these metabolites, and we found that lipid droplets stained by nile red [53] showed a striking difference between WT and ppc1-537 mutants (figure 7a). The number of lipid droplets was already relatively low in ppc1-537 mutant at the permissive temperature. At restrictive temperature, the number further decreased, while the number did not change in WT cells. Introduction of the plasmid carrying the ppc1+ gene into the mutant cells restored the number of lipid droplets (figure 7b). These results suggest that lipid droplet homeostasis requires proper CoA biosynthesis.Figure 7.


Impaired coenzyme A synthesis in fission yeast causes defective mitosis, quiescence-exit failure, histone hypoacetylation and fragile DNA.

Nakamura T, Pluskal T, Nakaseko Y, Yanagida M - Open Biol (2012)

Lipid homeostasis is impaired in ppc1-537 mutant. (a) WT and ppc1-537 cells grown at 26°C and at 36°C for 4 h were stained by nile red [53] to visualize lipid droplets. The number indicates the relative abundance of lipid droplets in WT and ppc1-537 cells. Scale bar, 10 μm. (b) Recovery of lipid droplets stained by nile red upon the transformation of ppc1+ gene. Scale bar, 10 μm. (c)(i) ppc1-537 mutant was sensitive to TSA at 26°C. The sensitivity was similar to that of clr6-1 mutant at 26°C. (ii) ppc1-537 was sensitive to nicotinamide (NA) at 26–30°C, while the WT cells normally grew at the same temperature.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3472395&req=5

RSOB120117F7: Lipid homeostasis is impaired in ppc1-537 mutant. (a) WT and ppc1-537 cells grown at 26°C and at 36°C for 4 h were stained by nile red [53] to visualize lipid droplets. The number indicates the relative abundance of lipid droplets in WT and ppc1-537 cells. Scale bar, 10 μm. (b) Recovery of lipid droplets stained by nile red upon the transformation of ppc1+ gene. Scale bar, 10 μm. (c)(i) ppc1-537 mutant was sensitive to TSA at 26°C. The sensitivity was similar to that of clr6-1 mutant at 26°C. (ii) ppc1-537 was sensitive to nicotinamide (NA) at 26–30°C, while the WT cells normally grew at the same temperature.
Mentions: The neutral lipids (triacylglycerol and cholesteryl esters) serve as an energy reserve, and these molecules are stored in lipid droplets. Acetyl-CoA is also required for the production of these metabolites, and we found that lipid droplets stained by nile red [53] showed a striking difference between WT and ppc1-537 mutants (figure 7a). The number of lipid droplets was already relatively low in ppc1-537 mutant at the permissive temperature. At restrictive temperature, the number further decreased, while the number did not change in WT cells. Introduction of the plasmid carrying the ppc1+ gene into the mutant cells restored the number of lipid droplets (figure 7b). These results suggest that lipid droplet homeostasis requires proper CoA biosynthesis.Figure 7.

Bottom Line: The mutant becomes auxotrophic to pantothenate at permissive temperature, displaying greatly decreased levels of CoA, acetyl-CoA and histone acetylation.Moreover, ppc1-537 mutant cells failed to restore proliferation from quiescence.Additionally, double-strand break repair is defective in the ppc1-537 mutant, producing fragile broken DNA, probably owing to diminished histone acetylation.

View Article: PubMed Central - PubMed

Affiliation: Okinawa Institute of Science and Technology Graduate University, Tancha 1919-1, Onna, Okinawa 904-0495, Japan.

ABSTRACT
Biosynthesis of coenzyme A (CoA) requires a five-step process using pantothenate and cysteine in the fission yeast Schizosaccharomyces pombe. CoA contains a thiol (SH) group, which reacts with carboxylic acid to form thioesters, giving rise to acyl-activated CoAs such as acetyl-CoA. Acetyl-CoA is essential for energy metabolism and protein acetylation, and, in higher eukaryotes, for the production of neurotransmitters. We isolated a novel S. pombe temperature-sensitive strain ppc1-537 mutated in the catalytic region of phosphopantothenoylcysteine synthetase (designated Ppc1), which is essential for CoA synthesis. The mutant becomes auxotrophic to pantothenate at permissive temperature, displaying greatly decreased levels of CoA, acetyl-CoA and histone acetylation. Moreover, ppc1-537 mutant cells failed to restore proliferation from quiescence. Ppc1 is thus the product of a super-housekeeping gene. The ppc1-537 mutant showed combined synthetic lethal defects with five of six histone deacetylase mutants, whereas sir2 deletion exceptionally rescued the ppc1-537 phenotype. In synchronous cultures, ppc1-537 cells can proceed to the S phase, but lose viability during mitosis failing in sister centromere/kinetochore segregation and nuclear division. Additionally, double-strand break repair is defective in the ppc1-537 mutant, producing fragile broken DNA, probably owing to diminished histone acetylation. The CoA-supported metabolism thus controls the state of chromosome DNA.

Show MeSH
Related in: MedlinePlus