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Vol. 18, Issue 9, 3556-3567, September 2007

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Yeast Cells Lacking the CIT1-encoded Mitochondrial Citrate Synthase Are Hypersusceptible to Heat- or Aging-induced Apoptosis

Yong Joo Lee^*, Kwang Lae Hoe, and Pil Jae Maeng^*

*Department of Microbiology, School of Bioscience and Biotechnology, Chungnam National University, 305-764 Daejeon, Korea; and Functional Genomics Research Center, Korea Research Institute of Bioscience and Biotechnology, 305-806 Daejeon, Korea

Submitted February 12, 2007; Revised June 22, 2007; Accepted June 25, 2007
Monitoring Editor: Peter Walter

In Saccharomyces cerevisiae, the initial reaction of the tricarboxylic acid cycle is catalyzed by the mitochondrial citrate synthase Cit1. The function of Cit1 has previously been studied mainly in terms of acetate utilization and metabolon construction. Here, we report the relationship between the function of Cit1 and apoptosis. Yeast cells with cit1 deletion showed a temperature-sensitive growth phenotype, and they displayed a rapid loss in viability associated with typical apoptotic hallmarks, i.e., reactive oxygen species (ROS) accumulation and nuclear fragmentation, DNA breakage, and phosphatidylserine translocation, when exposed to heat stress. On long-term cultivation, cit1 null strains showed increased potentials for both aging-induced apoptosis and adaptive regrowth. Activation of the metacaspase Yca1 was detected during heat- or aging-induced apoptosis in cit1 null strains, and accordingly, deletion of YCA1 suppressed the apoptotic phenotype caused by cit1 null mutation. Cells with cit1 deletion showed higher tendency toward glutathione (GSH) depletion and subsequent ROS accumulation than the wild type, which was rescued by exogenous GSH, glutamate, or glutathione disulfide (GSSG). These results led us to conclude that GSH deficiency in cit1 null cells is caused by an insufficient supply of glutamate necessary for biosynthesis of GSH rather than the depletion of reducing power required for reduction of GSSG to GSH.

Address correspondence to: Pil Jae Maeng (pjmaeng{at}cnu.ac.kr).

Abbreviations used: CFU, colony-forming unit; CS, citrate synthase; DCFH-DA, 2',7'-dichlorodihydrofluorescein diacetate; DCF, 2',7'-dichlorofluorescein diacetate; DTNB, 5,5'-dithiobis-(2-nitrobenzoic acid); GR, glutathione reductase; GSH, glutathione; GSSG, glutathione disulfide; ROS, reactive oxygen species; SCD, synthetic complete glucose; TCA, tricarboxylic acid; ts, temperature-sensitive; TUNEL, terminal deoxynucleotidyl transferase dUTP nick-end labeling.