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Starvation induced cell death in autophagy-defective yeast mutants is caused by mitochondria dysfunction.

Suzuki SW, Onodera J, Ohsumi Y - PLoS ONE (2011)

Bottom Line: We found that buffering of the starvation medium dramatically restored the viability of atg mutants.Consequently, autophagy-defective mutants accumulated the high level of ROS, leading to deficient respiratory function, resulting in the loss of mitochondria DNA (mtDNA).We also showed that mtDNA deficient cells are subject to cell death under low pH starvation conditions.

View Article: PubMed Central - PubMed

Affiliation: Frontier Research Center, Tokyo Institute of Technology, Yokohama, Japan.

ABSTRACT
Autophagy is a highly-conserved cellular degradation and recycling system that is essential for cell survival during nutrient starvation. The loss of viability had been used as an initial screen to identify autophagy-defective (atg) mutants of the yeast Saccharomyces cerevisiae, but the mechanism of cell death in these mutants has remained unclear. When cells grown in a rich medium were transferred to a synthetic nitrogen starvation media, secreted metabolites lowered the extracellular pH below 3.0 and autophagy-defective mutants mostly died. We found that buffering of the starvation medium dramatically restored the viability of atg mutants. In response to starvation, wild-type (WT) cells were able to upregulate components of the respiratory pathway and ROS (reactive oxygen species) scavenging enzymes, but atg mutants lacked this synthetic capacity. Consequently, autophagy-defective mutants accumulated the high level of ROS, leading to deficient respiratory function, resulting in the loss of mitochondria DNA (mtDNA). We also showed that mtDNA deficient cells are subject to cell death under low pH starvation conditions. Taken together, under starvation conditions non-selective autophagy, rather than mitophagy, plays an essential role in preventing ROS accumulation, and thus in maintaining mitochondria function. The failure of response to starvation is the major cause of cell death in atg mutants.

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Related in: MedlinePlus

Viability of respiratory deficient cells during nitrogen starvation.(A) WT and atg1Δ cells were transferred to non-buffered SD-N medium for the indicated time. Cell viability was determined by phloxine B staining. Nitrogen-starved cells were plated onto YEPD agar and overlaid with TTC agar to examine the respiratory competency of formed colonies. The black and gray areas indicate the percentage of viable cells that are respiratory competent or respiratory deficient, respectively. (B) WT, atg1Δ, rho0, and rho0 atg1Δ cells grown in YEPD medium were transferred to SD-N with or without 10 mM NAC for the indicated time. In the presence of NAC, medium pH was adjusted by using KOH. Cell viability and medium pH were examined by phloxine B staining and pH meter, respectively. These data represent the average of three independent experiments and bars indicate standard deviations.
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pone-0017412-g005: Viability of respiratory deficient cells during nitrogen starvation.(A) WT and atg1Δ cells were transferred to non-buffered SD-N medium for the indicated time. Cell viability was determined by phloxine B staining. Nitrogen-starved cells were plated onto YEPD agar and overlaid with TTC agar to examine the respiratory competency of formed colonies. The black and gray areas indicate the percentage of viable cells that are respiratory competent or respiratory deficient, respectively. (B) WT, atg1Δ, rho0, and rho0 atg1Δ cells grown in YEPD medium were transferred to SD-N with or without 10 mM NAC for the indicated time. In the presence of NAC, medium pH was adjusted by using KOH. Cell viability and medium pH were examined by phloxine B staining and pH meter, respectively. These data represent the average of three independent experiments and bars indicate standard deviations.

Mentions: In atg mutants, the time course of the appearance of the respiratory-deficient cells in buffered starvation medium temporally corresponded to the loss of cell viability seen in non-buffered starvation medium. We monitored the respiratory activity of autophagy-defective mutants in non-buffered starvation medium, and after 96 hours of starvation nearly all of the surviving atg1Δ cells exhibited respiratory activity (Figure 5A), indicating that respiratory-deficient cells were selectively eliminated.


Starvation induced cell death in autophagy-defective yeast mutants is caused by mitochondria dysfunction.

Suzuki SW, Onodera J, Ohsumi Y - PLoS ONE (2011)

Viability of respiratory deficient cells during nitrogen starvation.(A) WT and atg1Δ cells were transferred to non-buffered SD-N medium for the indicated time. Cell viability was determined by phloxine B staining. Nitrogen-starved cells were plated onto YEPD agar and overlaid with TTC agar to examine the respiratory competency of formed colonies. The black and gray areas indicate the percentage of viable cells that are respiratory competent or respiratory deficient, respectively. (B) WT, atg1Δ, rho0, and rho0 atg1Δ cells grown in YEPD medium were transferred to SD-N with or without 10 mM NAC for the indicated time. In the presence of NAC, medium pH was adjusted by using KOH. Cell viability and medium pH were examined by phloxine B staining and pH meter, respectively. These data represent the average of three independent experiments and bars indicate standard deviations.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3045454&req=5

pone-0017412-g005: Viability of respiratory deficient cells during nitrogen starvation.(A) WT and atg1Δ cells were transferred to non-buffered SD-N medium for the indicated time. Cell viability was determined by phloxine B staining. Nitrogen-starved cells were plated onto YEPD agar and overlaid with TTC agar to examine the respiratory competency of formed colonies. The black and gray areas indicate the percentage of viable cells that are respiratory competent or respiratory deficient, respectively. (B) WT, atg1Δ, rho0, and rho0 atg1Δ cells grown in YEPD medium were transferred to SD-N with or without 10 mM NAC for the indicated time. In the presence of NAC, medium pH was adjusted by using KOH. Cell viability and medium pH were examined by phloxine B staining and pH meter, respectively. These data represent the average of three independent experiments and bars indicate standard deviations.
Mentions: In atg mutants, the time course of the appearance of the respiratory-deficient cells in buffered starvation medium temporally corresponded to the loss of cell viability seen in non-buffered starvation medium. We monitored the respiratory activity of autophagy-defective mutants in non-buffered starvation medium, and after 96 hours of starvation nearly all of the surviving atg1Δ cells exhibited respiratory activity (Figure 5A), indicating that respiratory-deficient cells were selectively eliminated.

Bottom Line: We found that buffering of the starvation medium dramatically restored the viability of atg mutants.Consequently, autophagy-defective mutants accumulated the high level of ROS, leading to deficient respiratory function, resulting in the loss of mitochondria DNA (mtDNA).We also showed that mtDNA deficient cells are subject to cell death under low pH starvation conditions.

View Article: PubMed Central - PubMed

Affiliation: Frontier Research Center, Tokyo Institute of Technology, Yokohama, Japan.

ABSTRACT
Autophagy is a highly-conserved cellular degradation and recycling system that is essential for cell survival during nutrient starvation. The loss of viability had been used as an initial screen to identify autophagy-defective (atg) mutants of the yeast Saccharomyces cerevisiae, but the mechanism of cell death in these mutants has remained unclear. When cells grown in a rich medium were transferred to a synthetic nitrogen starvation media, secreted metabolites lowered the extracellular pH below 3.0 and autophagy-defective mutants mostly died. We found that buffering of the starvation medium dramatically restored the viability of atg mutants. In response to starvation, wild-type (WT) cells were able to upregulate components of the respiratory pathway and ROS (reactive oxygen species) scavenging enzymes, but atg mutants lacked this synthetic capacity. Consequently, autophagy-defective mutants accumulated the high level of ROS, leading to deficient respiratory function, resulting in the loss of mitochondria DNA (mtDNA). We also showed that mtDNA deficient cells are subject to cell death under low pH starvation conditions. Taken together, under starvation conditions non-selective autophagy, rather than mitophagy, plays an essential role in preventing ROS accumulation, and thus in maintaining mitochondria function. The failure of response to starvation is the major cause of cell death in atg mutants.

Show MeSH
Related in: MedlinePlus