Limits...
Deteriorated stress response in stationary-phase yeast: Sir2 and Yap1 are essential for Hsf1 activation by heat shock and oxidative stress, respectively.

Nussbaum I, Weindling E, Jubran R, Cohen A, Bar-Nun S - PLoS ONE (2014)

Bottom Line: However, the molecular processes underlying stress response of non-dividing cells are poorly understood, although deteriorated stress response is one of the hallmarks of aging.This response is orchestrated largely by the conserved transcription factor Hsf1, which in S. cerevisiae regulates expression of multiple genes in response to diverse stresses.Rather, factors that participate in Hsf1 activation appear to be compromised.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.

ABSTRACT
Stationary-phase cultures have been used as an important model of aging, a complex process involving multiple pathways and signaling networks. However, the molecular processes underlying stress response of non-dividing cells are poorly understood, although deteriorated stress response is one of the hallmarks of aging. The budding yeast Saccharomyces cerevisiae is a valuable model organism to study the genetics of aging, because yeast ages within days and are amenable to genetic manipulations. As a unicellular organism, yeast has evolved robust systems to respond to environmental challenges. This response is orchestrated largely by the conserved transcription factor Hsf1, which in S. cerevisiae regulates expression of multiple genes in response to diverse stresses. Here we demonstrate that Hsf1 response to heat shock and oxidative stress deteriorates during yeast transition from exponential growth to stationary-phase, whereas Hsf1 activation by glucose starvation is maintained. Overexpressing Hsf1 does not significantly improve heat shock response, indicating that Hsf1 dwindling is not the major cause for Hsf1 attenuated response in stationary-phase yeast. Rather, factors that participate in Hsf1 activation appear to be compromised. We uncover two factors, Yap1 and Sir2, which discretely function in Hsf1 activation by oxidative stress and heat shock. In Δyap1 mutant, Hsf1 does not respond to oxidative stress, while in Δsir2 mutant, Hsf1 does not respond to heat shock. Moreover, excess Sir2 mimics the heat shock response. This role of the NAD+-dependent Sir2 is supported by our finding that supplementing NAD+ precursors improves Hsf1 heat shock response in stationary-phase yeast, especially when combined with expression of excess Sir2. Finally, the combination of excess Hsf1, excess Sir2 and NAD+ precursors rejuvenates the heat shock response.

Show MeSH

Related in: MedlinePlus

Activation of Hsf1 by heat shock is restored in stationary-phase yeast by combination of excess Hsf1, excess Sir2 and NAD+ precursor.(A) Wild-type W303-1b cells harboring HSE2-LacZ plasmid were transformed with empty vectors (EVs) or with a combination of pHSF1+pSIR2 plasmids. Cells grown at 30°C, either exponentially (light blue bars) or to stationary-phase (dark blue bars), were incubated for 30 min with (+) or without (−) NR (10 µM) prior to the heat shock. Cells were either incubated further for 20 min at 30°C (−) or subjected to a 20 min heat shock (HS) at 42°C (+). Hsf1 activity was measured as β-galactosidase specific activity. Data are the mean plus standard error of 3 independent experiments. (B) At each treatment, the activity in stationary-phase cells was calculated as % of the activity in exponentially-growing cells.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4214751&req=5

pone-0111505-g008: Activation of Hsf1 by heat shock is restored in stationary-phase yeast by combination of excess Hsf1, excess Sir2 and NAD+ precursor.(A) Wild-type W303-1b cells harboring HSE2-LacZ plasmid were transformed with empty vectors (EVs) or with a combination of pHSF1+pSIR2 plasmids. Cells grown at 30°C, either exponentially (light blue bars) or to stationary-phase (dark blue bars), were incubated for 30 min with (+) or without (−) NR (10 µM) prior to the heat shock. Cells were either incubated further for 20 min at 30°C (−) or subjected to a 20 min heat shock (HS) at 42°C (+). Hsf1 activity was measured as β-galactosidase specific activity. Data are the mean plus standard error of 3 independent experiments. (B) At each treatment, the activity in stationary-phase cells was calculated as % of the activity in exponentially-growing cells.

Mentions: Our findings in Figures 5–7 suggested that both Sir2 and NAD+ were limiting in stationary-phase yeast. Yet, additional factors seemed to be limiting in the Hsf1 activation cascade, as the heat shock response in these cells fell short of that in exponentially-growing yeast (Figures 7 and S3). Moreover, while the effect of excess Sir2 was further augmented by NR, neither in exponentially-growing nor in stationary-phase yeast was the excess Sir2 (with or without NR) further increased by heat shock (Figures 7 and S3). A plausible candidate for such a limiting factor was Hsf1 itself, since excess Hsf1 improved Hsf1 activity in stationary-phase yeast (Figure 2). Indeed, overexpressing Hsf1 together with Sir2 and providing the cells with NR restored the heat shock response of stationary-phase yeast to nearly 70% of that of exponentially-growing cells (Figure 8). Thus, limiting levels of three factors appear to impair the ability of stationary-phase yeast to respond to heat shock: Hsf1, Sir2 and NAD+.


Deteriorated stress response in stationary-phase yeast: Sir2 and Yap1 are essential for Hsf1 activation by heat shock and oxidative stress, respectively.

Nussbaum I, Weindling E, Jubran R, Cohen A, Bar-Nun S - PLoS ONE (2014)

Activation of Hsf1 by heat shock is restored in stationary-phase yeast by combination of excess Hsf1, excess Sir2 and NAD+ precursor.(A) Wild-type W303-1b cells harboring HSE2-LacZ plasmid were transformed with empty vectors (EVs) or with a combination of pHSF1+pSIR2 plasmids. Cells grown at 30°C, either exponentially (light blue bars) or to stationary-phase (dark blue bars), were incubated for 30 min with (+) or without (−) NR (10 µM) prior to the heat shock. Cells were either incubated further for 20 min at 30°C (−) or subjected to a 20 min heat shock (HS) at 42°C (+). Hsf1 activity was measured as β-galactosidase specific activity. Data are the mean plus standard error of 3 independent experiments. (B) At each treatment, the activity in stationary-phase cells was calculated as % of the activity in exponentially-growing cells.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4214751&req=5

pone-0111505-g008: Activation of Hsf1 by heat shock is restored in stationary-phase yeast by combination of excess Hsf1, excess Sir2 and NAD+ precursor.(A) Wild-type W303-1b cells harboring HSE2-LacZ plasmid were transformed with empty vectors (EVs) or with a combination of pHSF1+pSIR2 plasmids. Cells grown at 30°C, either exponentially (light blue bars) or to stationary-phase (dark blue bars), were incubated for 30 min with (+) or without (−) NR (10 µM) prior to the heat shock. Cells were either incubated further for 20 min at 30°C (−) or subjected to a 20 min heat shock (HS) at 42°C (+). Hsf1 activity was measured as β-galactosidase specific activity. Data are the mean plus standard error of 3 independent experiments. (B) At each treatment, the activity in stationary-phase cells was calculated as % of the activity in exponentially-growing cells.
Mentions: Our findings in Figures 5–7 suggested that both Sir2 and NAD+ were limiting in stationary-phase yeast. Yet, additional factors seemed to be limiting in the Hsf1 activation cascade, as the heat shock response in these cells fell short of that in exponentially-growing yeast (Figures 7 and S3). Moreover, while the effect of excess Sir2 was further augmented by NR, neither in exponentially-growing nor in stationary-phase yeast was the excess Sir2 (with or without NR) further increased by heat shock (Figures 7 and S3). A plausible candidate for such a limiting factor was Hsf1 itself, since excess Hsf1 improved Hsf1 activity in stationary-phase yeast (Figure 2). Indeed, overexpressing Hsf1 together with Sir2 and providing the cells with NR restored the heat shock response of stationary-phase yeast to nearly 70% of that of exponentially-growing cells (Figure 8). Thus, limiting levels of three factors appear to impair the ability of stationary-phase yeast to respond to heat shock: Hsf1, Sir2 and NAD+.

Bottom Line: However, the molecular processes underlying stress response of non-dividing cells are poorly understood, although deteriorated stress response is one of the hallmarks of aging.This response is orchestrated largely by the conserved transcription factor Hsf1, which in S. cerevisiae regulates expression of multiple genes in response to diverse stresses.Rather, factors that participate in Hsf1 activation appear to be compromised.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel.

ABSTRACT
Stationary-phase cultures have been used as an important model of aging, a complex process involving multiple pathways and signaling networks. However, the molecular processes underlying stress response of non-dividing cells are poorly understood, although deteriorated stress response is one of the hallmarks of aging. The budding yeast Saccharomyces cerevisiae is a valuable model organism to study the genetics of aging, because yeast ages within days and are amenable to genetic manipulations. As a unicellular organism, yeast has evolved robust systems to respond to environmental challenges. This response is orchestrated largely by the conserved transcription factor Hsf1, which in S. cerevisiae regulates expression of multiple genes in response to diverse stresses. Here we demonstrate that Hsf1 response to heat shock and oxidative stress deteriorates during yeast transition from exponential growth to stationary-phase, whereas Hsf1 activation by glucose starvation is maintained. Overexpressing Hsf1 does not significantly improve heat shock response, indicating that Hsf1 dwindling is not the major cause for Hsf1 attenuated response in stationary-phase yeast. Rather, factors that participate in Hsf1 activation appear to be compromised. We uncover two factors, Yap1 and Sir2, which discretely function in Hsf1 activation by oxidative stress and heat shock. In Δyap1 mutant, Hsf1 does not respond to oxidative stress, while in Δsir2 mutant, Hsf1 does not respond to heat shock. Moreover, excess Sir2 mimics the heat shock response. This role of the NAD+-dependent Sir2 is supported by our finding that supplementing NAD+ precursors improves Hsf1 heat shock response in stationary-phase yeast, especially when combined with expression of excess Sir2. Finally, the combination of excess Hsf1, excess Sir2 and NAD+ precursors rejuvenates the heat shock response.

Show MeSH
Related in: MedlinePlus