Limits...
Genome-wide functional profiling identifies genes and processes important for zinc-limited growth of Saccharomyces cerevisiae.

North M, Steffen J, Loguinov AV, Zimmerman GR, Vulpe CD, Eide DJ - PLoS Genet. (2012)

Bottom Line: Our studies also indicated the critical role of macroautophagy in low zinc growth.Finally, as a result of our analysis, we discovered a previously unknown role for the ICE2 gene in maintaining ER zinc homeostasis.Thus, functional profiling has provided many new insights into genes and processes that are needed for cells to thrive under the stress of zinc deficiency.

View Article: PubMed Central - PubMed

Affiliation: Department of Nutritional Science and Toxicology, University of California Berkeley, Berkeley, California, USA.

ABSTRACT
Zinc is an essential nutrient because it is a required cofactor for many enzymes and transcription factors. To discover genes and processes in yeast that are required for growth when zinc is limiting, we used genome-wide functional profiling. Mixed pools of ∼4,600 deletion mutants were inoculated into zinc-replete and zinc-limiting media. These cells were grown for several generations, and the prevalence of each mutant in the pool was then determined by microarray analysis. As a result, we identified more than 400 different genes required for optimal growth under zinc-limiting conditions. Among these were several targets of the Zap1 zinc-responsive transcription factor. Their importance is consistent with their up-regulation by Zap1 in low zinc. We also identified genes that implicate Zap1-independent processes as important. These include endoplasmic reticulum function, oxidative stress resistance, vesicular trafficking, peroxisome biogenesis, and chromatin modification. Our studies also indicated the critical role of macroautophagy in low zinc growth. Finally, as a result of our analysis, we discovered a previously unknown role for the ICE2 gene in maintaining ER zinc homeostasis. Thus, functional profiling has provided many new insights into genes and processes that are needed for cells to thrive under the stress of zinc deficiency.

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Effects of zinc status on sensitivity to an ER stress inducer.Wild-type (BY4743) cells were inoculated into zinc-replete (LZM+100 µM ZnCl2, +Zn) or zinc-limiting (LZM+1 µM ZnCl2, −Zn) media containing a range of tunicamycin concentrations and grown overnight prior to measuring the culture optical densities at 600 nm (OD600). Hypersensitivity to tunicamycin was observed for zinc-limited cells at 0.1, 0.25, and 0.5 mM tunicamcyin. Data presented are the averages of triplicate cultures for each condition and the error bars indicate ±1 S.D. For most points, the symbols obscure the error bars.
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pgen-1002699-g004: Effects of zinc status on sensitivity to an ER stress inducer.Wild-type (BY4743) cells were inoculated into zinc-replete (LZM+100 µM ZnCl2, +Zn) or zinc-limiting (LZM+1 µM ZnCl2, −Zn) media containing a range of tunicamycin concentrations and grown overnight prior to measuring the culture optical densities at 600 nm (OD600). Hypersensitivity to tunicamycin was observed for zinc-limited cells at 0.1, 0.25, and 0.5 mM tunicamcyin. Data presented are the averages of triplicate cultures for each condition and the error bars indicate ±1 S.D. For most points, the symbols obscure the error bars.

Mentions: Many genes related to endoplasmic reticulum function were also identified (Table 3). These included MSC2, which encodes a zinc transporter protein that, along with Zrg17 (see above), transports zinc into the ER. HAC1 and IRE1, which encode the transcription factor and its controlling sensor that are active in the ER unfolded protein response (UPR), molecular co-chaperones (JEM1, SCJ1), and several genes involved in ER protein modification and trafficking were identified as growing poorly in low zinc. These results suggested that disruption of ER function in many different ways impairs the ability of cells to grow in low zinc. To test this hypothesis further, we examined the effects of tunicamycin, an inducer of ER stress [30], [31], on low zinc growth. Cells were grown in either low zinc (LZM+1 µM ZnCl2) or zinc-replete conditions (LZM+100 µM ZnCl2) with a range of tunicamycin concentrations. As shown in Figure 4, sensitivity to tunicamycin was greatly exacerbated by growth in low zinc medium relative to zinc-replete conditions. These results support the hypothesis that unperturbed ER function is critical to growth when zinc is scarce.


Genome-wide functional profiling identifies genes and processes important for zinc-limited growth of Saccharomyces cerevisiae.

North M, Steffen J, Loguinov AV, Zimmerman GR, Vulpe CD, Eide DJ - PLoS Genet. (2012)

Effects of zinc status on sensitivity to an ER stress inducer.Wild-type (BY4743) cells were inoculated into zinc-replete (LZM+100 µM ZnCl2, +Zn) or zinc-limiting (LZM+1 µM ZnCl2, −Zn) media containing a range of tunicamycin concentrations and grown overnight prior to measuring the culture optical densities at 600 nm (OD600). Hypersensitivity to tunicamycin was observed for zinc-limited cells at 0.1, 0.25, and 0.5 mM tunicamcyin. Data presented are the averages of triplicate cultures for each condition and the error bars indicate ±1 S.D. For most points, the symbols obscure the error bars.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3369956&req=5

pgen-1002699-g004: Effects of zinc status on sensitivity to an ER stress inducer.Wild-type (BY4743) cells were inoculated into zinc-replete (LZM+100 µM ZnCl2, +Zn) or zinc-limiting (LZM+1 µM ZnCl2, −Zn) media containing a range of tunicamycin concentrations and grown overnight prior to measuring the culture optical densities at 600 nm (OD600). Hypersensitivity to tunicamycin was observed for zinc-limited cells at 0.1, 0.25, and 0.5 mM tunicamcyin. Data presented are the averages of triplicate cultures for each condition and the error bars indicate ±1 S.D. For most points, the symbols obscure the error bars.
Mentions: Many genes related to endoplasmic reticulum function were also identified (Table 3). These included MSC2, which encodes a zinc transporter protein that, along with Zrg17 (see above), transports zinc into the ER. HAC1 and IRE1, which encode the transcription factor and its controlling sensor that are active in the ER unfolded protein response (UPR), molecular co-chaperones (JEM1, SCJ1), and several genes involved in ER protein modification and trafficking were identified as growing poorly in low zinc. These results suggested that disruption of ER function in many different ways impairs the ability of cells to grow in low zinc. To test this hypothesis further, we examined the effects of tunicamycin, an inducer of ER stress [30], [31], on low zinc growth. Cells were grown in either low zinc (LZM+1 µM ZnCl2) or zinc-replete conditions (LZM+100 µM ZnCl2) with a range of tunicamycin concentrations. As shown in Figure 4, sensitivity to tunicamycin was greatly exacerbated by growth in low zinc medium relative to zinc-replete conditions. These results support the hypothesis that unperturbed ER function is critical to growth when zinc is scarce.

Bottom Line: Our studies also indicated the critical role of macroautophagy in low zinc growth.Finally, as a result of our analysis, we discovered a previously unknown role for the ICE2 gene in maintaining ER zinc homeostasis.Thus, functional profiling has provided many new insights into genes and processes that are needed for cells to thrive under the stress of zinc deficiency.

View Article: PubMed Central - PubMed

Affiliation: Department of Nutritional Science and Toxicology, University of California Berkeley, Berkeley, California, USA.

ABSTRACT
Zinc is an essential nutrient because it is a required cofactor for many enzymes and transcription factors. To discover genes and processes in yeast that are required for growth when zinc is limiting, we used genome-wide functional profiling. Mixed pools of ∼4,600 deletion mutants were inoculated into zinc-replete and zinc-limiting media. These cells were grown for several generations, and the prevalence of each mutant in the pool was then determined by microarray analysis. As a result, we identified more than 400 different genes required for optimal growth under zinc-limiting conditions. Among these were several targets of the Zap1 zinc-responsive transcription factor. Their importance is consistent with their up-regulation by Zap1 in low zinc. We also identified genes that implicate Zap1-independent processes as important. These include endoplasmic reticulum function, oxidative stress resistance, vesicular trafficking, peroxisome biogenesis, and chromatin modification. Our studies also indicated the critical role of macroautophagy in low zinc growth. Finally, as a result of our analysis, we discovered a previously unknown role for the ICE2 gene in maintaining ER zinc homeostasis. Thus, functional profiling has provided many new insights into genes and processes that are needed for cells to thrive under the stress of zinc deficiency.

Show MeSH
Related in: MedlinePlus