NRG1 is required for glucose repression of the SUC2 and GAL genes of Saccharomyces cerevisiae - PubMed (original) (raw)

NRG1 is required for glucose repression of the SUC2 and GAL genes of Saccharomyces cerevisiae

H Zhou et al. BMC Genet. 2001.

Abstract

Background: Glucose repression of transcription in the yeast, Saccharomyces cerevisiae, has been shown to be controlled by several factors, including two repressors called Mig1 and Mig2. Past results suggest that other repressors may be involved in glucose repression.

Results: By a screen for factors that control transcription of the glucose-repressible SUC2 gene of S. cerevisiae, the NRG1 gene was identified. Analysis of an nrg1Delta mutant has demonstrated that mRNA levels are elevated at both the SUC2 and of the GAL genes of S. cerevisiae when cells are grown under normally glucose-repressing conditions. In addition, genetic interactions have been detected between nrg1Delta and other factors that control SUC2 transcription.

Conclusions: The analysis of nrg1Delta demonstrates that Nrg1 plays a role in glucose repression of the SUC2 and GAL genes of S. cerevisiae. Thus, three repressors, Nrg1, Mig1, and Mig2, are involved as the downstream targets of the glucose signaling in S. cerevisiae.

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Figures

Figure 1

Figure 1

Overexpressing a truncated clone of NRG1 suppresses snf2Δ. Yeast strains FY32 (snf2Δ1::HIS3 SUC2) and yHZ269 (snf2Δ1::HIS3 SUC2-36) were transformed with nrglΔZn or full-length NRG1 cloned in pRS426, as well as vector alone. Ura+ single colonies carrying each construct were resuspended in 200 μl sterile water, and spotted on SC-Ura plates containing glucose or raffinose as the carbon source. Plates were photographed on day 2.

Figure 2

Figure 2

Deletion of NRG1 partially abolishes glucose repression. nrg1Δ allows cells to grow on sucrose plates containing 2-deoxyglucose, and has additive effects with mig1Δ and mig2Δ. A single colony of each strain was inoculated into liquid YPD and grown to saturation (approx. 1 × 108 cells/ml). The cultures were then diluted 1:2 (upper panels) or 1:5 (lower panels) in sterile water, and spotted on YPD plates and YP sucrose plates with 200 μg/ml 2-deoxyglucose. Plates were photographed on after 1 and 2 days of incubation at 30°C.

Figure 3

Figure 3

Deletion of NRG1 causes defects in glucose repression. (A) A single colony of each strain was inoculated into YPD liquid with 2% glucose and grown to mid-log phase (approx. 1 × 107 cells/ml). The cells were harvested, and total RNA was isolated and analyzed by electrophoresis followed by hybridization with probes specific to SUC2 or SPT15. The intensities of each band was quantitated using phosphoimager and ImageQuant software. The amount of SUC2 mRNA in each strain was normalized to SPT15, and the result obtained for the wild-type strain was assigned the arbitrary unit of 1.0 and used to calculate the relative SUC2 mRNA levels in other strains. (B) Northern analysis of GAL1-10 mRNA in mutant strains. A single colony of each strain was inoculated into SD complete liquid with 2% glucose+2% galactose and grown to mid-log phase. The cells were harvested, and total RNA was isolated from each and analyzed by electrophoresis followed by hybridization with probes specific to GAL1, GAL10 or SPT15. Quantitation was carried out as for (A).

Figure 4

Figure 4

Mutations in SNF1 and SNF/SWI can be suppressed by both nrg1Δ and miglΔ. A single colony of each strain was inoculated into YPD liquid and grown over-night to saturation and adjusted in water to 1 × 108 cells/ml. The cultures were then diluted 1:2 in sterile water and spotted on YPD, YP galactose and YP sucrose plates, with uracil added to each plate to 80 μM. The first spot of each row represents a cell count of 5 × 107 cells/ml, which is diluted 1:4 for the second spot and 1:2 for each spot thereafter. YPD and YP sucrose plates were photographed after incubation at 30°C for 2 days, and YP galactose plates were photographed after 5 days.

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