PP1 phosphatase-binding motif in Reg1 protein of Saccharomyces cerevisiae is required for interaction with both the PP1 phosphatase Glc7 and the Snf1 protein kinase

Tabba, Shadi; Mangat, Simmanjeet; McCartney, Rhonda R.; Schmidt, Martin C.

doi:10.1016/j.cellsig.2010.02.003

Cited by 26 publications

(36 citation statements)

References 33 publications

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“…However, control of localization is not likely to be the regulatory mechanism directing Reg1-Glc7 to the Snf1 complex, since Reg1 is predominantly cytoplasmic (12), while the substrates of interest, the different isoforms of Snf1, show distinct localization patterns (50). Similarly, Reg1 abundance does not change appreciably in response to the carbon source (46,49), so Reg1 abundance is not likely to be the determining mechanism. Posttranslational modification of Reg1 is a possible mechanism controlling substrate targeting.…”

Section: Discussionmentioning

confidence: 99%

“…Extracts were prepared in triplicate from cells shifted to low glucose, since these conditions gave the strongest Reg1-Snf1 interaction (46). Snf1 was collected by immunoprecipitation, and the abundance of associated Reg1 was determined by quantitative Western blotting (Fig.…”

Section: Vol 10 2011mentioning

confidence: 99%

“…However, not all the data support this model. The same point mutations in the Reg1 protein that disrupt binding to Glc7 also disrupt binding to Snf1, suggesting that these proteins may compete for binding to the same surface of Reg1 (46). If Glc7 and Snf1 compete for binding to the same site on Reg1, it is hard to imagine how Reg1 could target Glc7 to Snf1.…”

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confidence: 99%

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Reg1 Protein Regulates Phosphorylation of All Three Snf1 Isoforms but Preferentially Associates with the Gal83 Isoform

et al. 2011

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The phosphorylation status of the Snf1 activation loop threonine is determined by changes in the rate of its dephosphorylation, catalyzed by the yeast PP1 phosphatase Glc7 in complex with the Reg1 protein. Previous studies have shown that Reg1 can associate with both Snf1 and Glc7, suggesting substrate binding as a mechanism for Reg1-mediated targeting of Glc7. In this study, the association of Reg1 with the three Snf1 isoforms was measured by two-hybrid analysis and coimmunoprecipitation. We found that Reg1 association with Snf1 occurred almost exclusively with the Gal83 isoform of the Snf1 complex. Nonetheless, Reg1 plays an important role in determining the phosphorylation status of all three Snf1 isoforms. We found that the rate of dephosphorylation for isoforms of Snf1 did not correlate with the amount of associated Reg1 protein. Functional chimeric ␤ subunits containing residues from Gal83 and Sip2 were used to map the residues needed to promote Reg1 association with the N-terminal 150 residues of Gal83. The Gal83 isoform of Snf1 is the only isoform capable of nuclear localization. A Gal83-Sip2 chimera containing the first 150 residues of Gal83 was able to associate with the Reg1 protein but did not localize to the nucleus. Therefore, nuclear localization is not required for Reg1 association. Taken together, these data indicate that the ability of Reg1 to promote the dephosphorylation of Snf1 is not directly related to the strength of its association with the Snf1 complex.

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Section: Discussionmentioning

confidence: 99%

Section: Vol 10 2011mentioning

confidence: 99%

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Reg1 Protein Regulates Phosphorylation of All Three Snf1 Isoforms but Preferentially Associates with the Gal83 Isoform

et al. 2011

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“…This phosphatase maintains dephosphorylated Hxk2 and Mig1 proteins, which in this phosphorylation state have a nuclear localization (28,30). Glc7-Reg1 also dephosphorylates and therefore maintains the inactivated kinase Snf1 (39,66). Thus, under high glucose conditions it formed a repressor complex in the SUC2 promoter, which repress the gene expression.…”

Section: Discussionmentioning

confidence: 99%

Hexokinase 2 Is an Intracellular Glucose Sensor of Yeast Cells That Maintains the Structure and Activity of Mig1 Protein Repressor Complex

Vega¹,

Riera²,

Fernández-Cid³

et al. 2016

Journal of Biological Chemistry

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Hexokinase 2 (Hxk2) from Saccharomyces cerevisiae is a bifunctional enzyme, being both a catalyst in the cytosol and an important regulator of the glucose repression signal in the nucleus. Despite considerable recent progress, little is known about the regulatory mechanism that controls nuclear Hxk2 association with the SUC2 promoter chromatin and how this association is necessary for SUC2 gene repression. Our data indicate that in the SUC2 promoter context, Hxk2 functions through a variety of structurally unrelated factors, mainly the DNA-binding Mig1 and Mig2 repressors and the regulatory Snf1 and Reg1 factors. Hxk2 sustains the repressor complex architecture maintaining transcriptional repression at the SUC2 gene. Using chromatin immunoprecipitation assays, we discovered that the Hxk2 in its open configuration, at low glucose conditions, leaves the repressor complex that induces its dissociation and promotes SUC2 gene expression. In high glucose conditions, Hxk2 adopts a close conformation that promotes Hxk2 binding to the Mig1 protein and the reassembly of the SUC2 repressor complex. Additional findings highlight the possibility that Hxk2 constitutes an intracellular glucose sensor that operates by changing its conformation in response to cytoplasmic glucose levels that regulate its interaction with Mig1 and thus its recruitment to the repressor complex of the SUC2 promoter. Thus, our data indicate that Hxk2 is more intimately involved in gene regulation than previously thought.

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“…Since many mutants have different growth rates, we plot cell growth as a percentage of growth relative to cultures with no 2DG. This assay yields a reproducible measurement of 2DG toxicity and resistance (Tabba et al 2010). Here, we examined the ability of 2DG to inhibit yeast cell growth in liquid cultures as a function of carbon source.…”

Section: Dg Toxicity Is Carbon Source Dependentmentioning

confidence: 99%

Genetic Analysis of Resistance and Sensitivity to 2-Deoxyglucose in Saccharomyces cerevisiae

McCartney¹,

Chandrashekarappa²,

Zhang³

et al. 2014

Genetics

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Aerobic glycolysis is a metabolic pathway utilized by human cancer cells and also by yeast cells when they ferment glucose to ethanol. Both cancer cells and yeast cells are inhibited by the presence of low concentrations of 2-deoxyglucose (2DG). Genetic screens in yeast used resistance to 2-deoxyglucose to identify a small set of genes that function in regulating glucose metabolism. A recent high throughput screen for 2-deoxyglucose resistance identified a much larger set of seemingly unrelated genes. Here, we demonstrate that these newly identified genes do not in fact confer significant resistance to 2-deoxyglucose. Further, we show that the relative toxicity of 2-deoxyglucose is carbon source dependent, as is the resistance conferred by gene deletions. Snf1 kinase, the AMPactivated protein kinase of yeast, is required for 2-deoxyglucose resistance in cells growing on glucose. Mutations in the SNF1 gene that reduce kinase activity render cells hypersensitive to 2-deoxyglucose, while an activating mutation in SNF1 confers 2-deoxyglucose resistance. Snf1 kinase activated by 2-deoxyglucose does not phosphorylate the Mig1 protein, a known Snf1 substrate during glucose limitation. Thus, different stimuli elicit distinct responses from the Snf1 kinase.

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PP1 phosphatase-binding motif in Reg1 protein of Saccharomyces cerevisiae is required for interaction with both the PP1 phosphatase Glc7 and the Snf1 protein kinase

Cited by 26 publications

References 33 publications

Reg1 Protein Regulates Phosphorylation of All Three Snf1 Isoforms but Preferentially Associates with the Gal83 Isoform

Reg1 Protein Regulates Phosphorylation of All Three Snf1 Isoforms but Preferentially Associates with the Gal83 Isoform

Hexokinase 2 Is an Intracellular Glucose Sensor of Yeast Cells That Maintains the Structure and Activity of Mig1 Protein Repressor Complex

Genetic Analysis of Resistance and Sensitivity to 2-Deoxyglucose in Saccharomyces cerevisiae

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