In Vivo Phosphorylation of Ser21 and Ser83 during Nutrient-induced Activation of the Yeast Protein Kinase A (PKA) Target Trehalase

Schepers, Wim; Zeebroeck, Griet Van; Pinkse, Martijn W. H.; Verhaert, Peter; Thevelein, Johan M.

doi:10.1074/jbc.m112.421503

Cited by 68 publications

(76 citation statements)

References 62 publications

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“…Holt et al found Nth1-Ser66 to be the third-most strongly CDK-responsive residue in the proteome. PKA activates Nth1 by phosphorylation on Ser20, 21, 60 and 83 (Schepers et al, 2012; Veisova et al, 2012), so it seemed plausible that phosphorylation of Ser66 in the middle of this cluster of PKA sites might also activate Nth1. Likewise, three phosphoproteomic studies (Albuquerque et al, 2008; Soulard et al, 2010; Swaney et al, 2013) showed that Ser19 of Gph1 ( S PHQ) is phosphorylated in vivo .…”

Section: Resultsmentioning

confidence: 99%

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Cyclin-Dependent Kinase Co-Ordinates Carbohydrate Metabolism and Cell Cycle in S. cerevisiae

et al. 2016

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Summary Cyclin Dependent Kinases (CDKs) control cell division in eukaryotes by phosphorylating proteins involved in division. But successful proliferation requires co-ordination between division, and cellular growth in mass. Previous proteomic studies suggested that metabolic proteins, as well as cell division proteins, could potentially be substrates of cyclin dependent kinases. Here we focus on two metabolic enzymes of the yeast S. cerevisiae, neutral trehalase (Nth1) and glycogen phosphorylase (Gph1) and show that their activities are likely directly controlled by CDK activity, thus allowing co-ordinate regulation of carbohydrate metabolism with cell division processes. In this case, co-ordinate regulation may optimize the decision to undertake a final cell division as nutrients are being exhausted. Co-regulation of cell division processes and metabolic processes by CDK activity may be a general phenomenon important for co-ordinating the cell cycle with growth.

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Section: Resultsmentioning

confidence: 99%

“…At the same time, high PKA activity leads to Gph1 phosphorylation on T31, activating the enzyme and glycogen breakdown (Lin et al, 1996). PKA may also activate neutral trehalase (Nth1) by phosphorylating S20, S21, S60 and S83, with S60 and S83 apparently most important (Schepers et al, 2012; Veisova et al, 2012). …”

Section: Introductionmentioning

confidence: 99%

Cyclin-Dependent Kinase Co-Ordinates Carbohydrate Metabolism and Cell Cycle in S. cerevisiae

et al. 2016

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“…We used the well-established downstream target trehalase as a direct read-out for PKA activity 1522. This showed that very low iron concentrations, as low as 100 nM, in the form of FeCl 3 or FeCl 2 were able to trigger a rapid, transient increase in trehalase activity in cells starved for two days for iron using the BPS chelator (Fig.…”

Section: Resultsmentioning

confidence: 99%

Identification of Ftr1 and Zrt1 as iron and zinc micronutrient transceptors for activation of the PKA pathway in Saccharomyces cerevisiae

Schothorst

Zeebroeck

Thevelein³

2017

Microb Cell

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Multiple types of nutrient transceptors, membrane proteins that combine a transporter and receptor function, have now been established in a variety of organisms. However, so far all established transceptors utilize one of the macronutrients, glucose, amino acids, ammonium, nitrate, phosphate or sulfate, as substrate. This is also true for the Saccharomyces cerevisiae transceptors mediating activation of the PKA pathway upon re-addition of a macronutrient to glucose-repressed cells starved for that nutrient, re-establishing a fermentable growth medium. We now show that the yeast high-affinity iron transporter Ftr1 and high-affinity zinc transporter Zrt1 function as transceptors for the micronutrients iron and zinc. We show that replenishment of iron to iron-starved cells or zinc to zinc-starved cells triggers within 1-2 minutes a rapid surge in trehalase activity, a well-established PKA target. The activation with iron is dependent on Ftr1 and with zinc on Zrt1, and we show that it is independent of intracellular iron and zinc levels. Similar to the transceptors for macronutrients, Ftr1 and Zrt1 are strongly induced upon iron and zinc starvation, respectively, and they are rapidly downregulated by substrate-induced endocytosis. Our results suggest that transceptor-mediated signaling to the PKA pathway may occur in all cases where glucose-repressed yeast cells have been starved first for an essential nutrient, causing arrest of growth and low activity of the PKA pathway, and subsequently replenished with the lacking nutrient to re-establish a fermentable growth medium. The broadness of the phenomenon also makes it likely that nutrient transceptors use a common mechanism for signaling to the PKA pathway.

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“…The yeast Nth1, compared with trehalases from prokaryotic and higher eukaryotic organisms, exhibits distinct domain structure and regulation, as its activity is triggered by association with the 14-3-3 protein, which recognizes two phosphorylated motifs within the N-terminal extension of Nth1 (20)(21)(22)(23). Previous studies have suggested an allosteric mechanism, whereby 14-3-3 protein binding affects the structure of the catalytic domain of pNth1 , with Nth1-CaBD (the EF-hand-like motif-containing domain) being the intermediary between 14-3-3 and the catalytic domain (37,38).…”

Section: Discussionmentioning

confidence: 99%

“…1A), indicating significantly different regulation (17)(18)(19). It was subsequently shown that S. cerevisiae Nth1 is indeed activated in a PKA-and calcium-dependent manner in a process that requires interaction with the 14-3-3 protein through motifs containing phosphoserines pS60 and pS83 (20)(21)(22)(23). Significance 14-3-3 proteins are conserved scaffolding proteins expressed in all eukaryotic cells, where they regulate the function of several hundreds of partner proteins by constraining their conformation.…”

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confidence: 99%

Molecular basis of the 14-3-3 protein-dependent activation of yeast neutral trehalase Nth1

Alblova

Smidova

Dočekal

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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The 14-3-3 proteins, a family of highly conserved scaffolding proteins ubiquitously expressed in all eukaryotic cells, interact with and regulate the function of several hundreds of partner proteins. Yeast neutral trehalases (Nth), enzymes responsible for the hydrolysis of trehalose to glucose, compared with trehalases from other organisms, possess distinct structure and regulation involving phosphorylation at multiple sites followed by binding to the 14-3-3 protein. Here we report the crystal structures of yeast Nth1 and its complex with Bmh1 (yeast 14-3-3 isoform), which, together with mutational and fluorescence studies, indicate that the binding of Nth1 by 14-3-3 triggers Nth1’s activity by enabling the proper 3D configuration of Nth1’s catalytic and calcium-binding domains relative to each other, thus stabilizing the flexible part of the active site required for catalysis. The presented structure of the Bmh1:Nth1 complex highlights the ability of 14-3-3 to modulate the structure of a multidomain binding partner and to function as an allosteric effector. Furthermore, comparison of the Bmh1:Nth1 complex structure with those of 14-3-3:serotonin N-acetyltransferase and 14-3-3:heat shock protein beta-6 complexes revealed similarities in the 3D structures of bound partner proteins, suggesting the highly conserved nature of 14-3-3 affects the structures of many client proteins.

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In Vivo Phosphorylation of Ser21 and Ser83 during Nutrient-induced Activation of the Yeast Protein Kinase A (PKA) Target Trehalase

Cited by 68 publications

References 62 publications

Cyclin-Dependent Kinase Co-Ordinates Carbohydrate Metabolism and Cell Cycle in S. cerevisiae

Cyclin-Dependent Kinase Co-Ordinates Carbohydrate Metabolism and Cell Cycle in S. cerevisiae

Identification of Ftr1 and Zrt1 as iron and zinc micronutrient transceptors for activation of the PKA pathway in Saccharomyces cerevisiae

Molecular basis of the 14-3-3 protein-dependent activation of yeast neutral trehalase Nth1

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