Snf1 Phosphorylates Adenylate Cyclase and Negatively Regulates Protein Kinase A-dependent Transcription in Saccharomyces cerevisiae

Nicastro, Raffaele; Tripodi, Farida; Gaggini, Marco; Castoldi, Andrea; Reghellin, Veronica; Nonnis, Simona; Tedeschi, Gioacchino; Coccetti, Paola

doi:10.1074/jbc.m115.658005

Cited by 56 publications

(53 citation statements)

References 59 publications

(70 reference statements)

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“…Of particular note, we identified direct phosphorylation sites on the well-known Snf1 targets Mig1 (63, 64), Hxk1/2 (65,66), and Cat8 (67, 68), including previously undocumented sites. We also observed phosphorylation on S241 of adenylate cyclase, which has recently been reported to be a direct Snf1 substrate (69,70). We found both direct (Sip1) and indirect (Sip1, Gal83, and Sip2) phosphorylation on all ␤-subunits of SNF1 complex, which play a role in determining Snf1 substrate specificity and subcellular localization (24).…”

Section: Inhibition Of Cdc28-as Results In Phosphorylation and Dephossupporting

confidence: 71%

Machine Learning of Global Phosphoproteomic Profiles Enables Discrimination of Direct versus Indirect Kinase Substrates

Kanshin

Giguère

Cheng

et al. 2017

Molecular & Cellular Proteomics

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Mass spectrometry allows quantification of tens of thousands of phosphorylation sites from minute amounts of cellular material. Despite this wealth of information, our understanding of phosphorylation-based signaling is limited, in part because it is not possible to deconvolute substrate phosphorylation that is directly mediated by a particular kinase phosphorylation that is mediated by downstream kinases. Here, we describe a framework for assignment of direct kinase substrates using a combination of selective chemical inhibition, quantitative phosphoproteomics, and machine learning techniques. Our workflow allows classification of phosphorylation events following inhibition of an analog-sensitive kinase into kinase-independent effects of the inhibitor, direct effects on cognate substrates, and indirect effects mediated by downstream kinases or phosphatases. We applied this method to identify many direct targets of Cdc28 and Snf1 kinases in the budding yeast Global phosphoproteome analysis of acute time-series demonstrated that dephosphorylation of direct kinase substrates occurs more rapidly compared with indirect substrates, both after inhibitor treatment and under a physiological nutrient shift in cells. Mutagenesis experiments revealed a high proportion of functionally relevant phosphorylation sites on Snf1 targets. For example, Snf1 itself was inhibited through autophosphorylation on Ser and new phosphosites were discovered that modulate the activity of the Reg1 regulatory subunit of the Glc7 phosphatase and the Gal83 β-subunit of SNF1 complex. This methodology applies to any kinase for which a functional analog sensitive version can be constructed to facilitate the dissection of the global phosphorylation network.

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Section: Inhibition Of Cdc28-as Results In Phosphorylation and Dephossupporting

confidence: 71%

Machine Learning of Global Phosphoproteomic Profiles Enables Discrimination of Direct versus Indirect Kinase Substrates

Kanshin

Giguère

Cheng

et al. 2017

Molecular & Cellular Proteomics

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“…This is supported by our GSEA analysis that indicated a significant similarity between the transcriptional profile of snf5 and mutant of the kinase Sak1 that regulates the activity of Snf1 [53] ( Figure 7A ). Furthermore, in S. cerevisiae , Cyr1 is phosphorylated by the Snf1 to promote ATP homeostasis [54], a mechanism that might be conserved in C. albicans . Together, this led to hypothesize that in C. albicans , Snf1 might be activated under hypoxia to accommodate the metabolic demand and to promote the flexibility of carbon utilization through Cyr1-Snf5 axis.…”

Section: Resultsmentioning

confidence: 99%

A novel genetic circuitry governing hypoxic metabolic flexibility, commensalism and virulence in the fungal pathogen Candida albicans

Burgain

Pic

Markey

et al. 2019

Preprint

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Inside the human host, the pathogenic yeast Candida albicans colonizes predominantly oxygen-poor niches such as the gastrointestinal and vaginal tracts, but also oxygen-rich environments such as cutaneous epithelial cells and oral mucosa. This suppleness requires an effective mechanism to reprogram reversibly the primary metabolism in response to oxygen variation. Here, we have uncovered that Snf5, a subunit of SWI/SNF chromatin remodeling complex, is a major transcriptional regulator that links oxygen status to the metabolic capacity of C. albicans. Snf5 and other subunits of SWI/SNF complex were required to activate genes of alternative carbon utilization and other carbohydrates related process specifically under hypoxia. snf5 mutant exhibited an altered metabolome reflecting that SWI/SNF plays an essential role in maintaining metabolic homeostasis and carbon flux in C. albicans under hypoxia. Snf5 was necessary to activate the transcriptional program linked to both commensal and invasive growth. Accordingly, snf5 was unable to maintain its growth in the stomach, the cecum and the colon of mice. snf5 was also avirulent as it was unable to invade Galleria larvae or to cause damage to human enterocytes and murine macrophages. Among candidates of signaling pathways in which Snf5 might operate, phenotypic analysis revealed that mutants of Ras1-cAMP-PKA pathway, as well as mutants of Yak1 and Yck2 kinases exhibited a similar carbon flexibility phenotype as did snf5 under hypoxia. Genetic interaction analysis indicated that the adenylate cyclase Cyr1, a key component of the Ras1-cAMP pathway, but not Ras1, interacted genetically with Snf5. Our study yielded unprecedented insight into the oxygen-sensitive regulatory circuit that control metabolic flexibility, stress, commensalism and virulence in C. albicans.Author SummaryA critical aspect of eukaryotic cell fitness is the ability to sense and adapt to variations in oxygen concentrations in their local environment. Hypoxia leads to a substantial remodeling of cell metabolism and energy homeostasis, and thus, organisms must develop an effective regulatory mechanism to cope with oxygen depletion. Candida albicans is an opportunistic yeast that is the most prevalent human fungal pathogens. This yeast colonizes diverse niches inside the human host with contrasting carbon sources and oxygen concentrations. While hypoxia is the predominant conditions that C. albicans encounters inside most of the niches, the impact of this condition on metabolic flexibility, a major determinant of fungal virulence, was completely neglected. Here, we uncovered that the chromatin remodelling complex SWI/SNF is master regulatory circuit that links oxygen status to a broad spectrum of carbon utilization routes. Snf5 was essential for the maintenance of C. albicans as a commensal and also for the expression of its virulence. The oxygen-sensitive regulators identified in this work provide a framework to comprehensively understand the virulence of human fungal pathogens and represent a therapeutic value to fight fungal infections.

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“…The dephosphorylation of Snf1 through G1/S may be important because when Snf1 is activated (like AMPK in mammals) it acts as a "brake pedal" slowing growth and energy consuming processes (Ghillebert et al, 2011;Coccetti et al, 2018). Thus, Snf1 inactivates many processes typically activated by PKA (Nicastro et al, 2015). During entry into the cell cycle at G1/S, phosphoregulation may shift the balance between PKA and Snf1 to enhance growth promoting pathways and rewire metabolism to turn storage compounds such as trehalose, glycogen, or lipid droplets into macromolecules that support cell cycle progression ( Figure 8B).…”

Section: Discussionmentioning

confidence: 99%

Multiple Layers of Phospho-Regulation Coordinate Metabolism and the Cell Cycle in Budding Yeast

Zhang

Winkler

Schlottmann

et al. 2019

Front. Cell Dev. Biol.

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Snf1 Phosphorylates Adenylate Cyclase and Negatively Regulates Protein Kinase A-dependent Transcription in Saccharomyces cerevisiae

Cited by 56 publications

References 59 publications

Machine Learning of Global Phosphoproteomic Profiles Enables Discrimination of Direct versus Indirect Kinase Substrates

Machine Learning of Global Phosphoproteomic Profiles Enables Discrimination of Direct versus Indirect Kinase Substrates

A novel genetic circuitry governing hypoxic metabolic flexibility, commensalism and virulence in the fungal pathogen Candida albicans

Multiple Layers of Phospho-Regulation Coordinate Metabolism and the Cell Cycle in Budding Yeast

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