The Saccharomyces cerevisiae AMPK, Snf1, Negatively Regulates the Hog1 MAPK Pathway in ER Stress Response

Mizuno, Tomoaki; Masuda, Y.; Irie, Kenji

doi:10.1371/journal.pgen.1005491

Cited by 49 publications

(76 citation statements)

References 40 publications

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“…and Piao et al . reported recently that ScSnf1 could modulate the activity of ScHog1 through its upstream kinase ScSsk1, which is independent of the classical osmosensing pathway [25,26]. In our screening, we identified PpSnf1 subunit PpGal83 ( PAS_chr1-4_0498 ) and PpHog1 ( PAS_chr1-3_0232 ) involved in positive and negative regulation of P AOX1 respectively.…”

Section: Discussionmentioning

confidence: 92%

Kinase Screening in Pichia pastoris Identified Promising Targets Involved in Cell Growth and Alcohol Oxidase 1 Promoter (PAOX1) Regulation

et al. 2016

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As one of the most commonly used eukaryotic recombinant protein expression systems, P. pastoris relies heavily on the AOX1 promoter (PAOX1), which is strongly induced by methanol but strictly repressed by glycerol and glucose. However, the complicated signaling pathways involved in PAOX1 regulation when supplemented with different carbon sources are poorly understood. Here we constructed a kinase deletion library in P. pastoris and identified 27 mutants which showed peculiar phenotypes in cell growth or PAOX1 regulation. We analyzed both annotations and possible functions of these 27 targets, and then focused on the MAP kinase Hog1. In order to locate its potential downstream components, we performed the phosphoproteome analysis on glycerol cultured WT and Δhog1 strains and identified 157 differentially phosphorylated proteins. Our results identified important kinases involved in P. pastoris cell growth and PAOX1 regulation, which could serve as valuable targets for further mechanistic studies.

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Section: Discussionmentioning

confidence: 92%

Kinase Screening in Pichia pastoris Identified Promising Targets Involved in Cell Growth and Alcohol Oxidase 1 Promoter (PAOX1) Regulation

et al. 2016

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“…1a). This reg1 defect could be significantly restored by snf1 mutation, while cells harboring snf1 single mutation only exhibited a mild defect in HAC1 mRNA splicing under unstressed conditions19. Thus, alteration of the kinetics of HAC1 mRNA splicing caused by reg1 mutation was expected to be highly sensitive to reduction of Snf1 function.…”

Section: Resultsmentioning

confidence: 96%

“…We have revealed Snf1 as a negative regulator of the UPR pathway and the Hog1 MAPK pathway in ER stress response19. The deletion of the SNF1 gene caused increased resistance to ER stress.…”

mentioning

confidence: 99%

Expression control of the AMPK regulatory subunit and its functional significance in yeast ER stress response

Kimura

Irie

Mizuno

2017

Sci Rep

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AMP-activated protein kinase (AMPK) is an evolutionarily conserved heterotrimeric kinase complex consisting of a catalytic subunit, α, and two regulatory subunits, β and γ. Previously, we demonstrated that Snf1, the Saccharomyces cerevisiae ortholog of AMPK, negatively regulates the unfolded protein response (UPR) pathway and the Hog1 MAP kinase pathway in ER stress response. However, it remains unclear how the alternate three β subunits, Sip1, Sip2, and Gal83, of the Snf1 complex participate in ER stress response. Here, we show that Gal83 plays a major role in Snf1-mediated downregulation of the UPR and Hog1 pathways. Gal83 is the most abundant β subunit in the normal state and further induced by ER stress. This induction is mediated via activation of the GAL83 promoter by the UPR. When expressed under the control of the GAL83 promoter, Sip2 exhibits potent functional activity equivalent to Gal83. Our results suggest that the functional significance of the β subunit of Snf1 AMPK in ER stress response is defined by modulation of the expression level through regulation of the promoter activity.

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“…We generated GST fusions to the Pho85 and Ssn3 cyclin dependent kinases and to the Hog1 MAP kinase. We chose these three kinases since each had documented genetic and/or biochemical interactions with Snf1 (34–37). For substrates, we purified from bacteria the unphosphorylated Snf1 heterotrimer with different beta subunits, including two deletions mutants of the Gal83 protein (Fig.…”

Section: Resultsmentioning

confidence: 99%

Activation and inhibition of Snf1 kinase activity by phosphorylation within the activation loop

McCartney

Garnar-Wortzel

Chandrashekarappa

et al. 2016

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

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The AMP-activated protein kinase is a metabolic regulator that transduces information about energy and nutrient availability. In yeast, the AMP-activated protein kinase, called Snf1, is activated when energy and nutrients are scarce. Earlier studies have demonstrated that activation of Snf1 requires the phosphorylation of the activation loop on threonine 210. Here we examined the regulation of Snf1 kinase activity in response to phosphorylation at other sites. Phosphoproteomic studies have identified numerous phosphorylation sites within the Snf1 kinase enzyme. We made amino acid substitutions in the Snf1 protein that were either non-phosphorylatable (serine to alanine) or phospho-mimetic (serine to glutamate) and examined the effects of these changes on Snf1 kinase function in vivo and on its catalytic activity in vitro. We found that changes to most of the phosphorylation sites had no effect on Snf1 kinase function. However, changes to serine 214, a site within the kinase activation loop, inhibited Snf1 kinase activity. Snf1-activating kinase 1 still phosphorylates Snf1-S214E on threonine 210 but the S214E enzyme is non-functional in vivo and catalytically inactive in vitro. We conclude that yeast have developed two distinct pathways for down-regulating Snf1 activity. The first is through direct dephosphorylation of the conserved activation loop threonine. The second is through phosphorylation of serine 214.

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The Saccharomyces cerevisiae AMPK, Snf1, Negatively Regulates the Hog1 MAPK Pathway in ER Stress Response

Cited by 49 publications

References 40 publications

Kinase Screening in Pichia pastoris Identified Promising Targets Involved in Cell Growth and Alcohol Oxidase 1 Promoter (PAOX1) Regulation

Kinase Screening in Pichia pastoris Identified Promising Targets Involved in Cell Growth and Alcohol Oxidase 1 Promoter (PAOX1) Regulation

Expression control of the AMPK regulatory subunit and its functional significance in yeast ER stress response

Activation and inhibition of Snf1 kinase activity by phosphorylation within the activation loop

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