Nitrogen Availability and TOR Regulate the Snf1 Protein Kinase in Saccharomyces cerevisiae

Department of Biological Sciences, University of Wisconsin-Milwaukee, 3209 N. Maryland Ave., Milwaukee, WI 53211, USA.
Eukaryotic Cell (Impact Factor: 3.18). 12/2006; 5(11):1831-7. DOI: 10.1128/EC.00110-06
Source: PubMed


In the yeast Saccharomyces cerevisiae, the Snf1 protein kinase of the Snf1/AMP-activated protein kinase (AMPK) family regulates a wide range of responses to stress caused by glucose deprivation. The stress signal is relayed via upregulation of Snf1, which depends on phosphorylation of its activation loop Thr210 residue by upstream kinases. Although Snf1 is also required for coping with various stresses unrelated to glucose deprivation, some evidence suggests a role for low-level basal activity of unphosphorylated Snf1, rather than a specific signaling function. We previously found that Snf1 is required for diploid pseudohyphal differentiation, a developmental response to nitrogen limitation. Here, we present evidence that Snf1 is directly involved in nitrogen signaling. First, genetic analyses suggest that pseudohyphal differentiation depends on the stimulatory phosphorylation of Snf1 at Thr210. Second, immunochemical data indicate that nitrogen limitation improves Thr210 phosphorylation. Analyses of pseudohyphal differentiation in cells with catalytically inactive and hyperactive Snf1 support the role of Snf1 activity. Finally, we show that Snf1 is negatively regulated by the rapamycin-sensitive TOR kinase which plays essential roles in signaling nitrogen and amino acid availability. This and other evidence implicate Snf1 in the integration of signals regarding nitrogen and carbon stress. TOR and Snf1/AMPK are highly conserved in evolution, and their novel functional interaction in yeast suggests similar mechanisms in other eukaryotes.

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    • "However , rapamycin treatment inhibits rather than stimulates filamentation, even at sublethal doses, suggesting at most an indirect role of TORC1 in filamenation. Finally, Snf1 T210 phosphorylation is stimulated not only by glucose limitation but also by nitrogen limitation, even in the presence of high levels of glucose (Orlova et al. 2006, 2010). Thus, Snf1 activation provides the most consistent connection between filamentation and the multiple forms of nutritional deprivation required for filamentation. "
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    ABSTRACT: Availability of key nutrients, such as sugars, amino acids, and nitrogen compounds, dictates the developmental programs and the growth rates of yeast cells. A number of overlapping signaling networks-those centered on Ras/protein kinase A, AMP-activated kinase, and target of rapamycin complex I, for instance-inform cells on nutrient availability and influence the cells' transcriptional, translational, posttranslational, and metabolic profiles as well as their developmental decisions. Here I review our current understanding of the structures of the networks responsible for assessing the quantity and quality of carbon and nitrogen sources. I review how these signaling pathways impinge on transcriptional, metabolic, and developmental programs to optimize survival of cells under different environmental conditions. I highlight the profound knowledge we have gained on the structure of these signaling networks but also emphasize the limits of our current understanding of the dynamics of these signaling networks. Moreover, the conservation of these pathways has allowed us to extrapolate our finding with yeast to address issues of lifespan, cancer metabolism, and growth control in more complex organisms.
    Genetics 09/2012; 192(1):73-105. DOI:10.1534/genetics.111.135731 · 5.96 Impact Factor
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    • "Interestingly the highest suppression of the 3-AT toxicity we observed in the yeast strain lacking SNF1 gene but the N-terminal 108 codons [48] growing on agar media containing 10 mM 3-AT and glycerol as a carbon source. This has a sense, because Snf1 kinase is, besides other stimuli, activated during amino acid and generally nitrogen starvation [57], [58] and during the shift from glucose to glycerol media, when its relocation to the nucleus was also reported [59]. Further, the N-terminal 108 amino acids contain almost complete a structurally isolated β-rich lobe of the Snf1 kinase domain comprising of β1-3 strands and a well conserved regulatory αC helix including ATP-binding site and conserved residues Lys84 and Glu103. "
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    ABSTRACT: Interleukin-1α (IL-1α) is a proinflammatory cytokine and a key player in host immune responses in higher eukaryotes. IL-1α has pleiotropic effects on a wide range of cell types, and it has been extensively studied for its ability to contribute to various autoimmune and inflammation-linked disorders, including rheumatoid arthritis, Alzheimer's disease, systemic sclerosis and cardiovascular disorders. Interestingly, a significant proportion of IL-1α is translocated to the cell nucleus, in which it interacts with histone acetyltransferase complexes. Despite the importance of IL-1α, little is known regarding its binding targets and functions in the nucleus. We took advantage of the histone acetyltransferase (HAT) complexes being evolutionarily conserved from yeast to humans and the yeast SAGA complex serving as an epitome of the eukaryotic HAT complexes. Using gene knock-out technique and co-immunoprecipitation of the IL-1α precursor with TAP-tagged subunits of the yeast HAT complexes, we mapped the IL-1α-binding site to the HAT/Core module of the SAGA complex. We also predicted the 3-D structure of the IL-1α N-terminal domain, and by employing structure similarity searches, we found a similar structure in the C-terminal regulatory region of the catalytic subunit of the AMP-activated/Snf1 protein kinases, which interact with HAT complexes both in mammals and yeast, respectively. This finding is further supported with the ability of the IL-1α precursor to partially rescue growth defects of snf1Δ yeast strains on media containing 3-Amino-1,2,4-triazole (3-AT), a competitive inhibitor of His3. Finally, the careful evaluation of our data together with other published data in the field allows us to hypothesize a new function for the ADA complex in SAGA complex assembly.
    PLoS ONE 08/2012; 7(8):e41801. DOI:10.1371/journal.pone.0041801 · 3.23 Impact Factor
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    • "When glucose is unavailable, alternative carbon sources are used for the production of metabolic energy [15]. Snf1 is activated by glucose limitation by phosphorylation of the threonine 210 and by inactivation of Tor kinases [16] [17]. Gln3 phosphorylation and subcellular localization are regulated by glucose availability via the SNF1/AMPK pathway [18]. "
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    ABSTRACT: Yeast cells are able to adapt their metabolism according to the quality of both carbon and nitrogen sources available in the environment. Saccharomyces cerevisiae UGA4 gene encodes a permease capable of transporting γ-aminobutyric acid (GABA) into the cells. Yeast uses this amino acid as a nitrogen source or as a carbon skeleton that enters the tricarboxylic acid cycle. The quality of the carbon source modulates UGA4 expression through two parallel pathways, each one acting on different regulatory elements, the UAS(GATA) and the UAS(GABA). In the presence of a fermentable carbon source, UGA4 expression is induced by GABA while in the presence of a non-fermentable carbon source this expression is GABA-independent. The aim of this work was to study the mechanisms responsible for the differences in the profiles of UGA4 expression in both growth conditions. We found that although the subcellular localization of Gln3 depends on the carbon source and UGA4 expression depends on Tor1 and Snf1, Gln3 localization does not depend on these kinases. We also found that the phosphorylation of Gln3 is mediated by two systems activated by a non-fermentable carbon source, involving the Snf1 kinase and an unidentified TORC1-regulated kinase. We also found that the activity of the main transcription factors responsible for UGA4 induction by GABA varies depending on the quality of the carbon source. In a fermentable carbon source such as glucose, the negative GATA factor Dal80 binds to UGA4 promoter; only after the addition of the inducer, the positive factors Uga3, Dal81 and Gln3 interact with the promoter removing Dal80 and leading to gene induction. In contrast, in the non-fermentable carbon source acetate the negative GATA factor remains bound to UGA4 promoter in the presence or absence of GABA, the positive factors are not detected bound in any of these conditions and in consequence, UGA4 is not induced.
    Biochemical and Biophysical Research Communications 04/2012; 421(3):572-7. DOI:10.1016/j.bbrc.2012.04.047 · 2.30 Impact Factor
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