1. Biochemistry and Chemical Biology
  2. Cell Biology

Down-regulation of TORC2-Ypk1 signaling promotes MAPK-independent survival under hyperosmotic stress

  1. Alexander Muir
  2. Françoise M Roelants
  3. Garrett Timmons
  4. Kristin L Leskoske
  5. Jeremy Thorner  Is a corresponding author
  1. Massachusetts Institute of Technology, United Kingdom
  2. University of California, Berkeley, United States
https://doi.org/10.7554/eLife.09336
Share this article
Cite this article
  1. Alexander Muir
  2. Françoise M Roelants
  3. Garrett Timmons
  4. Kristin L Leskoske
  5. Jeremy Thorner
(2015)
Down-regulation of TORC2-Ypk1 signaling promotes MAPK-independent survival under hyperosmotic stress
eLife 4:e09336.
https://doi.org/10.7554/eLife.09336
  2. Download BibTeX
  3. Download .RIS

Abstract

In eukaryotes, exposure to hypertonic conditions activates a MAPK (Hog1 in S.cerevisiae and ortholog p38 in human cells). In yeast, intracellular glycerol accumulates to counterbalance the high external osmolarity. To prevent glycerol efflux, Hog1 action impedes the function of the aquaglyceroporin Fps1, in part, by displacing channel co-activators (Rgc1/2). However, Fps1 closes upon hyperosmotic shock even in hog1∆ cells, indicating another mechanism to prevent Fps1-mediated glycerol efflux. In our prior proteome-wide screen, Fps1 was identified as a target of TORC2-dependent protein kinase Ypk1 (Muir et al., 2014). We show here that Fps1 is an authentic Ypk1 substrate and that the open channel state of Fps1 requires phosphorylation by Ypk1. Moreover, hyperosmotic conditions block TORC2-dependent Ypk1-mediated Fps1 phosphorylation, causing channel closure, glycerol accumulation, and enhanced survival under hyperosmotic stress. These events are all Hog1-independent. Our findings define the underlying molecular basis of a new mechanism for responding to hypertonic conditions.

Article and author information

Author details

  1. Alexander Muir

    Vander Heiden Lab, Department of Biology and Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.

  2. Françoise M Roelants

    Division of Biochemistry, Biophysics and Structural Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Garrett Timmons

    Department of Chemistry, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Kristin L Leskoske

    Division of Biochemistry, Biophysics and Structural Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Jeremy Thorner

    Division of Biochemistry, Biophysics and Structural Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    For correspondence
    jthorner@berkeley.edu
    Competing interests
    The authors declare that no competing interests exist.

Copyright

© 2015, Muir et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 2,411
    views
  • 564
    downloads
  • 46
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Alexander Muir
  2. Françoise M Roelants
  3. Garrett Timmons
  4. Kristin L Leskoske
  5. Jeremy Thorner
(2015)
Down-regulation of TORC2-Ypk1 signaling promotes MAPK-independent survival under hyperosmotic stress
eLife 4:e09336.
https://doi.org/10.7554/eLife.09336

Share this article

https://doi.org/10.7554/eLife.09336

Categories and tags

Research organism

Further reading

    1. Biochemistry and Chemical Biology
    2. Cell Biology

    TORC2-dependent protein kinase Ypk1 phosphorylates ceramide synthase to stimulate synthesis of complex sphingolipids

    Alexander Muir, Subramaniam Ramachandran ... Jeremy Thorner
    Research Article Updated

    Plasma membrane lipid composition must be maintained during growth and under environmental insult. In yeast, signaling mediated by TOR Complex 2 (TORC2)-dependent protein kinase Ypk1 controls lipid abundance and distribution in response to membrane stress. Ypk1, among other actions, alleviates negative regulation of L-serine:palmitoyl-CoA acyltransferase, upregulating production of long-chain base precursors to sphingolipids. To explore other roles for TORC2-Ypk1 signaling in membrane homeostasis, we devised a three-tiered genome-wide screen to identify additional Ypk1 substrates, which pinpointed both catalytic subunits of the ceramide synthase complex. Ypk1-dependent phosphorylation of both proteins increased upon either sphingolipid depletion or heat shock and was important for cell survival. Sphingolipidomics, other biochemical measurements and genetic analysis demonstrated that these modifications of ceramide synthase increased its specific activity and stimulated channeling of long-chain base precursors into sphingolipid end-products. Control at this branch point also prevents accumulation of intermediates that could compromise cell growth by stimulating autophagy.

    1. Biochemistry and Chemical Biology
    2. Cancer Biology

    The deubiquitinase Ubp3/Usp10 constrains glucose-mediated mitochondrial repression via phosphate budgeting

    Vineeth Vengayil, Shreyas Niphadkar ... Sunil Laxman
    Research Article

    Many cells in high glucose repress mitochondrial respiration, as observed in the Crabtree and Warburg effects. Our understanding of biochemical constraints for mitochondrial activation is limited. Using a Saccharomyces cerevisiae screen, we identified the conserved deubiquitinase Ubp3 (Usp10), as necessary for mitochondrial repression. Ubp3 mutants have increased mitochondrial activity despite abundant glucose, along with decreased glycolytic enzymes, and a rewired glucose metabolic network with increased trehalose production. Utilizing ∆ubp3 cells, along with orthogonal approaches, we establish that the high glycolytic flux in glucose continuously consumes free Pi. This restricts mitochondrial access to inorganic phosphate (Pi), and prevents mitochondrial activation. Contrastingly, rewired glucose metabolism with enhanced trehalose production and reduced GAPDH (as in ∆ubp3 cells) restores Pi. This collectively results in increased mitochondrial Pi and derepression, while restricting mitochondrial Pi transport prevents activation. We therefore suggest that glycolytic flux-dependent intracellular Pi budgeting is a key constraint for mitochondrial repression.

    1. Biochemistry and Chemical Biology

    Immunotherapy: The power lies in the glycans

    Luca Unione, Jesús Jiménez-Barbero
    Insight

    Glycans play an important role in modulating the interactions between natural killer cells and antibodies to fight pathogens and harmful cells.