Combining Deep Sequencing, Proteomics, Phosphoproteomics, and Functional Screens To Discover Novel Regulators of Sphingolipid Homeostasis

Lebesgue, Nicolas; Megyeri, Márton; Cristobal, Alba; Scholten, Arjen; Chuartzman, Silvia; Voichek, Yoav; Scheltema, Richard A.; Mohammed, Shabaz; Futerman, Anthony H.; Schuldiner, Maya; Heck, Albert J. R.; Lemeer, Simone

doi:10.1021/acs.jproteome.6b00691

“…Rtg1/3 likely also governs other steps in the production of sphingolipids because deletion of the rtg genes resulted in the alteration of the ratio of 42-and 44-carbon species in all complex sphingolipids ( Figure 1D). Most gene products known to be involved in sphingolipid biosynthesis appear to be regulated at post-translational levels in S. cerevisiae (Lebesgue et al, 2017). Consistent with this notion, the levels of the majority of C. albicans transcripts encoding sphingolipid biosynthesis enzymes remained unchanged after myriocin-induced sphingolipid depletion ( Figure 3A).…”

Section: Discussionsupporting

confidence: 69%

“…Our RNA-seq experiment comparing myriocin-treated versus untreated C. albicans cells identified 255 protein-coding transcripts that altered their expression in response to the drug (Àlog 10 p > 10 and expression changes >2-fold; Figure 3A; Table S3; 289 targets at p < 0.001 and expression changes >2-fold). Transcriptome studies in S. cerevisiae (Lebesgue et al, 2017;Liu et al, 2013) have found that myriocin does not cause major changes in the transcripts that encode enzymes mediating sphingolipid biosynthesis (it was in the phosphoproteome where relevant changes were more likely to occur; Lebesgue et al, 2017). Both Lebesgue et al (2017) and Liu et al, (2013) found that the predominant effect at the transcriptome level was in stress, ribosome, and RNA processing genes.…”

Section: Sphingolipid Depletion and Tor Inhibition Have Opposite Effementioning

confidence: 99%

“…Transcriptome studies in S. cerevisiae (Lebesgue et al, 2017;Liu et al, 2013) have found that myriocin does not cause major changes in the transcripts that encode enzymes mediating sphingolipid biosynthesis (it was in the phosphoproteome where relevant changes were more likely to occur; Lebesgue et al, 2017). Both Lebesgue et al (2017) and Liu et al, (2013) found that the predominant effect at the transcriptome level was in stress, ribosome, and RNA processing genes. We found that in C. albicans the only Gene Ontology (GO) terms overrepresented were mitosis and cell cycle (expression repressed by myriocin indicating that the drug causes cell cycle arrest) (p = 6.19 3 10 À18 ) ( Figure 3A).…”

Section: Sphingolipid Depletion and Tor Inhibition Have Opposite Effementioning

confidence: 99%

See 1 more Smart Citation

The Regulatory Proteins Rtg1/3 Govern Sphingolipid Homeostasis in the Human-Associated Yeast Candida albicans

Moreno-Velásquez

¹

,

Tint

²

,

Toledo

³

et al. 2020

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“…For example, proteomewide mass spectrometry studies have found that all three of the Gin4-related kinases are strongly hyperphosphorylated on multiple sites in rts1∆ cells, and that synthesis of ceramides influences Gin4 phosphorylation (Zapata et al 2014;Lebesgue et al 2017).…”

Section: Elm1 and The Gin4-related Kinases Do Not Influence Torc2 Sigmentioning

confidence: 99%

Modulation of TORC2 Signaling by a Conserved Lkb1 Signaling Axis in Budding Yeast

Alcaide-Gavilán

¹

,

Lucena

²

,

Schubert

³

et al. 2018

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Nutrient availability, growth rate and cell size are closely linked. For example, in budding yeast, the rate of cell growth is proportional to nutrient availability, cell size is proportional to growth rate, and growth rate is proportional to cell size. Thus, cells grow slowly in poor nutrients and are nearly half the size of cells growing in rich nutrients. Moreover, large cells grow faster than small cells. A signaling network that surrounds Tor kinase complex 2 (TORC2) plays an important role in enforcing these proportional relationships. Cells that lack components of the TORC2 network fail to modulate their growth rate or size in response to changes in nutrient availability. Here, we show that budding yeast homologs of the Lkb1 tumor suppressor kinase are required for normal modulation of TORC2 signaling in response to changes in carbon source. Lkb1 kinases activate Snf1/AMPK to initiate transcription of genes required for utilization of poor carbon sources. However, Lkb1 influences TORC2 signaling via a novel pathway that is independent of Snf1/AMPK. Of the three Lkb1 homologs in budding yeast, Elm1 plays the most important role in modulating TORC2. Elm1 activates a pair of related kinases called Gin4 and Hsl1. Previous work found that loss of Gin4 and Hsl1 causes cells to undergo unrestrained growth during a prolonged mitotic arrest, which suggests that they play a role in linking cell cycle progression to cell growth. We found that Gin4 and Hsl1 also control the TORC2 network. In addition, Gin4 and Hsl1 are themselves influenced by signals from the TORC2 network, consistent with previous work showing that the TORC2 network constitutes a feedback loop. Together, the data suggest a model in which the TORC2 network sets growth rate in response to carbon source, while also relaying signals via Gin4 and Hsl1 that set the critical amount of growth required for cell cycle progression. This kind of close linkage between control of cell growth and size would suggest a simple mechanistic explanation for the proportional relationship between cell size and growth rate.3

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“…Several observations suggest that the Gin4-related kinases are influenced by Rts1-dependent feedback signals from the TORC2 network. For example, proteomewide mass spectrometry studies have found that all three of the Gin4-related kinases are strongly hyperphosphorylated on multiple sites in rts1∆ cells, and that synthesis of ceramides influences Gin4 phosphorylation (Zapata et al 2014;Lebesgue et al 2017). In addition, the increased TORC2 signaling caused by rts1∆ is dependent upon the Gin4related kinases.…”

Section: Elm1 and The Gin4-related Kinases Do Not Influence Torc2 Sigmentioning

confidence: 99%

A conserved Lkb1 signaling axis modulates TORC2 signaling in budding yeast

Kellogg

¹

,

Lucena

²

,

Alcaide-Gavilán

³

et al. 2018

Preprint

1

0

View full text Add to dashboard Cite

Nutrient availability, growth rate and cell size are closely linked. For example, in budding yeast, the rate of cell growth is proportional to nutrient availability, cell size is proportional to growth rate, and growth rate is proportional to cell size. Thus, cells grow slowly in poor nutrients and are nearly half the size of cells growing in rich nutrients. Moreover, large cells grow faster than small cells. A signaling network that surrounds Tor kinase complex 2 (TORC2) plays an important role in enforcing these proportional relationships. Cells that lack components of the TORC2 network fail to modulate their growth rate or size in response to changes in nutrient availability. Here, we show that budding yeast homologs of the Lkb1 tumor suppressor kinase are required for normal modulation of TORC2 signaling and in response to changes in carbon source. Lkb1 kinases activate Snf1/AMPK to initiate transcription of genes required for utilization of poor carbon sources. However, Lkb1 influences TORC2 signaling via a novel pathway that is independent of Snf1/AMPK. Of the three Lkb1 homologs in budding yeast, Elm1 plays the most important role in modulating TORC2. Elm1 activates a pair of related kinases called Gin4 and Hsl1. Previous work found that loss of Gin4 and Hsl1 causes cells to undergo unrestrained growth during a prolonged mitotic arrest, which suggests that play a role in linking cell cycle progression to cell growth. We found that Gin4 and Hsl1 also control the TORC2 network. In addition, Gin4 and Hsl1 are themselves influenced by signals from the TORC2 network, consistent with previous work showing that the TORC2 network constitutes a feedback loop. Together, the data suggest a model in which the TORC2 network sets growth rate in response to carbon source, while also relaying signals via Gin4 and Hsl1 that set the critical amount of growth required for cell cycle progression. This kind of close linkage between control of cell growth and size would suggest a simple mechanistic explanation for the proportional relationship between cell size and growth rate. Growth is a defining feature of life, and always occurs at the level of cells. Mechanisms that control the rate, extent, location and timing of cell growth are responsible for the myriad forms of life that have been produced by evolution. Mechanisms that detect and limit growth during the cell cycle ultimately define the size of proliferating cells. Control of cell growth also requires mechanisms that coordinate the diverse processes of growth, including membrane growth and biogenesis, to ensure that the rate of each process is matched to the availability of building blocks.A number of observations have pointed to close relationships between nutrient availability, growth rate, and cell size. Limiting nutrients causes reduced growth rate as well as coordinated changes in the transcription of over a thousand genes, which suggests extensive coordination of growth-related processes (Brauer et al. 2008).

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Combining Deep Sequencing, Proteomics, Phosphoproteomics, and Functional Screens To Discover Novel Regulators of Sphingolipid Homeostasis

Cited by 12 publications

References 49 publications

The Regulatory Proteins Rtg1/3 Govern Sphingolipid Homeostasis in the Human-Associated Yeast Candida albicans

The Regulatory Proteins Rtg1/3 Govern Sphingolipid Homeostasis in the Human-Associated Yeast Candida albicans

Modulation of TORC2 Signaling by a Conserved Lkb1 Signaling Axis in Budding Yeast

A conserved Lkb1 signaling axis modulates TORC2 signaling in budding yeast

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