Rapid Glucose Depletion Immobilizes Active Myosin V on Stabilized Actin Cables

Xu, Li Chong; Bretscher, Anthony

doi:10.1016/j.cub.2014.09.017

Cited by 19 publications

(34 citation statements)

References 32 publications

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“…To compare our findings with published data on actin distribution in fixed 9 and live glucose-starving cells 2425 we employed image analyses of wild-type rho + (respiring) cells expressing established fluorescence markers of the two different F-actin structures patches and cables (Abp1-RFP and Abp140-GFP). Both the glucose-grown and the glucose-depleted cells were fixed with 3.7 % formaldehyde for 30 minutes, and changes in distribution of both markers Abp1-RFP and Abp140-GFP were analyzed (Fig.…”

Section: Resultsmentioning

confidence: 96%

Formaldehyde fixation is detrimental to actin cables in glucose-depleted S. cerevisiae cells

Vašicová¹,

Rinnerthaler²,

Haskova³

et al. 2016

MIC

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Actin filaments form cortical patches and emanating cables in fermenting cells of Saccharomyces cerevisiae. This pattern has been shown to be depolarized in glucose-depleted cells after formaldehyde fixation and staining with rhodamine-tagged phalloidin. Loss of actin cables in mother cells was remarkable. Here we extend our knowledge on actin in live glucose-depleted cells co-expressing the marker of actin patches (Abp1-RFP) with the marker of actin cables (Abp140-GFP). Glucose depletion resulted in appearance of actin patches also in mother cells. However, even after 80 min of glucose deprivation these cells showed a clear network of actin cables labeled with Abp140-GFP in contrast to previously published data. In live cells with a mitochondrial dysfunction (rho0 cells), glucose depletion resulted in almost immediate appearance of Abp140-GFP foci partially overlapping with Abp1-RFP patches in mother cells. Residual actin cables were clustered in patch-associated bundles. A similar overlapping “patchy” pattern of both actin markers was observed upon treatment of glucose-deprived rho+ cells with FCCP (the inhibitor of oxidative phosphorylation) and upon treatment with formaldehyde. While the formaldehyde-targeted process stays unknown, our results indicate that published data on yeast actin cytoskeleton obtained from glucose-depleted cells after fixation should be considered with caution.

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Section: Resultsmentioning

confidence: 96%

Formaldehyde fixation is detrimental to actin cables in glucose-depleted S. cerevisiae cells

Vašicová¹,

Rinnerthaler²,

Haskova³

et al. 2016

MIC

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“…First, we tested if the QUEEN ratio, indeed, reflects the cellular concentration of ATP in living yeast cells. A previous biochemical study has demonstrated that ATP levels in budding yeast cells dropped to 15% upon glucose depletion and gradually recovered to 50% of the original level within 20 min (Xu and Bretscher, 2014). Moreover, replacement of glucose in medium with 2-deoxy-Dglucose (2DG), which strongly inhibits glycolysis, reduced cellular ATP levels to <1% at least for 30 min (Serrano, 1977;Xu and Bretscher, 2014).…”

Section: Queen Is a Reliable Atp Biosensor In Budding Yeast Cellsmentioning

confidence: 99%

“…Second, we tested if QUEEN can reversibly monitor a change in ATP concentration. We took advantage of gph1Δ mutant cells that cannot catabolize glycogen and do not show any recovery of ATP after glucose depletion (Xu and Bretscher, 2014). The mean QUEEN ratios in glucose-depleted gph1Δ cells declined rapidly within 10-15 min, but re-feeding of glucose fully recovered these values within a minute ( Fig.…”

Section: Queen Is a Reliable Atp Biosensor In Budding Yeast Cellsmentioning

confidence: 99%

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Reliable imaging of ATP in living budding and fission yeast

Takaine

Ueno

Kitamura

et al. 2019

Journal of Cell Science

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Adenosine triphosphate (ATP) is a main metabolite essential for all living organisms. However, our understanding of ATP dynamics within a single living cell is very limited. Here, we optimized the ATP-biosensor QUEEN and monitored the dynamics of ATP with good spatial and temporal resolution in living yeasts. We found stable maintenance of ATP concentration in wild-type yeasts, regardless of carbon sources or cell cycle stages, suggesting that mechanism exists to maintain ATP at a specific concentration. We further found that ATP concentration is not necessarily an indicator of metabolic activity, as there is no clear correlation between ATP level and growth rates. During fission yeast meiosis, we found a reduction in ATP levels, suggesting that ATP homeostasis is controlled by differentiation. The use of QUEEN in yeasts offers an easy and reliable assay for ATP dynamicity and will answer several unaddressed questions about cellular metabolism in eukaryotes.

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“…The work of Yu et al revealed the different dynamics and motility of cables in polarized and unpolarized cells in the exponential phase (30). Additionally, in vivo observations of cables under conditions of acute glucose depletion revealed the stabilization of cables (31), albeit previous observations of fixed cells detected the persistence of depolarized patches only (32). This in vivo approach has not yet been used for monitoring of actin cables in post-diauxic and stationary phases.…”

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confidence: 99%

The Stationary-Phase Cells of Saccharomyces cerevisiae Display Dynamic Actin Filaments Required for Processes Extending Chronological Life Span

Vašicová

Lejskova

Malcová

et al. 2015

Molecular and Cellular Biology

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Stationary-growth-phase Saccharomyces cerevisiae yeast cultures consist of nondividing cells that undergo chronological aging. For their successful survival, the turnover of proteins and organelles, ensured by autophagy and the activation of mitochondria, is performed. Some of these processes are engaged in by the actin cytoskeleton. In S. cerevisiae stationary-phase cells, F actin has been shown to form static aggregates named actin bodies, subsequently cited to be markers of quiescence. Our in vivo analyses revealed that stationary-phase cultures contain cells with dynamic actin filaments, besides the cells with static actin bodies. The cells with dynamic actin displayed active endocytosis and autophagy and well-developed mitochondrial networks. Even more, stationary-phase cell cultures grown under calorie restriction predominantly contained cells with actin cables, confirming that the presence of actin cables is linked to successful adaptation to stationary phase. Cells with actin bodies were inactive in endocytosis and autophagy and displayed aberrations in mitochondrial networks. Notably, cells of the respiratory activity-deficient cox4⌬ strain displayed the same mitochondrial aberrations and actin bodies only. Additionally, our results indicate that mitochondrial dysfunction precedes the formation of actin bodies and the appearance of actin bodies corresponds to decreased cell fitness. We conclude that the F-actin status reflects the extent of damage that arises from exponential growth.O nce glucose and other external nutrients are exhausted, cell division stops and Saccharomyces cerevisiae cells are able to survive for extended periods of time. This period of survival has been termed chronological aging and has become a model for aging of postmitotic tissues (1, 2). The cells in these nondividing stationary-phase cell cultures are often termed quiescent (Q) cells (3, 4). Some authors claim that stationary-phase yeast cell populations are heterogeneous and only a portion of them have characteristics of quiescence (5, 6). The ability to survive the period of scarcity of external nutrition and reproduce again upon refeeding is influenced by several life span-extending genetic and environmental interventions. One of the most cited is calorie restriction (CR) (7).In general, cells that are in a nutrient-poor environment activate processes that help them to efficiently utilize inner resources and thus prolong the life span. A catabolic process that has a positive impact on chronological aging is autophagy, which provides nutrients by the vacuolar degradation of damaged or superfluous macromolecules and organelles (8, 9). In addition, Fabrizio et al. demonstrated that the deletion of several genes encoding endosomal functions also shortens the life span (8). Note that some of them have not been directly implicated in autophagy (8). The efficient utilization of resources is ensured by the activation of mitochondrial respiration. It has been proved that the utilization of carbohydrate stores by respiration instead...

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Rapid Glucose Depletion Immobilizes Active Myosin V on Stabilized Actin Cables

Cited by 19 publications

References 32 publications

Formaldehyde fixation is detrimental to actin cables in glucose-depleted S. cerevisiae cells

Formaldehyde fixation is detrimental to actin cables in glucose-depleted S. cerevisiae cells

Reliable imaging of ATP in living budding and fission yeast

The Stationary-Phase Cells of Saccharomyces cerevisiae Display Dynamic Actin Filaments Required for Processes Extending Chronological Life Span

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