Biology subjects

Ewald, J. C.

Publications and source records attributed to Ewald, J. C..

6 recordsLinked to original sources

bioRxiv2026

Swe1 delays cell cycle progression during impaired mitochondrial inheritance to promote mitochondrial homeostasis

Mitochondrial homeostasis is essential for cellular health. To maintain mitochondrial homeostasis in proliferating cells, mitochondrial inheritance to daughter cells must be tightly coordinated with growth and cell division. In budding yeast, mitochondrial inheritance is ensured by active transport into the growing bud. Although inheritance defects are well known to affect cell physiology, the regulatory mechanisms linking mitochondrial inheritance to cell cycle control remain unclear. Here, we use the inheritance defect of cells depleted of the mitochondrial inheritance adaptor Mmr1 to investigate this coordination. Time-resolved analysis of more than 8,000 budding events revealed that budding duration is directly linked to the time of mitochondrial inheritance. To understand the underlying mechanism, we performed a genetic interaction screen using the auxin-inducible degron library. Strains depleted of cell cycle regulators, particularly those involved in mitotic progression, were highly enriched in clones with synthetic growth defects. Among these, Swe1, the bud morphogenesis checkpoint kinase, emerged as a key candidate affecting budding duration in response to impaired mitochondrial inheritance. Co-depletion of Mmr1 and Swe1 strongly reduces the mitochondrial inheritance-dependent lengthening of budding duration, leading to a large fraction of daughter cells with insufficient mitochondrial content. Thus, Swe1-mediated mitotic delay contributes to slowing cell cycle progression in response to defective mitochondrial inheritance. We suggest calling this regulatory loop MIBA: mitochondrial inheritance-dependent budding adaptation. Together, our findings reveal a mechanism by which cells couple mitochondrial inheritance to cell cycle progression and highlight the importance of this coupling for mitochondrial homeostasis across the population.

cell biology↗

bioRxiv2025

When mitochondria fall apart: Unbalanced mitochondrial segregation triggers loss of mtDNA in the absence of mitochondrial fusion

Mitochondrial biogenesis and inheritance must be carefully regulated alongside cell division to ensure proper mitochondrial function and cell survival. The dynamics of the mitochondrial network, including fusion and fission, play a crucial role in mitochondrial inheritance by facilitating the distribution and quality control of mitochondria. In budding yeast, simultaneous inhibition of both fusion and fission leads to loss of mitochondrial DNA (mtDNA) integrity, resulting in an increased frequency of petite cells. Loss of mitochondrial fusion alone results in the complete loss of mtDNA. While the loss of mtDNA in the absence of mitochondrial fusion has been known for almost 30 years, the reason remained unclear. Here, we investigate the consequences of impaired mitochondrial fusion through depletion of the mitofusin Fzo1. We follow the emerging phenotype by live-cell imaging and the analysis of more than thirty thousand single cells across their cell cycle. Fzo1 depletion causes rapid mitochondrial fragmentation and a reduction in mitochondrial membrane potential, followed by a progressive decline of mtDNA content and cellular growth rate over several cell divisions. During division, Fzo1-depleted daughters obtain an unusually large amount of mitochondria, leaving the mother with too little. This results in a strong disbalance of mitochondrial mass in the population. Additionally, Fzo1-depleted cells lose the ability to adjust mtDNA synthesis to compensate for a low mitochondrial content. The combined effects of unequal distribution and reduced synthesis drive rapid mtDNA loss. These results show how fusion defects lead to mtDNA loss and mitochondrial dysfunction, contributing to understanding diseases linked to fusion defects.

cell biology↗

bioRxiv2024

Cytosolic peroxiredoxin Tsa1 influences acetic acid metabolism and pH homeostasis in wine yeasts

Acetic acid is a key metabolite in yeast fermentation, influencing wine quality through its role in volatile acidity. In Saccharomyces cerevisiae, acetic acid production involves aldehyde dehydrogenases, primarily Ald6 during fermentation and Ald4 under respiration. However, the regulatory mechanisms of these enzymes throughout fermentation and how they differ in commonly used strains remain partially unclear. This study explores the cytosolic peroxiredoxin Tsa1 as a novel regulator of acetic acid metabolism. TSA1 gene deletion revealed strain-dependent effects on acetic acid metabolism and tolerance, showing reduced production and enhanced consumption in laboratory media. Under respiration, Ald4-driven acetic acid production, which raises extracellular pH, was mitigated by the absence of Tsa1. During wine fermentation, TSA1 deletion decreased the initial acetic acid surge by downregulating ALD6 transcription and enzymatic activity. These findings position Tsa1 as a metabolic regulator and a potential target for modulating acetic acid levels to manage volatile acidity and wine quality.

microbiology↗

bioRxiv2021

When Yeast Cells Change their Mind: Cell Cycle 'Start' is Reversible under Starvation

Eukaryotic cells decide in late G1 whether to commit to another round of genome duplication and division. This point of irreversible cell cycle commitment is a molecular switch termed "Restriction Point" in mammals and "Start" in budding yeast. At Start, yeast cells integrate multiple signals such as pheromones, osmolarity, and nutrients. If sufficient nutrients are lacking, cells will not pass Start. However, how the cells respond to nutrient depletion after they have made the Start decision, remains poorly understood. Here, we analyze by live cell imaging how post-Start yeast cells respond to nutrient depletion. We monitor fluorescently labelled Whi5, the cell cycle inhibitor whose export from the nucleus determines Start. Surprisingly, we find that cells that have passed Start can re-import Whi5 back into the nucleus. This occurs when cells are faced with starvation up to 20 minutes after Start. In these cells, the positive feedback loop is interrupted, Whi5 re-binds DNA, and CDK activation occurs a second time once nutrients are replenished. Cells which re-import Whi5 also become sensitive to mating pheromone again, and thus behave like pre-Start cells. In summary, we show that upon starvation the commitment decision at Start can be reversed. We therefore propose that in yeast, as has been suggested for mammalian cells, cell cycle commitment is a multi-step process, where irreversibility in face of nutrient signaling is only reached approximately 20 minutes after CDK activation at Start.

cell biology↗

bioRxiv2020

Regulation of trehalase activity by multi-site phosphorylation and 14-3-3 interaction

Protein phosphorylation enables a rapid adjustment of cellular activities to diverse intracellular and environmental stimuli. Many phosphoproteins are targeted on more than one site, which allows the integration of multiple signals and the implementation of complex responses. However, the hierarchy and interplay between multiple phospho-sites are often unknown. Here, we study multi-site phosphorylation using the yeast trehalase Nth1 and its activator, the 14-3-3 protein Bmh1, as a model. Nth1 is known to be phosphorylated by the metabolic kinase PKA on four serine residues and by the cell cycle kinase CDK on one residue. However, how these five phospho-sites adjust Nth1 activity remains unclear. Using a novel reporter construct, we investigated the contribution of the individual sites for the regulation of the trehalase and its 14-3-3 interactor. In contrast to the constitutively phosphorylated S20 and S83, the weaker sites S21 and S60 are only phosphorylated by increased PKA activity. For binding Bmh1, S83 functions as the high-affinity "gatekeeper" site, but successful binding of the Bmh1 dimer and thus Nth1 activation requires S60 as a secondary site. Under nutrient-poor conditions with low PKA activity, S60 is not efficiently phosphorylated and the cell cycle dependent phosphorylation of S66 by Cdk1 contributes to Nth1 activity, likely by providing an alternative Bmh1 binding site. Additionally, the PKA sites S20 and S21 modulate the dephosphorylation of Nth1 on downstream Bmh1 sites. In summary, our results expand our molecular understanding of Nth1 regulation and provide a new aspect of the interaction of 14-3-3 proteins with their targets.

biochemistry↗

bioRxiv2019

Multiple layers of phospho-regulation coordinate metabolism and the cell cycle in budding yeast

The coordination of metabolism and growth with cell division is crucial for proliferation. While it has long been known that cell metabolism regulates the cell division cycle, it is becoming increasingly clear that the cell division cycle also regulates metabolism. In budding yeast, we previously showed that over half of all measured metabolites change concentration through the cell cycle indicating that metabolic fluxes are extensively regulated during cell cycle progression. However, how this regulation is achieved still remains poorly understood. Since both the cell cycle and metabolism are regulated to a large extent by protein phosphorylation, we here decided to measure the phosphoproteome through the budding yeast cell cycle. Specifically, we chose a cell cycle synchronisation strategy that avoids stress and nutrient-related perturbations of metabolism, and we grew the yeast on ethanol minimal medium to force cells to utilize their full biosynthetic repertoire. Using a tandem-mass-tagging approach, we found over 200 sites on metabolic enzymes and transporters to be phospho-regulated. These sites were distributed among many pathways including carbohydrate catabolism, lipid metabolism and amino acid synthesis and therefore likely contribute to changing metabolic fluxes through the cell cycle. Among all one thousand sites whose phosphorylation increases through the cell cycle, the CDK consensus motif and an arginine-directed motif were highly enriched. This arginine-directed R-R-x-S motif is associated with protein-kinase A, which regulates metabolism and promotes growth. Finally, we also found over one thousand sites that are dephosphorylated through the G1/S transition. We speculate that the phosphatase Glc7/ PP1, known to regulate both the cell cycle and carbon metabolism, may play an important role because its regulatory subunits are phospho-regulated in our data. In summary, our results identify extensive cell cycle dependent phosphorylation and dephosphorylation of metabolic enzymes and suggest multiple mechanisms through which the cell division cycle regulates metabolic signalling pathways to temporally coordinate biosynthesis with distinct phases of the cell division cycle.

cell biology↗