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Nature Metabolism Oct 2023Amino acid homeostasis is critical for many cellular processes. It is well established that amino acids are compartmentalized using pH gradients generated between...
Amino acid homeostasis is critical for many cellular processes. It is well established that amino acids are compartmentalized using pH gradients generated between organelles and the cytoplasm; however, the dynamics of this partitioning has not been explored. Here we develop a highly sensitive pH reporter and find that the major amino acid storage compartment in Saccharomyces cerevisiae, the lysosome-like vacuole, alkalinizes before cell division and re-acidifies as cells divide. The vacuolar pH dynamics require the uptake of extracellular amino acids and activity of TORC1, the v-ATPase and the cycling of the vacuolar specific lipid phosphatidylinositol 3,5-bisphosphate, which is regulated by the cyclin-dependent kinase Pho85 (CDK5 in mammals). Vacuolar pH regulation enables amino acid sequestration and mobilization from the organelle, which is important for mitochondrial function, ribosome homeostasis and cell size control. Collectively, our data provide a new paradigm for the use of dynamic pH-dependent amino acid compartmentalization during cell growth/division.
Topics: Animals; Vacuoles; Saccharomyces cerevisiae Proteins; Saccharomyces cerevisiae; Homeostasis; Amino Acids; Cell Division; Cell Cycle; Hydrogen-Ion Concentration; Mammals
PubMed: 37640943
DOI: 10.1038/s42255-023-00872-1 -
The New Phytologist Jan 2021The coordinated control of ion transport across the two major membranes of differentiated plant cells, the plasma and the vacuolar membranes, is fundamental in cell... (Review)
Review
The coordinated control of ion transport across the two major membranes of differentiated plant cells, the plasma and the vacuolar membranes, is fundamental in cell physiology. The stomata responses to the fluctuating environmental conditions are an illustrative example. Indeed, they rely on the coordination of ion fluxes between the different cell compartments. The cytosolic environment, which is an interface between intracellular compartments, and the activity of the ion transporters localised in the different membranes influence one each other. Here we analyse the molecular mechanisms connecting and modulating the transport processes at both the plasma and the vacuolar membranes of guard cells.
Topics: Arabidopsis; Biological Transport; Cell Membrane; Ion Transport; Vacuoles
PubMed: 33007120
DOI: 10.1111/nph.16983 -
Autophagy Oct 2022Small 30-nm vesicles containing the integral membrane protein Atg9 provide the initial membrane source for autophagy in yeast. Atg23 is an Atg9 binding protein that is...
Small 30-nm vesicles containing the integral membrane protein Atg9 provide the initial membrane source for autophagy in yeast. Atg23 is an Atg9 binding protein that is required for Atg9 vesicle trafficking but whose exact function is unknown. In our recent paper, we explored the function of Atg23 using an approach combining cellular biology and biochemistry on purified protein. We determined that Atg23 is an elongated dimer spanning 320 Å in length. We also demonstrated that Atg23 is a membrane-binding and -tethering protein. Furthermore, we identified a series of amino acids residing in a putative coiled-coil region that when mutated prevent Atg23 dimer formation resulting in a stable Atg23 monomer. Last, we demonstrated that when monomeric Atg23 is expressed in yeast lacking Atg23, this leads to a loss of Atg23 puncta, a reduction in Atg9 puncta, a reduction in nonselective autophagy and a complete block in the cytoplasm-to-vacuole targeting (Cvt) pathway.
Topics: Amino Acids; Autophagy; Autophagy-Related Proteins; Membrane Proteins; Protein Transport; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Vacuoles
PubMed: 35867625
DOI: 10.1080/15548627.2022.2105107 -
Microbiology Spectrum Mar 2019Bacteria of the genus colonize a wide variety of mammalian hosts, in which their infectious cycle and ability to cause disease predominantly rely on an intracellular... (Review)
Review
Bacteria of the genus colonize a wide variety of mammalian hosts, in which their infectious cycle and ability to cause disease predominantly rely on an intracellular lifestyle within phagocytes. Upon entry into host cells, organisms undergo a complex, multistage intracellular cycle in which they sequentially traffic through, and exploit functions of, the endocytic, secretory, and autophagic compartments via type IV secretion system (T4SS)-mediated delivery of bacterial effectors. These effectors modulate an array of host functions and machineries to first promote conversion of the initial endosome-like -containing vacuole (eBCV) into a replication-permissive organelle derived from the host endoplasmic reticulum (rBCV) and then to an autophagy-related vacuole (aBCV) that mediates bacterial egress. Here we detail and discuss our current knowledge of cellular and molecular events of the intracellular cycle. We discuss the importance of the endosomal stage in determining T4SS competency, the roles of autophagy in rBCV biogenesis and aBCV formation, and T4SS-driven mechanisms of modulation of host secretory traffic in rBCV biogenesis and bacterial egress.
Topics: Animals; Bacterial Proteins; Brucella; Brucellosis; Cytoplasm; Host-Pathogen Interactions; Humans; Life Cycle Stages; Phagocytes; Vacuoles
PubMed: 30848234
DOI: 10.1128/microbiolspec.BAI-0006-2019 -
Oncotarget Aug 2016Cytoplasmic vacuolization (also called cytoplasmic vacuolation) is a well-known morphological phenomenon observed in mammalian cells after exposure to bacterial or viral... (Review)
Review
Cytoplasmic vacuolization (also called cytoplasmic vacuolation) is a well-known morphological phenomenon observed in mammalian cells after exposure to bacterial or viral pathogens as well as to various natural and artificial low-molecular-weight compounds. Vacuolization often accompanies cell death; however, its role in cell death processes remains unclear. This can be attributed to studying vacuolization at the level of morphology for many years. At the same time, new data on the molecular mechanisms of the vacuole formation and structure have become available. In addition, numerous examples of the association between vacuolization and previously unknown cell death types have been reported. Here, we review these data to make a deeper insight into the role of cytoplasmic vacuolization in cell death and survival.
Topics: Animals; Bacterial Infections; Bacterial Proteins; Cell Death; Cell Survival; Cytoplasm; Endoplasmic Reticulum; Endoplasmic Reticulum-Associated Degradation; Humans; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; Necrosis; Vacuoles; Virus Diseases
PubMed: 27331412
DOI: 10.18632/oncotarget.10150 -
Cell Host & Microbe Sep 2017The replication vacuole of Legionella pneumophila makes contact with host mitochondria. In this issue of Cell Host & Microbe, Escoll et al. (2017) dissect the...
The replication vacuole of Legionella pneumophila makes contact with host mitochondria. In this issue of Cell Host & Microbe, Escoll et al. (2017) dissect the mechanisms of this interaction, the effect of the T4SS effector MitF on mitochondrial function, and the resultant metabolic reprogramming of infected cells to benefit the bacteria.
Topics: Bacterial Proteins; Humans; Legionella pneumophila; Mitochondria; Vacuoles
PubMed: 28910630
DOI: 10.1016/j.chom.2017.08.014 -
Small GTPases Sep 2018Small GTPases of the Rab protein family control intracellular vesicular trafficking to allow their communication and maintenance. It is a common strategy for... (Review)
Review
Small GTPases of the Rab protein family control intracellular vesicular trafficking to allow their communication and maintenance. It is a common strategy for intracellular bacteria to exploit these pathways to shape their respective niches for survival. The subversion of Rabs for the generation of an intracellular environment favoring the pathogen has been described almost exclusively for intracellular bacteria that reside within bacterial containing vacuoles (BCVs). However, less is known about Rab subversion for bacteria that rupture the BCV to reach the host cytoplasm. Here, we provide recent examples of Rab targeting by both groups of intracellular bacteria with a special focus on Shigella, the causative agent of bacillary dysentery. Shigella recruits Rab11, the hallmark of the perinuclear recycling compartment to in situ formed macropinosomes at the entry foci via the bacterial effector IpgD. This leads to efficient BCV rupture and cytosolic escape. We discuss the concept of diverted recycling through host Rab GTPases that emerges as a novel pathogen strategy.
Topics: Humans; Intracellular Space; Shigella; Vacuoles; rab GTP-Binding Proteins
PubMed: 27763815
DOI: 10.1080/21541248.2016.1240494 -
PloS One 2022Vacuoles in plants and fungi play critical roles in cell metabolism and osmoregulation. To support these functions, vacuoles change their morphology, e.g. they fragment...
Vacuoles in plants and fungi play critical roles in cell metabolism and osmoregulation. To support these functions, vacuoles change their morphology, e.g. they fragment when these organisms are challenged with draught, high salinity or metabolic stress (e.g. acetate accumulation). In turn, morphology reflects an equilibrium between membrane fusion and fission that determines size, shape and copy number. By studying Saccharomyces cerevisiae and its vacuole as models, conserved molecular mechanisms responsible for fusion have been revealed. However, a detailed understanding of vacuole fission and how these opposing processes respond to metabolism or osmoregulation remain elusive. Herein we describe a new fluorometric assay to measure yeast vacuole fission in vitro. For proof-of-concept, we use this assay to confirm that acetate, a metabolic stressor, triggers vacuole fission and show it blocks homotypic vacuole fusion in vitro. Similarly, hypertonic stress induced by sorbitol or glucose caused robust vacuole fission in vitro whilst inhibiting fusion. Using wortmannin to inhibit phosphatidylinositol (PI) -kinases or rGyp1-46 to inactivate Rab-GTPases, we show that acetate stress likely targets PI signaling, whereas osmotic stress affects Rab signaling on vacuole membranes to stimulate fission. This study sets the stage for further investigation into the mechanisms that change vacuole morphology to support cell metabolism and osmoregulation.
Topics: Acetates; Membrane Fusion; Osmotic Pressure; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Vacuoles
PubMed: 35834522
DOI: 10.1371/journal.pone.0271199 -
Autophagy Sep 2019Based on my reading, and on my own experience, I have come to realize that people learn in different ways, and this can include the use of different media. This is one...
Based on my reading, and on my own experience, I have come to realize that people learn in different ways, and this can include the use of different media. This is one reason I have worked with various artists to portray the topic of autophagy through paintings, music and dance. Indeed, comments from members of the audience who have attended one of my seminars often suggest that a particular artistic approach 'hit home' and added meaning to them about the topic. In this issue of the journal I describe another such project-'the-found-art vacuole'-that utilized the talents of an amazing watercolor painter, Scott Hartley. The object of Scott's painting is the only artophagy composition that I have 'made'-assembled is a more accurate word. Doing so was quite fun, but after examining many of the 'antique' items that form the 'found-art vacuole', I realized that it would be nice to have a painting that was done in exquisite detail. The requirement for detail immediately made me think of Scott, whose work I was familiar with from the Ann Arbor Art Fair. To quote a line from the Belleville News-Democrat describing Scott's taking first place in an art competition, 'He began by doing landscapes, and eventually found a different style for his work: the intricacies of urban architecture, of alleys and fire escapes in a city neighborhood.' This does describe the nature of Scott's work, but you have to see these paintings to appreciate the detail.
Topics: Autophagy; Music; Paintings; Saccharomyces cerevisiae; Vacuoles
PubMed: 31238790
DOI: 10.1080/15548627.2019.1630225 -
Biochimica Et Biophysica Acta.... May 2019Cellular ageing results in accumulating damage to various macromolecules and the progressive decline of organelle function. Yeast vacuoles as well as their counterpart... (Review)
Review
Cellular ageing results in accumulating damage to various macromolecules and the progressive decline of organelle function. Yeast vacuoles as well as their counterpart in higher eukaryotes, the lysosomes, emerge as central organelles in lifespan determination. These acidic organelles integrate enzymatic breakdown and recycling of cellular waste with nutrient sensing, storage, signalling and mobilization. Establishing physical contact with virtually all other organelles, vacuoles serve as hubs of cellular homeostasis. Studies in Saccharomyces cerevisiae contributed substantially to our understanding of the ageing process per se and the multifaceted roles of vacuoles/lysosomes in the maintenance of cellular fitness with progressing age. Here, we discuss the multiple roles of the vacuole during ageing, ranging from vacuolar dynamics and acidification as determinants of lifespan to the function of this organelle as waste bin, recycling facility, nutrient reservoir and integrator of nutrient signalling.
Topics: Lysosomes; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Time Factors; Vacuoles
PubMed: 30796938
DOI: 10.1016/j.bbamcr.2019.02.011