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Molecules (Basel, Switzerland) Feb 2023BF-azadipyrromethenes are highly versatile fluorophores used for cellular and in vivo imaging in the near-infrared and far-red regions of the spectrum. As of yet, their...
BF-azadipyrromethenes are highly versatile fluorophores used for cellular and in vivo imaging in the near-infrared and far-red regions of the spectrum. As of yet, their use in conjunction with super-resolution imaging methodologies has not been explored. In this report, a series of structurally related BF-azadipyrromethenes has been examined for their suitability for use with stimulated emission depletion (STED) nanoscopy. The potential for STED imaging was initially evaluated using aqueous solutions of fluorophores as an effective predictor of fluorophore suitability. For live cell STED imaging in both 2D and 3D, several far-red emitting BF-azadipyrromethenes were successfully employed. Image resolution below the diffraction limit of a confocal microscope was demonstrated through measurement of distinct intracellular features including the nuclear membrane, nuclear lamina invaginations, the endoplasmic reticulum, and vacuoles. As the STED ability of BF-azadipyrromethene fluorophores has now been established, their use with this super-resolution method may be expected to increase in the future.
Topics: Microscopy, Fluorescence; Fluorescent Dyes; Vacuoles
PubMed: 36771082
DOI: 10.3390/molecules28031415 -
Trends in Microbiology Aug 2022Invasive bacteria colonise their host tissues by establishing niches inside eukaryotic cells, where they grow either in the cytosol or inside a specialised vacuole.... (Review)
Review
Invasive bacteria colonise their host tissues by establishing niches inside eukaryotic cells, where they grow either in the cytosol or inside a specialised vacuole. These two distinct intracellular lifestyles both present benefits but also impose various constraints on pathogenic microorganisms, in terms of nutrient acquisition, space requirements, exposure to immune responses, and ability to disseminate. Here we review the major characteristics of cytosolic and vacuolar lifestyles and the strategies used by bacteria to overcome challenges specific to each compartment. Recent research providing evidence that these scenarios are not mutually exclusive is presented, with the dual lifestyles of two foodborne pathogens, Listeria monocytogenes and Salmonella Typhimurium, discussed in detail. Finally, we elaborate on the conceptual implications of polyvalence from the perspective of host-pathogen interactions.
Topics: Cytosol; Host-Pathogen Interactions; Listeria monocytogenes; Salmonella typhimurium; Vacuoles
PubMed: 35168833
DOI: 10.1016/j.tim.2022.01.015 -
MBio Oct 2023Inflammasomes are essential for host defense against intracellular bacterial pathogens like , as they activate caspases, which promote cytokine release and cell death to...
Inflammasomes are essential for host defense against intracellular bacterial pathogens like , as they activate caspases, which promote cytokine release and cell death to control infection. In mice, interferon (IFN) signaling promotes inflammasome responses against bacteria by inducing a family of IFN-inducible GTPases known as guanylate-binding proteins (GBPs). Within murine macrophages, IFN promotes the rupture of the -containing vacuole (LCV), while GBPs are dispensable for this process. Instead, GBPs facilitate the lysis of cytosol-exposed . In contrast, the functions of IFN and GBPs in human inflammasome responses to are poorly understood. We show that IFN-γ enhances inflammasome responses to in human macrophages. Human GBP1 is required for these IFN-γ-driven inflammasome responses. Furthermore, GBP1 co-localizes with and/or LCVs in a type IV secretion system (T4SS)-dependent manner and promotes damage to the LCV, which leads to increased exposure of the bacteria to the host cell cytosol. Thus, our findings reveal species- and pathogen-specific differences in how GBPs function to promote inflammasome responses.
Topics: Humans; Animals; Mice; Inflammasomes; Legionella; Vacuoles; Carrier Proteins; Signal Transduction; Legionella pneumophila; GTP-Binding Proteins
PubMed: 37737612
DOI: 10.1128/mbio.01707-23 -
Plant Physiology Mar 2022Stomatal movement is essential for plants to optimize transpiration and therefore photosynthesis. Rapid changes in the stomatal aperture are accompanied by adjustment of...
Stomatal movement is essential for plants to optimize transpiration and therefore photosynthesis. Rapid changes in the stomatal aperture are accompanied by adjustment of vacuole volume and morphology in guard cells (GCs). In Arabidopsis (Arabidopsis thaliana) leaf epidermis, stomatal development undergoes a cell-fate transition including four stomatal lineage cells: meristemoid, guard mother cell, young GC, and GC. Little is known about the mechanism underlying vacuole dynamics and vacuole formation during stomatal development. Here, we utilized whole-cell electron tomography (ET) analysis to elucidate vacuole morphology, formation, and development in different stages of stomatal lineage cells at nanometer resolution. The whole-cell ET models demonstrated that large vacuoles were generated from small vacuole stepwise fusion/maturation along stomatal development stages. Further ET analyses verified the existence of swollen intraluminal vesicles inside distinct vacuoles at certain developmental stages of stomatal lineage cells, implying a role of multivesicular body fusion in stomatal vacuole formation. Collectively, our findings demonstrate a mechanism mediating vacuole formation in Arabidopsis stomatal development and may shed light on the role of vacuoles in stomatal movement.
Topics: Arabidopsis; Arabidopsis Proteins; Electron Microscope Tomography; Plant Stomata; Vacuoles
PubMed: 35134219
DOI: 10.1093/plphys/kiac028 -
Virulence Dec 2021Bacteria of the genus are natural pathogens of amoebae that can cause a severe pneumonia in humans called Legionnaires' Disease. Human disease results from inhalation... (Review)
Review
Bacteria of the genus are natural pathogens of amoebae that can cause a severe pneumonia in humans called Legionnaires' Disease. Human disease results from inhalation of -contaminated aerosols and subsequent bacterial replication within alveolar macrophages. pathogenicity in humans has resulted from extensive co-evolution with diverse genera of amoebae. To replicate intracellularly, generates a replication-permissive compartment called the -containing vacuole (LCV) through the concerted action of hundreds of Dot/Icm-translocated effector proteins. In this review, we present a collective overview of pathogenicity including infection mechanisms, secretion systems, and translocated effector function. We also discuss innate and adaptive immune responses to , the implications of genome diversity and future avenues for the field.
Topics: Humans; Immunity; Legionella; Legionnaires' Disease; Vacuoles; Virulence
PubMed: 33843434
DOI: 10.1080/21505594.2021.1903199 -
Nature Communications Jun 2023Liquid droplets of biomolecules play key roles in organizing cellular behavior, and are also technologically relevant, yet physical studies of dynamic processes of such...
Liquid droplets of biomolecules play key roles in organizing cellular behavior, and are also technologically relevant, yet physical studies of dynamic processes of such droplets have generally been lacking. Here, we investigate and quantify the dynamics of formation of dilute internal inclusions, i.e., vacuoles, within a model system consisting of liquid droplets of DNA 'nanostar' particles. When acted upon by DNA-cleaving restriction enzymes, these DNA droplets exhibit cycles of appearance, growth, and bursting of internal vacuoles. Analysis of vacuole growth shows their radius increases linearly in time. Further, vacuoles pop upon reaching the droplet interface, leading to droplet motion driven by the osmotic pressure of restriction fragments captured in the vacuole. We develop a model that accounts for the linear nature of vacuole growth, and the pressures associated with motility, by describing the dynamics of diffusing restriction fragments. The results illustrate the complex non-equilibrium dynamics possible in biomolecular condensates.
Topics: Vacuoles; DNA
PubMed: 37328453
DOI: 10.1038/s41467-023-39175-0 -
International Journal of Molecular... Feb 2023Large vacuoles are a predominant cell organelle throughout the plant body. They maximally account for over 90% of cell volume and generate turgor pressure that acts as a... (Review)
Review
Large vacuoles are a predominant cell organelle throughout the plant body. They maximally account for over 90% of cell volume and generate turgor pressure that acts as a driving force of cell growth, which is essential for plant development. The plant vacuole also acts as a reservoir for sequestering waste products and apoptotic enzymes, thereby enabling plants to rapidly respond to fluctuating environments. Vacuoles undergo dynamic transformation through repeated enlargement, fusion, fragmentation, invagination, and constriction, eventually resulting in the typical 3-dimensional complex structure in each cell type. Previous studies have indicated that such dynamic transformations of plant vacuoles are governed by the plant cytoskeletons, which consist of F-actin and microtubules. However, the molecular mechanism of cytoskeleton-mediated vacuolar modifications remains largely unclear. Here we first review the behavior of cytoskeletons and vacuoles during plant development and in response to environmental stresses, and then introduce candidates that potentially play pivotal roles in the vacuole-cytoskeleton nexus. Finally, we discuss factors hampering the advances in this research field and their possible solutions using the currently available cutting-edge technologies.
Topics: Vacuoles; Cytoskeleton; Microtubules; Plants; Actin Cytoskeleton
PubMed: 36835552
DOI: 10.3390/ijms24044143 -
Proceedings of the National Academy of... Jun 2023Peptidoglycan is a critical component of the bacteria cell envelope. Remodeling of the peptidoglycan is required for numerous essential cellular processes and has been...
Peptidoglycan is a critical component of the bacteria cell envelope. Remodeling of the peptidoglycan is required for numerous essential cellular processes and has been linked to bacterial pathogenesis. Peptidoglycan deacetylases that remove the acetyl group of the -acetylglucosamine (NAG) subunit protect bacterial pathogens from immune recognition and digestive enzymes secreted at the site of infection. However, the full extent of this modification on bacterial physiology and pathogenesis is not known. Here, we identify a polysaccharide deacetylase of the intracellular bacterial pathogen and define a two-tiered role for this enzyme in pathogenesis. First, NAG deacetylation is important for the proper localization and function of the Type IVb secretion system, linking peptidoglycan editing to the modulation of host cellular processes through the action of secreted virulence factors. As a consequence, the vacuole mis-traffics along the endocytic pathway to the lysosome, preventing the formation of a replication permissive compartment. Second, within the lysosome, the inability to deacetylate the peptidoglycan renders the bacteria more sensitive to lysozyme-mediated degradation, resulting in increased bacterial death. Thus, the ability to deacetylate NAG is important for bacteria to persist within host cells and in turn, virulence. Collectively, these results expand the function of peptidoglycan deacetylases in bacteria, linking peptidoglycan editing, Type IV secretion, and the intracellular fate of a bacterial pathogen.
Topics: Humans; Legionella pneumophila; Peptidoglycan; Vacuoles; Legionella; Lysosomes; Bacterial Proteins; Legionnaires' Disease
PubMed: 37252954
DOI: 10.1073/pnas.2119658120 -
Genes Jan 2021Target of rapamycin complex 1 (TORC1), a serine/threonine-protein kinase complex highly conserved among eukaryotes, coordinates cellular growth and metabolism with... (Review)
Review
Target of rapamycin complex 1 (TORC1), a serine/threonine-protein kinase complex highly conserved among eukaryotes, coordinates cellular growth and metabolism with environmental cues, including nutrients and growth factors. Aberrant TORC1 signaling is associated with cancers and various human diseases, and TORC1 also plays a key role in ageing and lifespan, urging current active research on the mechanisms of TORC1 regulation in a variety of model organisms. Identification and characterization of the RAG small GTPases as well as their regulators, many of which are highly conserved from yeast to humans, led to a series of breakthroughs in understanding the molecular bases of TORC1 regulation. Recruitment of mammalian TORC1 (mTORC1) by RAGs to lysosomal membranes is a key step for mTORC1 activation. Interestingly, the RAG GTPases in fission yeast are primarily responsible for attenuation of TORC1 activity on vacuoles, the yeast equivalent of lysosomes. In this review, we summarize our current knowledge about the functions of TORC1 regulators on yeast vacuoles, and illustrate the conserved and divergent mechanisms of TORC1 regulation between yeasts and mammals.
Topics: Animals; Humans; Intracellular Membranes; Lysosomes; Mechanistic Target of Rapamycin Complex 1; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Transcription Factors; Vacuoles
PubMed: 33445779
DOI: 10.3390/genes12010088 -
The EMBO Journal Jul 2023The facultative intracellular pathogen Brucella abortus interacts with several organelles of the host cell to reach its replicative niche inside the endoplasmic...
The facultative intracellular pathogen Brucella abortus interacts with several organelles of the host cell to reach its replicative niche inside the endoplasmic reticulum. However, little is known about the interplay between the intracellular bacteria and the host cell mitochondria. Here, we showed that B. abortus triggers substantive mitochondrial network fragmentation, accompanied by mitophagy and the formation of mitochondrial Brucella-containing vacuoles during the late steps of cellular infection. Brucella-induced expression of the mitophagy receptor BNIP3L is essential for these events and relies on the iron-dependent stabilisation of the hypoxia-inducible factor 1α. Functionally, BNIP3L-mediated mitophagy appears to be advantageous for bacterial exit from the host cell as BNIP3L depletion drastically reduces the number of reinfection events. Altogether, these findings highlight the intricate link between Brucella trafficking and the mitochondria during host cell infection.
Topics: Brucella abortus; Mitophagy; Vacuoles; Endoplasmic Reticulum; Mitochondria
PubMed: 37232029
DOI: 10.15252/embj.2022112817