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MSphere Aug 2023is an obligate, intracellular parasite. Infection of a cell produces a unique niche for the parasite named the parasitophorous vacuole (PV) initially composed of host...
is an obligate, intracellular parasite. Infection of a cell produces a unique niche for the parasite named the parasitophorous vacuole (PV) initially composed of host plasma membrane invaginated during invasion. The PV and its membrane (parasitophorous vacuole membrane [PVM]) are subsequently decorated with a variety of parasite proteins allowing the parasite to optimally grow in addition to manipulate host processes. Recently, we reported a proximity-labeling screen at the PVM-host interface and identified host endoplasmic reticulum (ER)-resident motile sperm domain-containing protein 2 (MOSPD2) as being enriched at this location. Here we extend these findings in several important respects. First, we show that the extent and pattern of host MOSPD2 association with the PVM differ dramatically in cells infected with different strains of . Second, in cells infected with Type I RH strain, the MOSPD2 staining is mutually exclusive with regions of the PVM that associate with mitochondria. Third, immunoprecipitation and liquid chromatography tandem mass spectrometry (LC-MS/MS) with epitope-tagged MOSPD2-expressing host cells reveal strong enrichment of several PVM-localized parasite proteins, although none appear to play an essential role in MOSPD2 association. Fourth, most MOSPD2 associating with the PVM is newly translated after infection of the cell and requires the major functional domains of MOSPD2, identified as the CRAL/TRIO domain and tail anchor, although these domains were not sufficient for PVM association. Lastly, ablation of MOSPD2 results in, at most, a modest impact on growth . Collectively, these studies provide new insight into the molecular interactions involving MOSPD2 at the dynamic interface between the PVM and the host cytosol. IMPORTANCE is an intracellular pathogen that lives within a membranous vacuole inside of its host cell. This vacuole is decorated by a variety of parasite proteins that allow it to defend against host attack, acquire nutrients, and interact with the host cell. Recent work identified and validated host proteins enriched at this host-pathogen interface. Here, we follow up on one candidate named MOSPD2 shown to be enriched at the vacuolar membrane and describe it as having a dynamic interaction at this location depending on a variety of factors. Some of these include the presence of host mitochondria, intrinsic domains of the host protein, and whether translation is active. Importantly, we show that MOSPD2 enrichment at the vacuole membrane differs between strains indicating active involvement of the parasite with this phenotype. Altogether, these results shed light on the mechanism and role of protein associations in the host-pathogen interaction.
Topics: Male; Animals; Toxoplasma; Vacuoles; Chromatography, Liquid; Protozoan Proteins; Semen; Tandem Mass Spectrometry; Membrane Proteins
PubMed: 37341482
DOI: 10.1128/msphere.00670-22 -
Autophagy 2018Membrane protein recycling is a fundamental process from yeast to humans. The lysosome (or vacuole in yeast) receives membrane proteins from the secretory, endocytic,...
Membrane protein recycling is a fundamental process from yeast to humans. The lysosome (or vacuole in yeast) receives membrane proteins from the secretory, endocytic, and macroautophagy/autophagy pathways. Although some of these membrane proteins appear to be recycled, the molecular mechanisms underlying this retrograde trafficking are poorly understood. Our recent study revealed that the transmembrane autophagy protein Atg27 is recycled from the vacuole membrane using a 2-step recycling process. First, the Snx4 complex recycles Atg27 from the vacuole to the endosome. Then, the retromer complex mediates endosome-to-Golgi retrograde transport. Thus, 2 distinct protein complexes facilitate the sequential retrograde trafficking for Atg27. As far as we know, Atg27 is the first physiological substrate for the vacuole-to-endosome retrograde trafficking pathway.
Topics: Endosomes; Golgi Apparatus; Intracellular Membranes; Lysosomes; Membrane Fusion; Membrane Proteins; Models, Biological; Protein Transport; Vacuoles
PubMed: 29995558
DOI: 10.1080/15548627.2018.1496719 -
PLoS Pathogens Jul 2023A key element of Plasmodium biology and pathogenesis is the trafficking of ~10% of the parasite proteome into the host red blood cell (RBC) it infects. To cross the...
A key element of Plasmodium biology and pathogenesis is the trafficking of ~10% of the parasite proteome into the host red blood cell (RBC) it infects. To cross the parasite-encasing parasitophorous vacuole membrane, exported proteins utilise a channel-forming protein complex termed the Plasmodium translocon of exported proteins (PTEX). PTEX is obligatory for parasite survival, both in vitro and in vivo, suggesting that at least some exported proteins have essential metabolic functions. However, to date only one essential PTEX-dependent process, the new permeability pathways, has been described. To identify other essential PTEX-dependant proteins/processes, we conditionally knocked down the expression of one of its core components, PTEX150, and examined which pathways were affected. Surprisingly, the food vacuole mediated process of haemoglobin (Hb) digestion was substantially perturbed by PTEX150 knockdown. Using a range of transgenic parasite lines and approaches, we show that two major Hb proteases; falcipain 2a and plasmepsin II, interact with PTEX core components, implicating the translocon in the trafficking of Hb proteases. We propose a model where these proteases are translocated into the PV via PTEX in order to reach the cytostome, located at the parasite periphery, prior to food vacuole entry. This work offers a second mechanistic explanation for why PTEX function is essential for growth of the parasite within its host RBC.
Topics: Animals; Plasmodium falciparum; Vacuoles; Protein Transport; Protozoan Proteins; Erythrocytes; Parasites; Peptide Hydrolases
PubMed: 37523385
DOI: 10.1371/journal.ppat.1011006 -
Life Science Alliance Sep 2022Membrane contact sites are functional nodes at which organelles reorganize metabolic pathways and adapt to changing cues. In , the nuclear envelope subdomain surrounding...
Membrane contact sites are functional nodes at which organelles reorganize metabolic pathways and adapt to changing cues. In , the nuclear envelope subdomain surrounding the nucleolus, very plastic and prone to expansion, can establish contacts with the vacuole and be remodeled in response to various metabolic stresses. While using genotoxins with unrelated purposes, we serendipitously discovered a fully new remodeling event at this nuclear subdomain: the nuclear envelope partitions into its regular contact with the vacuole and a dramatic internalization within the nucleus. This leads to the nuclear engulfment of a globular, cytoplasmic portion. In spite of how we discovered it, the phenomenon is likely DNA damage-independent. We define lipids supporting negative curvature, such as phosphatidic acid and sterols, as bona fide drivers of this event. Mechanistically, we suggest that the engulfment of the cytoplasm triggers a suction phenomenon that enhances the docking of proton pump-containing vesicles with the vacuolar membrane, which we show matches a boost in autophagy. Thus, our findings unveil an unprecedented remodeling of the nucleolus-surrounding membranes with impact on metabolic adaptation.
Topics: Autophagy; Cytoplasm; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Vacuoles
PubMed: 35568434
DOI: 10.26508/lsa.202101160 -
Life Science Alliance Oct 2022The ribosomal DNA (rDNA) array of has served as a model to address chromosome organization. In cells arrested before anaphase (mid-M), the rDNA acquires a highly...
The ribosomal DNA (rDNA) array of has served as a model to address chromosome organization. In cells arrested before anaphase (mid-M), the rDNA acquires a highly structured chromosomal organization referred to as the rDNA loop, whose length can double the cell diameter. Previous works established that complexes such as condensin and cohesin are essential to attain this structure. Here, we report that the rDNA loop adopts distinct presentations that arise as spatial adaptations to changes in the nuclear morphology triggered during mid-M arrests. Interestingly, the formation of the rDNA loop results in the appearance of a space under the loop (SUL) which is devoid of nuclear components yet colocalizes with the vacuole. We show that the rDNA-associated nuclear envelope (NE) often reshapes into a ladle to accommodate the vacuole in the SUL, with the nucleus becoming bilobed and doughnut-shaped. Finally, we demonstrate that the formation of the rDNA loop and the SUL require TORC1, membrane synthesis and functional vacuoles, yet is independent of nucleus-vacuole junctions and rDNA-NE tethering.
Topics: Anaphase; DNA, Ribosomal; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Vacuoles
PubMed: 35961781
DOI: 10.26508/lsa.202101161 -
Plant Physiology Jun 2017Recent research on vacuolar ion channels, transporters, and pumps of Arabidopsis highlight their function and roles in stomatal opening and closure. (Review)
Review
Recent research on vacuolar ion channels, transporters, and pumps of Arabidopsis highlight their function and roles in stomatal opening and closure.
Topics: Arabidopsis; Arabidopsis Proteins; Ion Channels; Ion Transport; Plant Stomata; Vacuoles
PubMed: 28381500
DOI: 10.1104/pp.17.00130 -
Current Biology : CB Jan 2016Macroautophagy (hereafter autophagy) is an evolutionarily conserved process in which portions of the cytoplasm are engulfed, degraded, and subsequently recycled. The... (Review)
Review
Macroautophagy (hereafter autophagy) is an evolutionarily conserved process in which portions of the cytoplasm are engulfed, degraded, and subsequently recycled. The Atg17-Atg31-Atg29 complex translocates to the phagophore assembly site (PAS), where an autophagosome forms, at a very early stage of autophagy, playing a vital role in autophagy induction. Here, we identified a novel role of this complex in a late stage of autophagy where it coordinates with Atg11 to regulate autophagy-specific fusion with the vacuole. Atg17 and Atg11 interact with the vacuolar SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) Vam7 independently of each other. Several hydrophobic residues in helix 1 and helix 4 of Atg17 and the SNARE domain of Vam7 mediate the Atg17-Vam7 interaction. An F317D mutation of Atg17, which diminishes its interaction with Vam7 without affecting its interaction with Atg13 or Atg31, leads to a defect in the fusion of autophagosomes with the vacuole and decreased autophagy activity. These results provide the first demonstration that the Atg17-Atg31-Atg29 complex functions in both early and late stages of autophagy and also provide a mechanistic explanation for the coordination of autophagosome completion and fusion with the vacuole.
Topics: Animals; Autophagy; Autophagy-Related Proteins; Humans; Membrane Fusion; Phagosomes; Protein Binding; Saccharomyces cerevisiae Proteins; Vacuoles
PubMed: 26774783
DOI: 10.1016/j.cub.2015.11.054 -
International Journal of Molecular... Jan 2023Vacuolar processing enzymes (VPEs) are plant cysteine proteases that are subjected to autoactivation in an acidic pH. It is presumed that VPEs, by activating other... (Review)
Review
Vacuolar processing enzymes (VPEs) are plant cysteine proteases that are subjected to autoactivation in an acidic pH. It is presumed that VPEs, by activating other vacuolar hydrolases, are in control of tonoplast rupture during programmed cell death (PCD). Involvement of VPEs has been indicated in various types of plant PCD related to development, senescence, and environmental stress responses. Another pathway induced during such processes is autophagy, which leads to the degradation of cellular components and metabolite salvage, and it is presumed that VPEs may be involved in the degradation of autophagic bodies during plant autophagy. As both PCD and autophagy occur under similar conditions, research on the relationship between them is needed, and VPEs, as key vacuolar proteases, seem to be an important factor to consider. They may even constitute a potential point of crosstalk between cell death and autophagy in plant cells. This review describes new insights into the role of VPEs in plant PCD, with an emphasis on evidence and hypotheses on the interconnections between autophagy and cell death, and indicates several new research opportunities.
Topics: Apoptosis; Plants; Vacuoles; Autophagy; Cell Death
PubMed: 36674706
DOI: 10.3390/ijms24021198 -
Cell Reports Jan 2021Membrane contact sites facilitate the exchange of metabolites between organelles to support interorganellar communication. The nucleus-vacuole junctions (NVJs) establish...
Membrane contact sites facilitate the exchange of metabolites between organelles to support interorganellar communication. The nucleus-vacuole junctions (NVJs) establish physical contact between the perinuclear endoplasmic reticulum (ER) and the vacuole. Although the NVJ tethers are known, how NVJ abundance and composition are controlled in response to metabolic cues remains elusive. Here, we identify the ER protein Snd3 as central factor for NVJ formation. Snd3 interacts with NVJ tethers, supports their targeting to the contacts, and is essential for NVJ formation. Upon glucose exhaustion, Snd3 relocalizes from the ER to NVJs and promotes contact expansion regulated by central glucose signaling pathways. Glucose replenishment induces the rapid dissociation of Snd3 from the NVJs, preceding the slow disassembly of the junctions. In sum, this study identifies a key factor required for formation and regulation of NVJs and provides a paradigm for metabolic control of membrane contact sites.
Topics: Cell Nucleus; Glucose; Phosphate Transport Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Signal Transduction; Vacuoles
PubMed: 33472077
DOI: 10.1016/j.celrep.2020.108637 -
Journal of Cataract and Refractive... May 2020To develop an advanced test methodology for quantification of scattered light from intraocular lenses (IOLs) and to evaluate the correlation between IOL vacuole...
PURPOSE
To develop an advanced test methodology for quantification of scattered light from intraocular lenses (IOLs) and to evaluate the correlation between IOL vacuole characteristics and measured scattered light.
SETTING
U.S. Food and Drug Administration, Optical Therapeutics and Medical Nanophotonics Laboratory, Silver Spring, Maryland, USA.
DESIGN
Experimental and analytical study.
METHODS
Twenty-four IOLs containing vacuoles were evaluated using a digital microscopy approach for identifying and characterizing the vacuoles present. A scanning light scattering profiler (SLSP) was used to evaluate and quantify the amount of scattered light from each IOL and from a 25th control IOL without any vacuoles. A variety of IOLs and vacuoles were also modeled in a Zemax simulation of the SLSP, and the simulated scattered light was modeled.
RESULTS
The scattered light as measured with SLSP was well correlated with vacuole characteristics, specifically density and size, as measured under the digital microscope for the 24 vacuole-containing IOLs. Additional correlations were found between vacuole sizes, orientations, and the angle at which light was scattered most severely. These correlations were also present in the Zemax model.
CONCLUSIONS
Vacuole optical characteristics can be well correlated with measured scatter, demonstrating an ability to predict scattered light based solely on microscope evaluation. Furthermore, the quantitative amount of scatter predicted with Zemax simulations trended closely with the experimentally measured trends.
Topics: Humans; Lenses, Intraocular; Maryland; Scattering, Radiation; Vacuoles; Vision, Ocular
PubMed: 32358273
DOI: 10.1097/j.jcrs.0000000000000167