Spatiotemporal Quantification of Endosomal Acidification on the Viral Journey

Small ◽  
2021 ◽  
pp. 2104200
Author(s):  
Zhi‐Gang Wang ◽  
Liang Zhao ◽  
Lu‐Lu Chen ◽  
Hao‐Yang Liu ◽  
Lei Wang ◽  
...  
Keyword(s):  
Endosomal Acidification

scholarly journals Effects of Basic Amino Acids and Their Derivatives on SARS-CoV-2 and Influenza-A Virus Infection

Viruses ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1301
Author(s):  
Ivonne Melano ◽  
Li-Lan Kuo ◽  
Yan-Chung Lo ◽  
Po-Wei Sung ◽  
Ni Tien ◽  
...  
Keyword(s):  
Amino Acids ◽  
Virus Infection ◽  
Influenza A Virus ◽  
Influenza A ◽  
Enveloped Viruses ◽  
Virus Attachment ◽  
Basic Amino Acids ◽  
Ester Derivative ◽  
Endosomal Acidification

Amino acids have been implicated with virus infection and replication. Here, we demonstrate the effects of two basic amino acids, arginine and lysine, and their ester derivatives on infection of two enveloped viruses, SARS-CoV-2, and influenza A virus. We found that lysine and its ester derivative can efficiently block infection of both viruses in vitro. Furthermore, the arginine ester derivative caused a significant boost in virus infection. Studies on their mechanism of action revealed that the compounds potentially disturb virus uncoating rather than virus attachment and endosomal acidification. Our findings suggest that lysine supplementation and the reduction of arginine-rich food intake can be considered as prophylactic and therapeutic regimens against these viruses while also providing a paradigm for the development of broad-spectrum antivirals.

scholarly journals Proton-activated chloride channel PAC regulates endosomal acidification and transferrin receptor-mediated endocytosis

Cell Reports ◽  
2021 ◽  
Vol 34 (4) ◽  
pp. 108683 ◽  
Author(s):  
James Osei-Owusu ◽  
Junhua Yang ◽  
Ka Ho Leung ◽  
Zheng Ruan ◽  
Wei Lü ◽  
...  
Keyword(s):  
Chloride Channel ◽  
Transferrin Receptor ◽  
Receptor Mediated Endocytosis ◽  
Endosomal Acidification

scholarly journals Synthetic Histidine-Rich Peptides Inhibit Candida Species and Other Fungi In Vitro: Role of Endocytosis and Treatment Implications

2006 ◽  
Vol 50 (8) ◽  
pp. 2797-2805 ◽  
Author(s):  
Jingsong Zhu ◽  
Paul W. Luther ◽  
Qixin Leng ◽  
A. James Mixson
Keyword(s):  
Nucleic Acid ◽  
Antifungal Activity ◽  
Antifungal Agents ◽  
Fungal Growth ◽  
Nucleic Acid Delivery ◽  
Histatin 5 ◽  
Ph Buffering ◽  
Endosomal Acidification

ABSTRACT A family of histidine-rich peptides, histatins, is secreted by the parotid gland in mammals and exhibits marked inhibitory activity against a number of Candida species. We were particularly interested in the mechanism by which histidine-rich peptides inhibit fungal growth, because our laboratory has synthesized a variety of such peptides for drug and nucleic acid delivery. In contrast to naturally occurring peptides that are linear, peptides made on synthesizers can be varied with respect to their degrees of branching. Using this technology, we explored whether histidine-lysine (HK) polymers of different complexities and degrees of branching affect the growth of several species of Candida. Polymers with higher degrees of branching were progressively more effective against Candida albicans, with the four-branched polymer, H2K4b, most effective. Furthermore, H2K4b accumulated efficiently in C. albicans, which may indicate its ability to transport other antifungal agents intracellularly. Although H2K4b had greater antifungal activity than histatin 5, their mechanisms were similar. Toxicity in C. albicans induced by histatin 5 or branched HK peptides was markedly reduced by 4,4′-diisothiocyanato-stilbene-2,2′-disulfonate, an inhibitor of anion channels. We also determined that bafilomycin A1, an inhibitor of endosomal acidification, significantly decreased the antifungal activity of H2K4b. This suggests that the pH-buffering and subsequent endosomal-disrupting properties of histidine-rich peptides have a role in their antifungal activity. Moreover, the ability of the histidine component of these peptides to disrupt endosomes, which allows their escape from the lysosomal pathway, may explain why these peptides are both effective antifungal agents and nucleic acid delivery carriers.

β-Barrel outer membrane proteins suppress mTORC2 activation and induce autophagic responses

2018 ◽  
Vol 11 (558) ◽  
pp. eaat7493 ◽  
Author(s):  
Anu Chaudhary ◽  
Cassandra Kamischke ◽  
Mara Leite ◽  
Melissa A. Altura ◽  
Loren Kinman ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Epithelial Cells ◽  
Outer Membrane ◽  
Cellular Response ◽  
Tertiary Structure ◽  
Inflammatory Responses ◽  
Outer Membrane Proteins ◽  
Cell Surface Receptor ◽  
Mouse Bone Marrow ◽  
Endosomal Acidification

The outer membranes of Gram-negative bacteria and mitochondria contain proteins with a distinct β-barrel tertiary structure that could function as a molecular pattern recognized by the innate immune system. Here, we report that purified outer membrane proteins (OMPs) from different bacterial and mitochondrial sources triggered the induction of autophagy-related endosomal acidification, LC3B lipidation, and p62 degradation. Furthermore, OMPs reduced the phosphorylation and therefore activation of the multiprotein complex mTORC2 and its substrate Akt in macrophages and epithelial cells. The cell surface receptor SlamF8 and the DNA-protein kinase subunit XRCC6 were required for these OMP-specific responses in macrophages and epithelial cells, respectively. The addition of OMPs to mouse bone marrow–derived macrophages infected withSalmonellaTyphimurium facilitated bacterial clearance. These data identify a specific cellular response mediated by bacterial and mitochondrial OMPs that can alter inflammatory responses and influence the killing of pathogens.

scholarly journals Niclosamide inhibits SARS-CoV2 entry by blocking internalization through pH-dependent CLIC/GEEC endocytic pathway

2020 ◽  
Author(s):  
Chaitra Prabhakara ◽  
Rashmi Godbole ◽  
Parijat Sil ◽  
Sowmya Jahnavi ◽  
Thomas S van Zanten ◽  
...  
Keyword(s):  
Viral Infection ◽  
Receptor Binding ◽  
Endocytic Pathway ◽  
Host Cells ◽  
Entry Inhibitor ◽  
Receptor Binding Domain ◽  
Endosomal Acidification ◽  
Independent Mechanism ◽  
Ph Dependent ◽  
Endosomal Ph

AbstractMany viruses utilize the host endo-lysosomal network to infect cells. Tracing the endocytic itinerary of SARS-CoV2 can provide insights into viral trafficking and aid in designing new therapeutic targets. Here, we demonstrate that the receptor binding domain (RBD) of SARS-CoV2 is internalized via the clathrin and dynamin-independent, pH-dependent CLIC/GEEC (CG) endocytic pathway. Endosomal acidification inhibitors like BafilomycinA1 and NH4Cl, which inhibit the CG pathway, strongly block the uptake of RBD. Using transduction assays with SARS-CoV2 Spike-pseudovirus, we confirmed that these acidification inhibitors also impede viral infection. By contrast, Chloroquine neither affects RBD uptake nor extensively alters the endosomal pH, yet attenuates Spike-pseudovirus entry, indicating a pH-independent mechanism of intervention. We screened a subset of FDA-approved acidification inhibitors and found Niclosamide to be a potential SARS-CoV2 entry inhibitor. Niclosamide, thus, could provide broader applicability in subverting infection of similar category viruses entering host cells via this pH-dependent endocytic pathway.

scholarly journals Blocking Hepatitis C Virus Infection with Recombinant Form of Envelope Protein 2 Ectodomain

2009 ◽  
Vol 83 (21) ◽  
pp. 11078-11089 ◽  
Author(s):  
Jillian Whidby ◽  
Guaniri Mateu ◽  
Hannah Scarborough ◽  
Borries Demeler ◽  
Arash Grakoui ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Mammalian Cells ◽  
Expression System ◽  
Developed Countries ◽  
Viral Envelope ◽  
Hcv Infection ◽  
Size Exclusion ◽  
Amino Terminal ◽  
Endosomal Acidification

ABSTRACT More than 120 million people worldwide are chronically infected with hepatitis C virus (HCV), making HCV infection the leading cause of liver transplantation in developed countries. Treatment is limited, and efficacy depends upon the infecting strain and the initial viral load. The HCV envelope glycoproteins (E1 and E2) are involved in receptor binding, virus-cell fusion, and entry into the host cell. HCV infection proceeds by endosomal acidification, suggesting that fusion of the viral envelope with cellular membranes is a pH-triggered event. E2 consists of an amino-terminal ectodomain, an amphipathic helix that forms a stem region, and a carboxy-terminal membrane-associating segment. We have devised a novel expression system for the production of a secreted form of E2 ectodomain (eE2) from mammalian cells and performed a comprehensive biochemical and biophysical characterization. eE2 is properly folded, as determined by binding to human CD81, blocking of infection of cell culture-derived HCV, and recognition by antibodies from patients chronically infected with different genotypes of HCV. The glycosylation pattern, number of disulfide bonds, oligomerization state, and secondary structure of eE2 have been characterized using mass spectrometry, size exclusion chromatography, circular dichroism, and analytical ultracentrifugation. These results advance the understanding of E2 and may assist in the design of an HCV vaccine and entry inhibitor.

scholarly journals Microbial co-infection alters macrophage polarization, phagosomal escape, and microbial killing

2018 ◽  
Vol 24 (3) ◽  
pp. 152-162 ◽  
Author(s):  
Nikita H Trivedi ◽  
Jieh-Juen Yu ◽  
Chiung-Yu Hung ◽  
Richard P Doelger ◽  
Christopher S Navara ◽  
...  
Keyword(s):  
Bacterial Infection ◽  
Macrophage Activation ◽  
Secondary Infection ◽  
Macrophage Polarization ◽  
Innate Immune ◽  
No Production ◽  
Bacterial Killing ◽  
Macrophage Response ◽  
Endosomal Acidification ◽  
J774 Cells

Macrophages are important innate immune cells that respond to microbial insults. In response to multi-bacterial infection, the macrophage activation state may change upon exposure to nascent mediators, which results in different bacterial killing mechanism(s). In this study, we utilized two respiratory bacterial pathogens, Mycobacterium bovis (Bacillus Calmette Guẻrin, BCG) and Francisella tularensis live vaccine strain (LVS) with different phagocyte evasion mechanisms, as model microbes to assess the influence of initial bacterial infection on the macrophage response to secondary infection. Non-activated (M0) macrophages or activated M2-polarized cells (J774 cells transfected with the mouse IL-4 gene) were first infected with BCG for 24–48 h, subsequently challenged with LVS, and the results of inhibition of LVS replication in the macrophages was assessed. BCG infection in M0 macrophages activated TLR2-MyD88 and Mincle-CARD9 signaling pathways, stimulating nitric oxide (NO) production and enhanced killing of LVS. BCG infection had little effect on LVS escape from phagosomes into the cytosol in M0 macrophages. In contrast, M2-polarized macrophages exhibited enhanced endosomal acidification, as well as inhibiting LVS replication. Pre-infection with BCG did not induce NO production and thus did not further reduce LVS replication. This study provides a model for studies of the complexity of macrophage activation in response to multi-bacterial infection.

Kinetics of endosomal pH evolution in Dictyostelium discoideum amoebae. Study by fluorescence spectroscopy

1993 ◽  
Vol 105 (3) ◽  
pp. 861-866 ◽  
Author(s):  
L. Aubry ◽  
G. Klein ◽  
J.L. Martiel ◽  
M. Satre
Keyword(s):  
Dictyostelium Discoideum ◽  
Fluid Phase ◽  
Lag Phase ◽  
Nutritive Medium ◽  
Chase Period ◽  
Endosomal Acidification ◽  
Fluid Phase Endocytosis ◽  
Endosomal Ph ◽  
Efflux Kinetics ◽  
Secondary Lysosomes

The evolution of endo-lysosomal pH in Dictyostelium discoideum amoebae was examined during fluid-phase endocytosis. Pulse-chase experiments were conducted in nutritive medium or in non-nutritive medium using fluorescein labelled dextran (FITC-dextran) as fluid-phase marker and pH probe. In both conditions, efflux kinetics were characterized by an extended lag phase lasting for 45–60 min and corresponding to intracellular transit of FITC-dextran cohort. During the chase period, endosomal pH decreased during approximately 20 min from extracellular pH down to pH 4.6-5.0, then, it increased within the next 20–40 min to reach pH 6.0-6.2. It was only at this stage that FITC-dextran was released back into the medium with pseudo first-order kinetics. A vacuolar H(+)-ATPase is involved in endosomal acidification as the acidification process was markedly reduced in mutant strain HGR8, partially defective in vacuolar H(+)-ATPase and in parent type strain AX2 by bafilomycin A1, a selective inhibitor of this enzyme. Our data suggest that endocytic cargo is channeled from endosomes to secondary lysosomes that are actively linked to the plasma membrane via recycling vesicles.

Endocytosis and Ca2+ are required for endotoxin-stimulated TNF-alpha release by rat Kupffer cells

1996 ◽  
Vol 271 (5) ◽  
pp. G920-G928 ◽  
Author(s):  
S. N. Lichtman ◽  
J. Wang ◽  
C. Zhang ◽  
J. J. Lemasters
Keyword(s):  
Kupffer Cells ◽  
Cytochalasin B ◽  
De Novo ◽  
Tnf Alpha ◽  
Mrna Levels ◽  
Necrosis Factor Alpha ◽  
Endosomal Acidification ◽  
A Cell ◽  
Factor Alpha ◽  
Free Media

Endotoxin [lipopolysaccharide (LPS)] is a cell wall polymer derived from Gram-negative bacteria that stimulates macrophages to produce a variety of inflammatory mediators. In these studies, we examined LPS-stimulated formation of tumor necrosis factor-alpha (TNF-alpha) by cultured rat Kupffer cells. Cytochalasin B and methylpalmitate, blockers of endocytosis, decreased LPS-stimulated TNF-alpha release by > 92%. Bafilomycin A, monensin, and chloroquine, which prevent endosomal acidification, also blocked LPS-stimulated release of TNF-alpha by > 90%. Cytochalasin B and bafilomycin A decreased TNF-alpha mRNA levels by > 90% after LPS stimulation. Consistent with the requirement for LPS uptake and processing was the observation that Kupffer cells required 30 min of contact with LPS for maximal TNF-alpha release. LPS-stimulated TNF-alpha release was unaltered by incubation in Ca(2+)-free ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid medium, and A-23187, a Ca2+ ionophore, failed to stimulate TNF-alpha release in the absence of LPS. However, nisoldipine, a Ca2+ channel blocker, suppressed LPS-stimulated TNF-alpha release in cells cultured both in Ca(2+)-containing and Ca(2+)-free media. Although thapsigargin did not block TNF-alpha release, this depleter of intracellular Ca2+ stores blocked LPS-stimulated TNF-alpha synthesis in Ca(2+)-free medium and decreased TNF-alpha mRNA levels by 80%. Furthermore, LPS induced a late rise in intracellular free Ca2+ demonstrated by video microscopy of fura 2-loaded Kupffer cells. De novo protein and RNA synthesis were required, since cycloheximide and actinomycin D also inhibited LPS-stimulated TNF-alpha release. We compared free TNF-alpha secreted into culture supernatants with cell-associated TNF-alpha and found that cytochalasin B, bafilomycin A, chloroquine, monensin, and nisoldipine did not increase bound, cell-associated TNF-alpha. We conclude that endocytosis and endocytic processing may be necessary for LPS-stimulated TNF-alpha release from Kupffer cells. Ca2+ release, regulated by dihydropyridine-sensitive Ca2+ channels, also appears to be necessary for LPS-induced signaling and may arise from intracellular stores associated with the endosome/lysosome compartment.

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