scholarly journals Structural Investigation of Orf Virus Bcl-2 Homolog ORFV125 Interactions with BH3-Motifs from BH3-Only Proteins Puma and Hrk

Viruses ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1374
Author(s):  
Chathura D. Suraweera ◽  
Mark G. Hinds ◽  
Marc Kvansakul
Keyword(s):  
Cell Death ◽  
Structural Investigation ◽  
Potent Inhibitor ◽  
Host Cells ◽  
Orf Virus ◽  
Structural Basis ◽  
Signalling Pathways ◽  
Ionic Interaction ◽  
Host Cell Apoptosis ◽  
Binding Groove

Numerous viruses have evolved sophisticated countermeasures to hijack the early programmed cell death of host cells in response to infection, including the use of proteins homologous in sequence or structure to Bcl-2. Orf virus, a member of the parapoxviridae, encodes for the Bcl-2 homolog ORFV125, a potent inhibitor of Bcl-2-mediated apoptosis in the host. ORFV125 acts by directly engaging host proapoptotic Bcl-2 proteins including Bak and Bax as well as the BH3-only proteins Hrk and Puma. Here, we determined the crystal structures of ORFV125 bound to the BH3 motif of proapoptotic proteins Puma and Hrk. The structures reveal that ORFV125 engages proapoptotic BH3 motif peptides using the canonical ligand binding groove. An Arg located in the structurally equivalent BH1 region of ORFV125 forms an ionic interaction with the conserved Asp in the BH3 motif in a manner that mimics the canonical ionic interaction seen in host Bcl-2:BH3 motif complexes. These findings provide a structural basis for Orf virus-mediated inhibition of host cell apoptosis and reveal the flexibility of virus encoded Bcl-2 proteins to mimic key interactions from endogenous host signalling pathways.

scholarly journals Crystal structures of ORFV125 provide insight into orf virus-mediated inhibition of apoptosis

2020 ◽  
Vol 477 (23) ◽  
pp. 4527-4541 ◽  
Author(s):  
Chathura D. Suraweera ◽  
Mark G. Hinds ◽  
Marc Kvansakul
Keyword(s):  
Host Cell ◽  
Crystal Structures ◽  
B Cell Lymphoma ◽  
Orf Virus ◽  
Structural Basis ◽  
Inhibitory Protein ◽  
Affinity Ligand ◽  
Host Cell Apoptosis ◽  
Mechanistic Basis ◽  
Binding Groove

Premature apoptosis of cells is a strategy utilized by multicellular organisms to counter microbial threats. Orf virus (ORFV) is a large double-stranded DNA virus belonging to the poxviridae. ORFV encodes for an apoptosis inhibitory protein ORFV125 homologous to B-cell lymphoma 2 or Bcl-2 family proteins, which has been shown to inhibit host cell encoded pro-apoptotic Bcl-2 proteins. However, the structural basis of apoptosis inhibition by ORFV125 remains to be clarified. We show that ORFV125 is able to bind to a range of peptides spanning the BH3 motif of human pro-apoptotic Bcl-2 proteins including Bax, Bak, Puma and Hrk with modest to weak affinity. We then determined the crystal structures of ORFV125 alone as well as bound to the highest affinity ligand Bax BH3 motif. ORFV125 adopts a globular Bcl-2 fold comprising 7 α-helices, and utilizes the canonical Bcl-2 binding groove to engage pro-apoptotic host cell Bcl-2 proteins. In contrast with a previously predicted structure, ORFV125 adopts a domain-swapped dimeric topology, where the α1 helix from one protomer is swapped into a neighbouring unit. Furthermore, ORFV125 differs from the conserved architecture of the Bcl-2 binding groove and instead of α3 helix forming one of the binding groove walls, ORFV125 utilizes an extended α2 helix that comprises the equivalent region of helix α3. This results in a subtle variation of previously observed dimeric Bcl-2 architectures in other poxvirus and human encoded Bcl-2 proteins. Overall, our results provide a structural and mechanistic basis for orf virus-mediated inhibition of host cell apoptosis.

scholarly journals Crystal structures of the sheeppoxvirus encoded inhibitor of apoptosis SPPV14 bound to Hrk and Bax BH3 peptides

2020 ◽  
Author(s):  
Chathura D. Suraweera ◽  
Denis R. Burton ◽  
Mark G. Hinds ◽  
Marc Kvansakul
Keyword(s):  
Crystal Structures ◽  
Viral Infections ◽  
Structural Basis ◽  
Ionic Interaction ◽  
Apoptosis Inhibition ◽  
Programmed Death ◽  
Infected Cells ◽  
Binding Groove ◽  
Almost All ◽  
Multicellular Organisms

AbstractProgrammed death of infected cells is used by multicellular organisms to counter viral infections. Sheeppoxvirus encodes for SPPV14, a potent inhibitor of Bcl-2 mediated apoptosis. We reveal the structural basis of apoptosis inhibition by determining crystal structures of SPPV14 bound to BH3 motifs of proapoptotic Bax and Hrk. The structures show that SPPV14 engages BH3 peptides using the canonical ligand binding groove. Unexpectedly, Arg84 from SPPV14 forms an ionic interaction with the conserved Asp in the BH3 motif in a manner that replaces the canonical ionic interaction seen in almost all host Bcl-2:BH3 motif complexes. These results reveal the flexibility of virus encoded Bcl-2 proteins to mimic key interactions from endogenous host signalling pathways to retain BH3-binding and pro-survival functionality.

scholarly journals Vibrio vulnificus Hemolysin Induces TNF-α Independent ROS Production by Primary Murine Macrophages

2021 ◽  
Author(s):  
Yong-Liang Lou ◽  
Dan-Li Xie ◽  
Xian-Hui Huang ◽  
Meng-Meng Zheng ◽  
Ting Zhang ◽  
...  
Keyword(s):  
Cell Death ◽  
Vibrio Vulnificus ◽  
Opportunistic Pathogen ◽  
Cytolytic Activity ◽  
Host Cells ◽  
Ros Generation ◽  
Ros Production ◽  
Host Cell Apoptosis ◽  
Tnf Α ◽  
The Impact

Abstract BackgroundVibrio vulnificus (V. vulnificus) is a gram-negative opportunistic pathogen that causes lethal infections in humans. Vibrio vulnificus hemolysin (VVH) is a key virulence factor that exhibits strong hemolytic and cytolytic activities and contributes to the pathogen's invasion, vasodilatation, and septic shock. Most of the studies so far have focused on VVH's cytolytic activity against cell lines derived from host cells. However, the cytolytic activity of VVH on primary macrophages is still unclear. In addition, although it is known that VVH induces host cell apoptosis via triggering ROS production, the impact of VVH on host immune response has not been fully understood. This study aimed to investigate the role of VVH-induced TNF-α expression and ROS production in the absence of apoptosis of murine primary macrophages and related signaling pathways using FACS, DCFH-DA, real-time PCR, and western blotting. ResultsThe results showed that murine primary macrophages from different organs displayed differential sensitivities towards VVH-induced cell death. Liver Kupffer cells, splenic macrophages, and BMMfs were more sensitive to VVH-induced cytotoxicity, while alveolar macrophages, lung interstitial macrophages, and lung neutrophils were resistant to VVH-induced cell death. Besides, we found that a low dose of VVH, which did not induce apoptosis in murine primary macrophages, could induce apoptosis independent TNF-α expression and ROS generation. Such ROS production in macrophages could be further blocked by inhibiting p38-MAPKs or NFκB activation but was not affected by knockout of TNF-α. ConclusionsVVH produced cytotoxicity in macrophages, an apoptosis-independent TNF-α expression, and ROS production, which provides insight into the mechanism underlying the crosstalk between VVH-induced inflammation and oxidative stress.

Structural basis ofDeerpox virus-mediated inhibition of apoptosis

2015 ◽  
Vol 71 (8) ◽  
pp. 1593-1603 ◽  
Author(s):  
Denis R. Burton ◽  
Sofia Caria ◽  
Bevan Marshall ◽  
Michele Barry ◽  
Marc Kvansakul
Keyword(s):  
Structural Analysis ◽  
Ligand Binding ◽  
Structural Basis ◽  
Inhibitor Of Apoptosis ◽  
Defence Mechanism ◽  
Partial Obstruction ◽  
Host Cell Apoptosis ◽  
Infected Cells ◽  
Binding Groove ◽  
Innate Defence

Apoptosis is a key innate defence mechanism to eliminate virally infected cells. To counteract premature host-cell apoptosis, poxviruses have evolved numerous molecular strategies, including the use of Bcl-2 proteins, to ensure their own survival. Here, it is reported that theDeerpox virusinhibitor of apoptosis, DPV022, only engages a highly restricted set of death-inducing Bcl-2 proteins, including Bim, Bax and Bak, with modest affinities. Structural analysis reveals that DPV022 adopts a Bcl-2 fold with a dimeric domain-swapped topology and binds pro-death Bcl-2 proteinsviatwo conserved ligand-binding grooves found on opposite sides of the dimer. Structures of DPV022 bound to Bim, Bak and Bax BH3 domains reveal that a partial obstruction of the binding groove is likely to be responsible for the modest affinities of DPV022 for BH3 domains. These findings reveal that domain-swapped dimeric Bcl-2 folds are not unusual and may be found more widely in viruses. Furthermore, the modest affinities of DPV022 for pro-death Bcl-2 proteins suggest that two distinct classes of anti-apoptotic viral Bcl-2 proteins exist: those that are monomeric and tightly bind a range of death-inducing Bcl-2 proteins, and others such as DPV022 that are dimeric and only bind a very limited number of death-inducing Bcl-2 proteins with modest affinities.

Toxoplasma Gondii Bradyzoites Elicit Transcriptional Changes in Host Cells to Prevent IFNγ-Mediated Cell Death

2019 ◽  
Author(s):  
Simona Seizova ◽  
Alexandra L Garnham ◽  
Michael J Coffey ◽  
Lachlan W Whitehead ◽  
Kelly L Rogers ◽  
...  
Keyword(s):  
Cell Death ◽  
Toxoplasma Gondii ◽  
Host Cells ◽  
Transcriptional Changes

scholarly journals Pathogen induced subversion of NAD+ metabolism mediating host cell death: a target for development of chemotherapeutics

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Ayushi Chaurasiya ◽  
Swati Garg ◽  
Ashish Khanna ◽  
Chintam Narayana ◽  
Ved Prakash Dwivedi ◽  
...  
Keyword(s):  
Cell Death ◽  
Host Cell ◽  
Nicotinamide Adenine Dinucleotide ◽  
Malaria Parasite ◽  
Host Cells ◽  
Nicotinamide Adenine ◽  
Targeted Therapeutics ◽  
Nad Metabolism ◽  
Host Cell Death ◽  
Nanomolar Range

AbstractHijacking of host metabolic status by a pathogen for its regulated dissemination from the host is prerequisite for the propagation of infection. M. tuberculosis secretes an NAD+-glycohydrolase, TNT, to induce host necroptosis by hydrolyzing Nicotinamide adenine dinucleotide (NAD+). Herein, we expressed TNT in macrophages and erythrocytes; the host cells for M. tuberculosis and the malaria parasite respectively, and found that it reduced the NAD+ levels and thereby induced necroptosis and eryptosis resulting in premature dissemination of pathogen. Targeting TNT in M. tuberculosis or induced eryptosis in malaria parasite interferes with pathogen dissemination and reduction in the propagation of infection. Building upon our discovery that inhibition of pathogen-mediated host NAD+ modulation is a way forward for regulation of infection, we synthesized and screened some novel compounds that showed inhibition of NAD+-glycohydrolase activity and pathogen infection in the nanomolar range. Overall this study highlights the fundamental importance of pathogen-mediated modulation of host NAD+ homeostasis for its infection propagation and novel inhibitors as leads for host-targeted therapeutics.

scholarly journals Cell Death Signaling Pathway Induced by Cholix Toxin, a Cytotoxin and eEF2 ADP-Ribosyltransferase Produced by Vibrio cholerae

Toxins ◽  
2020 ◽  
Vol 13 (1) ◽  
pp. 12
Author(s):  
Kohei Ogura ◽  
Kinnosuke Yahiro ◽  
Joel Moss
Keyword(s):  
Cell Death ◽  
Protein Kinase ◽  
Vibrio Cholerae ◽  
Elongation Factor ◽  
Host Cells ◽  
Mitogen Activated Protein Kinase ◽  
Pseudomonas Exotoxin A ◽  
Tnf Α ◽  
Induced Apoptosis ◽  
Adp Ribosyltransferase

Pathogenic microorganisms produce various virulence factors, e.g., enzymes, cytotoxins, effectors, which trigger development of pathologies in infectious diseases. Cholera toxin (CT) produced by O1 and O139 serotypes of Vibrio cholerae (V. cholerae) is a major cytotoxin causing severe diarrhea. Cholix cytotoxin (Cholix) was identified as a novel eukaryotic elongation factor 2 (eEF2) adenosine-diphosphate (ADP)-ribosyltransferase produced mainly in non-O1/non-O139 V. cholerae. The function and role of Cholix in infectious disease caused by V. cholerae remain unknown. The crystal structure of Cholix is similar to Pseudomonas exotoxin A (PEA) which is composed of an N-terminal receptor-recognition domain and a C-terminal ADP-ribosyltransferase domain. The endocytosed Cholix catalyzes ADP-ribosylation of eEF2 in host cells and inhibits protein synthesis, resulting in cell death. In a mouse model, Cholix caused lethality with severe liver damage. In this review, we describe the mechanism underlying Cholix-induced cytotoxicity. Cholix-induced apoptosis was regulated by mitogen-activated protein kinase (MAPK) and protein kinase C (PKC) signaling pathways, which dramatically enhanced tumor necrosis factor-α (TNF-α) production in human liver, as well as the amount of epithelial-like HepG2 cancer cells. In contrast, Cholix induced apoptosis in hepatocytes through a mitochondrial-dependent pathway, which was not stimulated by TNF-α. These findings suggest that sensitivity to Cholix depends on the target cell. A substantial amount of information on PEA is provided in order to compare/contrast this well-characterized mono-ADP-ribosyltransferase (mART) with Cholix.

scholarly journals Structural basis for distinct inflammasome complex assembly by human NLRP1 and CARD8

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Qin Gong ◽  
Kim Robinson ◽  
Chenrui Xu ◽  
Phuong Thao Huynh ◽  
Kelvin Han Chung Chong ◽  
...  
Keyword(s):  
Cell Death ◽  
Inner Core ◽  
Inflammatory Diseases ◽  
Structural Features ◽  
Structural Basis ◽  
Cultured Human Cells ◽  
Structural Insight ◽  
Sensor Proteins ◽  
Insight Into

AbstractNod-like receptor (NLR) proteins activate pyroptotic cell death and IL-1 driven inflammation by assembling and activating the inflammasome complex. Closely related sensor proteins NLRP1 and CARD8 undergo unique auto-proteolysis-dependent activation and are implicated in auto-inflammatory diseases; however, their mechanisms of activation are not understood. Here we report the structural basis of how the activating domains (FIINDUPA-CARD) of NLRP1 and CARD8 self-oligomerize to assemble distinct inflammasome complexes. Recombinant FIINDUPA-CARD of NLRP1 forms a two-layered filament, with an inner core of oligomerized CARD surrounded by an outer ring of FIINDUPA. Biochemically, self-assembled NLRP1-CARD filaments are sufficient to drive ASC speck formation in cultured human cells—a process that is greatly enhanced by NLRP1-FIINDUPA which forms oligomers in vitro. The cryo-EM structures of NLRP1-CARD and CARD8-CARD filaments, solved here at 3.7 Å, uncover unique structural features that enable NLRP1 and CARD8 to discriminate between ASC and pro-caspase-1. In summary, our findings provide structural insight into the mechanisms of activation for human NLRP1 and CARD8 and reveal how highly specific signaling can be achieved by heterotypic CARD interactions within the inflammasome complexes.

Structural Basis for the Understanding of Entry Inhibitors Against SARS Viruses

2021 ◽  
Vol 28 ◽  
Author(s):  
Prem Kumar Kushwaha ◽  
Neha Kumari ◽  
Sneha Nayak ◽  
Keshav Kishor ◽  
Ashoke Sharon
Keyword(s):  
Viral Entry ◽  
Molecular Level ◽  
Virus Entry ◽  
Holistic Approach ◽  
Symptomatic Treatment ◽  
Host Cells ◽  
Structural Basis ◽  
Viral Fusion ◽  
Structural Studies ◽  
Entry Inhibitors

: Outbreaks due to Severe Acute Respiratory Syndrome-Corona virus 2 (SARS-CoV-2) initiated in Wuhan city, China, in December 2019 which continued to spread internationally, posing a pandemic threat as declared by WHO and as of March 10, 2021, confirmed cases reached 118 million along with 2.6 million deaths worldwide. In the absence of specific antiviral medication, symptomatic treatment and physical isolation remain the options to control the contagion. The recent clinical trials on antiviral drugs highlighted some promising compounds such as umifenovir (haemagglutinin-mediated fusion inhibitor), remdesivir (RdRp nucleoside inhibitor), and favipiravir (RdRp Inhibitor). WHO launched a multinational clinical trial on several promising analogs as a potential treatment to combat SARS infection. This situation urges a holistic approach to invent safe and specific drugs as a prophylactic and therapeutic cure for SARS-related-viral diseases, including COVID-19. : It is significant to note that researchers worldwide have been doing their best to handle the crisis and have produced an extensive and promising literature body. It opens a scope and allows understanding the viral entry at the molecular level. A structure-based approach can reveal the molecular-level understanding of viral entry interaction. The ligand profiling and non-covalent interactions among participating amino-acid residues are critical information to delineate a structural interpretation. The structural investigation of SARS virus entry into host cells will reveal the possible strategy for designing drugs like entry inhibitors. : The structure-based approach demonstrates details at the 3D molecular level. It shows specificity about SARS-CoV-2 spike interaction, which uses human angiotensin-converting enzyme 2 (ACE2) as a receptor for entry, and the human protease completes the process of viral fusion and infection. : The 3D structural studies reveal the existence of two units, namely S1 and S2. S1 is called a receptor-binding domain (RBD) and responsible for interacting with the host (ACE2), and the S2 unit participates in the fusion of viral and cellular membranes. TMPRSS2 mediates the cleavage at S1/S2 subunit interface in S-protein of SARS CoV-2, leading to viral fusion. Conformational difference associated with S1 binding alters ACE2 interaction and inhibits viral fusion. Overall, the detailed 3D structural studies help understand the 3D structural basis of interaction between viruses with host factors and available scope for the new drug discovery process targeting SARS-related virus entry into the host cell.

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