Emerging chikungunya virus variants at the E1-E1 inter-glycoprotein spike interface impact virus attachment and Inflammation

Chikungunya virus (CHIKV) is a re-emerging arthropod-borne alphavirus and a serious threat to human health. Therefore, efforts toward elucidating how this virus causes disease and the molecular mechanisms underlying steps of the viral replication cycle are crucial. Using an in vivo transmission system that allows intra-host evolution, we identified an emerging CHIKV variant carrying a mutation in the E1 glycoprotein (V156A) in the serum of mice and saliva of mosquitoes. E1 V156A has since emerged in humans during an outbreak in Brazil, co-occurring with a second mutation, E1 K211T, suggesting an important role for these residues in CHIKV biology. Given the emergence of these variants, we hypothesized that they function to promote CHIKV infectivity and subsequent disease. Here, we show that E1 V156A and E1 K211T modulate virus attachment and fusion and impact binding to heparin, a homolog of heparan sulfate, a key entry factor on host cells. These variants also exhibit differential neutralization by anti-glycoprotein monoclonal antibodies, suggesting structural impacts on the particle that may be responsible for altered interactions at the host membrane. Finally, E1 V156A and E1 K211T exhibit increased titers in an adult arthritic mouse model and induce increased foot-swelling at the site of injection. Taken together, this work has revealed new roles for E1 where discrete regions of the glycoprotein are able to modulate cell attachment and swelling within the host. IMPORTANCE Alphaviruses represent a growing threat to human health worldwide. The re-emerging alphavirus chikungunya virus (CHIKV) has rapidly spread to new geographic regions in the last several decades, causing overwhelming outbreaks of disease, yet there are no approved vaccines or therapeutics. The CHIKV glycoproteins are key determinants of CHIKV adaptation and virulence. In this study, we identify and characterize the emerging E1 glycoprotein variants, V156A and K211T, that have since emerged in nature. We demonstrate that E1 V156A and K211T function in virus attachment to cells, a role that until now has been only attributed to specific residues of the CHIKV E2 glycoprotein. We also demonstrate E1 V156A and K211T to increase foot-swelling of the ipsilateral foot in mice infected with these variants. Observing that these variants and other pathogenic variants occur at the E1-E1 inter-spike interface, we highlight this structurally important region as critical for multiple steps during CHIKV infection. Together, these studies further defines the function of E1 in CHIKV infection and can inform the development of therapeutic or preventative strategies.

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Structural basis of Chikungunya virus inhibition by monoclonal antibodies

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2008051117 ◽

2020 ◽

Vol 117 (44) ◽

pp. 27637-27645

Author(s):

Qun Fei Zhou ◽

Julie M. Fox ◽

James T. Earnest ◽

Thiam-Seng Ng ◽

Arthur S. Kim ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Chikungunya Virus ◽

Neutralizing Antibody ◽

Structural Basis ◽

Chikv Infection ◽

Receptor Binding Site ◽

Viral Pathogen ◽

Virus Attachment

Chikungunya virus (CHIKV) is an emerging viral pathogen that causes both acute and chronic debilitating arthritis. Here, we describe the functional and structural basis as to how two anti-CHIKV monoclonal antibodies, CHK-124 and CHK-263, potently inhibit CHIKV infection in vitro and in vivo. Our in vitro studies show that CHK-124 and CHK-263 block CHIKV at multiple stages of viral infection. CHK-124 aggregates virus particles and blocks attachment. Also, due to antibody-induced virus aggregation, fusion with endosomes and egress are inhibited. CHK-263 neutralizes CHIKV infection mainly by blocking virus attachment and fusion. To determine the structural basis of neutralization, we generated cryogenic electron microscopy reconstructions of Fab:CHIKV complexes at 4- to 5-Å resolution. CHK-124 binds to the E2 domain B and overlaps with the Mxra8 receptor-binding site. CHK-263 blocks fusion by binding an epitope that spans across E1 and E2 and locks the heterodimer together, likely preventing structural rearrangements required for fusion. These results provide structural insight as to how neutralizing antibody engagement of CHIKV inhibits different stages of the viral life cycle, which could inform vaccine and therapeutic design.

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Foreign body responses in mouse central nervous system mimic natural wound responses and alter biomaterial functions

Nature Communications ◽

10.1038/s41467-020-19906-3 ◽

2020 ◽

Vol 11 (1) ◽

Author(s):

Timothy M. OʼShea ◽

Alexander L. Wollenberg ◽

Jae H. Kim ◽

Yan Ao ◽

Timothy J. Deming ◽

...

Keyword(s):

Central Nervous System ◽

Nervous System ◽

Foreign Body ◽

Molecular Mechanisms ◽

Host Cells ◽

Peripheral Tissues ◽

Molecular Delivery ◽

Wound Responses ◽

The Central Nervous System

AbstractBiomaterials hold promise for therapeutic applications in the central nervous system (CNS). Little is known about molecular factors that determine CNS foreign body responses (FBRs) in vivo, or about how such responses influence biomaterial function. Here, we probed these factors in mice using a platform of injectable hydrogels readily modified to present interfaces with different physiochemical properties to host cells. We found that biomaterial FBRs mimic specialized multicellular CNS wound responses not present in peripheral tissues, which serve to isolate damaged neural tissue and restore barrier functions. We show that the nature and intensity of CNS FBRs are determined by definable properties that significantly influence hydrogel functions, including resorption and molecular delivery when injected into healthy brain or stroke injuries. Cationic interfaces elicit stromal cell infiltration, peripherally derived inflammation, neural damage and amyloid production. Nonionic and anionic formulations show minimal levels of these responses, which contributes to superior bioactive molecular delivery. Our results identify specific molecular mechanisms that drive FBRs in the CNS and have important implications for developing effective biomaterials for CNS applications.

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Complementation of the fadA Mutation in Fusobacterium nucleatum Demonstrates that the Surface-Exposed Adhesin Promotes Cellular Invasion and Placental Colonization

Infection and Immunity ◽

10.1128/iai.00209-09 ◽

2009 ◽

Vol 77 (7) ◽

pp. 3075-3079 ◽

Cited By ~ 48

Author(s):

Akihiko Ikegami ◽

Peter Chung ◽

Yiping W. Han

Keyword(s):

Host Cell ◽

Cell Attachment ◽

Adverse Pregnancy Outcome ◽

Fusobacterium Nucleatum ◽

Host Cells ◽

Mouse Placenta ◽

Host Cell Attachment ◽

Adverse Pregnancy

ABSTRACT Fusobacterium nucleatum is a gram-negative oral anaerobe implicated in periodontal disease and adverse pregnancy outcome. The organism colonizes the mouse placenta, causing localized infection and inflammation. The mechanism of placental colonization has not been elucidated. Previous studies identified a novel adhesin from F. nucleatum, FadA, as being involved in the attachment and invasion of host cells. The fadA deletion mutant F. nucleatum 12230 US1 was defective in host cell attachment and invasion in vitro, but it also exhibited pleiotropic effects with altered cell morphology and growth rate. In this study, a fadA-complementing clone, F. nucleatum 12230 USF81, was constructed. The expression of FadA on USF81 was confirmed by Western blotting and immunofluorescent labeling. USF81 restored host cell attachment and invasion activities. The ability of F. nucleatum 12230, US1, and USF81 to colonize the mouse placenta was examined. US1 was severely defective in placental colonization compared to the wild type and USF81. Thus, FadA plays an important role in F. nucleatum colonization in vivo. These results also represent the first complementation studies for F. nucleatum. FadA may be a therapeutic target for preventing F. nucleatum colonization of the host.

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Proteoglycans in host–pathogen interactions: molecular mechanisms and therapeutic implications

Expert Reviews in Molecular Medicine ◽

10.1017/s1462399409001367 ◽

2010 ◽

Vol 12 ◽

Cited By ~ 62

Author(s):

Allison H. Bartlett ◽

Pyong Woo Park

Keyword(s):

Cell Surface ◽

Virulence Factors ◽

Molecular Mechanisms ◽

Biological Significance ◽

Host Cells ◽

Microbial Pathogens ◽

Therapeutic Implications ◽

Core Proteins

Many microbial pathogens subvert proteoglycans for their adhesion to host tissues, invasion of host cells, infection of neighbouring cells, dissemination into the systemic circulation, and evasion of host defence mechanisms. Where studied, specific virulence factors mediate these proteoglycan–pathogen interactions, which are thus thought to affect the onset, progression and outcome of infection. Proteoglycans are composites of glycosaminoglycan (GAG) chains attached covalently to specific core proteins. Proteoglycans are expressed ubiquitously on the cell surface, in intracellular compartments, and in the extracellular matrix. GAGs mediate the majority of ligand-binding activities of proteoglycans, and many microbial pathogens elaborate cell-surface and secreted factors that interact with GAGs. Some pathogens also modulate the expression and function of proteoglycans through known virulence factors. Several GAG-binding pathogens can no longer attach to and invade host cells whose GAG expression has been reduced by mutagenesis or enzymatic treatment. Furthermore, GAG antagonists have been shown to inhibit microbial attachment and host cell entry in vitro and reduce virulence in vivo. Together, these observations underscore the biological significance of proteoglycan–pathogen interactions in infectious diseases.

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Sulfated Glycans and Related Digestive Enzymes in the Zika Virus Infectivity: Potential Mechanisms of Virus-Host Interaction and Perspectives in Drug Discovery

Interdisciplinary Perspectives on Infectious Diseases ◽

10.1155/2017/4894598 ◽

2017 ◽

Vol 2017 ◽

pp. 1-8 ◽

Cited By ~ 2

Author(s):

Vitor H. Pomin

Keyword(s):

Neural Development ◽

Zika Virus ◽

Molecular Mechanisms ◽

Surface Proteins ◽

Host Cells ◽

Future Research ◽

Virus Infectivity ◽

Virus Attachment ◽

Chemical Structures ◽

Host Interaction

As broadly reported, there is an ongoing Zika virus (ZIKV) outbreak in countries of Latin America. Recent findings have demonstrated that ZIKV causes severe defects on the neural development in fetuses in utero and newborns. Very little is known about the molecular mechanisms involved in the ZIKV infectivity. Potential therapeutic agents are also under investigation. In this report, the possible mechanisms of action played by glycosaminoglycans (GAGs) displayed at the surface proteoglycans of host cells, and likely in charge of interactions with surface proteins of the ZIKV, are highlighted. As is common for the most viruses, these sulfated glycans serve as receptors for virus attachment onto the host cells and consequential entry during infection. The applications of (1) exogenous sulfated glycans of different origins and chemical structures capable of competing with the virus attachment receptors (supposedly GAGs) and (2) GAG-degrading enzymes able to digest the virus attachment receptors on the cells may be therapeutically beneficial as anti-ZIKV. This communication attempts, therefore, to offer some guidance for the future research programs aimed to unveil the molecular mechanisms underlying the ZIKV infectivity and to develop therapeutics capable of decreasing the devastating consequences caused by ZIKV outbreak in the Americas.

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Role of F1C Fimbriae, Flagella, and Secreted Bacterial Components in the Inhibitory Effect of Probiotic Escherichia coli Nissle 1917 on Atypical Enteropathogenic E. coli Infection

Infection and Immunity ◽

10.1128/iai.01431-13 ◽

2014 ◽

Vol 82 (5) ◽

pp. 1801-1812 ◽

Cited By ~ 23

Author(s):

Sylvia Kleta ◽

Marcel Nordhoff ◽

Karsten Tedin ◽

Lothar H. Wieler ◽

Rafal Kolenda ◽

...

Keyword(s):

Escherichia Coli ◽

Molecular Mechanisms ◽

Inhibitory Effect ◽

Host Cells ◽

Single Infection ◽

Content Type ◽

E Coli ◽

Initial Adhesion

ABSTRACTEnteropathogenicEscherichia coli(EPEC) is recognized as an important intestinal pathogen that frequently causes acute and persistent diarrhea in humans and animals. The use of probiotic bacteria to prevent diarrhea is gaining increasing interest. The probioticE. colistrain Nissle 1917 (EcN) is known to be effective in the treatment of several gastrointestinal disorders. While bothin vitroandin vivostudies have described strong inhibitory effects of EcN on enteropathogenic bacteria, including pathogenicE. coli, the underlying molecular mechanisms remain largely unknown. In this study, we examined the inhibitory effect of EcN on infections of porcine intestinal epithelial cells with atypical enteropathogenicE. coli(aEPEC) with respect to single infection steps, including adhesion, microcolony formation, and the attaching and effacing phenotype. We show that EcN drastically reduced the infection efficiencies of aEPEC by inhibiting bacterial adhesion and growth of microcolonies, but not the attaching and effacing of adherent bacteria. The inhibitory effect correlated with EcN adhesion capacities and was predominantly mediated by F1C fimbriae, but also by H1 flagella, which served as bridges between EcN cells. Furthermore, EcN seemed to interfere with the initial adhesion of aEPEC to host cells by secretion of inhibitory components. These components do not appear to be specific to EcN, but we propose that the strong adhesion capacities enable EcN to secrete sufficient local concentrations of the inhibitory factors. The results of this study are consistent with a mode of action whereby EcN inhibits secretion of virulence-associated proteins of EPEC, but not their expression.

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Potent germline-like monoclonal antibodies: Rapid identification of promising candidates for antibody-based antiviral therapy

Antibody Therapeutics ◽

10.1093/abt/tbab008 ◽

2021 ◽

Author(s):

Xiaoyi Zhu ◽

Fei Yu ◽

Yanling Wu ◽

Tianlei Ying

Keyword(s):

Monoclonal Antibodies ◽

Rational Design ◽

Large Scale ◽

Molecular Mechanisms ◽

Viral Infections ◽

Somatic Hypermutation ◽

Affinity Maturation ◽

Human Mabs

Abstract Recent years, fully human monoclonal antibodies (mAbs) are making up an increasing share of the pharmaceutical market. However, to improve affinity and efficacy of antibodies, many somatic hypermutation could be introduced during affinity maturation, which cause several issues including safety and efficacy and limit their application in clinic. Here, we propose a special class of human mAbs with limited level of somatic mutations, referred to as germline-like mAbs. Remarkably, germline-like mAbs could have high affinity and potent neutralizing activity in vitro and in various animal models, despite lacking of extensive affinity maturation. Furthermore, the germline nature of these mAbs implies that they exhibit lower immunogenicity and can be elicited relatively fast in vivo compared with highly somatically mutated antibodies. In this review, we summarize germline-like mAbs with strong therapeutic and protection activity against various viruses that caused large-scale outbreaks in the last decade, including influenza virus H7N9, Zika virus (ZIKV), Dengue virus (DENV), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We also illustrate underlying molecular mechanisms of these germline-like antibodies against viral infections from the structural and genetic perspective, thus providing insight into further development as therapeutic agents for treatment of infectious diseases and implication for rational design of effective vaccines.

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The dual function monoclonal antibodies VIR-7831 and VIR-7832 demonstrate potent in vitro and in vivo activity against SARS-CoV-2

10.1101/2021.03.09.434607 ◽

2021 ◽

Cited By ~ 7

Author(s):

Andrea L. Cathcart ◽

Colin Havenar-Daughton ◽

Florian A. Lempp ◽

Daphne Ma ◽

Michael Schmid ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Severe Acute Respiratory Syndrome ◽

Host Cells ◽

Infection Model ◽

Dual Function ◽

Resistance Mutations ◽

Viral Respiratory Infection ◽

Dual Action

ABSTRACTVIR-7831 and VIR-7832 are dual action monoclonal antibodies (mAbs) targeting the spike glycoprotein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). VIR-7831 and VIR-7832 were derived from a parent antibody (S309) isolated from memory B cells of a 2003 severe acute respiratory syndrome coronavirus (SARS-CoV) survivor. Both mAbs contain an “LS” mutation in the Fc region to prolong serum half-life and potentially enhance distribution to the respiratory mucosa. In addition, VIR-7832 encodes an Fc GAALIE mutation that has been shown previously to evoke CD8+ T-cells in the context of an in vivo viral respiratory infection. VIR-7831 and VIR-7832 potently neutralize live wild-type SARS-CoV-2 in vitro as well as pseudotyped viruses encoding spike protein from the B.1.1.7, B.1.351 and P.1 variants. In addition, they retain activity against monoclonal antibody resistance mutations that confer reduced susceptibility to currently authorized mAbs. The VIR-7831/VIR-7832 epitope does not overlap with mutational sites in the current variants of concern and continues to be highly conserved among circulating sequences consistent with the high barrier to resistance observed in vitro. Furthermore, both mAbs can recruit effector mechanisms in vitro that may contribute to clinical efficacy via elimination of infected host cells. In vitro studies with these mAbs demonstrated no enhancement of infection. In a Syrian Golden hamster proof-of concept wildtype SARS-CoV-2 infection model, animals treated with VIR-7831 had less weight loss, and significantly decreased total viral load and infectious virus levels in the lung compared to a control mAb. Taken together, these data indicate that VIR-7831 and VIR-7832 are promising new agents in the fight against COVID-19.

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Efficient inhibition of SARS-CoV-2 strains by a novel ACE2-IgG4-Fc fusion protein with a stabilized hinge region

10.1101/2020.12.06.413443 ◽

2020 ◽

Author(s):

Hristo L. Svilenov ◽

Julia Sacherl ◽

Alwin Reiter ◽

Lisa Wolff ◽

Cho-Chin Chen ◽

...

Keyword(s):

Receptor Activation ◽

Fc Receptor ◽

Host Cells ◽

Fusion Partner ◽

The Novel ◽

Single Digit ◽

Angiotensin Converting Enzyme 2 ◽

Host Membrane ◽

Pharmaceutical Properties

AbstractThe novel severe acute respiratory syndrome (SARS)-like coronavirus (SARS-CoV-2) enters its host cells after binding to the angiotensin-converting enzyme 2 (ACE2) via its spike glycoprotein. This interaction is critical for virus entry and virus-host membrane fusion. Soluble ACE2 ectodomains bind and neutralize the virus but the short in vivo half-lives of soluble ACE2 limits its therapeutic use. Fusion of the fragment crystallizable (Fc) part of human immunoglobulin G (IgG) to the ACE2 ectodomain can prolong the in vivo half-life but bears the risk of unwanted Fc-receptor activation and antibody-dependent disease enhancement. Here, we describe optimized ACE2-Fc fusion constructs that avoid Fc-receptor binding by using IgG4-Fc as a fusion partner. The engineered ACE2-IgG4-Fc fusion proteins described herein exhibit promising pharmaceutical properties and a broad antiviral activity at single-digit nanomolar concentration. In addition, they allow to maintain the beneficial enzymatic activity of ACE2 and thus are very promising candidate antivirals broadly acting against coronaviruses.

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