Structure of Reovirus σ1 in Complex with Its Receptor Junctional Adhesion Molecule-A

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Many viruses invade mucosal surfaces to establish infection in the host. Some viruses are restricted to mucosal surfaces, whereas others disseminate to sites of secondary replication. Studies of strain-specific differences in reovirus mucosal infection and systemic dissemination have enhanced an understanding of viral determinants and molecular mechanisms that regulate viral pathogenesis. After peroral inoculation, reovirus strain type 1 Lang replicates to high titers in the intestine and spreads systemically, whereas strain type 3 Dearing (T3D) does not. These differences segregate with the viral S1 gene segment, which encodes attachment protein 1 and nonstructural protein 1s. In this study, we define genetic determinants that regulate reovirus-induced pathology following intranasal inoculation and respiratory infection. We report that two laboratory isolates of T3D, T3D C and T3D F , differ in the capacity to replicate in the respiratory tract and spread systemically; the T3D C isolate replicates to higher titers in the lungs and disseminates, while T3D F does not. Two nucleotide polymorphisms in the S1 gene influence these differences, and both S1 gene products are involved. T3D C amino acid polymorphisms in the tail and head domains of 1 protein influence the sensitivity of virions to protease-mediated loss of infectivity. The T3D C polymorphism at nucleotide 77, which leads to coding changes in both S1 gene products, promotes systemic dissemination from the respiratory tract. A 1s-null virus produces lower titers in the lung after intranasal inoculation and disseminates less efficiently to sites of secondary replication. These findings provide new insights into mechanisms underlying reovirus replication in the respiratory tract and systemic spread from the lung. Many viruses enter host organisms by invading mucosal surfaces, including those that line the respiratory tract. Infection by some pneumotropic viruses is restricted to the respiratory tract, whereas others replicate in the lung and disseminate to sites of secondary replication. Mammalian orthoreoviruses (reoviruses) naturally infect both the respiratory and gastrointestinal tracts (1). Reovirus strains differ in the capacity to replicate at mucosal sites and disseminate systemically. Studies of strain-specific differences in reovirus mucosal infection and systemic spread have enhanced an understanding of viral determinants and molecular mechanisms that regulate reovirus pathogenesis. For example, after peroral or intratracheal inoculation, reovirus strain type 1 Lang (T1L) replicates to higher titers than does strain type 3 Dearing (T3D) (2). This difference in replication efficiency at the site of primary replication segregates with the reovirus S1 gene segment (2, 3). Like T1L, reassortant virus 3HA1, with nine gene segments from T3D and an S1 gene from T1L, replicates to high titers in mucosal tissues (2). In contrast, reassortant virus 1HA3, with nine gene segments from T1L and an S1 gene from T3D, fails to replicate to high titers at those sit...

Section: Discussionmentioning

confidence: 89%

Genetic Determinants of Reovirus Pathogenesis in a Murine Model of Respiratory Infection

Nygaard

Lahti

Ikizler

et al. 2013

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“…The binding site for 1 is located on the most membrane-distal Ig-like domain of JAM-A, which extends from the cell by about 80 to 90 Å. JAM-A is engaged by residues near the bottom of the 1 head domain, adjacent to the ␤-spiral repeat region of the tail. Modeling indicates that reovirus binding to JAM-A on the cell surface would bring the top of the 1 head domain into close proximity with the cell membrane (26). When 1 is fully ligated with 5C6 or 9BG5 antibodies, which bind near the top and middle of the head domain, respectively, the virus almost certainly could not engage JAM-A; the membrane-anchored receptor could not reach its binding site on the 1 surface (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…The JAM-A-binding site is conserved between the reovirus serotypes and located at the lower part of the 1 head domain (26,29). The 9BG5 epitope is located closer to the JAM-A-binding site than is the 5C6 epitope.…”

Section: Blockade Of Reovirus Infectivity By 5c6 and 9bg5 Mabs And Fabsmentioning

confidence: 99%

Structural Insights into Reovirus σ1 Interactions with Two Neutralizing Antibodies

Dietrich

Ogden

Katen

et al. 2017

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Reovirus attachment protein 1 engages glycan receptors and junctional adhesion molecule-A (JAM-A) and is thought to undergo a conformational change during the proteolytic disassembly of virions to infectious subvirion particles (ISVPs) that accompanies cell entry. The 1 protein is also the primary target of neutralizing antibodies. Here, we present a structural and functional characterization of two neutralizing antibodies that target 1 of serotype 1 (T1) and serotype 3 (T3) reoviruses. The crystal structures revealed that each antibody engages its cognate 1 protein within the head domain via epitopes distinct from the JAM-A-binding site. Surface plasmon resonance and cell-binding assays indicated that both antibodies likely interfere with JAM-A engagement by steric hindrance. To define the interplay between the carbohydrate receptor and antibody binding, we conducted hemagglutination inhibition assays using virions and ISVPs. The glycan-binding site of T1 1 is located in the head domain and is partly occluded by the bound Fab in the crystal structure. The T1-specific antibody inhibited hemagglutination by virions and ISVPs, probably via direct interference with glycan engagement. In contrast to T1 1, the carbohydrate-binding site of T3 1 is located in the tail domain, distal to the antibody epitope. The T3-specific antibody inhibited hemagglutination by T3 virions but not ISVPs, indicating that the antibody-and glycan-binding sites in 1 are in closer spatial proximity on virions than on ISVPs. Our results provide direct evidence for a structural rearrangement of 1 during virion-to-ISVP conversion and contribute new information about the mechanisms of antibody-mediated neutralization of reovirus.IMPORTANCE Virus attachment proteins mediate binding to host cell receptors, serve critical functions in cell and tissue tropism, and are often targeted by the neutralizing antibody response. The structural investigation of antibody-antigen complexes can provide valuable information for understanding the molecular basis of virus neutralization. Studies with enveloped viruses, such as HIV and influenza virus, have helped to define sites of vulnerability and guide vaccination strategies. By comparison, less is known about antibody binding to nonenveloped viruses. Here, we structurally investigated two neutralizing antibodies that bind the attachment protein 1 of reovirus. Furthermore, we characterized the neutralization efficiency, the binding affinity for 1, and the effect of the antibodies on reovirus receptor engagement. Our analysis defines reovirus interactions with two neutralizing antibodies, allows us to propose a mechanism by which they block virus infection, and provides evidence for a conformational change in the 1 protein during viral cell entry.

“…Though the T1L and T3D 1 polypeptide chains have low sequence homology, they fold into nearly identical trimeric fibers (10,33). The 1 proteins of both the T1L and T3D prototype reovirus strains engage JAM-A via their head domains (13,33,34). However, the T1L and T3D 1 proteins engage distinct glycan receptors via disparate portions of the 1 trimer (13,14).…”

Section: Generation Of Isvps In Vitromentioning

confidence: 99%

Reovirus μ1 Protein Affects Infectivity by Altering Virus-Receptor Interactions

Thete

Snyder

Mainou

et al. 2016

Proteins that form the reovirus outer capsid play an active role in the entry of reovirus into host cells. Among these, the 1 protein mediates attachment of reovirus particles to host cells via interaction with cell surface glycans or the proteinaceous receptor junctional adhesion molecule A (JAM-A). The 1 protein functions to penetrate the host cell membrane to allow delivery of the genome-containing viral core particle into the cytoplasm to initiate viral replication. We demonstrate that a reassortant virus that expresses the M2 gene-encoded 1 protein derived from prototype strain T3D in an otherwise prototype T1L background (T1L/T3DM2) infects cells more efficiently than parental T1L. Unexpectedly, the enhancement in infectivity of T1L/T3DM2 is due to its capacity to attach to cells more efficiently. We present genetic data implicating the central region of 1 in altering the cell attachment property of reovirus. Our data indicate that the T3D 1-mediated enhancement in infectivity of T1L is dependent on the function of 1 and requires the expression of JAM-A. We also demonstrate that T1L/T3DM2 utilizes JAM-A more efficiently than T1L. These studies revealed a previously unknown relationship between two nonadjacent reovirus outer capsid proteins, 1 and 1. IMPORTANCEHow reovirus attaches to host cells has been extensively characterized. Attachment of reovirus to host cells is mediated by the 1 protein, and properties of 1 influence the capacity of reovirus to target specific host tissues and produce disease. Here, we present new evidence indicating that the cell attachment properties of 1 are influenced by the nature of 1, a capsid protein that does not physically interact with 1. These studies could explain the previously described role for 1 in influencing reovirus pathogenesis. These studies are also of broader significance because they highlight an example of how genetic reassortment between virus strains could produce phenotypes that are distinct from those of either parent.A ttachment of virus is the first step in the infection of host cells. Cell attachment occurs via interactions of viral attachment factors with host cell receptors. For enveloped viruses, viral glycoproteins embedded in the lipid membrane serve as attachment factors (1). For nonenveloped viruses, specific structural features on the capsid or sequences within the exposed portion of the viral structural proteins bind host receptors (1). Mutations within the receptor-binding site can alter the efficiency with which virus attaches to host cells and consequently modulate the capacity of the virus to establish infection. In viral systems where capsids are formed from multiple structural proteins, these proteins fit together in a precise geometric arrangement. Thus, changes to the properties of one capsid protein can influence the function of other capsid proteins. In this report, we highlight one such example by demonstrating a previously unknown functional relationship between two nonadjacent viral capsid proteins of mammalian orthoreo...