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Molecular analysis of the interaction of LCMV with its cellular receptor [alpha]-dystroglycan.

Kunz S, Sevilla N, McGavern DB, Campbell KP, Oldstone MB - J. Cell Biol. (2001)

Bottom Line: In the present study, we characterized the binding of LCMV to alpha-DG and addressed the role of alpha-DG-associated host-derived proteins in virus infection.We found that the COOH-terminal region of alpha-DG's first globular domain and the NH2-terminal region of the mucin-related structures of alpha-DG together form the binding site for LCMV.This competition of the virus with ECM molecules for receptor binding likely explains the recently found correlation between the affinity of LCMV binding to alpha-DG, tissue tropism, and pathological potential.

View Article: PubMed Central - PubMed

Affiliation: The Scripps Research Institute, Division of Virology, Department of Neuropharmacology, La Jolla, CA 92037, USA.

ABSTRACT
alpha-Dystroglycan (DG) has been identified as the cellular receptor for lymphocytic choriomeningitis virus (LCMV) and Lassa fever virus (LFV). This subunit of DG is a highly versatile cell surface molecule that provides a molecular link between the extracellular matrix (ECM) and a beta-DG transmembrane component, which interacts with the actin-based cytoskeleton. In addition, DG exhibits a complex pattern of interaction with a wide variety of ECM and cellular proteins. In the present study, we characterized the binding of LCMV to alpha-DG and addressed the role of alpha-DG-associated host-derived proteins in virus infection. We found that the COOH-terminal region of alpha-DG's first globular domain and the NH2-terminal region of the mucin-related structures of alpha-DG together form the binding site for LCMV. The virus-alpha-DG binding unlike ECM alpha-DG interactions was not dependent on divalent cations. Despite such differences in binding, LCMV and laminin-1 use, in part, an overlapping binding site on alpha-DG, and the ability of an LCMV isolate to compete with laminin-1 for receptor binding is determined by its binding affinity to alpha-DG. This competition of the virus with ECM molecules for receptor binding likely explains the recently found correlation between the affinity of LCMV binding to alpha-DG, tissue tropism, and pathological potential. LCMV strains and variants with high binding affinity to alpha-DG but not low affinity binders are able to infect CD11c+ dendritic cells, which express alpha-DG at their surface. Infection followed by dysfunction of these antigen-presenting cells contributes to immunosuppression and persistent viral infection in vivo.

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Blocking of LCMV ARM53b infection of ES cells with soluble laminin-1. (A) Infection of ES cells cultivated on fibronectin (Fn) and laminin-1 (Ln) substrates is mediated by α-DG. DG−/− ES cells (black bars) and DG+/− ES cells (white bars) were cultivated on Fn or Ln for 20 h before infection with LCMV clone-13 (cl13) and ARM53b (ARM) using an MOI of 1. After 16 h, infection levels were assessed by staining with a LCMV-NP–specific monoclonal antibody. The sequences of the NPs of LCMV ARM53b and clone-13 are identical and therefore recognized equivalently by this antibody (Salvato et al., 1988). Triplicate samples were counted (500 cells each for DG−/− ES and 200 cells each for DG+/− cells; n = 3, ± SD). (B) Blocking of LCMV infection of ES cells with soluble laminin-1. DG+/− ES cells were cultivated on fibronectin in absence of laminin-1(Fn/0) or on laminin in the absence (Ln/0) or in the presence of laminin-1 at 5 (Ln/5), 12.5 (Ln/12.5), or 50 (Ln/50) μg/ml soluble laminin-1. 24 h after the addition of laminin-1, cells were infected with LCMV ARM53b (black bars) or clone-13 (white bars) at an MOI of 1 for 16 h. Infection levels were assessed by detecting LCMV-NP. In ARM53b-infected specimens, 500 cells were examined per sample; in clone-13–infected specimens, 200 cells were examined per sample. LCMV-NP–positive cells were scored (n = 4, ± SD).
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fig7: Blocking of LCMV ARM53b infection of ES cells with soluble laminin-1. (A) Infection of ES cells cultivated on fibronectin (Fn) and laminin-1 (Ln) substrates is mediated by α-DG. DG−/− ES cells (black bars) and DG+/− ES cells (white bars) were cultivated on Fn or Ln for 20 h before infection with LCMV clone-13 (cl13) and ARM53b (ARM) using an MOI of 1. After 16 h, infection levels were assessed by staining with a LCMV-NP–specific monoclonal antibody. The sequences of the NPs of LCMV ARM53b and clone-13 are identical and therefore recognized equivalently by this antibody (Salvato et al., 1988). Triplicate samples were counted (500 cells each for DG−/− ES and 200 cells each for DG+/− cells; n = 3, ± SD). (B) Blocking of LCMV infection of ES cells with soluble laminin-1. DG+/− ES cells were cultivated on fibronectin in absence of laminin-1(Fn/0) or on laminin in the absence (Ln/0) or in the presence of laminin-1 at 5 (Ln/5), 12.5 (Ln/12.5), or 50 (Ln/50) μg/ml soluble laminin-1. 24 h after the addition of laminin-1, cells were infected with LCMV ARM53b (black bars) or clone-13 (white bars) at an MOI of 1 for 16 h. Infection levels were assessed by detecting LCMV-NP. In ARM53b-infected specimens, 500 cells were examined per sample; in clone-13–infected specimens, 200 cells were examined per sample. LCMV-NP–positive cells were scored (n = 4, ± SD).

Mentions: In the light of the competition between LCMV and laminin-1 found, we questioned whether α-DG–associated laminin-1 clusters at the surface of ES cells may interfere with virus infection. Since ES cells express little if any laminin-1, the formation of laminin-1 clusters can be induced in both DG+/+ and DG+/− but not in DG−/− ES cells by addition of purified soluble laminin-1 (Henry and Campbell, 1998). We cultivated DG+/− and DG−/− ES cells either on fibronectin, a substratum that does not involve α-DG for cell adhesion, or on laminin-1. Regardless of substrate, DG+/− ES cells were infected efficiently with LCMV ARM53b and clone-13, indicating the presence of sufficient amounts of α-DG at the apical surface to mediate viral entry (Fig. 7 A). As demonstrated previously (Henry and Campbell, 1998; Fig. 6), the addition of soluble laminin-1 produced laminin-1 clusters at the surfaces of DG+/− but not DG−/− ES cells. These laminin clusters significantly reduced subsequent infection with LCMV ARM53b but not clone-13 (Fig. 7 B).


Molecular analysis of the interaction of LCMV with its cellular receptor [alpha]-dystroglycan.

Kunz S, Sevilla N, McGavern DB, Campbell KP, Oldstone MB - J. Cell Biol. (2001)

Blocking of LCMV ARM53b infection of ES cells with soluble laminin-1. (A) Infection of ES cells cultivated on fibronectin (Fn) and laminin-1 (Ln) substrates is mediated by α-DG. DG−/− ES cells (black bars) and DG+/− ES cells (white bars) were cultivated on Fn or Ln for 20 h before infection with LCMV clone-13 (cl13) and ARM53b (ARM) using an MOI of 1. After 16 h, infection levels were assessed by staining with a LCMV-NP–specific monoclonal antibody. The sequences of the NPs of LCMV ARM53b and clone-13 are identical and therefore recognized equivalently by this antibody (Salvato et al., 1988). Triplicate samples were counted (500 cells each for DG−/− ES and 200 cells each for DG+/− cells; n = 3, ± SD). (B) Blocking of LCMV infection of ES cells with soluble laminin-1. DG+/− ES cells were cultivated on fibronectin in absence of laminin-1(Fn/0) or on laminin in the absence (Ln/0) or in the presence of laminin-1 at 5 (Ln/5), 12.5 (Ln/12.5), or 50 (Ln/50) μg/ml soluble laminin-1. 24 h after the addition of laminin-1, cells were infected with LCMV ARM53b (black bars) or clone-13 (white bars) at an MOI of 1 for 16 h. Infection levels were assessed by detecting LCMV-NP. In ARM53b-infected specimens, 500 cells were examined per sample; in clone-13–infected specimens, 200 cells were examined per sample. LCMV-NP–positive cells were scored (n = 4, ± SD).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2198839&req=5

fig7: Blocking of LCMV ARM53b infection of ES cells with soluble laminin-1. (A) Infection of ES cells cultivated on fibronectin (Fn) and laminin-1 (Ln) substrates is mediated by α-DG. DG−/− ES cells (black bars) and DG+/− ES cells (white bars) were cultivated on Fn or Ln for 20 h before infection with LCMV clone-13 (cl13) and ARM53b (ARM) using an MOI of 1. After 16 h, infection levels were assessed by staining with a LCMV-NP–specific monoclonal antibody. The sequences of the NPs of LCMV ARM53b and clone-13 are identical and therefore recognized equivalently by this antibody (Salvato et al., 1988). Triplicate samples were counted (500 cells each for DG−/− ES and 200 cells each for DG+/− cells; n = 3, ± SD). (B) Blocking of LCMV infection of ES cells with soluble laminin-1. DG+/− ES cells were cultivated on fibronectin in absence of laminin-1(Fn/0) or on laminin in the absence (Ln/0) or in the presence of laminin-1 at 5 (Ln/5), 12.5 (Ln/12.5), or 50 (Ln/50) μg/ml soluble laminin-1. 24 h after the addition of laminin-1, cells were infected with LCMV ARM53b (black bars) or clone-13 (white bars) at an MOI of 1 for 16 h. Infection levels were assessed by detecting LCMV-NP. In ARM53b-infected specimens, 500 cells were examined per sample; in clone-13–infected specimens, 200 cells were examined per sample. LCMV-NP–positive cells were scored (n = 4, ± SD).
Mentions: In the light of the competition between LCMV and laminin-1 found, we questioned whether α-DG–associated laminin-1 clusters at the surface of ES cells may interfere with virus infection. Since ES cells express little if any laminin-1, the formation of laminin-1 clusters can be induced in both DG+/+ and DG+/− but not in DG−/− ES cells by addition of purified soluble laminin-1 (Henry and Campbell, 1998). We cultivated DG+/− and DG−/− ES cells either on fibronectin, a substratum that does not involve α-DG for cell adhesion, or on laminin-1. Regardless of substrate, DG+/− ES cells were infected efficiently with LCMV ARM53b and clone-13, indicating the presence of sufficient amounts of α-DG at the apical surface to mediate viral entry (Fig. 7 A). As demonstrated previously (Henry and Campbell, 1998; Fig. 6), the addition of soluble laminin-1 produced laminin-1 clusters at the surfaces of DG+/− but not DG−/− ES cells. These laminin clusters significantly reduced subsequent infection with LCMV ARM53b but not clone-13 (Fig. 7 B).

Bottom Line: In the present study, we characterized the binding of LCMV to alpha-DG and addressed the role of alpha-DG-associated host-derived proteins in virus infection.We found that the COOH-terminal region of alpha-DG's first globular domain and the NH2-terminal region of the mucin-related structures of alpha-DG together form the binding site for LCMV.This competition of the virus with ECM molecules for receptor binding likely explains the recently found correlation between the affinity of LCMV binding to alpha-DG, tissue tropism, and pathological potential.

View Article: PubMed Central - PubMed

Affiliation: The Scripps Research Institute, Division of Virology, Department of Neuropharmacology, La Jolla, CA 92037, USA.

ABSTRACT
alpha-Dystroglycan (DG) has been identified as the cellular receptor for lymphocytic choriomeningitis virus (LCMV) and Lassa fever virus (LFV). This subunit of DG is a highly versatile cell surface molecule that provides a molecular link between the extracellular matrix (ECM) and a beta-DG transmembrane component, which interacts with the actin-based cytoskeleton. In addition, DG exhibits a complex pattern of interaction with a wide variety of ECM and cellular proteins. In the present study, we characterized the binding of LCMV to alpha-DG and addressed the role of alpha-DG-associated host-derived proteins in virus infection. We found that the COOH-terminal region of alpha-DG's first globular domain and the NH2-terminal region of the mucin-related structures of alpha-DG together form the binding site for LCMV. The virus-alpha-DG binding unlike ECM alpha-DG interactions was not dependent on divalent cations. Despite such differences in binding, LCMV and laminin-1 use, in part, an overlapping binding site on alpha-DG, and the ability of an LCMV isolate to compete with laminin-1 for receptor binding is determined by its binding affinity to alpha-DG. This competition of the virus with ECM molecules for receptor binding likely explains the recently found correlation between the affinity of LCMV binding to alpha-DG, tissue tropism, and pathological potential. LCMV strains and variants with high binding affinity to alpha-DG but not low affinity binders are able to infect CD11c+ dendritic cells, which express alpha-DG at their surface. Infection followed by dysfunction of these antigen-presenting cells contributes to immunosuppression and persistent viral infection in vivo.

Show MeSH
Related in: MedlinePlus