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The cytomegalovirus m155 gene product subverts natural killer cell antiviral protection by disruption of H60-NKG2D interactions.

Lodoen MB, Abenes G, Umamoto S, Houchins JP, Liu F, Lanier LL - J. Exp. Med. (2004)

Bottom Line: Treatment with the proteasome inhibitors lactacystin or epoxomicin reversed m155 down-regulation of H60.An MCMV mutant virus lacking m155 was severely attenuated in BALB/c mice; however, treatment with neutralizing anti-NKG2D monoclonal antibody or with NK-depleting anti-asialo GM1 antisera restored virulence of the mutant virus.Thus, down-regulation of H60 by m155 is a powerful mechanism of inhibiting NKG2D-mediated antiviral function.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Cancer Research Institute, University of California, San Francisco, San Francisco, CA 94143, USA.

ABSTRACT
Natural killer (NK) cells are an important early mediator of host immunity to murine cytomegalovirus (MCMV) infection. However, MCMV has evolved mechanisms to elude recognition and clearance by NK cells. We have identified an MCMV immune evasion protein that impairs NKG2D-mediated NK cell antiviral activity. Infection of BALB/c 3T3 cells with the Smith strain of MCMV resulted in strong down-regulation of H60, a high affinity ligand for NKG2D, from the surface of virus-infected cells. The MCMV m155 protein specifically down-regulated H60 without affecting expression of the other known NKG2D ligands, RAE-1 and MULT-1. Treatment with the proteasome inhibitors lactacystin or epoxomicin reversed m155 down-regulation of H60. An MCMV mutant virus lacking m155 was severely attenuated in BALB/c mice; however, treatment with neutralizing anti-NKG2D monoclonal antibody or with NK-depleting anti-asialo GM1 antisera restored virulence of the mutant virus. Thus, down-regulation of H60 by m155 is a powerful mechanism of inhibiting NKG2D-mediated antiviral function.

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Depletion of NK cells or blockade of NKG2D restores virulence of Dm155. (A) BALB/c mice were injected i.p. with control rat IgG, anti-asialo GM1 antisera, or anti-NKG2D mAb (CX5). After antibody treatment, mice were infected with 105 PFUs (A) or 2 × 105 PFUs (B) of Smith, Rqm155 (revertant), or Rvm155 (Dm155). 3 d after infection, spleens and livers were harvested. Plaque assays were performed on organ homogenates to determine viral titers. In each experiment, three mice were used in each experimental group and error bars represent the standard deviation of the viral titers from these three mice. Three independent experiments were performed and data from two experiments using 2 × 105 PFUs were combined for presentation in B. The limit of detection was 1 PFU/ml organ homogenate.
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fig4: Depletion of NK cells or blockade of NKG2D restores virulence of Dm155. (A) BALB/c mice were injected i.p. with control rat IgG, anti-asialo GM1 antisera, or anti-NKG2D mAb (CX5). After antibody treatment, mice were infected with 105 PFUs (A) or 2 × 105 PFUs (B) of Smith, Rqm155 (revertant), or Rvm155 (Dm155). 3 d after infection, spleens and livers were harvested. Plaque assays were performed on organ homogenates to determine viral titers. In each experiment, three mice were used in each experimental group and error bars represent the standard deviation of the viral titers from these three mice. Three independent experiments were performed and data from two experiments using 2 × 105 PFUs were combined for presentation in B. The limit of detection was 1 PFU/ml organ homogenate.

Mentions: We hypothesized that NK cell recognition of virus-infected cells may be impaired by m155. Therefore, we examined the significance of H60 down-regulation during MCMV infection in vivo. Before infection of BALB/c mice, which express functional H60 (14), we treated mice with a control rat IgG mAb, a neutralizing anti-NKG2D (CX5) mAb, or an NK cell–depleting anti-asialo GM1 antisera. CX5 mAb blocks the binding of NKG2D to its ligands and modulates the NKG2D receptor from the surface of NK cells, but does not deplete NK cells (22), thereby allowing us to directly examine the role of the NKG2D receptor in the immune response to MCMV. To investigate the total contribution of NK cells to the MCMV immune response, we depleted NK cells using anti-asialo GM1 antisera. After antibody treatment, mice were infected i.p. with 105 PFUs of virus per mouse (Fig. 4 A) or with 2 × 105 PFUs (Fig. 4 B) per mouse of wild-type Smith, Dm155, or the m155 revertant virus. Because previous studies have implicated NK cells in the control of MCMV predominantly in the liver and spleen and early during infection, viral titers in these organs were determined on day 3 after infection.


The cytomegalovirus m155 gene product subverts natural killer cell antiviral protection by disruption of H60-NKG2D interactions.

Lodoen MB, Abenes G, Umamoto S, Houchins JP, Liu F, Lanier LL - J. Exp. Med. (2004)

Depletion of NK cells or blockade of NKG2D restores virulence of Dm155. (A) BALB/c mice were injected i.p. with control rat IgG, anti-asialo GM1 antisera, or anti-NKG2D mAb (CX5). After antibody treatment, mice were infected with 105 PFUs (A) or 2 × 105 PFUs (B) of Smith, Rqm155 (revertant), or Rvm155 (Dm155). 3 d after infection, spleens and livers were harvested. Plaque assays were performed on organ homogenates to determine viral titers. In each experiment, three mice were used in each experimental group and error bars represent the standard deviation of the viral titers from these three mice. Three independent experiments were performed and data from two experiments using 2 × 105 PFUs were combined for presentation in B. The limit of detection was 1 PFU/ml organ homogenate.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2211837&req=5

fig4: Depletion of NK cells or blockade of NKG2D restores virulence of Dm155. (A) BALB/c mice were injected i.p. with control rat IgG, anti-asialo GM1 antisera, or anti-NKG2D mAb (CX5). After antibody treatment, mice were infected with 105 PFUs (A) or 2 × 105 PFUs (B) of Smith, Rqm155 (revertant), or Rvm155 (Dm155). 3 d after infection, spleens and livers were harvested. Plaque assays were performed on organ homogenates to determine viral titers. In each experiment, three mice were used in each experimental group and error bars represent the standard deviation of the viral titers from these three mice. Three independent experiments were performed and data from two experiments using 2 × 105 PFUs were combined for presentation in B. The limit of detection was 1 PFU/ml organ homogenate.
Mentions: We hypothesized that NK cell recognition of virus-infected cells may be impaired by m155. Therefore, we examined the significance of H60 down-regulation during MCMV infection in vivo. Before infection of BALB/c mice, which express functional H60 (14), we treated mice with a control rat IgG mAb, a neutralizing anti-NKG2D (CX5) mAb, or an NK cell–depleting anti-asialo GM1 antisera. CX5 mAb blocks the binding of NKG2D to its ligands and modulates the NKG2D receptor from the surface of NK cells, but does not deplete NK cells (22), thereby allowing us to directly examine the role of the NKG2D receptor in the immune response to MCMV. To investigate the total contribution of NK cells to the MCMV immune response, we depleted NK cells using anti-asialo GM1 antisera. After antibody treatment, mice were infected i.p. with 105 PFUs of virus per mouse (Fig. 4 A) or with 2 × 105 PFUs (Fig. 4 B) per mouse of wild-type Smith, Dm155, or the m155 revertant virus. Because previous studies have implicated NK cells in the control of MCMV predominantly in the liver and spleen and early during infection, viral titers in these organs were determined on day 3 after infection.

Bottom Line: Treatment with the proteasome inhibitors lactacystin or epoxomicin reversed m155 down-regulation of H60.An MCMV mutant virus lacking m155 was severely attenuated in BALB/c mice; however, treatment with neutralizing anti-NKG2D monoclonal antibody or with NK-depleting anti-asialo GM1 antisera restored virulence of the mutant virus.Thus, down-regulation of H60 by m155 is a powerful mechanism of inhibiting NKG2D-mediated antiviral function.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Cancer Research Institute, University of California, San Francisco, San Francisco, CA 94143, USA.

ABSTRACT
Natural killer (NK) cells are an important early mediator of host immunity to murine cytomegalovirus (MCMV) infection. However, MCMV has evolved mechanisms to elude recognition and clearance by NK cells. We have identified an MCMV immune evasion protein that impairs NKG2D-mediated NK cell antiviral activity. Infection of BALB/c 3T3 cells with the Smith strain of MCMV resulted in strong down-regulation of H60, a high affinity ligand for NKG2D, from the surface of virus-infected cells. The MCMV m155 protein specifically down-regulated H60 without affecting expression of the other known NKG2D ligands, RAE-1 and MULT-1. Treatment with the proteasome inhibitors lactacystin or epoxomicin reversed m155 down-regulation of H60. An MCMV mutant virus lacking m155 was severely attenuated in BALB/c mice; however, treatment with neutralizing anti-NKG2D monoclonal antibody or with NK-depleting anti-asialo GM1 antisera restored virulence of the mutant virus. Thus, down-regulation of H60 by m155 is a powerful mechanism of inhibiting NKG2D-mediated antiviral function.

Show MeSH
Related in: MedlinePlus