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Efficient BST2 antagonism by Vpu is critical for early HIV-1 dissemination in humanized mice.

Dave VP, Hajjar F, Dieng MM, Haddad É, Cohen ÉA - Retrovirology (2013)

Bottom Line: Interestingly, we also find that efficient HIV-1 release and dissemination are directly related to functional strength of Vpu in antagonizing BST2.Thus, reduced antagonism of BST2 due to β-TrCP binding domain mutations results in decreased plasma viremia and frequency of infected T cells, highlighting the importance of Vpu-mediated β-TrCP-dependent BST-2 degradation for optimal initial viral propagation.Overall, our findings suggest that BST2 antagonism by Vpu is critical for efficient early viral expansion and dissemination during acute infection and as such is likely to confer HIV-1 increased transmission fitness.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Human Retrovirology, Institut de Recherches Cliniques de Montréal (IRCM), 110 Pine avenue west, Montreal, QC H2W 1R7, Canada. eric.cohen@ircm.qc.ca.

ABSTRACT

Background: Vpu is a multifunctional accessory protein that enhances the release of HIV-1 by counteracting the entrapment of nascent virions on infected cell surface mediated by BST2/Tetherin. Vpu-mediated BST2 antagonism involves physical association with BST2 and subsequent mislocalization of the restriction factor to intracellular compartments followed by SCF(β-TrCP) E3 ligase-dependent lysosomal degradation. Apart from BST2 antagonism, Vpu also induces down regulation of several immune molecules, including CD4 and SLAMF6/NTB-A, to evade host immune responses and promote viral dissemination. However, it should be noted that the multiple functions of Vpu have been studied in cell-based assays, and thus it remains unclear how Vpu influences the dynamic of HIV-1 infection in in vivo conditions.

Results: Using a humanized mouse model of acute infection as well as CCR5-tropic HIV-1 that lack Vpu or encode WT Vpu or Vpu with mutations in the β-TrCP binding domain, we provide evidence that Vpu-mediated BST2 antagonism plays a crucial role in establishing early plasma viremia and viral dissemination. Interestingly, we also find that efficient HIV-1 release and dissemination are directly related to functional strength of Vpu in antagonizing BST2. Thus, reduced antagonism of BST2 due to β-TrCP binding domain mutations results in decreased plasma viremia and frequency of infected T cells, highlighting the importance of Vpu-mediated β-TrCP-dependent BST-2 degradation for optimal initial viral propagation.

Conclusions: Overall, our findings suggest that BST2 antagonism by Vpu is critical for efficient early viral expansion and dissemination during acute infection and as such is likely to confer HIV-1 increased transmission fitness.

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Related in: MedlinePlus

Infection of hu-mice with supra physiological dose of HIV-1-∆Vpu does not overcome BST2 restriction on early viral propagation. Hu-mice were infected with an inoculum containing 100-fold more (compared to low dose) infectious HIV-1-WT or HIV-1-∆Vpu and bled at 3, 7, 14 and 21-dpi. (A) shows RNA copy number/ml of plasma (log10 values) at indicated time points in hu-mice infected with HIV-1-WT or HIV-1 ∆Vpu virus (n = 5). The horizontal broken line depicts the detection limit of the viral load assay as in the Figure 1. (B) Frequency of p24+ T cells at 21-dpi in spleen of hu-mice infected with HIV-1-WT or HIV-1-∆Vpu (n ≥ 4). (C) Impact of Vpu on BST2, CD4 and NTB-A levels on p24- and p24+ T cells from individual hu-mouse infected with the indicated HIV-1 virus. (D) Comparison of relative BST2 levels on p24+ and p24- T cells from spleen of hu-mice inoculated with the indicated HIV-1 virus at 21 dpi (MFI on p24- T cells = 100%; n ≥ 4). (E) Bar graph for relative NTB-A down regulation on p24+ T cells compared to p24- T cells from hu-mice infected with the indicated HIV-1 virus at 21dpi. Error bars represent SD; *, p ≤ 0.05; N.S.: not significant.
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Figure 3: Infection of hu-mice with supra physiological dose of HIV-1-∆Vpu does not overcome BST2 restriction on early viral propagation. Hu-mice were infected with an inoculum containing 100-fold more (compared to low dose) infectious HIV-1-WT or HIV-1-∆Vpu and bled at 3, 7, 14 and 21-dpi. (A) shows RNA copy number/ml of plasma (log10 values) at indicated time points in hu-mice infected with HIV-1-WT or HIV-1 ∆Vpu virus (n = 5). The horizontal broken line depicts the detection limit of the viral load assay as in the Figure 1. (B) Frequency of p24+ T cells at 21-dpi in spleen of hu-mice infected with HIV-1-WT or HIV-1-∆Vpu (n ≥ 4). (C) Impact of Vpu on BST2, CD4 and NTB-A levels on p24- and p24+ T cells from individual hu-mouse infected with the indicated HIV-1 virus. (D) Comparison of relative BST2 levels on p24+ and p24- T cells from spleen of hu-mice inoculated with the indicated HIV-1 virus at 21 dpi (MFI on p24- T cells = 100%; n ≥ 4). (E) Bar graph for relative NTB-A down regulation on p24+ T cells compared to p24- T cells from hu-mice infected with the indicated HIV-1 virus at 21dpi. Error bars represent SD; *, p ≤ 0.05; N.S.: not significant.

Mentions: To investigate if exposing hu-mice to a supra-physiological dose of infectious HIV-1-∆Vpu virus could overcome the BST2 restriction on early HIV-1 expansion, we infected a cohort of hu-mice with a 100-fold more virus (~500,000 TCID50 of HIV-1-WT or -∆Vpu virus). The infected hu-mice were bled at 3, 7, 14 and 21-days post infection (dpi). Hu-mice challenged with high dose of HIV-1-WT or -∆Vpu showed detectable amounts of virus by 3-dpi. However, compared to HIV-1-∆Vpu infected animals, HIV-1-WT infected animals showed rapid increase in plasma viremia at subsequent time points. In fact, by 14- and 21-dpi there was an ~9- and ~15-fold increase, respectively, in the average amounts of virus in plasma of HIV-1-WT infected animals compared to HIV-1-∆Vpu infected animals (Figure 3A). In agreement with higher plasma viral load, average frequency of p24+ T cells at 21-dpi was ~ four-fold higher in spleen of hu-mice infected with HIV-1-WT than those challenged with Vpu-deleted HIV-1 (Figure 3B-C), and the infected T cells from HIV-1-WT but not -∆Vpu challenged hu-mice showed significant down regulation of BST2 (Figure 3C-D). That lower plasma viral load in HIV-1-∆Vpu infected hu-mice was largely due to impaired BST2 down regulation is supported by the fact that Vpu-deficient and proficient virus down regulated CD4 (Figure 3C) or NTB-A (Figure 3C and 3E) to a comparable extent. These data strongly suggest that in the absence of Vpu, infection of hu-mice with high dose of HIV-1 still fails to completely overcome BST2 restriction, thus highlighting the critical barrier that BST2 restriction might represent during acute infection in vivo. Collectively, our results suggest that Vpu-mediated BST-2 antagonism is critical for HIV-1 replication and propagation in vivo, especially at early times post-infection.


Efficient BST2 antagonism by Vpu is critical for early HIV-1 dissemination in humanized mice.

Dave VP, Hajjar F, Dieng MM, Haddad É, Cohen ÉA - Retrovirology (2013)

Infection of hu-mice with supra physiological dose of HIV-1-∆Vpu does not overcome BST2 restriction on early viral propagation. Hu-mice were infected with an inoculum containing 100-fold more (compared to low dose) infectious HIV-1-WT or HIV-1-∆Vpu and bled at 3, 7, 14 and 21-dpi. (A) shows RNA copy number/ml of plasma (log10 values) at indicated time points in hu-mice infected with HIV-1-WT or HIV-1 ∆Vpu virus (n = 5). The horizontal broken line depicts the detection limit of the viral load assay as in the Figure 1. (B) Frequency of p24+ T cells at 21-dpi in spleen of hu-mice infected with HIV-1-WT or HIV-1-∆Vpu (n ≥ 4). (C) Impact of Vpu on BST2, CD4 and NTB-A levels on p24- and p24+ T cells from individual hu-mouse infected with the indicated HIV-1 virus. (D) Comparison of relative BST2 levels on p24+ and p24- T cells from spleen of hu-mice inoculated with the indicated HIV-1 virus at 21 dpi (MFI on p24- T cells = 100%; n ≥ 4). (E) Bar graph for relative NTB-A down regulation on p24+ T cells compared to p24- T cells from hu-mice infected with the indicated HIV-1 virus at 21dpi. Error bars represent SD; *, p ≤ 0.05; N.S.: not significant.
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Figure 3: Infection of hu-mice with supra physiological dose of HIV-1-∆Vpu does not overcome BST2 restriction on early viral propagation. Hu-mice were infected with an inoculum containing 100-fold more (compared to low dose) infectious HIV-1-WT or HIV-1-∆Vpu and bled at 3, 7, 14 and 21-dpi. (A) shows RNA copy number/ml of plasma (log10 values) at indicated time points in hu-mice infected with HIV-1-WT or HIV-1 ∆Vpu virus (n = 5). The horizontal broken line depicts the detection limit of the viral load assay as in the Figure 1. (B) Frequency of p24+ T cells at 21-dpi in spleen of hu-mice infected with HIV-1-WT or HIV-1-∆Vpu (n ≥ 4). (C) Impact of Vpu on BST2, CD4 and NTB-A levels on p24- and p24+ T cells from individual hu-mouse infected with the indicated HIV-1 virus. (D) Comparison of relative BST2 levels on p24+ and p24- T cells from spleen of hu-mice inoculated with the indicated HIV-1 virus at 21 dpi (MFI on p24- T cells = 100%; n ≥ 4). (E) Bar graph for relative NTB-A down regulation on p24+ T cells compared to p24- T cells from hu-mice infected with the indicated HIV-1 virus at 21dpi. Error bars represent SD; *, p ≤ 0.05; N.S.: not significant.
Mentions: To investigate if exposing hu-mice to a supra-physiological dose of infectious HIV-1-∆Vpu virus could overcome the BST2 restriction on early HIV-1 expansion, we infected a cohort of hu-mice with a 100-fold more virus (~500,000 TCID50 of HIV-1-WT or -∆Vpu virus). The infected hu-mice were bled at 3, 7, 14 and 21-days post infection (dpi). Hu-mice challenged with high dose of HIV-1-WT or -∆Vpu showed detectable amounts of virus by 3-dpi. However, compared to HIV-1-∆Vpu infected animals, HIV-1-WT infected animals showed rapid increase in plasma viremia at subsequent time points. In fact, by 14- and 21-dpi there was an ~9- and ~15-fold increase, respectively, in the average amounts of virus in plasma of HIV-1-WT infected animals compared to HIV-1-∆Vpu infected animals (Figure 3A). In agreement with higher plasma viral load, average frequency of p24+ T cells at 21-dpi was ~ four-fold higher in spleen of hu-mice infected with HIV-1-WT than those challenged with Vpu-deleted HIV-1 (Figure 3B-C), and the infected T cells from HIV-1-WT but not -∆Vpu challenged hu-mice showed significant down regulation of BST2 (Figure 3C-D). That lower plasma viral load in HIV-1-∆Vpu infected hu-mice was largely due to impaired BST2 down regulation is supported by the fact that Vpu-deficient and proficient virus down regulated CD4 (Figure 3C) or NTB-A (Figure 3C and 3E) to a comparable extent. These data strongly suggest that in the absence of Vpu, infection of hu-mice with high dose of HIV-1 still fails to completely overcome BST2 restriction, thus highlighting the critical barrier that BST2 restriction might represent during acute infection in vivo. Collectively, our results suggest that Vpu-mediated BST-2 antagonism is critical for HIV-1 replication and propagation in vivo, especially at early times post-infection.

Bottom Line: Interestingly, we also find that efficient HIV-1 release and dissemination are directly related to functional strength of Vpu in antagonizing BST2.Thus, reduced antagonism of BST2 due to β-TrCP binding domain mutations results in decreased plasma viremia and frequency of infected T cells, highlighting the importance of Vpu-mediated β-TrCP-dependent BST-2 degradation for optimal initial viral propagation.Overall, our findings suggest that BST2 antagonism by Vpu is critical for efficient early viral expansion and dissemination during acute infection and as such is likely to confer HIV-1 increased transmission fitness.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Human Retrovirology, Institut de Recherches Cliniques de Montréal (IRCM), 110 Pine avenue west, Montreal, QC H2W 1R7, Canada. eric.cohen@ircm.qc.ca.

ABSTRACT

Background: Vpu is a multifunctional accessory protein that enhances the release of HIV-1 by counteracting the entrapment of nascent virions on infected cell surface mediated by BST2/Tetherin. Vpu-mediated BST2 antagonism involves physical association with BST2 and subsequent mislocalization of the restriction factor to intracellular compartments followed by SCF(β-TrCP) E3 ligase-dependent lysosomal degradation. Apart from BST2 antagonism, Vpu also induces down regulation of several immune molecules, including CD4 and SLAMF6/NTB-A, to evade host immune responses and promote viral dissemination. However, it should be noted that the multiple functions of Vpu have been studied in cell-based assays, and thus it remains unclear how Vpu influences the dynamic of HIV-1 infection in in vivo conditions.

Results: Using a humanized mouse model of acute infection as well as CCR5-tropic HIV-1 that lack Vpu or encode WT Vpu or Vpu with mutations in the β-TrCP binding domain, we provide evidence that Vpu-mediated BST2 antagonism plays a crucial role in establishing early plasma viremia and viral dissemination. Interestingly, we also find that efficient HIV-1 release and dissemination are directly related to functional strength of Vpu in antagonizing BST2. Thus, reduced antagonism of BST2 due to β-TrCP binding domain mutations results in decreased plasma viremia and frequency of infected T cells, highlighting the importance of Vpu-mediated β-TrCP-dependent BST-2 degradation for optimal initial viral propagation.

Conclusions: Overall, our findings suggest that BST2 antagonism by Vpu is critical for efficient early viral expansion and dissemination during acute infection and as such is likely to confer HIV-1 increased transmission fitness.

Show MeSH
Related in: MedlinePlus