Limits...
HIV-1 evades innate immune recognition through specific cofactor recruitment.

Rasaiyaah J, Tan CP, Fletcher AJ, Price AJ, Blondeau C, Hilditch L, Jacques DA, Selwood DL, James LC, Noursadeghi M, Towers GJ - Nature (2013)

Bottom Line: In each case, suppressed replication is rescued by IFN-receptor blockade, demonstrating a role for IFN in restriction.IFN production is dependent on viral reverse transcription but not integration, indicating that a viral reverse transcription product comprises the HIV-1 pathogen-associated molecular pattern.Finally, we show that we can pharmacologically induce wild-type HIV-1 infection to stimulate IFN secretion and an antiviral state using a non-immunosuppressive cyclosporine analogue.

View Article: PubMed Central - PubMed

Affiliation: University College London, Medical Research Council Centre for Medical Molecular Virology, Division of Infection and Immunity, University College London, 90 Gower Street, London WC1E 6BT, UK.

ABSTRACT
Human immunodeficiency virus (HIV)-1 is able to replicate in primary human macrophages without stimulating innate immunity despite reverse transcription of genomic RNA into double-stranded DNA, an activity that might be expected to trigger innate pattern recognition receptors. We reasoned that if correctly orchestrated HIV-1 uncoating and nuclear entry is important for evasion of innate sensors then manipulation of specific interactions between HIV-1 capsid and host factors that putatively regulate these processes should trigger pattern recognition receptors and stimulate type 1 interferon (IFN) secretion. Here we show that HIV-1 capsid mutants N74D and P90A, which are impaired for interaction with cofactors cleavage and polyadenylation specificity factor subunit 6 (CPSF6) and cyclophilins (Nup358 and CypA), respectively, cannot replicate in primary human monocyte-derived macrophages because they trigger innate sensors leading to nuclear translocation of NF-κB and IRF3, the production of soluble type 1 IFN and induction of an antiviral state. Depletion of CPSF6 with short hairpin RNA expression allows wild-type virus to trigger innate sensors and IFN production. In each case, suppressed replication is rescued by IFN-receptor blockade, demonstrating a role for IFN in restriction. IFN production is dependent on viral reverse transcription but not integration, indicating that a viral reverse transcription product comprises the HIV-1 pathogen-associated molecular pattern. Finally, we show that we can pharmacologically induce wild-type HIV-1 infection to stimulate IFN secretion and an antiviral state using a non-immunosuppressive cyclosporine analogue. We conclude that HIV-1 has evolved to use CPSF6 and cyclophilins to cloak its replication, allowing evasion of innate immune sensors and induction of a cell-autonomous innate immune response in primary human macrophages.

Show MeSH

Related in: MedlinePlus

HIV-1 CypA binding mutant CA P90A is restricted in MDM due to induction of Type-I IFN(a) Replication of WT HIV-1 or CA mutant P90A in MDM. (b) IFN-β levels in supernatants from (a). (c) Replication of HIV-1 CA P90A with IFNAR2 or control antibody (cAb). (d) as in Figure 1d. (e) Replication of WT or WT plus CA P90A. Mean data and regression lines are shown for biological replicates in c-e. P values (2-way ANOVA) are given for (c-d) IFNAR2 blockade and (e) co-infection with CA mutant P90A. (f) Infection of MDM by HIV-1 measured at 48h (g) GAPDH normalized IP10 RNA levels expressed as fold change over untreated cells after infection with WT or HIV-1 mutants (Mean of 3 technical replicates ±SEM, f-g).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3928559&req=5

Figure 3: HIV-1 CypA binding mutant CA P90A is restricted in MDM due to induction of Type-I IFN(a) Replication of WT HIV-1 or CA mutant P90A in MDM. (b) IFN-β levels in supernatants from (a). (c) Replication of HIV-1 CA P90A with IFNAR2 or control antibody (cAb). (d) as in Figure 1d. (e) Replication of WT or WT plus CA P90A. Mean data and regression lines are shown for biological replicates in c-e. P values (2-way ANOVA) are given for (c-d) IFNAR2 blockade and (e) co-infection with CA mutant P90A. (f) Infection of MDM by HIV-1 measured at 48h (g) GAPDH normalized IP10 RNA levels expressed as fold change over untreated cells after infection with WT or HIV-1 mutants (Mean of 3 technical replicates ±SEM, f-g).

Mentions: Since HIV-1 N74D is unable to appropriately utilize nuclear pore components and has retargeted integration properties1-3,11, we hypothesized that the HIV-1 CA mutant P90A, which fails to interact with the cyclophilins CypA and nuclear pore component Nup358, and also has retargeted integration2, might also trigger innate sensors. Indeed, HIV-1 P90A infection of MDM induced IFN-β production and an antiviral state in both replication and single-round infectivity assays, which was rescued by IFN receptor antibody (Fig. 3a-f, Extended Data Fig. 2, 3c,d). We find MDM infection by HIV-1 N74D, P90A or WT were equally increased by SIVmac VLP encoding Vpx, suggesting that mutant viruses were not specifically Vpx sensitive (Extended Data Fig. 5h). Quantitative RT-PCR and whole genome profiling demonstrated ISG induction after HIV-1 P90A infection (Extended Data Fig. 4b-d). Consistently, double mutation of P90A and RT D185E, but not IN D116N, suppressed IP10 induction (Fig. 3g). We hypothesise that viral DNA produced by reverse transcription is the target for innate sensing of both HIV-1 CA mutants N74D and P90A in MDM.


HIV-1 evades innate immune recognition through specific cofactor recruitment.

Rasaiyaah J, Tan CP, Fletcher AJ, Price AJ, Blondeau C, Hilditch L, Jacques DA, Selwood DL, James LC, Noursadeghi M, Towers GJ - Nature (2013)

HIV-1 CypA binding mutant CA P90A is restricted in MDM due to induction of Type-I IFN(a) Replication of WT HIV-1 or CA mutant P90A in MDM. (b) IFN-β levels in supernatants from (a). (c) Replication of HIV-1 CA P90A with IFNAR2 or control antibody (cAb). (d) as in Figure 1d. (e) Replication of WT or WT plus CA P90A. Mean data and regression lines are shown for biological replicates in c-e. P values (2-way ANOVA) are given for (c-d) IFNAR2 blockade and (e) co-infection with CA mutant P90A. (f) Infection of MDM by HIV-1 measured at 48h (g) GAPDH normalized IP10 RNA levels expressed as fold change over untreated cells after infection with WT or HIV-1 mutants (Mean of 3 technical replicates ±SEM, f-g).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3928559&req=5

Figure 3: HIV-1 CypA binding mutant CA P90A is restricted in MDM due to induction of Type-I IFN(a) Replication of WT HIV-1 or CA mutant P90A in MDM. (b) IFN-β levels in supernatants from (a). (c) Replication of HIV-1 CA P90A with IFNAR2 or control antibody (cAb). (d) as in Figure 1d. (e) Replication of WT or WT plus CA P90A. Mean data and regression lines are shown for biological replicates in c-e. P values (2-way ANOVA) are given for (c-d) IFNAR2 blockade and (e) co-infection with CA mutant P90A. (f) Infection of MDM by HIV-1 measured at 48h (g) GAPDH normalized IP10 RNA levels expressed as fold change over untreated cells after infection with WT or HIV-1 mutants (Mean of 3 technical replicates ±SEM, f-g).
Mentions: Since HIV-1 N74D is unable to appropriately utilize nuclear pore components and has retargeted integration properties1-3,11, we hypothesized that the HIV-1 CA mutant P90A, which fails to interact with the cyclophilins CypA and nuclear pore component Nup358, and also has retargeted integration2, might also trigger innate sensors. Indeed, HIV-1 P90A infection of MDM induced IFN-β production and an antiviral state in both replication and single-round infectivity assays, which was rescued by IFN receptor antibody (Fig. 3a-f, Extended Data Fig. 2, 3c,d). We find MDM infection by HIV-1 N74D, P90A or WT were equally increased by SIVmac VLP encoding Vpx, suggesting that mutant viruses were not specifically Vpx sensitive (Extended Data Fig. 5h). Quantitative RT-PCR and whole genome profiling demonstrated ISG induction after HIV-1 P90A infection (Extended Data Fig. 4b-d). Consistently, double mutation of P90A and RT D185E, but not IN D116N, suppressed IP10 induction (Fig. 3g). We hypothesise that viral DNA produced by reverse transcription is the target for innate sensing of both HIV-1 CA mutants N74D and P90A in MDM.

Bottom Line: In each case, suppressed replication is rescued by IFN-receptor blockade, demonstrating a role for IFN in restriction.IFN production is dependent on viral reverse transcription but not integration, indicating that a viral reverse transcription product comprises the HIV-1 pathogen-associated molecular pattern.Finally, we show that we can pharmacologically induce wild-type HIV-1 infection to stimulate IFN secretion and an antiviral state using a non-immunosuppressive cyclosporine analogue.

View Article: PubMed Central - PubMed

Affiliation: University College London, Medical Research Council Centre for Medical Molecular Virology, Division of Infection and Immunity, University College London, 90 Gower Street, London WC1E 6BT, UK.

ABSTRACT
Human immunodeficiency virus (HIV)-1 is able to replicate in primary human macrophages without stimulating innate immunity despite reverse transcription of genomic RNA into double-stranded DNA, an activity that might be expected to trigger innate pattern recognition receptors. We reasoned that if correctly orchestrated HIV-1 uncoating and nuclear entry is important for evasion of innate sensors then manipulation of specific interactions between HIV-1 capsid and host factors that putatively regulate these processes should trigger pattern recognition receptors and stimulate type 1 interferon (IFN) secretion. Here we show that HIV-1 capsid mutants N74D and P90A, which are impaired for interaction with cofactors cleavage and polyadenylation specificity factor subunit 6 (CPSF6) and cyclophilins (Nup358 and CypA), respectively, cannot replicate in primary human monocyte-derived macrophages because they trigger innate sensors leading to nuclear translocation of NF-κB and IRF3, the production of soluble type 1 IFN and induction of an antiviral state. Depletion of CPSF6 with short hairpin RNA expression allows wild-type virus to trigger innate sensors and IFN production. In each case, suppressed replication is rescued by IFN-receptor blockade, demonstrating a role for IFN in restriction. IFN production is dependent on viral reverse transcription but not integration, indicating that a viral reverse transcription product comprises the HIV-1 pathogen-associated molecular pattern. Finally, we show that we can pharmacologically induce wild-type HIV-1 infection to stimulate IFN secretion and an antiviral state using a non-immunosuppressive cyclosporine analogue. We conclude that HIV-1 has evolved to use CPSF6 and cyclophilins to cloak its replication, allowing evasion of innate immune sensors and induction of a cell-autonomous innate immune response in primary human macrophages.

Show MeSH
Related in: MedlinePlus