Complex Interplay between HIV-1 Capsid and MX2-Independent Alpha Interferon-Induced Antiviral Factors

Abstract

Type I interferons (IFNs), including IFN-α, upregulate an array of IFN-stimulated genes (ISGs) and potently suppress Human immunodeficiency virus type 1 (HIV-1) infectivity in CD4+ T cells, monocyte-derived macrophages, and dendritic cells. Recently, we and others identified ISG myxovirus resistance 2 (MX2) as an inhibitor of HIV-1 nuclear entry. However, additional antiviral blocks exist upstream of nuclear import, but the ISGs that suppress infection, e.g., prior to (or during) reverse transcription, remain to be defined. We show here that the HIV-1 CA mutations N74D and A105T, both of which allow escape from inhibition by MX2 and the truncated version of cleavage and polyadenylation specific factor 6 (CPSF6), as well as the cyclophilin A (CypA)-binding loop mutation P90A, all increase sensitivity to IFN-α-mediated inhibition. Using clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 technology, we demonstrate that the IFN-α hypersensitivity of these mutants in THP-1 cells is independent of MX2 or CPSF6. As expected, CypA depletion had no additional effect on the behavior of the P90A mutant but modestly increased the IFN-α sensitivity of wild-type virus. Interestingly, the infectivity of wild-type or P90A virus could be rescued from the MX2-independent IFN-α-induced blocks in THP-1 cells by treatment with cyclosporine (Cs) or its nonimmunosuppressive analogue SDZ-NIM811, indicating that Cs-sensitive host cell cyclophilins other than CypA contribute to the activity of IFN-α-induced blocks. We propose that cellular interactions with incoming HIV-1 capsids help shield the virus from recognition by antiviral effector mechanisms. Thus, the CA protein is a fulcrum for the dynamic interplay between cell-encoded functions that inhibit or promote HIV-1 infection.IMPORTANCE HIV-1 is the causative agent of AIDS. During acute HIV-1 infection, numerous proinflammatory cytokines are produced, including type I interferons (IFNs). IFNs can limit HIV-1 replication by inducing the expression of a set of antiviral genes that inhibit HIV-1 at multiple steps in its life cycle, including the postentry steps of reverse transcription and nuclear import. This is observed in cultured cell systems, as well as in clinical trials in HIV-1-infected patients. The identities of the cellular antiviral factors, their viral targets, and the underpinning mechanisms are largely unknown. We show here that the HIV-1 Capsid protein plays a central role in protecting the virus from IFN-induced inhibitors that block early postentry steps of infection. We further show that host cell cyclophilins play an important role in regulating these processes, thus highlighting the complex interplay between antiviral effector mechanisms and viral survival.

GILT restricts the cellular entry mediated by the envelope glycoproteins of SARS-CoV, Ebola virus and Lassa fever virus

“…Similar to previous report [33], THP-1 cells were spininfected with concentrated LentiCRISPRv2/GILT pseudovirus in the presence of 40 μg/ml DEAE-Dextran. Forty-eight hours after transduction, the transduced THP-1 cells were selected with 1 μg/ml of puromycin for two weeks.…”

“…By using the template of pcDNA3/ ACE2-C9 reported previously [32], human ACE2 with C-terminally C9 tag was subcloned into retroviral vector pCX 4 bsr vector between BamH I and Not I sites. LentiCRISPRv2/gilt plasmids were constructed using methods previously described [33]. Briefly, Guide RNA encoding oligonucleotides were annealed and cloned into BsmBI-linearized plentiCRISPRv2.…”

Capsid-Dependent Host Factors in HIV-1 Infection

“…For example, CypA knockout impaired viral infectivity in Jurkat cells but not in other cell types [23–25]. Non-immunosuppressive drugs that block the CA-CypA interaction appear to specifically perturb HIV-1 in primary cells [26, 27].…”

“…ISG products known to target the HIV-1 capsid include TRIM5α and MX2 [130]. However, as yet identified capsid-dependent ISGs likely contribute, because IFNs appear more potent with certain CA mutants to block uncoating and reverse transcription [25, 93, 128]. Innate immune pathways are activated by recognition of pathogen-associated molecular patterns (PAMPs) through pathogen recognition receptors.…”

TRIM34 acts with TRIM5 to restrict HIV and SIV capsids

“…Single amino acid mutations in the HIV-1 capsid protein, for example N74D for CPSF6 and P90A for CypA, abrogate binding to these host factors (Lee et al, 2010; Schaller et al, 2011). Both capsid mutants have been demonstrated to infect cells less efficiently than wild type in some cell types, including primary cell such as CD4+ T cells and monocyte-derived macrophages (MDMs) (Ambrose et al, 2012; Bulli et al, 2016; Kim et al, 2019; Schaller et al, 2011). Further, both the P90A and N74D capsid mutants have been shown to be hypersensitive to the effects of IFN (Bulli et al, 2016), suggesting that one or more interferon-induced restriction factors block infection of these capsid mutant viruses.…”