T cells detect intracellular DNA but fail to induce type I IFN responses: implications for restriction of HIV replication.

HIV infects key cell types of the immune system, most notably macrophages and CD4+ T cells. Whereas macrophages represent an important viral reservoir, activated CD4+ T cells are the most permissive cell types supporting high levels of viral replication. In recent years, it has been appreciated that the innate immune system plays an important role in controlling HIV replication, e.g. via interferon (IFN)-inducible restriction factors. Moreover, innate immune responses are involved in driving chronic immune activation and the pathogenesis of progressive immunodeficiency. Several pattern recognition receptors detecting HIV have been reported, including Toll-like receptor 7 and Retinoic-inducible gene-I, which detects viral RNA. Here we report that human primary T cells fail to induce strong IFN responses, despite the fact that this cell type does express key molecules involved in DNA signaling pathways. We demonstrate that the DNA sensor IFI16 migrates to sites of foreign DNA localization in the cytoplasm and recruits the signaling molecules stimulator of IFN genes and Tank-binding kinase, but this does not result in expression of IFN and IFN-stimulated genes. Importantly, we show that cytosolic DNA fails to affect HIV replication. However, exogenous treatment of activated T cells with type I IFN has the capacity to induce expression of IFN-stimulated genes and suppress HIV replication. Our data suggest the existence of an impaired DNA signaling machinery in T cells, which may prevent this cell type from activating cell-autonomous anti-HIV responses. This phenomenon could contribute to the high permissiveness of CD4+ T cells for HIV-1.

IFI16 senses DNA forms of the lentiviral replication cycle and controls HIV-1 replication.

Replication of lentiviruses generates different DNA forms, including RNA:DNA hybrids, ssDNA, and dsDNA. Nucleic acids stimulate innate immune responses, and pattern recognition receptors detecting dsDNA have been identified. However, sensors for ssDNA have not been reported, and the ability of RNA:DNA hybrids to stimulate innate immune responses is controversial. Using ssDNAs derived from HIV-1 proviral DNA, we report that this DNA form potently induces the expression of IFNs in primary human macrophages. This response was stimulated by stem regions in the DNA structure and was dependent on IFN-inducible protein 16 (IFI16), which bound immunostimulatory DNA directly and activated the stimulator of IFN genes -TANK-binding kinase 1 - IFN regulatory factors 3/7 (STING-TBK1-IRF3/7) pathway. Importantly, IFI16 colocalized and associated with lentiviral DNA in the cytoplasm in macrophages, and IFI16 knockdown in this cell type augmented lentiviral transduction and also HIV-1 replication. Thus, IFI16 is a sensor for DNA forms produced during the lentiviral replication cycle and regulates HIV-1 replication in macrophages.

Characterization of HIV-1 infection and innate sensing in different types of primary human monocyte-derived macrophages.

Macrophages play an important role in human immunodeficiency virus (HIV) pathogenesis and contribute to establishment of a viral reservoir responsible for continuous virus production and virus transmission to T cells. In this study, we investigated the differences between various monocyte-derived macrophages (MDMs) generated through different differentiation protocols and evaluated different cellular, immunological, and virological properties. We found that elevated and persistent HIV-1 pWT/BaL replication could be obtained only in MDMs grown in RPMI containing macrophage colony-stimulating factor (M-CSF). Interestingly, this MDM type was also most responsive to toll-like receptor stimulation. By contrast, all MDM types were activated to a comparable extent by intracellular DNA, and the macrophage serum-free medium-(Mac-SFM-)differentiated MDMs responded strongly to membrane fusion through expression of CXCL10. Finally, we found that HIV infection of RPMI/M-CSF-differentiated MDMs induced low-grade expression of two interferon-stimulated genes in some donors. In conclusion, our study demonstrates that the differentiation protocol used greatly influences the ability of MDMs to activate innate immune reactions and support HIV-1 replication. Paradoxically, the data show that the MDMs with the strongest innate immune response were also the most permissive for HIV-1 replication.

Genomic HIV RNA induces innate immune responses through RIG-I-dependent sensing of secondary-structured RNA.

BACKGROUND: Innate immune responses have recently been appreciated to play an important role in the pathogenesis of HIV infection. Whereas inadequate innate immune sensing of HIV during acute infection may contribute to failure to control and eradicate infection, persistent inflammatory responses later during infection contribute in driving chronic immune activation and development of immunodeficiency. However, knowledge on specific HIV PAMPs and cellular PRRs responsible for inducing innate immune responses remains sparse. METHODS/PRINCIPAL FINDINGS: Here we demonstrate a major role for RIG-I and the adaptor protein MAVS in induction of innate immune responses to HIV genomic RNA. We found that secondary structured HIV-derived RNAs induced a response similar to genomic RNA. In primary human peripheral blood mononuclear cells and primary human macrophages, HIV RNA induced expression of IFN-stimulated genes, whereas only low levels of type I IFN and tumor necrosis factor α were produced. Furthermore, secondary structured HIV-derived RNA activated pathways to NF-κB, MAP kinases, and IRF3 and co-localized with peroxisomes, suggesting a role for this organelle in RIG-I-mediated innate immune sensing of HIV RNA. CONCLUSIONS/SIGNIFICANCE: These results establish RIG-I as an innate immune sensor of cytosolic HIV genomic RNA with secondary structure, thereby expanding current knowledge on HIV molecules capable of stimulating the innate immune system.