Antiretroviral function of primate and murine PYHIN proteins and evasion by HIV-2

Abstract

PYHIN proteins are only found in mammals and play key roles in immune system against various pathogens. The mammalian pyhin gene locus shows variable deletion and expansion ranging from 4 in humans to a maximum of 13 in mice. Even tough initially thought to act as cytosolic DNA sensors that recognize foreign nucleic acids, increasing evidence suggests that PYHIN proteins also inhibit viral pathogens by more direct mechanisms. Human nuclear PYHIN proteins γ-IFN-Inducible protein 16 (IFI16), IFN-Inducible protein X (IFIX) and Myeloid Nuclear Differentiation Antigen (MNDA) inhibit lentiviral transcription of Human Immunodeficiency Virus 1 (HIV-1) by reducing the availability of host transcription factor Specificity protein 1 (Sp1), a process independent of their sensing function. However, it has remained unclear whether there are differences in the context of HIV-2 and its precursor Simian Immunodeficiency Virus from sooty mangabey monkeys (SIVsmm). My thesis aimed to explore whether and how PYHIN proteins affect the replication of HIV-2 and SIVsmm. My results revealed that HIV-2 strains show reduced susceptibility to PYHIN proteins IFI16, MNDA, and IFIX compared to SIVsmm. Additionally, endogenous IFI16 in primary Clusters of differentiation 4 (CD4) + T cells significantly lowered SIVsmm virus production, while epidemic HIV-2 strains exhibited almost complete resistance, suggesting that IFI16 was a potent barrier for SIVsmm transmission and spread in humans. Mutational analyses showed that the factors determining the susceptibility of SIVsmm to PYHIN protein restriction are fundamentally different from those in HIV-1 subtype B. The determinants of differential susceptibility map 3` half of the viral genome, with a particular emphasis on the nef coding region. This suggests that HIV-2 employs a distinct mechanism to evade restriction by nuclear human PYHIN proteins. Inhibition of HIV-1 requires both pyrin domain and the linker region. I show that inhibition of SIVsmm shares similarities with HIV-1 and requires nuclear localization of IFI16. However, both pyrin domain and HIN domains are dispensable for antiviral activity against SIVsmm. Here, I also examined the antiviral activity of all 13 murine PYHIN proteins to inhibit HIV-1 and murine leukemia virus (MLV). I show that overexpression of p203, p204, p205, p208, p209, p210, p211, and p212 inhibits production of infectious HIV-1; In contrast, p202, p207, and p213 showed little to no effect, while p206 and p214 demonstrated moderate activity. Inhibition of infectious HIV-1 production significantly correlated with the suppression of reporter gene expression by a proviral Moloney MLV-eGFP construct and HIV-1 and Friend MLV LTR luciferase reporter constructs. Overall, my results reveal that the antiviral functions of PYHIN proteins are preserved in both mice and humans, emphasizing their critical role in innate antiviral defense mechanisms. In conclusion, during adaptation to humans, HIV-2 group A evolved resistance to nuclear PYHIN proteins, and this most likely facilitated its epidemic spread. The mechanisms underlying the evasion of this restriction by HIV-2 are distinct from HIV-1 and map to the nef gene. PYHIN proteins are more enigmatic than previously anticipated because neither the pyrin nor the HIN domains are required for inhibition of SIVsmm. The determinants underlying the antiviral function as well as evasion of this restriction by HIV-2 are both distinct from HIV-1. Furthermore, I show that various mouse PYHIN proteins inhibit human and mouse retroviruses, similar to their human counterparts, and suggest a conserved mode of action across species and confirms evidence of overlapping functions but also remarkable diversification of mouse PYHIN proteins.