Defective Influenza A Virus RNA Products Mediate MAVS-Dependent Upregulation of Human Leukocyte Antigen Class I Proteins

Abstract

Influenza A virus (IAV) increases the presentation of class I human leukocyte antigen (HLA) proteins that limit antiviral responses mediated by natural killer (NK) cells, but molecular mechanisms for these processes have not yet been fully elucidated. We observed that infection with A/Fort Monmouth/1/1947(H1N1) IAV significantly increased the presentation of HLA-B, -C, and -E on lung epithelial cells. Virus entry was not sufficient to induce HLA upregulation because UV-inactivated virus had no effect. Aberrant internally deleted viral RNAs (vRNAs) known as mini viral RNAs (mvRNAs) and defective interfering RNAs (DI RNAs) expressed from an IAV minireplicon were sufficient for inducing HLA upregulation. These defective RNAs bind to retinoic acid-inducible gene I (RIG-I) and initiate mitochondrial antiviral signaling (MAVS) protein-dependent antiviral interferon (IFN) responses. Indeed, MAVS was required for HLA upregulation in response to IAV infection or ectopic mvRNA/DI RNA expression. The effect was partially due to paracrine signaling, as we observed that IAV infection or mvRNA/DI RNA-expression stimulated production of IFN-β and IFN-λ1 and conditioned media from these cells elicited a modest increase in HLA surface levels in naive epithelial cells. HLA upregulation in response to aberrant viral RNAs could be prevented by the Janus kinase (JAK) inhibitor ruxolitinib. While HLA upregulation would seem to be advantageous to the virus, it is kept in check by the viral nonstructural 1 (NS1) protein; we determined that NS1 limits cell-intrinsic and paracrine mechanisms of HLA upregulation. Taken together, our findings indicate that aberrant IAV RNAs stimulate HLA presentation, which may aid viral evasion of innate immunity. IMPORTANCE Human leukocyte antigens (HLAs) are cell surface proteins that regulate innate and adaptive immune responses to viral infection by engaging with receptors on immune cells. Many viruses have evolved ways to evade host immune responses by modulating HLA expression and/or processing. Here, we provide evidence that aberrant RNA products of influenza virus genome replication can trigger retinoic acid-inducible gene I (RIG-I)/mitochondrial antiviral signaling (MAVS)-dependent remodeling of the cell surface, increasing surface presentation of HLA proteins known to inhibit the activation of an immune cell known as a natural killer (NK) cell. While this HLA upregulation would seem to be advantageous to the virus, it is kept in check by the viral nonstructural 1 (NS1) protein, which limits RIG-I activation and interferon production by the infected cell.

A defective viral genome strategy elicits broad protective immunity against respiratory viruses

“…Our data indicate that eTIP1s are an effective antiviral approach to harness natural immunity to fight infection in a balanced, safe, and controlled manner, utilizing the regulatory circuits that evolved to ensure protection from disease without causing self-afflicted damage (Rahim et al, 2020;Vignuzzi and Lo ´pez, 2019). DVG-based antiviral therapy may offer greater benefits and safety than conventional therapies, given its broad-spectrum capacity that may prevent rapid emergence of resistant variants and its short and long-lasting protective activity.…”

Pre-existing immunity to influenza viruses through infection and/or vaccination leads to viral mutational signatures associated with unique immune responses during a subsequent infection

“…Interestingly, these mutations were not only abundant on the HA and NA genes as we hypothesized but were also found on the NP and PB1 genes. Specifically, the naïve-vaccinated animals had mutations concentrated in the RBD of HA including W150, L151P, and S183P (65, 66). Another mutation on the RBD mutation, P182Q, was found in the preimmune-mock vaccinated animals only, indicating a mutational pattern distinct from naïve-vaccinated animals.…”

Transient RNA structures cause aberrant influenza virus replication and innate immune activation

“…DVGs and mvRNAs bind RIG-I and activate the MAVS signaling cascade (9,11), but it is unclear what determines whether an IAV mvRNA is an inducer of the innate immune response. To systematically investigate if the sequence or secondary structure of an mvRNA can affect IAV RNA polymerase activity and innate immune activation, we engineered five segment 5-derived mvRNA templates.…”

“…This phenomenon underlies the lethal pathology of infections with 1918 H1N1 pandemic or highly pathogenic avian IAV (7,8). Various viral and host factors have been implicated in causing immunopathology, including the products of aberrant viral replication (9)(10)(11).…”

“…Such defective viral genomes (DVGs) (18) and mini viral RNAs (mvRNA) (9) are >180 nt and 56-125 nt long aberrant viral RNA molecules, respectively, that contain internal deletions between the conserved 5ʹ and 3ʹ termini (9,(19)(20)(21)(22). Both DVGs and mvRNAs can bind RIG-I and activate innate immune responses (9,11). It is presently not fully understood what determines the ability of DVGs and mvRNAs to activate RIG-I or how they are made.…”