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Antigenic diversity shapes immunity in distinct and unexpected ways. This is particularly true of the humoral response generated against influenza A viruses. While it is known that immunological memory developed against previously-encountered influenza A virus strains impacts the outcome of subsequent infections, exactly how sequential exposures to antigenically variant viruses shape the humoral immune response in humans remains poorly understood. To address this important question, a longitudinal analysis of antibody titers against various pandemic and seasonal strains of influenza virus spanning a 20-year period (1987–2008) was performed using samples from 40 individuals (d.o.b. 1917–1952) obtained from the Framingham Heart Study. Longitudinal increases in neutralizing antibody titers were observed against previously-encountered pandemic H2N2, H3N2 and H1N1 influenza A virus strains. Antibody titers against seasonal strains encountered later in life also increased longitudinally at a rate similar to that against their pandemic predecessors. Titers of cross-reactive antibodies specific to the hemagglutinin stalk domain were also investigated, since they are known to be influenced by exposure to antigenically diverse influenza A viruses. These titers rose modestly over time, even in the absence of major antigenic shifts. No sustained increase in neutralizing antibody titers against an antigenically more stable virus (human cytomegalovirus) was observed. The results herein describe a role for antigenic variation in shaping the humoral immune compartment, and provide a rational basis for the hierarchical nature of antibody titers against influenza A viruses in humans.

Significance Virus infections must be combated at a cellular level. The strategies used to inhibit virus differ dramatically when comparing plants and insects to mammals. Here, we identify an evolutionary conserved antiviral response that is independent of these known defenses. We demonstrate that an RNA nuclease called Drosha is repurposed during virus infection to cleave viral RNA and modulate the cellular environment as a means of inhibiting virus replication.

In contrast to the DNA-based viruses in prokaryotes, the emergence of eukaryotes provided the necessary compartmentalization and membranous environment for RNA viruses to flourish, creating the need for an RNA-targeting antiviral system1,2. Present day eukaryotes employ at least two main defense strategies that emerged as a result of this viral shift, namely antiviral RNA interference (RNAi) and the interferon (IFN) system2. Here, we demonstrate that Drosha and related RNase III ribonucleases from all three domains of life, also elicit RNA-targeting antiviral activity. Systemic evolution of ligands by exponential enrichment (SELEX) on this class of proteins illustrates the recognition of unbranched RNA stem loops. Biochemical analyses reveal that in this context, Drosha functions as an antiviral clamp, conferring steric hindrance on the RNA dependent RNA polymerases (RdRps) of diverse positive stranded RNA viruses. We present evidence for cytoplasmic translocation of RNase III nucleases in response to virus in diverse eukaryotes including: plants, arthropods, invertebrate chordates, and fish. These data implicate RNase III recognition of viral RNA as an antiviral defense that is independent of, and possibly predates, other known eukaryotic antiviral systems.

SUMMARY With the capacity to fine-tune protein expression via sequence-specific interactions, microRNAs (miRNAs) help regulate cell maintenance and differentiation. While some studies have also implicated miRNAs as regulators of the antiviral response, others have found that the RISC complex that facilitates miRNA-mediated silencing is rendered non-functional during cellular stress, including virus infection. To determine the global role of miRNAs in the cellular response to virus infection, we generated a vector that rapidly eliminates total cellular miRNA populations in terminally differentiated primary cultures. Loss of miRNAs has a negligible impact on both innate sensing of and immediate response to acute viral infection. In contrast, miRNA depletion specifically enhances cytokine expression, providing a post-translational mechanism for immune cell activation during cellular stress. This work highlights the physiological role of miRNAs during the antiviral response and suggests their contribution is limited to chronic infections and the acute activation of the adaptive immune response.