Abstract
The interferon-regulated antiviral responses are essential for the induction of both innate and adaptive immunity in mammals. Production of virus-derived small-interfering RNAs (vsiRNAs) to restrict virus infection by RNA interference (RNAi) is a recently identified mammalian immune response to several RNA viruses, which cause important human diseases such as influenza and Zika virus. However, little is known about Dicer processing of viral double-stranded RNA replicative intermediates (dsRNA-vRIs) in mammalian somatic cells. Here we show that infected somatic cells produced more influenza vsiRNAs than cellular microRNAs when both were produced by human Dicer expressed de novo, indicating that dsRNA-vRIs are not poor Dicer substrates as previously proposed according to in vitro Dicer processing of synthetic long dsRNA. We report the first evidence both for canonical vsiRNA production during wild-type Nodamura virus infection and direct vsiRNA sequestration by its RNAi suppressor protein B2 in two strains of suckling mice. Moreover, Sindbis virus (SINV) accumulation in vivo was decreased by prior production of SINV-targeting vsiRNAs triggered by infection and increased by heterologous expression of B2 in cis from SINV genome, indicating an antiviral function for the induced RNAi response. These findings reveal that unlike artificial long dsRNA, dsRNA-vRIs made during authentic infection of mature somatic cells are efficiently processed by Dicer into vsiRNAs to direct antiviral RNAi. Interestingly, Dicer processing of dsRNA-vRIs into vsiRNAs was inhibited by LGP2 (laboratory of genetics and physiology 2), which was encoded by an interferon-stimulated gene (ISG) shown recently to inhibit Dicer processing of artificial long dsRNA in cell culture. Our work thus further suggests negative modulation of antiviral RNAi by a known ISG from the interferon response.
Highlights
Dicer enzymes in the RNase III family mediate the biogenesis of microRNAs and small interfering RNAs in plants and animals [1,2]
We show that the double-stranded RNA (dsRNA) precursors of influenza virus-derived small-interfering RNAs (vsiRNAs) were processed more efficiently than cellular precursor microRNA hairpins by wild-type human Dicer expressed de novo in Dicer-knockout somatic cells
We found that infection of two strains of suckling mice with wild-type Nodamura virus (NoV) was associated with production of silencing-active vsiRNAs and direct sequestration of duplex vsiRNAs by its RNA interference (RNAi) suppressor protein B2
Summary
Dicer enzymes in the RNase III family mediate the biogenesis of microRNAs (miRNAs) and small interfering RNAs (siRNAs) in plants and animals [1,2]. The RNAi pathway functions as a potent antiviral immunity in plants and invertebrates because these hosts produce highly abundant virus-derived siRNAs (vsiRNAs) by Dicer from viral dsRNA precursors to guide RISC-dependent clearance of virus RNAs [6]. The type I interferon (IFN) response is a major first line of defense against virus infection before the activation of adaptive immunity [9,10]. The IFN antiviral response is frequently initiated by cytoplasmic sensing of viral RNA ligands by retinoic acidinducible gene I (RIG-I) or melanoma differentiation factor 5 (MDA5). Upon RNA binding, these RIG-I-like receptors (RLRs) interact with mitochondrial antiviral-signaling protein (MAVS, known as VISA, IPS-1 or Cardif) to activate RLR signal transduction, leading to transcriptional induction of the genes encoding type I IFN and other genes in the nucleus [11]. Not essential for the induction of the IFN response, LGP2 can modulate antiviral
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