Hay, B. N. | Search bioRxiv

bioRxiv2025

Ribosome biogenesis mediates the translational increase of non-optimal codon transcripts during IFN stimulation

The interferon response is a signaling pathway unique to vertebrates that links the innate and adaptive immune responses. Interferons signal through a cascade of factors including the JAK-STAT pathway to induce the transcription of hundreds of interferon-stimulated genes (ISGs). Although the main interferon signal transduction pathways and ISGs have been elucidated, translational regulation of ISG transcripts is not fully understood. Prior work demonstrated that ribosomal protein RPL28 negatively regulates a subset of ISGs; however, we find that this effect may be due to a reduction in overall ribosome availability. Multi-omics analysis of RNA-seq and LC-MS/MS data reveal proteins, including several ISGs, that are translationally up-regulated in IFN-{beta}-stimulated cells depleted of ribosome biogeneis factor BOP1. Analysis of codon usage demonstrates a significant reduction in codon optimality for proteins that are translationally up-regulated during BOP1 knockdown and IFN-{beta} stimulation. Using reporter constructs, we demonstrate that codon non-optimal reporters are translated more than codon-optimized reporters in BOP1-depleted IFN-{beta} cells. We propose that ribosome biogenesis regulates translational fine-tuning of integral protein production to ensure optimal interferon responses.

systems biology↗

bioRxiv2022

Cleavage of 14-3-3ϵ by the enteroviral 3C protease dampens RIG-I mediated antiviral signaling

Viruses have evolved diverse strategies to evade the host innate immune response and promote infection. The RIG-I-like-receptors RIG-I and MDA5 (RLRs) are antiviral factors that sense viral RNA and signal downstream via mitochondrial antiviral-signaling protein (MAVS) to activate type I interferon (IFN) expression. 14-3-3{varepsilon} is a key component of the RIG-I translocon complex that interacts with MAVS at the mitochondrial membrane; however, the exact role of 14-3-3{varepsilon} in this pathway is not well understood. In this study, we demonstrate that 14-3-3{varepsilon} is a direct substrate of both the poliovirus and coxsackievirus B3 (CVB3) 3C proteases (3Cpro), and that it is cleaved at Q236{downarrow}G237, resulting in the generation of N- and C-terminal fragments of 27.0 and 2.1 kDa, respectively. Expression of the N-terminal cleavage fragment in cells reduces IFNB mRNA production during poly(I:C) stimulation, thus suggesting an antagonistic effect in the presence of the endogenous 14-3-3{varepsilon} protein. The N-terminal 14-3-3{varepsilon} fragment does not interact with RIG-I in co-immunoprecipitation assays, nor can it facilitate RIG-I translocation to the mitochondria. Probing the intrinsically disordered C-terminal region identifies key residues responsible for RIG-I signaling. Finally, overexpression of the N-terminal fragment promotes CVB3 infection and influenza A virus (H1N1) RNA production and reduces IFNB mRNA production during infection. The strategic enterovirus 3Cpro-mediated cleavage of 14-3-3{varepsilon} antagonizes RIG-I signaling by disrupting critical interactions within the RIG-I translocon complex, thus contributing to evasion of the host antiviral response. Author SummaryHost antiviral factors work to sense virus infection through various mechanisms, including a complex signaling pathway known as the RIG-I like receptor (RLR) pathway. This pathway drives the production of antiviral molecules known as interferons, which are necessary to establish an antiviral state in the cellular environment. Key to this antiviral signaling pathway is the small chaperone protein 14-3-3{varepsilon}, which facilitates the delivery of a viral sensor protein, RIG-I, to the mitochondria. In this study, we show that the enteroviral 3C protease cleaves 14-3-3{varepsilon} during infection, rendering it incapable of facilitating this antiviral response. We also find that the cleavage fragment inhibits RIG-I signaling and promotes virus infection. Our findings reveal a novel viral strategy that restricts the antiviral host response and provides insights into the mechanisms underlying 14-3-3{varepsilon} function in RIG-I antiviral signaling.

microbiology↗