Investigating the effect of ribosome perturbation on the translational regulation of the interferon response

The work in this thesis details how changes in ribosome composition and abundance enhance the protein production of interferon-stimulated genes (ISGs), a critical component of innate immunity, ultimately revealing a novel layer of translational control exerted through codon non-optimality. We hypothesize that reducing ribosome biogenesis will facilitate the translation of transcripts of reduced codon optimality, particularly during times of additional stress such as interferon-β (IFN-β) stimulation. We first investigate the potential effects of ribosome heterogeneity through the knockdown of ribosomal proteins (RPs) during IFN-β stimulation; however, we find that targeted RP knockdowns resulted in a broad reduction of all RP levels. We demonstrate that this RP reduction resulted in an increase in ISGs and correlated with a reduction in ribosome biogenesis proteins. To further clarify the role of the ribosome in the translational increase in ISGs, we profiled the transcriptome and proteome under ribosome biogenesis protein BOP1 knockdown and IFN-β stimulation. An alignment of these data revealed that ISGs are translationally up-regulated during BOP1 knockdown and find that a shared commonality amongst all translationally up-regulated proteins is reduced codon optimality. When tested in a reporter assay, we found that non-optimal transcripts were shown to be more translationally up-regulated during BOP1 knockdown and IFN-β stimulation. We propose that this phenomenon represents a translational fine-tuning of key proteins required for the IFN response. We further probe the implications of this work by profiling the proteome of cells depleted of BOP1 and infected with murine respirovirus, with an emphasis on early and middle timepoints of infection. These findings show that reducing ribosome biogenesis through BOP1 knockdown mediates a shift towards the translation of transcripts with low codon optimality during the IFN response, suggesting a novel mechanism of translational control during the IFN response. Ultimately, these insights further shape our understanding of the IFN response and may impact the development of RNA therapeutics and novel antivirals.