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The genomes of positive-sense (+) single-stranded RNA (ssRNA) viruses are believed to be subjected to a wide range of RNA modifications. In this study, we focused on the chikungunya virus (CHIKV) as a model (+) ssRNA virus to study the landscape of viral RNA modification in infected human cells. Among the 32 distinct RNA modifications analysed by mass spectrometry, inosine was found enriched in the genomic CHIKV RNA. However, orthogonal validation by Illumina RNA-seq analyses did not identify any inosine modification along the CHIKV RNA genome. Moreover, CHIKV infection did not alter the expression of ADAR1 isoforms, the enzymes that catalyse the adenosine to inosine conversion. Together, this study highlights the importance of a multidisciplinary approach to assess the presence of RNA modifications in viral RNA genomes.

Despite the nuclear localization of the m 6 A machinery, the genomes of multiple exclusively-cytoplasmic RNA viruses, such as chikungunya (CHIKV) and dengue (DENV), are reported to be extensively m 6 A-modified. However, these findings are mostly based on m 6 A-Seq, an antibody-dependent technique with a high rate of false positives. Here, we address the presence of m 6 A in CHIKV and DENV RNAs. For this, we combine m 6 A-Seq and the antibody-independent SELECT and nanopore direct RNA sequencing techniques with functional, molecular, and mutagenesis studies. Following this comprehensive analysis, we find no evidence of m 6 A modification in CHIKV or DENV transcripts. Furthermore, depletion of key components of the host m 6 A machinery does not affect CHIKV or DENV infection. Moreover, CHIKV or DENV infection has no effect on the m 6 A machinery's localization. Our results challenge the prevailing notion that m 6 A modification is a general feature of cytoplasmic RNA viruses and underscore the importance of validating RNA modifications with orthogonal approaches., (© 2024. The Author(s).)

Ample evidence indicates that codon usage bias regulates gene expression. How viruses, such as the emerging mosquito-borne Chikungunya virus (CHIKV), express their genomes at high levels despite an enrichment in rare codons remains a puzzling question. Using ribosome footprinting, we analyze translational changes that occur upon CHIKV infection. We show that CHIKV infection induces codon-specific reprogramming of the host translation machinery to favor the translation of viral RNA genomes over host mRNAs with an otherwise optimal codon usage. This reprogramming was mostly apparent at the endoplasmic reticulum, where CHIKV RNAs show high ribosome occupancy. Mechanistically, it involves CHIKV-induced overexpression of KIAA1456, an enzyme that modifies the wobble U34 position in the anticodon of tRNAs, which is required for proper decoding of codons that are highly enriched in CHIKV RNAs. Our findings demonstrate an unprecedented interplay of viruses with the host tRNA epitranscriptome to adapt the host translation machinery to viral production., (© 2022. The Author(s).)