Respiratory syncytial virus (RSV) enhances translation of virus-resembling AU-rich host transcripts

Viral infection often triggers eukaryotic initiator factor 2a (eIF2a) phosphorylation, leading to global 5'-cap-dependent translation inhibition. RSV encodes messenger RNAs (mRNAs) mimicking 5'-cap structures of host mRNAs and thus inhibition of cap-dependent translation initiation would likely also reduce viral translation. We confirmed that RSV limits widespread translation initiation inhibition and unexpectedly found that the fraction of ribosomes within polysomes increases during infection, indicating higher ribosome loading on mRNAs during infection. We found that AU-rich host transcripts that are less efficiently translated under normal conditions become more efficient at recruiting ribosomes, similar to RSV transcripts. Viral transcripts are transcribed in cytoplasmic inclusion bodies, where the viral AU-rich binding protein M2-1 has been shown to bind viral transcripts and shuttle them into the cytoplasm. We further demonstrated that M2-1 is found on polysomes, and that M2-1 might deliver host AU-rich transcripts for translation. Overall design: To determine which viral and host transcripts are associated with polysomes during RSV infection, total and polysome-associated RNA from mock- and RSV-infected cells was collected, Trizol extracted and sequenced after poly(A) (A+) enrichment. RNA-sequencing samples contain biological triplicates for (1) mock-infected total RNA, (2) RSV-infected total RNA, (3) mock-infected polysome-associated RNA, and (4) RSV-infected polysome-associated RNA.