The translational landscape of HIV-1 infected cells reveals key gene regulatory principles
Human immunodeficiency virus-1 (HIV-1) uses a number of strategies to modulate viral and host gene expression during its life cycle. To characterize the transcriptional and translational landscape of HIV-1 infected cells, we used a combination of ribosome profiling, disome sequencing and RNA sequencing. We show that HIV-1 messenger RNAs are efficiently translated at all stages of infection, despite evidence for a substantial decrease in the translational efficiency of host genes that are implicated in host cell translation. Our data identify upstream open reading frames in the HIV-1 5'-untranslated region as well as internal open reading frames in the Vif and Pol coding domains. We also observed ribosomal collisions in Gag-Pol upstream of the ribosome frameshift site that we attributed to an RNA structural fold using RNA structural probing and functional analysis. Antisense oligonucleotides designed to alter the base of this structure decreased frameshift efficiency. Overall, our data highlight the complexity of HIV-1 gene regulation and provide a key resource for decoding of host–pathogen interactions upon HIV-1 infection. Furthermore, we provide evidence for a RNA structural fold including the frameshift site that could serve as a target for antiviral therapy. Overall design: To investigate the translational landscape of HIV-1 infected T-cells, we performed ribosome profiling experiments (RiboSeq, DisomeSeq) treated either with cycloheximide or harringtonine, as well as RNASeq at different timepoints of infection (mock, 8 hpi, 16 hpi, 24 hpi). Furthermore, we analyzed the HIV transcriptome present in the cell with Nanopore sequencing. Lastly, to investigate the structure of the ribosome frameshift site, we employed RNA structural probing (DMS-MaPseq).