biostudies-literatureBertrand LSidaction1707https://www.ebi.ac.uk/biostudies/studies/S-EPMC11836469biostudies-literatureUnknown16(1)The development of ribosomal profiling (Riboseq) revealed the immense coding capacity of human and viral genomes. Here, we used Riboseq to delineate the translatome of HIV-1 in infected CD4⁺ T cells. In addition to canonical viral protein coding sequences (CDSs), we identify 98 alternative open reading frames (ARFs), corresponding to small Open Reading Frames (sORFs) that are distributed across the HIV genome including the UTR regions. Using a database of HIV genomes, we observe that most ARF amino-acid sequences are likely conserved among clade B and C of HIV-1, with 8 ARF-encoded amino-acid sequences being more conserved than the overlapping CDSs. Using T cell-based assays and mass spectrometry-based immunopeptidomics, we demonstrate that ARFs encode viral polypeptides. In theNature communicationsUnveiling conserved HIV-1 open reading frames encoding T cell antigens using ribosome profiling.PMC1183646917-1-AEQ-11102-1Decimo DBet AZiegler MCNamy OVinh JJeger-Madiot RBeauclair GHatin IFrancois PArbes HGuiguettaz LEsclatine ARamirez BCVerdier YRicci EPGregoire SMoris AWalz JCardinaud SGouttefangeas CAutran BPereira MBertrand LHocqueloux LLambotte OAltfeld MGross ARammensee HGGraff-Dubois SLabaronne ENelde ASamri ADemais SfalseUnveiling conserved HIV-1 open reading frames encoding T cell antigens using ribosome profiling.The development of ribosomal profiling (Riboseq) revealed the immense coding capacity of human and viral genomes. Here, we used Riboseq to delineate the translatome of HIV-1 in infected CD4⁺ T cells. In addition to canonical viral protein coding sequences (CDSs), we identify 98 alternative open reading frames (ARFs), corresponding to small Open Reading Frames (sORFs) that are distributed across the HIV genome including the UTR regions. Using a database of HIV genomes, we observe that most ARF amino-acid sequences are likely conserved among clade B and C of HIV-1, with 8 ARF-encoded amino-acid sequences being more conserved than the overlapping CDSs. Using T cell-based assays and mass spectrometry-based immunopeptidomics, we demonstrate that ARFs encode viral polypeptides. In the2025-01-01T00:00:00Z2025 Feb2025-04-20T01:49:50.155Z2025-04-20T01:49:50.155ZS-EPMC118364693996634010.1038/s41467-025-56773-2