Unveiling novel conserved HIV-1 open reading frames encoding T-cell antigens using ribosome profiling.

Alternative reading frames (ARFs) were described in several viral genomes using ribosome profiling such as HCMV, KSV, and SARS-CoV-2 but no demonstration has been made on HIV genome. Using ribosome profiling, we uncovered 98 conserved HIV ARFs distributed across the genome, with high conservation among HIV clade B and C isolates. Our analysis revealed that at least 42 ARFs encode viral polypeptides, as demonstrated by T-cell responses targeting 45 ARF-derived peptides in patients under treatment or naturally controlling the infection. At the same time, we identified a ligand of HLA-A*0201 derived from an identified ARF on primary infected cells. These responses were mediated by polyfunctional CD4+ T-cells secreting at least 3 cytokines simultaneously. Our discovery expands the list of conserved viral polypeptides that might be potential targets for vaccination strategies. Overall design: RiboSeq of supT1 cells infected by the virus HIV NL4-3 XCS. Cells are harvested 20h post-infection. Two independant infections have been done, which were sequenced in triplicate

交替阅读框（Alternative reading frames, ARFs）已通过核糖体图谱（ribosome profiling）技术在多种病毒基因组中被报道，涵盖HCMV、KSV及SARS-CoV-2，但目前尚未在HIV基因组中得到证实。本研究借助核糖体图谱技术，在HIV全基因组范围内发现了98个保守型HIV交替阅读框，且这些阅读框在HIV B亚型与C亚型分离株中呈现高度保守性。分析结果显示，至少42个交替阅读框可编码病毒多肽，这一结论得到了临床治疗组及自然感染控制组患者体内靶向45个交替阅读框衍生肽段的T细胞应答数据的验证。同时，本研究在原代感染细胞中鉴定出一个来自已验证交替阅读框的人类白细胞抗原（Human Leukocyte Antigen, HLA）-A*0201配体。此类应答由同时分泌至少3种细胞因子的多功能CD4+ T细胞所介导。本研究的发现扩充了可作为疫苗研发潜在靶点的保守型病毒多肽列表。实验整体设计：对感染HIV NL4-3 XCS毒株的supT1细胞开展核糖体测序（ribosome sequencing, RiboSeq）。于感染后20小时收集细胞，共完成两次独立感染实验，每份样本均进行三次重复测序。