The Dark Viral Proteome
The past decade has revolutionized our understanding of the coding capacity of genomes, unveiling a new universe of microproteins encoded by non-canonical ORFs, i.e., ORFs initiated by non-AUG codons or those spanning less than 100 amino acids. Illuminating previously unknown viral proteins will deepen our understanding of the viral life cycle and antiviral immunity. However, the landscape of coding sequences remains unknown. I have developed Massively Parallel Ribosome Profiling (MPRP), a pan-viral approach for the measurements of translated regions in ~700 human viruses, and identified more than 4,000 new ORFs. This study exposed a new realm of viral antigens, expanding our view of interactions between T cells and infected cells. Furthermore, I identified hundreds of upstream ORFs (uORFs) that regulate the translation levels of canonical viral proteins encoded from the same transcripts.​
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Unveiling immune targets from 'hidden' viral proteins
T cell-mediated immunity plays a crucial role in controlling viral infections. While current vaccine design strategies primarily focus on T cell epitopes derived from well-known viral proteins, the significance of microproteins originating from non-canonical open reading frames (ORFs) in T cell immunity remains largely unexplored. Unveiling this 'hidden source' of T cell epitopes will enhance our understanding of the immune response to viruses and provide novel targets for vaccine development. During my postdoctoral research, I utilized immunopeptidome profiling to identify viral peptides that are naturally processed and presented on human leukocyte antigen (HLA) molecules to T cells in infected cells. I adapted this protocol to study high-containment viruses and led a multi-institutional team, providing the first comprehensive view of SARS-CoV-2 HLA-I and HLA-II peptides. Our untargeted approach revealed a remarkable enrichment of peptides derived from viral non-canonical ORFs bound to the HLA-I complex. Notably, some of these peptides elicited stronger T cell responses than the most immunogenic T cell epitopes known to date, underscoring the importance of considering this previously overlooked source as potential viral immune targets.​​
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