Leveraging Signaling and Antigen-presenting Bifunctional Receptors (SABRs) with an expanded genetic code to interrogate the immunogenicity of post-translational modifications 3559

Abstract Description T cells survey cellular health by recognizing peptides presented on MHC through their T cell receptor (TCR), with TCR-pMHC specificity and affinity determining protective or pathogenic roles. While high-throughput sequencing has revealed TCR repertoire evolution in disease, identifying the antigens driving these changes remains challenging. Post-translational modifications (PTMs) introduce diversity into the antigenic landscape by modulating antigen processing, presentation, or TCR recognition and are linked to disease when dysregulated. However, PTMs remain underexplored as immune targets due to the inability of genetically encoded libraries to incorporate PTMs. To address this, we built upon our lab’s success with SABRs and developed SCEPTRE (SABRs with Codon Expanded Post-TRanslationally Modified Epitopes), a platform for high-throughput screening of PTM immunogenicity. SCEPTRE uses genetic code expansion with an orthogonal tRNA/aminoacyl-tRNA synthetase system to incorporate PTMs site-specifically into pMHC libraries, enabling PTM antigen discovery. We first demonstrated the ability to incorporate Lysine derivatives and Citrulline into SCEPTREs. We further utilized SCEPTREs to demonstrate that PTM position and type significantly influence MHC binding and TCR activation patterns. SCEPTRE offers a novel method to explore PTM-driven antigenicity, advancing the discovery of antigenic targets in autoimmune diseases and cancer, where PTM dysregulation plays a critical role. Funding Sources Supported by JDRF/3-SRA-2023-1354-S-B. Topic Categories Technological Innovations in Immunology (TECH)