Post-transcriptional modifications on tRNA fragments confer functional changes to high-density lipoproteins in atherosclerosis

SUMMARY Epitranscriptomic modifications on RNA play critical roles in stability, processing, and function, partly by influencing interactions with RNA-binding proteins and receptors. The role of post-transcriptional RNA modifications on cell-free non-coding small RNA (sRNA) remains poorly understood in disease contexts. High-density lipoproteins (HDL), which transport sRNAs, can lose their beneficial properties in atherosclerosis cardiovascular disease (ASCVD). We hypothesize that changes to regulatory modifications on HDL-sRNAs contribute to this dysfunction. To assess changes in HDL-sRNA modification status, HDL-derived RNA from healthy subjects and those with atherosclerotic lesion development were analyzed using LC-MS/MS and AlkB-facilitated RNA (de)Methylation Sequencing. ASVD-HDL showed an enrichment in modified nucleosides including m 1 A tRNA-derived sRNAs (tDRs), particularly tDR-ArgACG-1. Functional studies revealed that ASCVD-HDL induced cell adhesion genes, including TMEM123, in primary macrophages. Recombinant HDL loaded with m 1 A-tDR-ArgACG-1 induced immune signaling, and similarly upregulated TMEM123. These findings suggest HDL-delivered-m 1 A-tDR-ArgACG-1 act on adhesion genes and immune pathways, promoting macrophage activation.