When a host is infected with pathogenic microbes, it has two essential arms of defense to eliminate them: the innate immune system and the adaptive immune system. Toll-like receptors (TLRs) play a crucial role in the innate immune response. TLRs and other pathogen recognition receptors detect pathogen-associated molecular patterns (PAMPs) and initiate protective responses. Among the 10 TLRs characterized in humans, TLR1, -2, -4, -5, -6, and -10 localize to the cell surface (surface TLRs), while TLR3, -7, -8, and -9 localize to intracellular compartments such as endosomes (endosomal TLRs). Although endosomal TLR7 recognizes single-stranded RNAs, their endogenous RNA ligands have not been fully explored.
Here, we report 5′-tRNA half molecules (a half fragment of tRNA) as abundant activators of TLR7. Mycobacterial infection and accompanying surface TLR activation by lipopolysaccharide (LPS) and peptidoglycan (PGN) upregulate the expression of 5′-tRNA half molecules in human monocyte-derived macrophages (HMDMs). The abundant accumulation of 5′-tRNA halves also occur in HMDM-secreted extracellular vehicles (EVs). Further, absolute quantification of RNA revealed the abundant presence of EV-5′-tRNAHisGUG half molecules which is >200-fold higher than that of the most abundant EV-microRNA.
Sequence identification of the 5′-tRNA halves using cP-RNA-seq revealed abundant and selective packaging of specific 5′-tRNA half species into EVs. The EV-5′-tRNAHisGUG half was experimentally demonstrated to be delivered into endosomes in recipient cells and to activate endosomal TLR7. This activation of TLR7 was studied by quantifying the upregulated mRNA of cytokines as well as secreted pro-inflammatory cytokines. Upregulation of the 5′-tRNA half molecules was also observed in the plasma of patients infected with Mycobacterium tuberculosis, verifying that the observed phenomena occur not only in cell culture systems but also in actual pathological situations.
These results unveil a novel tRNA-engaged pathway in the innate immune response and assign the role of “immune activators” to 5′-tRNA half molecules.
A proposed model for 5′-tRNA half-mediated immune response
References
- Pawar K, Shigematsu M, Sharbati S, Kirino Y. Infection-induced 5′-half molecules of tRNAHisGUG activate Toll-like receptor 7. PLoS Biol. 2020 Dec 17;18(12):e3000982. doi: 10.1371/journal.pbio.3000982. PubMed PMID:33332353.