Profiling tRNA-derived fragments in LPS-induced microglia and their influence on immune response

The resident macrophages of the brain, microglia, react to immunological responses and preserve homeostasis in the central nervous system. Microglia play a critical role in neuroinflammation, which occurs in neurological and neurodegenerative diseases. Microglial responses are associated with neurodegenerative diseases, autoimmune diseases, and cancer, and therefore require strict regulation. Transfer RNA-derived fragments (tRFs) are small non-coding regulatory RNA molecules generated from the cleavage of transfer RNAs. tRFs have recently emerged as one of the key regulators of gene expression and cellular function in various biological processes. Understanding the content of tRFs altered in microglia due to LPS exposure may provide insights into the mechanisms involved in disease pathogenesis, where neuroinflammation plays a role. We used next-generation sequencing to determine tRF profiles of three different fractions (lysate, extracellular vesicles, and extracellular vesicles-free supernatant) of LPS-induced microglial cells. A total of 345 tRFs were differentially expressed across these fractions after LPS stimulation, including 13 DE-tRFs shared by all three compartments. Gene Ontology analysis of these 13 tRFs suggests their potential involvement in the inflammatory responses of microglia. Among the 13 consistently dysregulated DE-tRFs, tDR-1:33-Glu-CTC-1-M2 was selected for preliminary candidate-level assessment based on its consistent differential expression across all three fractions in our NGS data and previous reports linking this fragment to neuroinflammatory contexts.Increased expression pattern of tDR-1:33-Glu-CTC-1-M2 was further confirmed in lysate samples of LPS-induced microglial cells via RT-qPCR. Inhibition of tDR-1:33-Glu-CTC-1-M2 attenuated LPS-induced cytokine expression and release in microglia, supporting an association between this candidate tRF and inflammatory readouts in this model.