Identification And Characterization Of An Ancient Class Of Small RNAs Enriched In Serum Associating With Active Infection

Serum microRNAs (miRNAs) have recently emerged as promising biomarkers for a variety of diseases including cancer and metabolic disorders. There are too many RNase that it is difficult for RNA existing stably in serum. To data, serum miRNAs are mostly reported to be protected from rapide degradation though either encapsulated in the serum microvesicles (exosomes) or binding with serum proteins. However, despite the increasing category of small RNAs, the existence and diagnostic value of other serum small RNAs are surprisingly few.Transfer RNA (tRNA) is an accient class of RNA which expandly exist in the whole creature kingdom. Active aminoacyl tRNA deliver correct amino acid to the right site of the translating pepitides by precisiely complementing to the mRNA code with their anticode.tRNA play a essential role in genetic information translation. But expect of the role of protein translation, the other fuctions of tRNA still unclear. Previous studies at cellular level have demonstrated that tRNA derived small RNAs (tsRNAs) could be upregulated under various stresses. However, whether the tsRNA enriches in animal organism under physiological conditions is still unkown.Our lab recently identified a novel class of tsRNAs that were derived from the5’half of tRNAs (29-34nt) and highly concentrated in mature mouse sperm under physiological condition. Following this clue, we screened multiple mouse organs by small RNA deep sequencing (18-40nt) to reveal potential existence of tsRNAs in other tissues. With the next following study, we discovered an ancient class of tRNA-derived small RNAs (tsRNAs) abundantly, conservatively existed in the serum of a wide range of vertebrate species (from fish to human). Thus we revealed the evolutionary conservatism of tsRNAs along the evolution tree (from fish to human), establishing tsRNAs as an even more ancient, conservative and prevailing form of serum small RNA across vertebrate kingdom than miRNAs. We further found that tsRNAs are stably existed in the serum, free from serum microvesicles (exosomes). We next separated serum contents by centrifugal filters with different molecular weight cut-offs, and found that the tsRNAs were highly concentrated in serum complex with molecular weight>100kDa, suggesting their co-existence with serum protein complexes. We systematically examined the stability of serum tsRNAs by comparing serum miRNAs and synthesized tsRNAs, and get a novel conclusion that serum tsRNAs use the nucleotide modifications inheriting from tRNA predecessors to increase their stability. Thus we revealed the stabilization mechanism of serum tsRNAs in an RNase-rich blood environment involves at least two layers of protection: through binding and co-existed with serum protein complexes, as well as by nucleotide modifications inheriting from their tRNA predecessors. Most importantly, we found that serum tsRNAs showed surge up regulation during LPS-induced acute inflammation in mouse and monkey, as well as in human patients under virus infection (HBV replication phase), suggesting their active involvements in infection-induced defensive response.Overall, our data unveiled another hidden layer of serum small RNAs linking with disease condition, opening future avenues for the development of novel biomarker approaches based on analyzing serum tsRNAs.