RNase L produces tRNA-derived RNAs that contribute to translation inhibition.

Ribonuclease L (RNase L) is an RNase which is activated by viral double-stranded RNAs (dsRNAs). RNase L cleaves not only viral RNAs but also host RNAs, including mRNAs and tRNAs, which contributes to innate immune defense against viruses. While it has been reported that RNase L-mediated bulk mRNA cleavage induces rapid translation repression independently of the integrated stress response, the significance of RNase L-mediated tRNA cleavage remains largely unknown. Here we show that RNase L cleaves various tRNA species in the anticodon loops, generating transfer RNA-derived RNAs (tDRs) similar to tRNA-derived stress-induced RNAs (tiRNAs) that are generated by a stress-responsive RNase angiogenin (ANG). Three tRNA species (tRNA^Leu, tRNA^SeC _, and tRNA^Ser) were cleaved within the variable loops as well as in the anticodon loops by RNase L, generating noncanonical tDRs. As RNase L-induced 5'-tDR^Ala/Cys were similar in length to 5'-tiRNA^Ala/Cys that possess a translation inhibitory effect, we examined whether RNase L-induced 5'-tDR^Ala also inhibited translation. In vitro translation analysis showed that RNase L-induced 5'-tDR^Ala significantly inhibits mRNA translation like 5'-tiRNA^Ala, suggesting that the production of 5'-tDR^Ala may be involved in the mechanism of RNase L-mediated stress response during viral infection. Our data shed new light on the potential roles of tDRs in innate immunity against viral infection.