Sensing self and nonself circular RNAs

Circular RNAs (circRNAs) are single-stranded RNAs that are joined head to tail. Initially discovered as pathogen genomes such as hepatitis D virus (HDV) and plant viroids, circRNAs are recently recognized as a pervasive class of noncoding RNAs in eukaryotic cells, generated through back splicing. circRNAs have been postulated to function in cell-to-cell information transfer or memory due to their extraordinary stability. Whether and how circRNAs trigger immune recognition is not known. Here we show that exogenous circular RNAs potently stimulate immune signaling, and mammalian cells sense self vs. nonself circRNAs via circRNA biogenesis. Transfection of purified in vitro spliced circRNA into mammalian cells led to potent induction of innate immunity genes. The nucleic acid sensor RIG-I is necessary and sufficient to sense foreign circRNA, and RIG-I and foreign circRNA co-aggregate in cytoplasmic foci. CircRNA activation of innate immunity is independent of 5’ triphosphate, double-stranded RNA structure, or primary sequence of the foreign circRNA. Instead, self-nonself discrimination depends on the intron that programs the circRNA. Use of a human intron to express a foreign circRNA sequence abrogates immune activation, and the mature human circRNA is associated with diverse RNA binding proteins reflecting its endogenous splicing and biogenesis. These results reveal innate immune sensing of circRNA, a prevalent class of host and pathogen RNAs, and highlight introns—the predominant output of mammalian transcription—as unexpected arbiters of self-nonself identity in the RNA world.