The circular RNA circ_ATP8B regulates ROS production and antiviral immunity [circRNA-Seq]
Circular RNAs (circRNAs) are widely expressed in eukaryotes. However, only a subset has been functionally characterized. We have identified and validated a collection of circular RNAs in Drosophila melanogaster. We show that the circular RNA circ_ATP8B is induced by viral infection and that depletion of circ_ATP8B, but not its linear sibling, compromises viral infection both in cultured cells and in vivo. In addition, our analyses reveal that circ_ATP8B is enriched in the fly gut and that gut-specific depletion of circ_ATP8B attenuates viral replication in an oral infection model. Furthermore, we find circ_ATP8B-depletion resulted in increased levels of reactive oxygen species (ROS) and enhanced expression of Duox (Dual oxidase), which produces ROS. Genetic and pharmacological manipulation of circ_ATP8B-depleted flies that reduce ROS levels rescue the viral replication defects elicited by circ_ATP8B depletion. Notably, circ_ATP8B and Duox associate with each other, and that expression of various versions of circ_ATP8B that are competent in binding Duox, but not a mutant circ_ATP8B that is incapable of binding Duox, restores physiological levels of ROS in circ_ATP8B-depleted cells. Lastly, our data show that Gaq, a subunit of G protein required for optimal Duox activity, acts downstream of circ_ATP8B to regulate Duox activity. We conclude that circ_ATP8B regulates anti-viral immunity by modulating Duox-dependent ROS production. Overall design: This study aims to identify circular RNAs that are differentially expressed upon viral infection. Total RNAs were extracted from cultured Drosophila SL2 cells that were either uninfected or infected with Flock House virus. Ribosomal RNAs and linear RNAs were removed and the enriched circular RNA fraction was subjected to RNA-seq to identify circular RNAs.