Spatio-temporal regulation of circular RNA expression during porcine embryonic brain development

Background: Recently, thousands of circular RNAs (circRNAs) have been discovered in various tissues and cell types from human, mouse, fruit fly and nematodes. However, expression of circRNAs across mammalian brain development has never been examined. Results: Here we profile the expression of circRNA in five brain tissues at up to six time-points during fetal porcine development, constituting the first report of circRNA in the brain development of a large animal. An unbiased analysis reveals a highly complex regulation pattern of thousands of circular RNAs, with a distinct spatio-temporal expression profile. The amount and complexity of circRNA expression was most pronounced in cortex at day 60 of gestation. At this time-point we find 4634 unique circRNAs expressed from 2195 genes out of a total of 13,854 expressed genes. Approximately 20 % of the porcine splice sites involved in circRNA production are functionally conserved between mouse and human. Furthermore, we observe that "hot-spot" genes produce multiple circRNA isoforms, which are often differentially expressed across porcine brain development. A global comparison of porcine circRNAs reveals that introns flanking circularized exons are longer than average and more frequently contain proximal complementary SINEs, which potentially can facilitate base pairing between the flanking introns. Finally, we report the first use of RNase R treatment in combination with in situ hybridization to show dynamic subcellular localization of circRNA during development. Conclusions: These data demonstrate that circRNAs are highly abundant and dynamically expressed in a spatio-temporal manner in porcine fetal brain, suggesting important functions during mammalian brain development.