Identification of pathological pathways centered on circRNA dysregulation associated with irreversible progression of Alzheimer’s disease [5xFADHippocampus]

Background Circular RNAs (circRNAs) are highly stable regulatory molecules, often accumulated in the mammalian brain and thought to serve as “memory molecules” that govern the long process of aging. Mounting evidence demonstrated circRNA dysregulation in postmortem cortical regions of Alzheimer’s disease (AD) patients. However, whether and how circRNA dysregulation underlies AD progression remains unexplored. Methods Poly(A)-tailing/RNase R digestion was coupled with our recently published computational algorithm CARP to identify genome-wide dysregulation of circRNAs and their downstream pathways in the 5xFAD mouse cerebral cortex between 5- and 7-month of age, a critical window that marks the transition from reversible to irreversible pathogenic progression. Irreversibly dysregulated circRNAs and pathways in 5xFAD cortex were systematically compared with a large human AD cohort study. A top ranked circRNA conserved and commonly affected in AD patients and 5xFAD mice was depleted in cultured cells to confirm its causal roles in AD-relevant molecular and cellular changes. Results We discovered genome-wide circRNA alterations specifically in the 5xFAD cortex associated with AD progression and found numerous circRNAs also consistently affected in AD patient postmortem cortical areas. Among such circRNAs, circGigyf2 showed the highest net reduction starting from 7-month of age, which is highly conserved and negatively correlated with human AD severity. Mechanistically, we found multiple AD-affected splicing factors that are essential for circGigyf2 biogenesis. Functionally, we identified and experimentally validated the conserved roles of circGigyf2 in sponging AD-relevant miRNAs and AD-associated RNA binding proteins (RBPs) including the cleavage and polyadenylation factor 6 (CPSF6). CircGigyf2 downregulation in AD exert silencing activities of its sponged miRNAs and aberrantly enhanced the polyadenylation efficiency of many CPSF6 targets. These post-transcriptionally tuned mRNAs by circGigyf2 were enriched in apoptotic pathways. Furthermore, circGigyf2 depletion in a mouse immortalized neuronal cell line dysregulated circGigyf2-miRNA and circGigyf2-CPSF6 axes and potentiated neuronal responses to apoptotic insults, which strongly support the causative role of circGigyf2 deficiency in AD disease progression. Conclusions Together, our results unveiled pathological alterations of the brain circRNA landscape associated with irreversible disease stage in an AD mouse model and identified novel molecular mechanisms underlying dysregulation of conserved circRNA pathways that contribute to AD pathogenesis.