Comparative analysis of gammaherpesvirus circRNA repertoires: conserved and unique viral circRNAs.

Abstract

Recent studies have identified circular RNAs (circRNAs) expressed from the Epstein Barr virus (EBV) and Kaposi’s sarcoma herpesvirus (KSHV) human DNA tumor viruses. To gain initial insights into the potential relevance of EBV circRNAs in virus biology and disease, we assessed the circRNAome of the interspecies homologue, rhesus macaque lymphocryptovirus (rLCV) in a naturally occurring lymphoma from a simian immunodeficiency (SIV) virus infected rhesus macaque. This analysis revealed rLCV orthologs of the latency-associated EBV circular RNAs, circRPMS1_E4_E3a and circEBNA_U. Also identified in two samples displaying unusually high lytic gene expression was a novel rLCV circRNA that contains both conserved and rLCV-specific RPMS1 exons and whose backsplice junctions flank an rLCV lytic origin of replication (OriLyt). Analysis of a lytic infection model for the murid herpesvirus 68 (MHV68) rhadinovirus identified a cluster of circRNAs near an MHV68 lytic origin of replication with the most abundant of these, circM11_ORF69 spanning the OriLyt. Lastly, analysis of KSHV latency and reactivation models revealed the latency associated circRNA originating from the vIRF4 gene as the predominant viral circRNA. Together, this study broadens our appreciation for circRNA repertoires in the lymphocryptovirus and rhadinovirus genera of gammaherpesviruses and provides evolutionary support for viral circRNA functions in latency and viral replication.

IMPORTANCE

Infection with oncogenic gammaherpesviruses leads to long-term viral persistence through a dynamic interplay between the virus and the host immune system. Critical for remodeling of the host cell environment after the immune responses are viral non-coding RNAs that modulate host signaling pathways without attracting adaptive immune recognition. Despite the importance of non-coding RNAs in persistent infection, the circRNA class of non-coding RNAs has only recently been identified in gammaherpesviruses. Accordingly, their roles in virus infection and associated oncogenesis are unknown. Here we report evolutionary conservation of EBV encoded circRNAs by assessing the circRNAome in rLCV infected lymphomas from an SIV infected rhesus macaque and we report latent and lytic circRNAs from KSHV and MHV68. These experiments demonstrate utilization of the circular RNA class of RNAs across 4 members of the gammaherpesvirus subfamily and they identify orthologs and potential homoplastic circRNAs, implying conserved circRNA functions in virus biology and associated malignancies.