High-throughput sequencing reveals circular substrates for an archaeal RNA ligase

Abstract

It is only recently that the abundant presence of circular RNAs (circRNAs) in all kingdoms of Life, including the hyperthermophilic archaeon Pyrococcus abyssi, has emerged. This led us to investigate the physiologic significance of a previously observed weak intramolecular ligation activity of Pab1020 RNA ligase. Here we demonstrate that this enzyme, despite sharing significant sequence similarity with DNA ligases, is indeed an RNA-specific polynucleotide ligase efficiently acting on physiologically significant substrates. Using a combination of RNA immunoprecipitation assays and RNA-seq, our genome-wide studies revealed 133 individual circRNA loci in P. abyssi. The large majority of these loci interacted with Pab1020 in cells and circularization of selected C/D Box and 5S rRNA transcripts was confirmed biochemically. Altogether these studies revealed that Pab1020 is required for RNA circularization. Our results further suggest the functional speciation of an ancestral NTase domain and/or DNA ligase toward RNA ligase activity and prompt for further characterization of the widespread functions of circular RNAs in prokaryotes. Detailed insight into the cellular substrates of Pab1020 may facilitate the development of new biotechnological applications e.g. in ligation of preadenylated adaptors to RNA molecules.

RNA stabilization in hyperthermophilic archaea

“…70,71 An additional way to protect RNA termini against exonucleolytic activity is the ligation of available 5ʹ and 3ʹ termini, resulting in the formation of circular RNA species. 13,72,73 The described circular T. tenax SRP RNA exemplifies this case (Fig. 2B).…”

“…The translation initiation factor a/eIF2 can bind mRNAs with 5ʹppp ends (but not RNAs with 5ʹp or 5ʹpp ends) to provide protection from Sso‐aCPSF2 degradation, and in vivo experiments showed that the overproduction of a/eIF2(γ) increases the stability of different mRNAs . An additional way to protect RNA termini against exonucleolytic activity is the ligation of available 5ʹ and 3ʹ termini, resulting in the formation of circular RNA species . The described circular T. tenax SRP RNA exemplifies this case (Fig.…”

The 23S Ribosomal RNA From Pyrococcus furiosus Is Circularly Permuted

“…Circularization of RNA is widespread in archaea ( Danan et al, 2012 ) and P. furiosus appears to be particularly prone to RNA circularization, e.g., with abundant circularization of box C/D RNAs ( Starostina et al, 2004 ). Recently, an RNA ligase that ligates 5´-P and 3´-OH ends was characterized from Pyrococcus abyssi (the Pab 1,020 RNA ligase) and proposed to circularize many different cellular RNAs ( Becker et al, 2017 ). The excision of H98 in P. furiosus curiously resembles a step in rRNA processing in chloroplasts.…”

Circular RNA expression profiles reveal that hsa_circ_0018289 is up-regulated in cervical cancer and promotes the tumorigenesis

“…Presently, the role and mechanism of circRNAs in tumor progression are still not well identified [ 15 ]. Although circRNAs have been discovered for decades, the significance of circRNAs is recognized with the developing of high-throughput RNA-sequencing (RNA-Seq) and bioinformatics analysis [ 16 , 17 ]. Emerging evidences have illustrated that circRNAs play important role in multiple physiological and pathological functions, involving cardiovascular system disease, nervous system disease and series of tumors [ 18 – 20 ].…”