Millions of years of mammalian ribozyme activity mapped with a phylogenetic fitness landscape. Bendixsen_etal_CPEB3Phylo

The CPEB3 ribozyme is a catalytic RNA element that is highly conserved in mammals. The high conservation of the ribozyme sequence as well as the genomic location within the CPEB3 gene indicate that this ribozyme may serve important functions in mammals. However, little is known about the origins and evolution of this ribozyme during mammalian evolution. Here we use a combination of phylogenetic approaches and high-throughput sequencing-based ribozyme assays to resurrect the evolutionary pathways of this ribozyme over 100 million of years of mammalian evolution. We find that the predicted sequence at the root of placental mammals and marsupials is highly active, and this activity has been conserved in many lineages. The reduction or loss of ribozyme activity appears sporadically throughout the mammalian tree of life, which may indicate lineage specific events. Our data also enables the evaluation of an RNA fitness landscape that includes all the parsimonious pathways between ancient and extant sequences as well as the numerous mutational pathways that were avoided. Analysis of this landscape further supports that the sequence changes in the majority of these ribozymes are non-random indicating purifying selection to preserve this ribozyme function. The instances of reduced ribozyme activity may indicate cellular specific interactions that are required for ribozyme function or reduce the importance of the ribozyme activity.