Evolution of Nova-Dependent Splicing Regulation in the Brain

Abstract

A large number of alternative exons are spliced with tissue-specific patterns, but little is known about how such patterns have evolved. Here, we study the conservation of the neuron-specific splicing factors Nova1 and Nova2 and of the alternatively spliced exons they regulate in mouse brain. Whereas Nova RNA binding domains are 94% identical across vertebrate species, Nova-dependent splicing silencer and enhancer elements (YCAY clusters) show much greater divergence, as less than 50% of mouse YCAY clusters are conserved at orthologous positions in the zebrafish genome. To study the relation between the evolution of tissue-specific splicing and YCAY clusters, we compared the brain-specific splicing of Nova-regulated exons in zebrafish, chicken, and mouse. The presence of YCAY clusters in lower vertebrates invariably predicted conservation of brain-specific splicing across species, whereas their absence in lower vertebrates correlated with a loss of alternative splicing. We hypothesize that evolution of Nova-regulated splicing in higher vertebrates proceeds mainly through changes in cis-acting elements, that tissue-specific splicing might in some cases evolve in a single step corresponding to evolution of a YCAY cluster, and that the conservation level of YCAY clusters relates to the functions encoded by the regulated RNAs. Alternative splicing generates different mRNA isoforms from a single gene and thus increases the number of proteins a cell can produce. This is particularly important in the brain, which possesses a number of brain-specific splicing factors. In this study, we have looked at evolution of brain-specific splicing regulation by one such factor, Nova. Previous studies have identified ∼100 alternative exons that are regulated by Nova in mouse brain. We find that the Nova protein sequence changed little during vertebrate evolution from fish to human, whereas the RNA targets themselves have evolved significantly. Interestingly, the presence of conserved Nova binding elements in an RNA transcript in most cases correlates with conservation of brain-specific splicing. In addition, the evolution of Nova-dependent splicing relates to the functions encoded by the target RNAs, such that Nova-regulated splicing of RNAs encoding core roles such as synaptic adhesion, ion channel, and cytoskeletal proteins is on average more conserved than splicing of the RNAs encoding regulatory roles, such as transmembrane receptor and signal transduction proteins.