[S18]: RNA regulation and modular control of the neuronal synapse

The brain has unique systems for regulating RNA metabolism, and these are often targeted in human neurologic disease. One such protein, Nova, is associated with POMA, a paraneoplastic disorder. To better understand the role of Nova and other RNA binding proteins, we have tried to identify the array of RNAs that these proteins regulate, using RNA library selection, biochemical purification, CLIP (cross-linking immunoprecipitation), and development of a splicing microarray. We have found that Nova regulates neuron-specific splicing of 6–7% of alternative exons in the brain, with splicing changes ranging from 6to >100-fold in KO mice. Nova RNA targets appear to encode a biologically coherent set of proteins. Over 50 Nova splicing target RNAs validated in the brains of KO animals encode proteins that function at the neuronal synapse. Experiments to examine the physiologic consequence of these RNA-protein interactions are beginning to confirm the relevance of Nova to neuronal synaptic function. We have also been defining a map of the sequence and spatial distribution of Nova binding sites that confer splicing regulation. This RNA map successfully predicts new Nova targets from a genomewide screen and moreover, predicts with 100% accuracy whether Nova mediates up or down-regulation of alternative exon inclusion. These results suggest that combining bioinformatic approaches with genetics and biochemistry is able to identify likely mechanisms underlying how a tissue-specific factor regulates alternative splicing.