Prader–Willi locus Snord116 RNA processing requires an active endogenous allele and neuron-specific splicing by Rbfox3/NeuN

摘要

Prader–Willi syndrome (PWS), an imprinted neurodevelopmental disorder characterized by metabolic, sleep and neuropsychiatric features, is caused by the loss of paternal containing only non-coding RNAs (ncRNAs). The primary transcript is processed into small nucleolar RNAs (snoRNAs), which localize to nucleoli, and their spliced host gene , which is retained at its site of transcription. While functional complementation of the ncRNAs is a desirable goal for treating PWS, the mechanistic requirements of RNA processing are poorly understood. Here we developed and tested a novel transgenic mouse which ubiquitously expresses on both a wild-type and a paternal deletion ( ) background. Interestingly, while the transgene was ubiquitously expressed in multiple tissues, splicing of the transgene and production of snoRNAs was limited to brain tissues. Knockdown of encoding neuron-specific splicing factor neuronal nuclei (NeuN) in derived neurons, reduced splicing of the transgene in neurons. RNA fluorescence hybridization for revealed a single significantly larger signal in transgenic mice, demonstrating colocalization of transgenic and endogenous RNAs. Similarly, significantly increased snoRNA levels were detected in transgenic neuronal nucleoli, indicating that transgenic snoRNAs were effectively processed and localized. In contrast, neither transgenic nor snoRNAs were detectable in either non-neuronal tissues or neurons. Together, these results demonstrate that exogenous expression and neuron-specific splicing of the locus are insufficient to rescue the genetic deficiency of paternal deletion. Elucidating the mechanisms regulating processing and localization is essential to develop effective gene replacement therapies for PWS.