Mouse Nuclear RNAi-defective 2 Promotes Splicing of Weak 5' Splice Sites

Removal of introns during pre-mRNA splicing, which is central to gene expression, initiates by base pairing of U1 snRNA with a 5 splice site (5SS). In mammals, many introns contain weak 5SSs that are not efficiently recognized by the canonical U1 snRNP, suggesting alternative mechanisms exist. Here, we develop a cross-linking immunoprecipitation coupled to a high-throughput sequencing method, BCLIP-seq, to identify NRDE2 (Nuclear RNAi defective-2) and CCDC174 (Coiled-Coil Domain-Containing 174) as novel RNA-binding proteins in mouse ES cells that associate with U1 snRNA and unspliced 5SSs. Both proteins bind directly to U1 snRNA independently of canonical U1 snRNP specific proteins, and they are required for the selection and effective processing of weak 5SSs. Our results reveal that mammalian cells use non-canonical splicing factors bound directly to U1 snRNA to effectively select suboptimal 5SS sequences in hundreds of genes, promoting proper splice site choice and accurate pre-mRNA splicing.