Elucidating the Role of snRNA Modifications in Pre-mRNA Splicing Efficiency

The intricate process of RNA splicing is crucial for the generation of mature messenger RNA (mRNA), yet the influence of small nuclear RNA (snRNA) modifications on splicing efficiency remains poorly understood. This study aims to explore the impact of specific chemical modifications of snRNA on the splicing mechanism. Using a combination of CRISPR-Cas9 mediated gene editing and high-throughput sequencing, we introduced targeted modifications to U1 and U2 snRNAs in HeLa cells. Our results show that 2'-O-methylation at position 126 of U1 snRNA leads to a 15% increase in splicing efficiency, as evidenced by RT-qPCR analysis (p<0.05). Additionally, pseudouridylation at position 91 of U2 snRNA was found to enhance the interaction with pre-mRNA substrates, increasing catalytic activity by 12% (p<0.01). These modifications were further validated using RNA immunoprecipitation assays, confirming their role in stabilizing spliceosomal complexes. Our findings suggest that snRNA modifications are not merely auxiliary but are pivotal in optimizing spliceosomal function. These insights contribute to our understanding of post-transcriptional regulation and may have implications for therapeutic strategies targeting splicing abnormalities.