A novel role for nucleolin in splice site selection

Area of Science:

• Molecular Biology
• RNA Splicing
• Cellular Quality Control

Background:

• Latent 5' splice sites in human introns are activated during stress and cancer, producing aberrant nonsense mRNAs.
• A previously identified splicing suppression mechanism involves initiator-tRNA, independent of protein translation and NMD.
• The precise molecular players and interactions in this nuclear suppression pathway remain incompletely understood.

Purpose of the Study:

• To identify nuclear proteins that directly bind to initiator-tRNA.
• To elucidate the role of initiator-tRNA binding proteins in regulating latent splice site selection.
• To characterize a novel nuclear quality control mechanism for splice site selection.

Main Methods:

• UV-crosslinking to identify nuclear proteins interacting with initiator-tRNA.
• Co-immunoprecipitation and Western blotting to confirm protein-RNA interactions.
• Nucleolin knockdown experiments to assess functional impact on latent splicing.

Main Results:

• Nucleolin (NCL) was identified as a nuclear protein that directly and specifically binds initiator-tRNA.
• Initiator-tRNA and NCL were found to associate with pre-mRNA.
• Initiator-tRNA-mediated suppression of latent splicing requires NCL, and NCL depletion activates latent splicing in numerous transcripts.

Conclusions:

• Nucleolin acts as a nuclear quality control factor, binding initiator-tRNA to regulate splice site selection.
• This NCL-initiator-tRNA interaction suppresses latent splicing, preventing the generation of defective mRNAs.
• The findings reveal a novel mechanism protecting cells from the detrimental effects of aberrant splicing.