snoCLASH Reveals Extensive snoRNA-mRNA Interaction Networks

Area of Science:

• Molecular Biology
• RNA Biology
• Genomics

Background:

• Small nucleolar RNAs (snoRNAs) primarily guide ribosomal RNA (rRNA) modifications.
• Emerging evidence indicates snoRNAs interact with non-rRNA targets, but discovery is challenging due to rRNA abundance and technical hurdles.
• Systematic identification of these non-canonical interactions is crucial for understanding broader RNA regulation.

Purpose of the Study:

• To develop and validate a scalable method for identifying transcriptome-wide snoRNA-non-rRNA interactions.
• To discover novel mRNA targets of snoRNAs and their associated regulatory functions.
• To establish a platform for exploring non-canonical snoRNA functions beyond rRNA modification.

Main Methods:

• Development of snoCLASH, an optimized RNA binding protein (RBP)-based crosslinking, ligation, and sequencing framework.
• Integration of techniques including phenol-toluol extraction, polyA enrichment, nuclear fractionation, rRNA depletion, and dual-reference chimeric read analysis.
• Application of snoCLASH for transcriptome-scale identification of snoRNA-mRNA interactions.

Main Results:

• Identification of thousands of snoRNA-associated mRNA regions, including coding and regulatory elements.
• Enrichment of identified RBPs linked to epitranscriptomic regulation among snoRNA-associated regions.
• Functional validation of a snoRNA-mRNA interaction demonstrating snoRNA-dependent 2'-O-methylation impacting protein expression.

Conclusions:

• snoCLASH is a scalable platform for discovering and validating non-canonical snoRNA targets across the transcriptome.
• This method expands the known functions of snoRNAs beyond rRNA modification.
• The findings highlight a significant role for snoRNAs in regulating gene expression through mRNA interactions and epitranscriptomic modifications.