Mapping RNA-RNA Interactions Globally Using Biotinylated Psoralen

Area of Science:

• Molecular Biology
• Genomics
• Biochemistry

Background:

• Understanding RNA-RNA interactions is crucial for deciphering RNA-based gene regulation.
• Existing methods often focus on specific RNA or protein interactions, limiting a comprehensive view.
• The full scope of RNA interactions within cells remains largely unexplored.

Purpose of the Study:

• To present a novel, unbiased, genome-wide method for capturing RNA-RNA interactions in vivo.
• To enable the study of both intramolecular and intermolecular RNA base-pairing globally.
• To facilitate research into the dynamics of RNA organization across various cellular contexts and organisms.

Main Methods:

• Sequencing of Psoralen crosslinked, Ligated, and Selected Hybrids (SPLASH) utilizes in vivo crosslinking to capture interacting RNAs.
• Proximity ligation is employed to join crosslinked RNA fragments.
• High-throughput sequencing is used to identify and map these RNA interactions genome-wide.

Main Results:

• SPLASH allows for the unbiased, genome-wide identification of intramolecular and intermolecular RNA interactions in vivo.
• The method has been successfully applied to bacteria, yeast, and human cells.
• It captures a global view of RNA base-pairing partners.

Conclusions:

• SPLASH is a powerful tool for comprehensively studying RNA-RNA interactions across diverse biological systems.
• This method advances our understanding of RNA-based gene regulation and cellular organization.
• The protocol is efficient, with experimental and computational workflows completed in approximately 12 days.