Cell-type-resolved RNP topologies reveal dynamic structural mechanisms of splicing and therapeutic targets

Area of Science:

• Molecular Biology
• Structural Biology
• Genomics

Background:

• Resolving RNA conformations within native ribonucleoprotein (RNP) complexes is a significant challenge in molecular biology.
• Understanding RNA structure is crucial for deciphering gene regulation and disease mechanisms.

Purpose of the Study:

• To introduce a novel method, spatial hydroxyl acylation reversible crosslinking with immunoprecipitation (SHARCLIP), for capturing RNA interactions in cells.
• To create a comprehensive structural atlas of RNA conformations and their role in gene regulation.

Main Methods:

• SHARCLIP was employed to simultaneously capture RNA-RNA, RNA-protein, and protein-protein contacts within intact cells.
• Profiling of heterogeneous nuclear ribonucleoprotein C (HNRNPC)-associated RNA conformations was performed.

Main Results:

• A global, phased map of ribonucleosomes was established, resolving long-standing questions about hnRNP assembly.
• A dynamic structural atlas of over 10,000 RNAs across seven cell lineages was generated, revealing millions of dynamic loops, steric blockers, and conformational switches.
• The structural basis of mutually exclusive exons (MXEs) was deciphered, enabling the rational design of antisense oligonucleotides (ASOs).

Conclusions:

• SHARCLIP provides an unprecedented view of cellular RNA structural biology.
• The study demonstrates the potential of structure-guided ASO design for therapeutic applications, including isoform swapping in genes linked to genetic disorders.