Extracellular Vesicle-Mediated U1 snRNA Delivery Restores Aberrant Pre-mRNA Splicing in Human Cells

Area of Science:

• Molecular Biology
• Genetics
• Biotechnology

Background:

• Splicing defects are a major cause of genetic disorders, but effective correction strategies are limited.
• The U1 small nuclear RNA (snRNA) is crucial for accurate pre-messenger RNA splicing, and its disruption leads to disease.
• Extracellular vesicles (EVs) offer a potential delivery system for therapeutic molecules into cells.

Purpose of the Study:

• To investigate the potential of engineered extracellular vesicles (EVs) to deliver U1 snRNA for correcting genetic splicing defects.
• To establish a novel RNA-based therapeutic approach for genetic diseases stemming from aberrant splice-site recognition.

Main Methods:

• U1 snRNA-overexpressing HEK293T cells were used to generate U1 snRNA-enriched EVs.
• EVs were characterized for purity and U1 snRNA content using Western blots and RT-qPCR.
• HeLa cells with a β-thalassemia-like splice-site mutation were treated with U1-snRNA-enriched EVs to assess splicing correction.

Main Results:

• U1 snRNA-enriched EVs were successfully generated and characterized.
• Treatment with these EVs corrected up to 60% of normal exon-intron junction recognition in a dose-dependent manner.
• Correction efficacy was dependent on intact vesicular RNA, as heat or RNase treatment abolished the effect.

Conclusions:

• Extracellular vesicles can effectively transport spliceosomal snRNAs to recipient cells.
• This study demonstrates a novel therapeutic strategy using EVs to deliver U1 snRNA for correcting splicing defects.
• Engineered EVs represent a promising new class of RNA-based therapeutics for genetic disorders.