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Related Experiment Video

Updated: May 31, 2026

A Reporter Based Cellular Assay for Monitoring Splicing Efficiency

A Reporter Based Cellular Assay for Monitoring Splicing Efficiency

Published on: September 15, 2021

Functionally important structural elements of U12 snRNA.

Kavleen Sikand¹, Girish C Shukla

¹Center for Gene Regulation in Health and Disease, Cleveland State University, 2121 Euclid Avenue, Cleveland, OH 44115, USA.

Nucleic Acids Research

|July 9, 2011

Summary

U12 small nuclear RNA (snRNA) structures outside the branch site are crucial for U12-dependent splicing. Stem-loop IIa and III are essential, while stem-loop IIb is not required for this vital cellular process.

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Area of Science:

Molecular Biology
RNA Biology
Genetics

Background:

U12 snRNA is vital for U12-dependent introns in metazoans, analogous to U2 snRNA.
The branch site interaction region of U12 snRNA is known, but other structural elements' roles remain unclear.
U12 snRNA possesses a complex predicted secondary structure with stem-loops and single-stranded regions.

Purpose of the Study:

To investigate the in vivo requirement of U12 snRNA structural elements beyond the branch site.
To determine the necessity of specific stem-loops in U12-dependent splicing.
To validate predicted secondary structures of U12 snRNA through functional analysis.

Main Methods:

Utilized a branch site genetic suppression assay.
Introduced second-site mutations into suppressor U12 snRNA.
Assessed the impact of mutations on in vivo U12-dependent splicing.

Main Results:

Stem-loop IIa of U12 snRNA was found to be essential for in vivo splicing.
Stem-loop IIb, despite evolutionary conservation, was dispensable for splicing.
Stem-loop III, involved in binding to the p65 protein of the U11-U12 di.snRNP complex, is essential for splicing.

Conclusions:

The study experimentally validates the existence of proposed stem-loops in U12 snRNA.
Provides direct evidence for the functional importance of specific secondary structures in U12-dependent splicing.
Highlights differential requirements for structural elements within U12 snRNA during splicing.