Research article

Ligand-induced sequestering of branchpoint sequence allows conditional control of splicing

Dong-Suk Kim^1,2 , Veronica Gusti¹ , Kenneth J Dery^1,3 and Rajesh K Gaur^1,4

¹Division of Molecular Biology, Beckman Research Institute of the City of Hope, Duarte, CA 91010, USA

²MCDB Program, Iowa State University, Ames, IA 50011, USA

³Division of Immunology, Beckman Research Institute of the City of Hope, Duarte, CA 91010, USA

⁴Graduate School of Biological Sciences, Beckman Research Institute of the City of Hope, Duarte, CA 91010, USA

author email corresponding author email

BMC Molecular Biology 2008, 9:23doi:10.1186/1471-2199-9-23

Published:	12 February 2008

Abstract

Background

Despite tremendous progress in understanding the mechanisms of constitutive and alternative splicing, an important and widespread step along the gene expression pathway, our ability to deliberately regulate gene expression at this step remains rudimentary. The present study was performed to investigate whether a theophylline-dependent "splice switch" that sequesters the branchpoint sequence (BPS) within RNA-theophylline complex can regulate alternative splicing.

Results

We constructed a series of pre-mRNAs in which the BPS was inserted within theophylline aptamer. We show that theophylline-induced sequestering of BPS inhibits pre-mRNA splicing both in vitro and in vivo in a dose-dependent manner. Several lines of evidence suggest that theophylline-dependent inhibition of splicing is highly specific, and thermodynamic stability of RNA-theophylline complex as well as the location of BPS within this complex affects the efficiency of splicing inhibition. Finally, we have constructed an alternative splicing model pre-mRNA substrate in which theophylline caused exon skipping both in vitro and in vivo, suggesting that a small molecule-RNA interaction can modulate alternative splicing.

Conclusion

These findings provide the ability to control splicing pattern at will and should have important implications for basic, biotechnological, and biomedical research.