Human Molecular Genetics Advance Access originally published online on August 27, 2004
Human Molecular Genetics 2004 13(20):2409-2420; doi:10.1093/hmg/ddh272
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Human Molecular Genetics, Vol. 13, No. 20 © Oxford University Press 2004; all rights reserved
Correction of aberrant FGFR1 alternative RNA splicing through targeting of intronic regulatory elements
1Department of Endocrine Neoplasia and Hormonal Disorders, The University of Texas M. D. Anderson Cancer Center, Houston, TX 77030, USA and 2Program of Human and Molecular Genetics, The University of Texas Graduate School of Biomedical Sciences at Houston, Houston, TX 77030, USA
Received June 26, 2004; Accepted August 18, 2004
Alternative RNA splicing is now known to be pervasive throughout the genome and a target of human disease. We evaluated if targeting intronic splicing regulatory sequences with antisense oligonucleotides could be used to correct aberrant exon skipping. As a model, we targeted the intronic silencing sequence (ISS) elements flanking the alternatively spliced 
-exon of the endogenous fibroblast growth factor receptor 1 (FGFR1) gene, which is aberrantly skipped in human glioblastoma. Antisense morpholino oligonucleotides targeting either upstream or downstream ISS elements increased -exon inclusion from 10% up to 70% in vivo. The effect was dose dependent, sequence specific and reproducible in several human cell lines, but did not necessarily correlate with blocking of protein association in vitro. Simultaneous targeting of the ISS elements had no additive effect, suggesting that splicing regulation occurred through a shared mechanism. Broad applicability of this approach was demonstrated by similar targeting of the ISS elements of the human hnRNPA1 gene. The correction of FGFR1 gene splicing to >90% -exon inclusion in glioblastoma cells had no discernable effect on cell growth in culture, but was associated with an increase in unstimulated caspase-3 and -7 activity. The ability to manipulate endogenously expressed mRNA variants allows exploration of their functional relevance under normal and diseased physiological states.
* To whom correspondence should be addressed at: Department of Endocrine Neoplasia and Hormonal Disorders, Unit 435, The University of Texas M. D. Anderson Career Center, 1515 Holcombe Boulevard, Houston, TX 77030, USA. Tel: +1 7137422841; Fax: +1 7137945252; Email: gcote{at}mdanderson.org
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