Modulation of 5' splice site selection using tailed oligonucleotides carrying splicing signals.
BACKGROUND
We previously described the use of tailed oligonucleotides as a means of reprogramming alternative pre-mRNA splicing in vitro and in vivo. The tailed oligonucleotides that were used interfere with splicing because they contain a portion complementary to sequences immediately upstream of the target 5' splice site combined with a non-hybridizing 5' tail carrying binding sites for the hnRNP A1/A2 proteins. In the present study, we have tested the inhibitory activity of RNA oligonucleotides carrying different tail structures.


RESULTS
We show that an oligonucleotide with a 5' tail containing the human beta-globin branch site sequence inhibits the use of the 5' splice site of Bcl-xL, albeit less efficiently than a tail containing binding sites for the hnRNP A1/A2 proteins. A branch site-containing tail positioned at the 3' end of the oligonucleotide also elicited splicing inhibition but not as efficiently as a 5' tail. The interfering activity of a 3' tail was improved by adding a 5' splice site sequence next to the branch site sequence. A 3' tail carrying a Y-shaped branch structure promoted similar splicing interference. The inclusion of branch site or 5' splice site sequences in the Y-shaped 3' tail further improved splicing inhibition.


CONCLUSIONS
Our in vitro results indicate that a variety of tail architectures can be used to elicit splicing interference at low nanomolar concentrations, thereby broadening the scope and the potential impact of this antisense technology.
Gendron
Daniel
D
Département de Microbiologie et d'Infectiologie, Faculté de Médecine et des Sciences de la Santé, Université de Sherbrooke, Sherbrooke, Québec, Canada. gendron_daniel@yahoo.fr
Carriero
Sandra
S
Garneau
Daniel
D
Villemaire
Jonathan
J
Klinck
Roscoe
R
Elela
Sherif Abou
SA
Damha
Masad J
MJ
Chabot
Benoit
B
D017398:Alternative Splicing / Q000235:genetics*; D005786:Gene Expression Regulation / Q000235:genetics; D020868:Gene Silencing; D005818:Genetic Engineering / Q000379:methods*; D014644:Genetic Variation / Q000235:genetics; D016376:Oligonucleotides, Antisense / Q000235:genetics*; D022821:RNA Splice Sites / Q000235:genetics*
D016376:Oligonucleotides, Antisense; D022821:RNA Splice Sites