Blocking of an intronic splicing silencer completely rescues IKBKAP exon 20 splicing in familial dysautonomia patient cells

Nucleic Acids Research
Gitte H BruunBrage S Andresen

Abstract

Familial dysautonomia (FD) is a severe genetic disorder causing sensory and autonomic dysfunction. It is predominantly caused by a c.2204+6T>C mutation in the IKBKAP gene. This mutation decreases the 5' splice site strength of IKBKAP exon 20 leading to exon 20 skipping and decreased amounts of full-length IKAP protein. We identified a binding site for the splicing regulatory protein hnRNP A1 downstream of the IKBKAP exon 20 5'-splice site. We show that hnRNP A1 binds to this splicing regulatory element (SRE) and that two previously described inhibitory SREs inside IKBKAP exon 20 are also bound by hnRNP A1. Knockdown of hnRNP A1 in FD patient fibroblasts increases IKBKAP exon 20 inclusion demonstrating that hnRNP A1 is a negative regulator of IKBKAP exon 20 splicing. Furthermore, by mutating the SREs in an IKBKAP minigene we show that all three SREs cause hnRNP A1-mediated exon repression. We designed splice switching oligonucleotides (SSO) that blocks the intronic hnRNP A1 binding site, and demonstrate that this completely rescues splicing of IKBKAP exon 20 in FD patient fibroblasts and increases the amounts of IKAP protein. We propose that this may be developed into a potential new specific treatment of FD.

References

Feb 17, 2001·American Journal of Human Genetics·S A SlaugenhauptJ F Gusella
Feb 17, 2001·American Journal of Human Genetics·S L AndersonB Y Rubin
Feb 11, 2003·American Journal of Human Genetics·Math P CuajungcoSusan A Slaugenhaupt
Oct 3, 2003·Biochemical and Biophysical Research Communications·Sylvia L AndersonBerish Y Rubin
Jan 8, 2004·Human Molecular Genetics·Susan A SlaugenhauptJames F Gusella
Aug 3, 2004·Journal of Computational Biology : a Journal of Computational Molecular Cell Biology·Gene Yeo, Christopher B Burge
Sep 12, 2006·Human Mutation·El Chérif IbrahimRobin Reed
Jan 9, 2007·Journal of Molecular Medicine : Official Organ of the Gesellschaft Deutscher Naturforscher Und Ärzte·Matthew M HimsSusan A Slaugenhaupt
Jul 24, 2007·Genomics·Matthew M HimsSusan A Slaugenhaupt
Mar 29, 2008·American Journal of Human Genetics·Yimin HuaAdrian R Krainer
Aug 12, 2009·Molecular and Cellular Biology·Hazeem L Okunola, Adrian R Krainer
Nov 26, 2009·European Journal of Human Genetics : EJHG·Eva ProsConxi Lázaro
Jun 21, 2011·Proceedings of the National Academy of Sciences of the United States of America·Kian Huat LimWilliam G Fairbrother
Jul 14, 2011·Genome Research·Timothy Sterne-WeilerJeremy R Sanford
Jul 22, 2011·Pediatric Research·Felicia B AxelrodSusan A Slaugenhaupt
Aug 9, 2011·Human Molecular Genetics·Ranjit S ShettySusan A Slaugenhaupt
Jul 19, 2013·Nucleic Acids Research·Serena BonomiGiuseppe Biamonti
Sep 26, 2013·International Journal of Molecular Sciences·Jacques Jean-PhilippeMassimo Caputi
Nov 1, 2013·Proceedings of the National Academy of Sciences of the United States of America·Lynn GeorgeFrances Lefcort
Aug 13, 2014·Nature Reviews. Genetics·Xiang-Dong Fu, Manuel Ares
Feb 13, 2015·Proceedings of the National Academy of Sciences of the United States of America·Mayumi YoshidaMasatoshi Hagiwara
Nov 11, 2015·Human Mutation·Susana IgrejaMargarida D Amaral
Jun 12, 2016·Nucleic Acids Research·Mallory A Havens, Michelle L Hastings
Jul 7, 2016·BMC Biology·Gitte H BruunBrage S Andresen

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Citations

Jun 20, 2020·Nucleic Acids Research·Alexander Gulliver Bjørnholt GrønningBrage Storstein Andresen
Nov 26, 2020·Nature Reviews. Genetics·Fátima GebauerMatthias W Hentze
May 1, 2021·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Zhichao TangJingxin Wang

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Datasets Mentioned

BETA
GMO2343
GMO4663
GMO4899
GMO4589
GMO2912

Methods Mentioned

BETA
SMA
immunoprecipitation
affinity purifications
Surface Plasmon Resonance
chip
Transfection
electrophoresis
ISS
PCR
iCLIP

Software Mentioned

MaxEntScan
ggplot2
pbm package
Ensembl
SPRINTX
iCLIP
IDT
BLAST
R

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