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Published (Version of Record)CC BY V4.0, Open Access
Journal article
Antisense oligonucleotide induction of progerin in human myogenic cells
PLoS ONE, Vol.9(6)
2014
Abstract
We sought to use splice-switching antisense oligonucleotides to produce a model of accelerated ageing by enhancing expression of progerin, translated from a mis-spliced lamin A gene ( LMNA) transcript in human myogenic cells. The progerin transcript (LMNA Δ150) lacks the last 150 bases of exon 11, and is translated into a truncated protein associated with the severe premature ageing disease, Hutchinson-Gilford progeria syndrome (HGPS). HGPS arises from de novo mutations that activate a cryptic splice site in exon 11 of LMNA and result in progerin accumulation in tissues of mesodermal origin. Progerin has also been proposed to play a role in the 'natural' ageing process in tissues. We sought to test this hypothesis by producing a model of accelerated muscle ageing in human myogenic cells. A panel of splice-switching antisense oligonucleotides were designed to anneal across exon 11 of the LMNA pre-mRNA, and these compounds were transfected into primary human myogenic cells. RT-PCR showed that the majority of oligonucleotides were able to modify LMNA transcript processing. Oligonucleotides that annealed within the 150 base region of exon 11 that is missing in the progerin transcript, as well as those that targeted the normal exon 11 donor site induced the LMNA Δ150 transcript, but most oligonucleotides also generated variable levels of LMNA transcript missing the entire exon 11. Upon evaluation of different oligomer chemistries, the morpholino phosphorodiamidate oligonucleotides were found to be more efficient than the equivalent sequences prepared as oligonucleotides with 2′-O-methyl modified bases on a phosphorothioate backbone. The morpholino oligonucleotides induced nuclear localised progerin, demonstrated by immunostaining, and morphological nuclear changes typical of HGPS cells. We show that it is possible to induce progerin expression in myogenic cells using splice-switching oligonucleotides to redirect splicing of LMNA. This may offer a model to investigate the role of progerin in premature muscle ageing.
Details
- Title
- Antisense oligonucleotide induction of progerin in human myogenic cells
- Authors/Creators
- Y-B Luo (Author/Creator) - Xiangya Hospital Central South UniversityC. Mitrpant (Author/Creator) - Mahidol UniversityA.M. Adams (Author/Creator) - The University of Western AustraliaR.D. Johnsen (Author/Creator) - The University of Western AustraliaS. Fletcher (Author/Creator) - The University of Western AustraliaF.L. Mastaglia (Author/Creator) - Centre for Neuromuscular and Neurological Disorders, Australian Neuro-Muscular Research Institute, University of Western Australia, Perth, Australia; Institute for Immunology & Infectious Diseases, Murdoch University, Perth, Australia.S.D. Wilton (Author/Creator) - The University of Western Australia
- Publication Details
- PLoS ONE, Vol.9(6)
- Publisher
- Public Library of Science
- Identifiers
- 991005546032307891
- Copyright
- © 2014 Luo et al.
- Murdoch Affiliation
- Centre for Comparative Genomics; Institute for Immunology and Infectious Diseases
- Language
- English
- Resource Type
- Journal article
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