These authors contributed equally to this work.
Drug-induced readthrough of premature stop codons leads to the stabilization of laminin α2 chain mRNA in CMD myotubes
Article first published online: 12 DEC 2007
Copyright © 2007 John Wiley & Sons, Ltd.
The Journal of Gene Medicine
Volume 10, Issue 2, pages 217–224, February 2008
How to Cite
Allamand, V., Bidou, L., Arakawa, M., Floquet, C., Shiozuka, M., Paturneau-Jouas, M., Gartioux, C., Butler-Browne, G. S., Mouly, V., Rousset, J.-P., Matsuda, R., Ikeda, D. and Guicheney, P. (2008), Drug-induced readthrough of premature stop codons leads to the stabilization of laminin α2 chain mRNA in CMD myotubes. J. Gene Med., 10: 217–224. doi: 10.1002/jgm.1140
- Issue published online: 28 JAN 2008
- Article first published online: 12 DEC 2007
- Manuscript Accepted: 19 OCT 2007
- Manuscript Revised: 17 SEP 2007
- Manuscript Received: 6 JUL 2007
- Fondation pour la Recherche Médicale
- Association Française contre les Myopathies
- Association pour la Recherche sur le Cancer. Grant Number: 3849
- INSERM and Université Paris 6 Pierre et Marie Curie
- Japan Foundation for Aging and Health
- congenital muscular dystrophy;
- laminin α2 chain;
- premature termination codon;
- antibiotic-mediated readthrough;
- nonsense-mediated mRNA decay
The most common form of congenital muscular dystrophy is caused by a deficiency in the α2 chain of laminin-211, a protein of the extracellular matrix. A wide variety of mutations, including 20 to 30% of nonsense mutations, have been identified in the corresponding gene, LAMA2. A promising approach for the treatment of genetic disorders due to premature termination codons (PTCs) is the use of drugs to force stop codon readthrough.
Here, we analyzed the effects of two compounds on a PTC in the LAMA2 gene that targets the mRNA to nonsense-mediated RNA decay, in vitro using a dual reporter assay, as well as ex vivo in patient-derived myotubes.
We first showed that both gentamicin and negamycin promote significant readthrough of this PTC. We then demonstrated that the mutant mRNAs were strongly stabilized in patient-derived myotubes after administration of negamycin, but not gentamicin. Nevertheless, neither treatment allowed re-expression of the laminin α2-chain protein, pointing to problems that may have arisen at the translational or post-translational levels.
Taken together, our results emphasize that achievement of a clinical benefit upon treatment with novel readthrough-inducing agents would require several favourable conditions including PTC nucleotide context, intrinsic and induced stability of mRNA and correct synthesis of a full-length active protein. Copyright © 2007 John Wiley & Sons, Ltd.