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Mutation-directed studies on the function of the dystrophin ZZ domain and challenges to skipping of duplicated exons in DMD

Research Scholar

Adeline Chaffiol, Children's Hospital- Pediatrics (France)
Kevin Flanigan, Faculty Mentor

Biography

Adeline Chaffiol obtained her Ph.D. in Paris in 2005. The title of her thesis was “therapeutic strategies for Duchenne Muscular Dystrophy: evaluation of exon-skipping and regulation of myostatin in the mouse and canine models of DMD.” During her Ph.D., she met with Dr. Kevin Flanigan of Nationwide Children's Hospital, and he offered her a post-doctoral research position in his lab. After completing post-doctoral research in France, Chaffiol came to Columbus to work with Dr. Flanigan in April 2010.

What is the issue or problem addressed in your research?

Duchenne Muscular Dystrophy (DMD) is the most common and the most lethal of the genetic disorders that affect muscles throughout the body. The disease is due to mutations in the large dystrophin gene, carried on the X-chromosome. The mutation can be inherited from a parent or can be spontaneous and the severity of the disease depends on the type of mutations and how the gene is altered. DMD is typically associated with the loss of dystrophin, which plays an important role in skeletal and cardiac muscle fiber integrity via interactions with others proteins. The milder Becker Muscular Dystrophy is typically due to mutations that result in the presence of some functional dystrophin.

What methodology did you use in your research?

Chaffoil is developing two projects. The first one is the study of a particular domain of the dystrophin, called ZZ because of its structure, in which missense mutations make the dystrophin non-functional despite sometimes a significant amount of protein. This domain has been implicated in forming a stable interaction between dystrophin and several known and candidate binding partners. To investigate new partners, we're using a protein complex immunoprecipitation by targeting different dystrophin constructs (with three of the known missense mutations) with an antibody in order to pull-down the entire protein complex and thereby identify unknown members of the complex, notably by mass spectrometry.

The second project is the skipping of duplicated exons in DMD. Among the different type of mutations, the duplication of a part of the dystrophin gene represents about 10% of the mutations. Skipping of exon duplications would be expected to result in wild-type transcripts, expressing full-length dystrophin, and our preliminary results in the setting of single or double exon duplications suggest this is highly feasible. Then, as no animal model is available for duplications, we are generating an exon2-duplication transgenic mouse.

What are the purpose/rationale and implications of your research?

The first project is more fundamental research: missense mutations in DMD are quite rare, accounting for only 1.4% of all dystrophinopathy mutations. 11 point mutations have been identified in the ZZ domain but their consequence is not well studied and with discrepant results. In some ZZ missense patients, a sizeable amount of dystrophin localizes to the membrane, suggesting the dystrophin-associated glycoprotein complex may be intact, and interactions with other proteins mediate disease.

The second one is a therapeutic approach. The skipping strategy is already in clinical trials by using antisense oligonucleotides but the mouse we are generating will be the first animal model for testing the skipping of a duplicate exon in vivo.