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PhD Defense - Marilyne MALBOUYRES

When Nov 12, 2018
from 02:00 PM to 05:00 PM
Where 1 place de l'école
Contact Name
Contact Phone 04 2673 1355
Attendees Dr. Julie Batut, CR CNRS, Université de Toulouse, France. Rapporteure
Dr. Krzystof Jagla, DR INSERM, Université de Clermont Ferrand, France. Rapporteur
Dr. Christel Lefrançois, MCF-HDR, Université de La Rochelle, France. Examinatrice.
Pr. Ulrich Valcourt, PU, Université de Lyon, France. Examinateur.
Dr. Florence Ruggiero, DR CNRS, Université de Lyon, France. Directrice de thèse.

On Monday November 12th 2018, Marilyne MALBOUYRES will defend her thesis 

"Le collagène XXII dans la formation et la fonctionnalité de la jonction myotendineuse chez le poisson zèbre, de l'embryon à l'âge adulte"

Summary

The myotendinous junction (MTJ) consists of a specialized basement membrane that allows the transmission of the contractile muscle forces to the skeleton. My thesis project aimed to characterize the function of collagen XXII (COLXXII), a marker of the MTJ. With this end, we opted for an integrative biology approach using the zebrafish model. Temporal expression pattern of transcript and protein showed that col22a1 is first expressed throughout the somite at 22 hours post-fertilization (hpf) to be then strictly restricted to the muscle fiber ends at the level of the MTJ, where is deposited the protein. Using morpholino(MO)-knockdown, we demonstrated that the resulting col22a1 morphants develop a dystrophic-like phenotype, characterized by muscle fibers detachment from the MTJ. Muscle contractile activity was not affected in the morphants but because the myosepta (tendon) are distended and contain sparse collagen fibrils, the force transmission was greatly reduced. Synergistic interactions suggested that COLXXII interacts with the α7β1 integrin-dependent linkage complex. Because the window of MO effectiveness is limited to the first few days of development, we decided to generate two CRISPR/Cas9 lines designed to invalidate the expression of col22a1. While one-third of the col22a1-/-  larvae from the 2 mutant lines displayed a severe phenotype resulting in their death at around 2 weeks, the rest of larvae showed no apparent phenotype and reached adulthood. Phenotypic analysis of col22a1-/- larvae showed that the phenotypic traits are similar to those described in the morphants but with a lesser degree of severity in the mutants without apparent phenotype. Muscle weakness observed in 2wpf individuals with severe phenotype led to disabling posture defects while individuals with no apparent phenotype did show, at adulthood, diminished swimming performance with increased O2 consumption compared to wildtype. Such expressivity variability of phenotype could be explained by expression of modifier genes. However, we cannot exclude a possible gene compensation as suggested by a candidate gene approach. In conclusion, we showed that COLXXII is an important player of the development and function of the musculoskeletal system. In that, we identified COL22A1 as a potential candidate gene for unresolved cases of human myopathies. This study also revealed a variable expressivity of the col22a1 phenotype, a phenomenon frequently observed in human inherited disorders and brought an additional evidence of the reliability of zebrafish as a model of human diseases.

Key words: zebrafish, collagen XXII, myotendinous junction, morpholinos, CRIPSR-Cas9, muscular dystrophy, swimming performance, behavior.