Résumé :
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High-resolution cellular and molecular data from developing organisms has driven many of the advances in developmental biology. These same approaches, applied to muscle biology, should enable a better understanding of the key events of dynamics of muscles formation, maintenance, and degeneration. Applying high-resolution tools to muscular dystrophies cases will define the earliest differences between the development of muscle in normal animals and those with muscular dystrophies, offering both a better understanding and the development of new therapeutic strategies. Our studies strive to attain this goal by combining several approaches: 1) in vivo imaging of the cellular events of zebrafish somitogenesis and myogenesis, 2) a FlipTrap genetic screen to isolate lines with Citrine fusion protein expression in muscles, and 3) a new high-resolution in situ hybridization technique (ISH) known as Hybridization Chain Reaction (HCR) to decipher the co-expression of a variety of mRNA species in muscle and their corresponding proteins as the muscles are forming. Our work has analyzed the dynamics of several key molecules, at both the mRNA and the protein level, during somitogenesis and myogenesis in wildtype, mutant and morphant zebrafish embryos. We have established a dozen new transgenic lines, fusing fluorescent proteins into the normal protein encoding locus, enabling high resolution studies of endogeneous protein dynamics at normal levels of expression during muscle development in live animals. Our multiplex molecular techniques detect nascent dystrophin mRNA transcripts in the nuclei and follow their export and accumulation at the somite borders as the embryo is developing. These molecular and imaging studies are providing unprecedented details on the cellular dynamics and the expression of key players, such as dystrophin, during normal and dystrophic muscle development.
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