Novel Drosophila model of myotonic dystrophy type 1: phenotypic characterization and genome-wide view of altered gene expression.

Myotonic dystrophy type 1 (DM1) is a multisystemic RNA-dominant disorder characterized by myotonia and muscle degeneration. In DM1 patients, the mutant DMPK transcripts containing expanded CUG repeats form nuclear foci and sequester the Muscleblind-like 1 splicing factor, resulting in mis-splicing of its targets. However, several pathological defects observed in DM1 and their link with disease progression remain poorly understood. In an attempt to fill this gap, we generated inducible transgenic Drosophila lines with increasing number of CTG repeats. Targeting the expression of these repeats to the larval muscles recapitulated in a repeat-size-dependent manner the major DM1 symptoms such as muscle hypercontraction, splitting of muscle fibers, reduced fiber size or myoblast fusion defects. Comparative transcriptional profiling performed on the generated DM1 lines and on the muscleblind (mbl)-RNAi line revealed that nuclear accumulation of toxic CUG repeats can affect gene expression independently of splicing or Mbl sequestration. Also, in mblRNAi contexts, the largest portion of deregulated genes corresponded to single-transcript genes, revealing an unexpected impact of the indirect influence of mbl on gene expression. Among the single-transcript Mbl targets is Muscle protein 20 involved in myoblast fusion and causing the reduced number of nuclei in muscles of mblRNAi larvae. Finally, by combining in silico prediction of Mbl targets with mblRNAi microarray data, we found the calcium pump dSERCA as a Mbl splice target and show that the membrane dSERCA isoform is sufficient to rescue a DM1-induced hypercontraction phenotype in a Drosophila model.

Bruno-3 regulates sarcomere component expression and contributes to muscle phenotypes of myotonic dystrophy type 1.

Steinert disease, or myotonic dystrophy type 1 (DM1), is a multisystemic disorder caused by toxic noncoding CUG repeat transcripts, leading to altered levels of two RNA binding factors, MBNL1 and CELF1. The contribution of CELF1 to DM1 phenotypes is controversial. Here, we show that the Drosophila CELF1 family member, Bru-3, contributes to pathogenic muscle defects observed in a Drosophila model of DM1. Bru-3 displays predominantly cytoplasmic expression in muscles and its muscle-specific overexpression causes a range of phenotypes also observed in the fly DM1 model, including affected motility, fiber splitting, reduced myofiber length and altered myoblast fusion. Interestingly, comparative genome-wide transcriptomic analyses revealed that Bru-3 negatively regulates levels of mRNAs encoding a set of sarcomere components, including Actn transcripts. Conversely, it acts as a positive regulator of Actn translation. As CELF1 displays predominantly cytoplasmic expression in differentiating C2C12 myotubes and binds to Actn mRNA, we hypothesize that it might exert analogous functions in vertebrate muscles. Altogether, we propose that cytoplasmic Bru-3 contributes to DM1 pathogenesis in a Drosophila model by regulating sarcomeric transcripts and protein levels.

Muscle development and regeneration in normal and pathological conditions: learning from Drosophila.

The recent demonstration that, throughout evolution, many molecular mechanisms have been highly conserved is fundamental to the advancement of our knowledge on muscle development and regeneration. Research has provided new insights into genetic cascades governing early steps of embryonic myogenesis and the regeneration of adult muscle in normal and pathological conditions, thus revealing significant similarity of both processes. Here we provide a current view on genetic mechanisms underlying muscle regeneration with a special focus on regeneration processes that take place in diseased and aging human muscle. Through examples of Drosophila models of human muscular diseases, we discuss potential impact they might have on uncovering molecular bases and identifying new treatments of muscle disorders. Taking advantage of evolutionarily conserved aspects of muscle development and the relative ease by which molecular pathways can be uncovered and dissected in a simple animal model, the fruit fly, we provide a comprehensive analysis of muscle development in Drosophila. Importantly, identification of muscle stem cell like adult muscle precursors in Drosophila makes fruit fly an attractive model system for studying muscle stem cell biology and muscle regeneration. In support of this assumption, recent studies in our laboratory provide arguments that important insights into the biology of vertebrate muscle stem cells can be gained from genetic analysis in Drosophila.

The impact of acute caloric restriction on the metabolic phenotype in male C57BL/6 and DBA/2 mice.

Caloric restriction (CR) extends healthy lifespan in many organisms. DBA/2 mice, unlike C57BL/6 mice, are reported to be unresponsive to CR. To investigate potential differences underlying the CR response in male DBA/2 and C57BL/6 mice, we examined several metabolic parameters following acute (1-5 weeks) 30% CR. Acute CR decreased body mass (BM) in both strains, with lean and fat mass decreasing in proportion to BM. Resting metabolic rate (RMR) was unaltered by CR, following appropriate corrections for BM differences, although RMR was higher in DBA/2 compared to C57BL/6 mice. Acute CR decreased fed blood glucose levels in both strains, decreased fasting blood glucose in C57BL/6 mice but increased fasting levels in DBA/2 mice. Glucose tolerance improved after 1 week of CR in C57BL/6 mice but improved only after 4 weeks in DBA/2 mice. Acute CR had no effect on insulin levels, but lowered insulin sensitivity and decreased insulin-like growth factor-1 (IGF-1) levels in both strains. DBA/2 mice were hyperinsulinaemic and insulin resistant compared to C57BL/6 mice. These strain-specific differences in glucose homeostatic parameters may underlie the reported unresponsiveness of DBA/2 mice to CR. We also demonstrate delineation in the response of insulin and IGF-1 to acute CR in mice.