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Articles by Mariya M. Kucherenko in JoVE
JoVE General
Парафин и замороженные Разделы Drosophila Мышцы взрослых
Gene Expression and Signaling Research Group, Max Planck Institute for Biophysical Chemistry
Определение механизмов, лежащих повреждение мышц имеет решающее значение. Здесь мы представляем гистологической техники для подготовки парафин и замороженных участков Drosophila грудной мышцы. Это позволяет анализировать морфологию мышц и локализация белка и других компонентов клетки мышц.
Other articles by Mariya M. Kucherenko on PubMed
A Putative Src Homology 3 Domain Binding Motif but Not the C-terminal Dystrophin WW Domain Binding Motif is Required for Dystroglycan Function in Cellular Polarity in Drosophila
The conserved dystroglycan-dystrophin (Dg.Dys) complex connects the extracellular matrix to the cytoskeleton. In humans as well as Drosophila, perturbation of this complex results in muscular dystrophies and brain malformations and in some cases cellular polarity defects. However, the regulation of the Dg.Dys complex is poorly understood in any cell type. We now find that in loss-of-function and overexpression studies more than half (34 residues) of the Dg proline-rich conserved C-terminal regions can be truncated without significantly compromising its function in regulating cellular polarity in Drosophila. Notably, the truncation eliminates the WW domain binding motif at the very C terminus of the protein thought to mediate interactions with dystrophin, suggesting that a second, internal WW binding motif can also mediate this interaction. We confirm this hypothesis by using a sensitive fluorescence polarization assay to show that both WW domain binding sites of Dg bind to Dys in humans (K(d) = 7.6 and 81 microM, respectively) and Drosophila (K(d) = 16 and 46 microM, respectively). In contrast to the large deletion mentioned above, a single proline to an alanine point mutation within a predicted Src homology 3 domain (SH3) binding site abolishes Dg function in cellular polarity. This suggests that an SH3-containing protein, which has yet to be identified, functionally interacts with Dg.
Genetic Modifier Screens Reveal New Components That Interact with the Drosophila Dystroglycan-dystrophin Complex
The Dystroglycan-Dystrophin (Dg-Dys) complex has a capacity to transmit information from the extracellular matrix to the cytoskeleton inside the cell. It is proposed that this interaction is under tight regulation; however the signaling/regulatory components of Dg-Dys complex remain elusive. Understanding the regulation of the complex is critical since defects in this complex cause muscular dystrophy in humans. To reveal new regulators of the Dg-Dys complex, we used a model organism Drosophila melanogaster and performed genetic interaction screens to identify modifiers of Dg and Dys mutants in Drosophila wing veins. These mutant screens revealed that the Dg-Dys complex interacts with genes involved in muscle function and components of Notch, TGF-beta and EGFR signaling pathways. In addition, components of pathways that are required for cellular and/or axonal migration through cytoskeletal regulation, such as Semaphorin-Plexin, Frazzled-Netrin and Slit-Robo pathways show interactions with Dys and/or Dg. These data suggest that the Dg-Dys complex and the other pathways regulating extracellular information transfer to the cytoskeletal dynamics are more intercalated than previously thought.
Stress and Muscular Dystrophy: a Genetic Screen for Dystroglycan and Dystrophin Interactors in Drosophila Identifies Cellular Stress Response Components
In Drosophila, like in humans, Dystrophin Glycoprotein Complex (DGC) deficiencies cause a life span shortening disease, associated with muscle dysfunction. We performed the first in vivo genetic interaction screen in ageing dystrophic muscles and identified genes that have not been shown before to have a role in the development of muscular dystrophy and interact with dystrophin and/or dystroglycan. Mutations in many of the found interacting genes cause age-dependent morphological and heat-induced physiological defects in muscles, suggesting their importance in the tissue. Majority of them is phylogenetically conserved and implicated in human disorders, mainly tumors and myopathies. Functionally they can be divided into three main categories: proteins involved in communication between muscle and neuron, and interestingly, in mechanical and cellular stress response pathways. Our data show that stress induces muscle degeneration and accelerates age-dependent muscular dystrophy. Dystrophic muscles are already compromised; and as a consequence they are less adaptive and more sensitive to energetic stress and to changes in the ambient temperature. However, only dystroglycan, but not dystrophin deficiency causes extreme myodegeneration induced by energetic stress suggesting that dystroglycan might be a component of the low-energy pathway and act as a transducer of energetic stress in normal and dystrophic muscles.
New Dystrophin/Dystroglycan Interactors Control Neuron Behavior in Drosophila Eye
The Dystrophin Glycoprotein Complex (DGC) is a large multi-component complex that is well known for its function in muscle tissue. When the main components of the DGC, Dystrophin (Dys) and Dystroglycan (Dg) are affected cognitive impairment and mental retardation in addition to muscle degeneration can occur. Previously we performed an array of genetic screens using a Drosophila model for muscular dystrophy in order to find novel DGC interactors aiming to elucidate the signaling role(s) in which the complex is involved. Since the function of the DGC in the brain and nervous system has not been fully defined, we have here continued to analyze the DGC modifiers' function in the developing Drosophila brain and eye.
