Nestin : filament formation and expression in myogenesis and muscle disorders
Author: Sjöberg, Gunnar
Date: 1997-10-03
Location: Kemisalen, Doktorsringen 22B, Karolinska Institutet
Time: 9.30
Department: Inst för cell- och molekylärbiologi / Dept of Cell and Molecular Biology
Abstract
Intermediate filaments (IF), together with microfilaments and microtubules, are the major components of the cytoskeleton of eukaryotic cells. In myogenic cells three IF proteins are expressed; desmin, vimentin and nestin. The present investigation analyzes filament assembly and expression of these IFs in myogenesis and muscular disorders, with special focus on the IF protein nestin.
Nestin was shown to colocalize with desmin and vimentin throughout the cytoplasm of human fetal myoblasts and myotubes. This indicated a co-assembly of nestin with the other IFs, and when a full length nestin construct was transfected into an IF-free cell line, nestin was unable to form filaments by itself. However, in occasional cells expressing vimentin, and in cells previously transfected with a desmun construct, nestin is included in the respective IF network. In a yeast two-hybrid system, where protein-protein interactions are analyzed, nestin interacted strongly with vimentin and desmin, but weakly with itself. The inability of nestin to form homopolymers is therefore most likely due to an inability to form homodimers.
Nestin has been shown to have a transient expression pattern in skeletal muscle development. To further characterize nestin expression in developing muscle, we analyzed limb bud and heart at different stages of development. In the developing limb bud of mice, nestin expression was detected early, at low levels, in undifferentiated mesenchymal cells, but as differentiation proceeded, nestin was upregulated in cells undergoing myogenesis and downregulated in cells undergoing chondrogenesis. Nestin expression could be modified by the addition of growth factors affecting myogenic differentiation. A differential localization of nestin mRNA and protein was seen, where the mRNA was localized to the ends of the myofibers, while the protein was more uniformly distributed. In the developing heart, nestin expression was found in the atria and in a subset of cells, with a subendocardiac location, in the ventricles. In the adult heart nestin is confined to a small population of cells, the type of which needs further characterlzation.
Certain pathological disorders are known to be associated with a reexpression of developmentally downregulated proteins. This has shown to be useful in the pathological diagnosis of these disorders. Nestin, having a tissue restricted and developmentally regulated expression pattern, was analysed for expression in regenerative muscle disorders and rhabdomyosarcomas (RMS). In material from patients with muscular dystrophies or myositis, nestin was detected in regenerating muscle fibers. In RMS, nestin was shown to be expressed in all RMS also expressing desmin. In addition, one tumor, presumably a RMS, showed nestin, but not desmin expression, indicating that nestin may be a more sensitive marker for RMS than desmin. Finally, nestin expression in a case of desminopathy was found and compared to expression of desmin and vimentin.
In conclusion, nestin has been shown to be an obligate heterodimer in filament formation, and vimentin or desmin are the dimerizing partners in muscle cells. In the developing embryo nestin expression is restricted to certain tissues in a developmentally regulated program. We have also shown that the reappearance of nestin expression can be of potential use in histopathological diagnosis of certain neuromuscular disorders.
Nestin was shown to colocalize with desmin and vimentin throughout the cytoplasm of human fetal myoblasts and myotubes. This indicated a co-assembly of nestin with the other IFs, and when a full length nestin construct was transfected into an IF-free cell line, nestin was unable to form filaments by itself. However, in occasional cells expressing vimentin, and in cells previously transfected with a desmun construct, nestin is included in the respective IF network. In a yeast two-hybrid system, where protein-protein interactions are analyzed, nestin interacted strongly with vimentin and desmin, but weakly with itself. The inability of nestin to form homopolymers is therefore most likely due to an inability to form homodimers.
Nestin has been shown to have a transient expression pattern in skeletal muscle development. To further characterize nestin expression in developing muscle, we analyzed limb bud and heart at different stages of development. In the developing limb bud of mice, nestin expression was detected early, at low levels, in undifferentiated mesenchymal cells, but as differentiation proceeded, nestin was upregulated in cells undergoing myogenesis and downregulated in cells undergoing chondrogenesis. Nestin expression could be modified by the addition of growth factors affecting myogenic differentiation. A differential localization of nestin mRNA and protein was seen, where the mRNA was localized to the ends of the myofibers, while the protein was more uniformly distributed. In the developing heart, nestin expression was found in the atria and in a subset of cells, with a subendocardiac location, in the ventricles. In the adult heart nestin is confined to a small population of cells, the type of which needs further characterlzation.
Certain pathological disorders are known to be associated with a reexpression of developmentally downregulated proteins. This has shown to be useful in the pathological diagnosis of these disorders. Nestin, having a tissue restricted and developmentally regulated expression pattern, was analysed for expression in regenerative muscle disorders and rhabdomyosarcomas (RMS). In material from patients with muscular dystrophies or myositis, nestin was detected in regenerating muscle fibers. In RMS, nestin was shown to be expressed in all RMS also expressing desmin. In addition, one tumor, presumably a RMS, showed nestin, but not desmin expression, indicating that nestin may be a more sensitive marker for RMS than desmin. Finally, nestin expression in a case of desminopathy was found and compared to expression of desmin and vimentin.
In conclusion, nestin has been shown to be an obligate heterodimer in filament formation, and vimentin or desmin are the dimerizing partners in muscle cells. In the developing embryo nestin expression is restricted to certain tissues in a developmentally regulated program. We have also shown that the reappearance of nestin expression can be of potential use in histopathological diagnosis of certain neuromuscular disorders.
Issue date: 1997-09-12
Publication year: 1997
ISBN: 91-628-2687-5
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