Focal Adhesion Kinase Signaling Regulates the Expression of Caveolin 3 and ␤1 Integrin, Genes Essential for Normal Myoblast Fusion

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Focal Adhesion Kinase Signaling Regulates the Expression of Caveolin 3 and ␤1 Integrin, Genes Essential for Normal Myoblast Fusion

Authors
  • Navaline Quach1
  • Stefano Biressi
  • Louis F. Reichardt
  • Charles Keller
  • Thomas A. Rando
Type
Published Article
Journal
Molecular Biology of the Cell
Publisher
American Society for Cell Biology (ASCB)
Publication Date
Jul 11, 2009
Volume
20
Issue
14
Pages
3422–3435
Identifiers
DOI: 10.1091/mbc.e09-02-0175
PMID: 19458188
PMCID: PMC2710835
Source
AIMS
License
Green

Abstract

An essential phase of skeletal myogenesis is the fusion of mononucleated myoblasts to form multinucleated myotubes. Many cell adhesion proteins, including integrins, have been shown to be important for myoblast fusion in vertebrates, but the mechanisms by which these proteins regulate cell fusion remain mostly unknown. Here, we focused on the role of focal adhesion kinase (FAK), an important nonreceptor protein tyrosine kinase involved in integrin signaling, as a potential mediator by which integrins may regulate myoblast fusion. To test this hypothesis in vivo, we generated mice in which the Fak gene was disrupted specifically in muscle stem cells (" satellite cells ") and we found that this resulted in impaired myotube formation during muscle regeneration after injury. To examine the role of FAK in the fusion of myogenic cells, we examined the expression of FAK and the effects of FAK deletion on the differentiation of myoblasts in vitro. Differentiation of mouse primary myoblasts was accompanied by a rapid and transient increase of phosphory-lated FAK. To investigate the requirement of FAK in myoblast fusion, we used two loss-of-function approaches (a dominant-negative inhibitor of FAK and FAK small interfering RNA [siRNA]). Inhibition of FAK resulted in markedly impaired fusion but did not inhibit other biochemical measures of myogenic differentiation, suggesting a specific role of FAK in the morphological changes of cell fusion as part of the differentiation program. To examine the mechanisms by which FAK may be regulating fusion, we used microarray analysis to identify the genes that failed to be normally regulated in cells that were fusion defective due to FAK inhibition. Several genes that have been implicated in myoblast fusion were aberrantly regulated during differentiation when FAK was inhibited. Intriguingly, the normal increases in the transcript of caveolin 3 as well as an integrin subunit, the ␤1D isoform, were suppressed by FAK inhibition. We confirmed this also at the protein level and show that direct inhibition of ␤1D subunit expression by siRNA inhibited myotube formation with a prominent effect on secondary fusion. These data suggest that FAK regulation of profusion genes, including caveolin 3 and the ␤1D integrin subunit, is essential for morphological muscle differentiation.

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