Postel R, Vakeel P, Topczewski J, Knoll R, Bakkers J.. Zebrafish integrin-linked kinase is required in skeletal muscles for strengthening the integrin-ECM adhesion complex. Dev Biol 318: 92-101

Hubrecht Institute and University Medical Centre Utrecht, The Netherlands.
Developmental Biology (Impact Factor: 3.55). 07/2008; 318(1):92-101. DOI: 10.1016/j.ydbio.2008.03.024
Source: PubMed


Mechanical instability of skeletal muscle cells is the major cause of congenital muscular dystrophy. Here we show that the zebrafish lost-contact mutant, that lacks a functional integrin-linked kinase (ilk) gene, suffers from mechanical instability of skeletal muscle fibres. With genetic and morpholino knock-down experiments we demonstrate that: 1) laminin, itgalpha7, Ilk and beta-parvin are all critical for mechanical stability in skeletal muscles. 2) Ilk acts redundantly with the dystrophin/dystroglycan adhesion complex in maintaining mechanical stability of skeletal muscles. 3) Ilk protein is recruited to the myotendinous junctions, which requires the ECM component laminin and the presence of itgalpha7 in the sarcolemma. 4) Ilk, unexpectedly, is dispensable for formation of the adhesion complex. Ilk, however, is required for strengthening the adhesion of the muscle fibre with the ECM and this activity requires the presence of a functional kinase domain in Ilk. 5) We identified a novel interaction between Ilk and the mechanical stretch sensor protein MLP. Thus, Ilk is an essential intracellular component downstream of laminin and itgalpha7, providing strengthening of skeletal muscle fibre adhesion with the ECM and therefore qualified as a novel candidate gene for congenital muscular dystrophy.

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Available from: Jeroen Bakkers
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    • "Unfertilized eggs were removed at 3 hpf and 50 to 200 embryos were then kept for gross phenotype analysis and remaining eggs were processed as described below. For gene interaction assays, the integrin α7-ATG morpholinos (MO-itga7) (Postel et al., 2008), laminin-α2-MO (MO-lama2) (Pollard et al., 2006), dystroglycan-MO (MO-dag1) (Parsons et al., 2002) were used. Subphenotypic doses determined for each morpholino (0.0625 pmol for MO- itga7, 0.125 pmol for MO22a, 0.25 pmol for MO-dag1 and MO-lama2) were used for co-injection into one-cell stage embryos. "
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    ABSTRACT: The myotendinous junction (MTJ) is the major site of force transfer in skeletal muscle, and defects in its structure correlate with a subset of muscular dystrophies. Col22a1 encodes the MTJ component collagen XXII, the function of which remains unknown. Here, we have cloned and characterized the zebrafish col22a1 gene and conducted morpholino-based loss-of-function studies in developing embryos. We showed that col22a1 transcripts localize at muscle ends when the MTJ forms and that COLXXII protein integrates the junctional extracellular matrix. Knockdown of COLXXII expression resulted in muscular dystrophy-like phenotype, including swimming impairment, curvature of embryo trunk/tail, strong reduction of twitch-contraction amplitude and contraction-induced muscle fiber detachment, and provoked significant activation of the survival factor Akt. Electron microscopy and immunofluorescence studies revealed that absence of COLXXII caused a strong reduction of MTJ folds and defects in myoseptal structure. These defects resulted in reduced contractile force and susceptibility of junctional extracellular matrix to rupture when subjected to repeated mechanical stress. Co-injection of sub-phenotypic doses of morpholinos against col22a1 and genes of the major muscle linkage systems showed a synergistic gene interaction between col22a1 and itga7 (α7β1 integrin) that was not observed with dag1 (dystroglycan). Finally, pertinent to a conserved role in humans, the dystrophic phenotype was rescued by microinjection of recombinant human COLXXII. Our findings indicate that COLXXII contributes to the stabilization of myotendinous junctions and strengthens skeletal muscle attachments during contractile activity.
    Full-text · Article · Oct 2013 · Development
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    • "However, a histological analysis of skeletal muscle revealed typical symptoms of a progressive muscular dystrophy starting soon after birth, but with a distinct variability in different muscle types [42]. The knock-down of zebrafish Itga7 results in muscle fiber detachments similar to those observed in lama2 and lama4-deficient embryos [43]. The human deficiency in integrin α7 causes a mild disorder that is best characterized as congenital myopathy. "
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    ABSTRACT: We report an Italian family in which the proband showed a severe phenotype characterized by the association of congenital fiber type disproportion (CFTD) with a left ventricular non-compaction cardiomyopathy (LVNC). This study was focused on the identification of the responsible gene/s.Methods and results: Using the whole-exome sequencing approach, we identified the proband homozygous missense mutations in two genes, the myosin heavy chain 7B (MYH7B) and the integrin alpha 7 (ITGA7). Both genes are expressed in heart and muscle tissues, and both mutations were predicted to be deleterious and were not found in the healthy population.The R890C mutation in the MYH7B gene segregated with the LVNC phenotype in the examined family. It was also found in one unrelated patient affected by LVNC, confirming a causative role in cardiomyopathy.The E882K mutation in the ITGA7 gene, a key component of the basal lamina of muscle fibers, was found only in the proband, suggesting a role in CFTD. This study identifies two novel disease genes. Mutation in MYH7B causes a classical LVNC phenotype, whereas mutation in ITGA7 causes CFTD. Both phenotypes represent alterations of skeletal and cardiac muscle maturation and are usually not severe. The severe phenotype of the proband is most likely due to a synergic effect of these two mutations.This study provides new insights into the genetics underlying Mendelian traits and demonstrates a role for digenic inheritance in complex phenotypes.
    Full-text · Article · Jun 2013 · Orphanet Journal of Rare Diseases
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    • "In zebrafish, morpholino-based knockdown of itga7 induces a dystrophic phenotype similar to the lost-contact (loc) mutant, which carries an ilk null allele that leads to loss of integrin function (Knöll et al., 2007; Postel et al., 2008). In both loc mutants and itga7 morphants, myofibres detach from the myosepta, leading to progressive paralysis (Postel et al., 2008). Importantly, the zebrafish-based study of Knöll and colleagues demonstrates an epistatic regulation between ilk and lama4, which led to the identification of previously unknown ILK and LAMA4 mutations in individuals with dilated cardiomyopathy (Knöll et al., 2007). "
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    ABSTRACT: Muscular dystrophies are a group of genetic disorders that specifically affect skeletal muscle and are characterized by progressive muscle degeneration and weakening. To develop therapies and treatments for these diseases, a better understanding of the molecular basis of muscular dystrophies is required. Thus, identification of causative genes mutated in specific disorders and the study of relevant animal models are imperative. Zebrafish genetic models of human muscle disorders often closely resemble disease pathogenesis, and the optical clarity of zebrafish embryos and larvae enables visualization of dynamic molecular processes in vivo. As an adjunct tool, morpholino studies provide insight into the molecular function of genes and allow rapid assessment of candidate genes for human muscular dystrophies. This unique set of attributes makes the zebrafish model system particularly valuable for the study of muscle diseases. This review discusses how recent research using zebrafish has shed light on the pathological basis of muscular dystrophies, with particular focus on the muscle cell membrane and the linkage between the myofibre cytoskeleton and the extracellular matrix.
    Full-text · Article · Nov 2012 · Disease Models and Mechanisms
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