Laminin-alpha4 and integrin-linked kinase mutations cause human cardiomyopathy via simultaneous defects in cardiomyocytes and endothelial cells

Circulation. 2007 Jul 31;116(5):515-25. doi: 10.1161/CIRCULATIONAHA.107.689984. Epub 2007 Jul 23.

Abstract

Background: Extracellular matrix proteins, such as laminins, and endothelial cells are known to influence cardiomyocyte performance; however, the underlying molecular mechanisms remain poorly understood.

Methods and results: We used a forward genetic screen in zebrafish to identify novel genes required for myocardial function and were able to identify the lost-contact (loc) mutant, which encodes a nonsense mutation in the integrin-linked kinase (ilk) gene. This loc/ilk mutant is associated with a severe defect in cardiomyocytes and endothelial cells that leads to severe myocardial dysfunction. Additional experiments revealed the epistatic regulation between laminin-alpha4 (Lama4), integrin, and Ilk, which led us to screen for mutations in the human ILK and LAMA4 genes in patients with severe dilated cardiomyopathy. We identified 2 novel amino acid residue-altering mutations (2828C>T [Pro943Leu] and 3217C>T [Arg1073X]) in the integrin-interacting domain of the LAMA4 gene and 1 mutation (785C>T [Ala262Val]) in the ILK gene. Biacore quantitative protein/protein interaction data, which have been used to determine the equilibrium dissociation constants, point to the loss of integrin-binding capacity in case of the Pro943Leu (Kd=5+/-3 micromol/L) and Arg1073X LAMA4 (Kd=1+/-0.2 micromol/L) mutants compared with the wild-type LAMA4 protein (Kd=440+/-20 nmol/L). Additional functional data point to the loss of endothelial cells in affected patients as a direct consequence of the mutant genes, which ultimately leads to heart failure.

Conclusions: This is the first report on mutations in the laminin, integrin, and ILK system in human cardiomyopathy, which has consequences for endothelial cells as well as for cardiomyocytes, thus providing a new genetic basis for dilated cardiomyopathy in humans.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Amino Acid Substitution
  • Animals
  • COS Cells
  • Cardiomyopathy, Dilated / genetics*
  • Cardiomyopathy, Dilated / metabolism
  • Cardiomyopathy, Dilated / pathology
  • Cell Adhesion
  • Chlorocebus aethiops
  • Chromosome Mapping
  • Codon, Nonsense
  • DNA Mutational Analysis
  • Embryo, Nonmammalian / pathology
  • Endothelial Cells / pathology*
  • Epigenesis, Genetic
  • Extracellular Matrix / metabolism
  • Extracellular Matrix / pathology
  • Female
  • Heart / embryology
  • Heart Failure / etiology
  • Heart Failure / pathology
  • Humans
  • Integrins / metabolism
  • Laminin / genetics*
  • Laminin / physiology
  • Male
  • Middle Aged
  • Models, Molecular
  • Mutation, Missense*
  • Myocardium / pathology
  • Myocytes, Cardiac / pathology*
  • Oligonucleotides, Antisense / toxicity
  • Pedigree
  • Point Mutation*
  • Protein Binding
  • Protein Conformation
  • Protein Interaction Mapping
  • Protein Structure, Tertiary
  • Protein-Serine-Threonine Kinases / genetics*
  • Protein-Serine-Threonine Kinases / physiology
  • Transfection
  • Zebrafish / embryology
  • Zebrafish / genetics
  • Zebrafish Proteins / genetics
  • Zebrafish Proteins / physiology

Substances

  • Codon, Nonsense
  • Integrins
  • LAMA4 protein, human
  • LAMA4 protein, zebrafish
  • Laminin
  • Oligonucleotides, Antisense
  • Zebrafish Proteins
  • integrin-linked kinase
  • ILK protein, zebrafish
  • Protein-Serine-Threonine Kinases