title: Laminin-alpha4 and integrin-linked kinase mutations cause human cardiomyopathy via simultaneous defects in cardiomyocytes and endothelial cells.
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.
  
  
  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.
authors.affiliation: Institute of Molecular Medicine, University of California at San Diego, La Jolla, Calif, USA. rknoell@med.uni-goettingen.de
authors.forename: Ralph
authors.initials: R
authors.lastname: Knöll
authors.forename: Ruben
authors.initials: R
authors.lastname: Postel
authors.forename: Jianming
authors.initials: J
authors.lastname: Wang
authors.forename: Ralph
authors.initials: R
authors.lastname: Krätzner
authors.forename: Gerrit
authors.initials: G
authors.lastname: Hennecke
authors.forename: Andrei M
authors.initials: AM
authors.lastname: Vacaru
authors.forename: Padmanabhan
authors.initials: P
authors.lastname: Vakeel
authors.forename: Cornelia
authors.initials: C
authors.lastname: Schubert
authors.forename: Kenton
authors.initials: K
authors.lastname: Murthy
authors.forename: Brinda K
authors.initials: BK
authors.lastname: Rana
authors.forename: Dieter
authors.initials: D
authors.lastname: Kube
authors.forename: Gudrun
authors.initials: G
authors.lastname: Knöll
authors.forename: Katrin
authors.initials: K
authors.lastname: Schäfer
authors.forename: Takeharu
authors.initials: T
authors.lastname: Hayashi
authors.forename: Torbjorn
authors.initials: T
authors.lastname: Holm
authors.forename: Akinori
authors.initials: A
authors.lastname: Kimura
authors.forename: Nicholas
authors.initials: N
authors.lastname: Schork
authors.forename: Mohammad Reza
authors.initials: MR
authors.lastname: Toliat
authors.forename: Peter
authors.initials: P
authors.lastname: Nürnberg
authors.forename: Heinz-Peter
authors.initials: HP
authors.lastname: Schultheiss
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Schaper
authors.forename: Jutta
authors.initials: J
authors.lastname: Schaper
authors.forename: Erik
authors.initials: E
authors.lastname: Bos
authors.forename: Jeroen
authors.initials: J
authors.lastname: Den Hertog
authors.forename: Fredericus J M
authors.initials: FJ
authors.lastname: van Eeden
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Peters
authors.forename: Gerd
authors.initials: G
authors.lastname: Hasenfuss
authors.forename: Kenneth R
authors.initials: KR
authors.lastname: Chien
authors.forename: Jeroen
authors.initials: J
authors.lastname: Bakkers
mesh_terms: D000328:Adult; D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002311:Cardiomyopathy, Dilated / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002448:Cell Adhesion; D002522:Chlorocebus aethiops; D002874:Chromosome Mapping; D018389:Codon, Nonsense; D004252:DNA Mutational Analysis; D004625:Embryo, Nonmammalian / Q000473:pathology; D042783:Endothelial Cells / Q000473:pathology*; D044127:Epigenesis, Genetic; D005109:Extracellular Matrix / Q000378:metabolism / Q000473:pathology; D005260:Female; D006321:Heart / Q000196:embryology; D006333:Heart Failure / Q000209:etiology / Q000473:pathology; D006801:Humans; D016023:Integrins / Q000378:metabolism; D007797:Laminin / Q000235:genetics* / Q000502:physiology; D008297:Male; D008875:Middle Aged; D008958:Models, Molecular; D020125:Mutation, Missense; D009206:Myocardium / Q000473:pathology; D032383:Myocytes, Cardiac / Q000473:pathology*; D016376:Oligonucleotides, Antisense / Q000633:toxicity; D010375:Pedigree; D017354:Point Mutation; D011485:Protein Binding; D011487:Protein Conformation; D025941:Protein Interaction Mapping; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000502:physiology; D017434:Protein Structure, Tertiary; D014162:Transfection; D015027:Zebrafish / Q000196:embryology / Q000235:genetics; D029961:Zebrafish Proteins / Q000235:genetics / Q000502:physiology; D000099200:Scaffold Protein ILK
chemical_list: D018389:Codon, Nonsense; D016023:Integrins; D007797:Laminin; D016376:Oligonucleotides, Antisense; D017346:Protein Serine-Threonine Kinases; D029961:Zebrafish Proteins; D000099200:Scaffold Protein ILK; C501369:LAMA4 protein, human; C522179:LAMA4 protein, zebrafish; C522180:ILK protein, zebrafish