Scaffold-forming and Adhesive Contributions of Synthetic Laminin-binding Proteins to Basement Membrane Assembly

J Biol Chem. 2009 Mar 27;284(13):8984-94. doi: 10.1074/jbc.M809719200. Epub 2009 Feb 2.

Abstract

Laminins that possess three short arms contribute to basement membrane assembly by anchoring to cell surfaces, polymerizing, and binding to nidogen and collagen IV. Although laminins containing the alpha4 and alpha5 subunits are expressed in alpha2-deficient congenital muscular dystrophy, they may be ineffective substitutes because they bind weakly to cell surfaces and/or because they lack the third arm needed for polymerization. We asked whether linker proteins engineered to bind to deficient laminins that provide such missing activities would promote basement membrane assembly in a Schwann cell model. A chimeric fusion protein (alphaLNNd) that adds a short arm terminus to laminin through the nidogen binding locus was generated and compared with the dystrophy-ameliorating protein miniagrin (mAgrin) that binds to the laminin coiled-coil dystroglycan and sulfatides. alphaLNNd was found to mediate laminin binding to collagen IV, to bind to galactosyl sulfatide, and to selectively convert alpha-short arm deletion-mutant laminins LmDeltaalphaLN and LmDeltaalphaLN-L4b into polymerizing laminins. This protein enabled polymerization-deficient laminin but not an adhesion-deficient laminin lacking LG domains (LmDeltaLG) to assemble an extracellular matrix on Schwann cell surfaces. mAgrin, on the other hand, enabled LmDeltaLG to form an extracellular matrix on cell surfaces without increasing accumulation of non-polymerizing laminins. These gain-of-function studies reveal distinct polymerization and anchorage contributions to basement membrane assembly in which the three different LN domains mediate the former, and the LG domains provide primary anchorage with secondary contributions from the alphaLN domain. These findings may be relevant for an understanding of the pathogenesis and treatment of laminin deficiency states.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Agrin / chemistry
  • Agrin / genetics
  • Agrin / metabolism
  • Amino Acid Sequence
  • Animals
  • Basement Membrane / chemistry*
  • Basement Membrane / metabolism
  • Collagen Type IV / chemistry
  • Collagen Type IV / genetics
  • Collagen Type IV / metabolism
  • Dystroglycans / chemistry
  • Dystroglycans / genetics
  • Dystroglycans / metabolism
  • Humans
  • Laminin / chemistry*
  • Laminin / genetics
  • Laminin / metabolism
  • Membrane Glycoproteins / chemistry
  • Membrane Glycoproteins / genetics
  • Membrane Glycoproteins / metabolism
  • Mice
  • Models, Biological*
  • Muscular Dystrophies / genetics
  • Muscular Dystrophies / metabolism
  • Protein Binding
  • Protein Subunits / chemistry
  • Protein Subunits / genetics
  • Protein Subunits / metabolism
  • Recombinant Fusion Proteins / chemistry*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Schwann Cells / metabolism
  • Sequence Deletion

Substances

  • Agrin
  • Collagen Type IV
  • Laminin
  • Membrane Glycoproteins
  • Protein Subunits
  • Recombinant Fusion Proteins
  • miniagrin protein, mouse
  • nidogen
  • Dystroglycans