Fibulin-2 binds to the short arms of laminin-5 and laminin-1 via conserved amino acid sequences.
Epithelial cell-specific laminin-5, consisting of three chains, alpha3, beta3, and gamma2, is a component of the anchoring filament that traverses the lamina lucida beneath the hemidesmosomes of epidermal cells and functions to link these cells to the basement membrane. We have studied the molecular interaction between laminin-5 and extracellular matrix proteins using recombinant proteins and synthetic peptides. Affinity chromatography assays with recombinant fragments of the laminin gamma2 short arm identified a 195-kDa binding protein in the conditioned media from the mouse epidermal cell line Pam 212 and from primary dermal fibroblasts. This molecule was identified by Western blotting as fibulin-2, a recently identified extracellular matrix protein. Using deletion mutants and various synthetic peptides in competition assays, the 9-amino acid sequence SADFSVHKI (residues 199-207) in domain IV of the gamma2 chain was defined as a critical site for fibulin-2 binding. An anti-gamma2 antibody co-immunoprecipitated fibulin-2 from the conditioned media, further confirming the interaction of fibulin-2 with laminin-5. Fibulin-2 was also found to interact with laminin-1 (alpha1beta1gamma1) through a region (residues 654-665) of the alpha1 chain short arm whose sequence is similar to that of the fibulin-2 binding site of the gamma2 chain. Together these results suggest that fibulin-2 functions to bridge laminin-1 and laminin-5 with other extracellular matrix proteins, providing a linkage between the cell surface and the basement membrane.
Utani
A
A
Laboratory of Developmental Biology, NIDR, National Institutes of Health, Bethesda, Maryland 20892, USA.
Nomizu
M
M
Yamada
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D000831:Animals, Newborn; D001665:Binding Sites; D001667:Binding, Competitive; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D002294:Carcinoma, Squamous Cell; D015815:Cell Adhesion Molecules / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D017124:Conserved Sequence; D016326:Extracellular Matrix Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D051379:Mice; D008815:Mice, Inbred Strains; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry / Q000494:pharmacology; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012867:Skin / Q000378:metabolism*; D000097794:Kalinin
D002135:Calcium-Binding Proteins; D015815:Cell Adhesion Molecules; D016326:Extracellular Matrix Proteins; D009842:Oligopeptides; D010446:Peptide Fragments; D011994:Recombinant Proteins; D000097794:Kalinin; C084612:fibulin 2