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, alpha 3, beta 3, and gamma 2, 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 gamma 2 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 g-amino acid sequence SADFSVHKI (residues 199-207) in domain IV of the gamma 2 chain was defined as a critical site for fibulin-2 binding. An anti-gamma 2 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 (alpha 1 beta 1 gamma 1) through a region (residues 654-665) of the alpha 1 chain short arm whose sequence is similar to that of the fibulin-2 binding site of the gamma 2 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.