FUNCTIONAL REEXPRESSION OF LAMININ-5 IN LAMININ-GAMMA-2-DEFICIENT HUMAN KERATINOCYTES MODIFIES CELL MORPHOLOGY, MOTILITY, AND ADHESION

Herlitz junctional epidermolysis bullosa (H-JEB) is characterized by a reduced adherence of keratinocytes consequent to deficient expression of the extracellular adhesive ligand laminin-5, To complement the gen etic defect causing H-JEB, we transferred an eukaryotic cassette expre ssing the cDNA for the gamma 2 chain of laminin-5 into H-JEB keratinoc ytes in which the expression of the polypeptide is hampered by a homoz ygous mutation generating a premature termination codon. Transfection using adenovirus-polylysin-transferrin DNA complexes resulted in a tra nsient synthesis of the recombinant laminin gamma 2 chain that associa ted with the endogenous alpha 3 and beta 3 chains to form laminin-5 mo lecules readily deposited on the tissue culture substrate, Furthermore , retroviral mediated transduction of the gamma 2 cDNA yielded persist ent expression and polarized secretion of laminin-5, The protein incor porated into the basement membrane produced by the revertant cells ino culated subcutaneously in nude mice. In these transfectants, re-expres sion of laminin-5 induced changes in cell morphology and reorganizatio n of focal adhesions that assumed the shape and distribution of the co unterparts detected in normal keratinocytes. These observations correl ated with an enhanced cell-substrate adhesion and a reduced motility o f the transfected cells, Our results demonstrate that a restored expre ssion of laminin-5 induces a phenotypic reversion of genetically alter ed H-JEB keratinocytes and open new perspectives to the analysis of th e mechanisms regulating adhesion of epithelial cells.