Functional Re-expression of laminin-5 in laminin-gamma2-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 genetic defect causing H-JEB, we transferred an eukaryotic cassette expressing the cDNA for the gamma2 chain of laminin-5 into H-JEB keratinocytes in which the expression of the polypeptide is hampered by a homozygous mutation generating a premature termination codon. Transfection using adenovirus-polylysin-transferrin-DNA complexes resulted in a transient synthesis of the recombinant laminin gamma2 chain that associated with the endogenous alpha3 and beta3 chains to form laminin-5 molecules readily deposited on the tissue culture substrate. Furthermore, retroviral-mediated transduction of the gamma2 cDNA yielded persistent expression and polarized secretion of laminin-5. The protein incorporated into the basement membrane produced by the revertant cells inoculated subcutaneously in nude mice. In these transfectants, re-expression of laminin-5 induced changes in cell morphology and reorganization of focal adhesions that assumed the shape and distribution of the counterparts detected in normal keratinocytes. These observations correlated with an enhanced cell-substrate adhesion and a reduced motility of the transfected cells. Our results demonstrate that a restored expression of laminin-5 induces a phenotypic reversion of genetically altered H-JEB keratinocytes and open new perspectives to the analysis of the mechanisms regulating adhesion of epithelial cells.
Gagnoux-Palacios
L
L
U385 INSERM, Faculté de Médecine, 06107 Nice Cedex 2, France.
Vailly
J
J
Durand-Clement
M
M
Wagner
E
E
Ortonne
J P
JP
Meneguzzi
G
G
D000818:Animals; D002448:Cell Adhesion / Q000235:genetics; D015815:Cell Adhesion Molecules / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D002465:Cell Movement / Q000235:genetics; D048429:Cell Size / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D016109:Epidermolysis Bullosa, Junctional / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D015603:Keratinocytes / Q000166:cytology* / Q000378:metabolism*; D007797:Laminin / Q000172:deficiency*; D051379:Mice; D008819:Mice, Nude; D010641:Phenotype; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D000097794:Kalinin
D015815:Cell Adhesion Molecules; D018076:DNA, Complementary; D007797:Laminin; D011994:Recombinant Proteins; D000097794:Kalinin