Tyrosine phosphorylation: a critical component in the formation of hemidesmosomes.
Our goal was to evaluate the role of tyrosine phosphorylation in the complete formation of hemidesmosomes that occurs during development or during remodeling after injury. A corneal organ culture system was used to study hemidesmosome formation as it would occur in an intact tissue. Phosphorylation of the integrin subunit beta 4 and bullous pemphigoid antigen-1 (BPAG-1) was examined, as these proteins are known to play a role in linking the electron-dense plaques along the basal surface with the intermediate filaments to complete the formation of hemidesmosomes. Corneal epithelial sheets were placed on substrata that contained an intact basal lamina or basal laminae that had been either modified or removed. These constructs were incubated for up to 18 h, and hemidesmosome formation was evaluated by using transmission electron microscopy. When epithelial sheets were placed on intact basal laminae and incubated in the presence of the tyrosine kinase inhibitor genistein (200 microM), hemidesmosome formation was impaired. The formation of electron-dense regions was delayed, and no association of intermediate filaments was detected. Results were confirmed by biochemical studies. When the epithelium and underlying proteins were extracted and immunoprecipitated with beta 4 or BPAG-1, tyrosine phosphorylation decreased in the presence of genistein. In addition, the phosphorylation of beta 4 decreased when epithelial sheets were incubated on substrata from which the basal lamina had been removed or altered. Thus, a reduction in phosphorylation of tyrosine residues impairs the formation of mature hemidesmosomes, and substrata that fail to support hemidesmosome formation also demonstrate decreased phosphorylation of tyrosine residues.
J
J
Payne
H
H
Gong
V
V
Trinkaus-Randall
D000468:Alkalies / Q000494:pharmacology; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism / Q000502:physiology; D001324:Autoantigens / Q000378:metabolism; D001485:Basement Membrane / Q000187:drug effects / Q000378:metabolism; D002352:Carrier Proteins; D003094:Collagen / Q000378:metabolism; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D019573:Epithelium, Corneal / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D019833:Genistein / Q000494:pharmacology; D022002:Hemidesmosomes / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D007150:Immunohistochemistry; D039663:Integrin beta4; D008854:Microscopy, Electron; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D009924:Organ Culture Techniques; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000187:drug effects / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011817:Rabbits; D000097805:Collagen Type XVII
D000468:Alkalies; D015703:Antigens, CD; D001324:Autoantigens; D002352:Carrier Proteins; D003094:Collagen; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D019833:Genistein; D039663:Integrin beta4; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases; D000097805:Collagen Type XVII; C095667:Dst protein, mouse