Gene Correction of Integrin beta sub(4)-dependent Pyloric Atresia-Junctional Epidermolysis Bullosa Keratinocytes Establishes a Role for beta sub(4) Tyrosines 1422 and 1440 in Hemidesmosome Assembly

The cytoplasmic domain of beta sub(4) integrin contains two pairs of fibronectin-like repeats separated by a connecting segment. The connecting segment harbors a putative tyrosine activation motif in which tyrosines 1422 and 1440 are phosphorylated in response to alpha sub(6) beta sub(4) binding to...

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Bibliographic Details
Published in:The Journal of biological chemistry 2001-11, Vol.276 (44), p.41336-41342
Main Authors: Dellambra, E, Prislei, S, Salvati, AL, Madeddu, M L, Golisano, O, Siviero, E, Bondanza, S, Cicuzza, S, Orecchia, A, Giancotti, F G, Zambruno, G, De Luca, M
Format: Article
Language:English
Subjects:
hemidesmosomes
pyloric atresia
tyrosine
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Summary:The cytoplasmic domain of beta sub(4) integrin contains two pairs of fibronectin-like repeats separated by a connecting segment. The connecting segment harbors a putative tyrosine activation motif in which tyrosines 1422 and 1440 are phosphorylated in response to alpha sub(6) beta sub(4) binding to laminin-5. Primary beta sub(4)-null keratinocytes, obtained from a newborn suffering from lethal junctional epidermolysis bullosa, were stably transduced with retroviruses carrying a full-length beta sub(4) cDNA or a beta sub(4) cDNA with phenylalanine substitutions at Tyr-1422 and Tyr-1440. Hemidesmosome assembly was evaluated on organotypic skin cultures. beta sub(4)-corrected keratinocytes were indistinguishable from normal cells in terms of alpha sub(6) beta sub(4) expression, the localization of hemidesmosome components, and hemidesmosome structure and density, suggesting full genetic and functional correction of beta sub(4)-null keratinocytes. In cultures generated from beta sub(4) super(Y1422F/Y1440F) keratinocytes, beta sub(4) mutants as well as alpha sub(6) integrin, HD1/plectin, and BP180 were not concentrated at the dermal-epidermal junction. Furthermore, the number of hemidesmosomes was strikingly reduced as compared with beta sub(4)-corrected keratinocytes. The rare hemidesmosomes detected in beta sub(4) super(Y1422F/Y1440) cells were devoid of sub-basal dense plates and of inner cytoplasmic plaques with keratin filament insertion. Collectively, our data demonstrate that the beta sub(4) tyrosine activation motif is not required for the localization of alpha sub(6) beta sub(4) at the keratinocyte plasma membrane but is essential for optimal assembly of bona fide hemidesmosomes.
ISSN:0021-9258