Structural basis of the interaction between integrin ¿6ß4 and the bullous pemphigoid antigen BP230 in hemidesmosomes

Abstract

Hemidesmosomes are junctional complexes that mediate the stable adhesion of basal epithelial cells to the basement membrane by linking the extracellular matrix to the intermediate filament system of the cytoskeleton. Disorders that target hemidesmosomal proteins cause severe skin blistering diseases. Type I hemidesmosomes, present in the epidermis, contain three transmembrane proteins integrin ¿6ß4, bullous pemphigoid antigen 180 (BP180 or BPAG2), and tetraspanin CD151, and the cytoplasmic proteins plectin, and BP230 (BPAG1e). Integrin ¿6ß4 is connected to intermediate filaments via plectin and BP230, which bind to the cyto-domain of the ß4 subunit. To unravel the molecular basis of the BP230-ß4 interaction, we first mapped their mutual binding sites and subsequently solved the crystal structure of a human BP230-ß4 complex, which was refined to 2.05 ¿ resolution. A ~25-residues long segment of the N-terminal region of BP230 binds to the third and fourth fibronectin type III domains (FnIII-3,4) of ß4. The initial part of the BP230 site contacts the FnIII-4 domain, while the final part is inserted in a cleft between the FnIII-3 and FnIII-4 domains, which in turn form an inter-domain ionic clasp required for binding. Using double electron-electron resonance spectroscopy (DEER), we show that BP230-binding induces closure of the two FnIII domains of ß4. Disruption of the BP230-ß4 interface prevents the recruitment of BP230 to hemidesmosomes in keratinocytes in culture, revealing a key role of the BP230-ß4 interaction for the assembly of hemidesmosomes. Phosphomimetic substitutions of T1663 of ß4, and T39 and S46 of BP230, disrupt binding, suggesting that the BP230-ß4 interaction might be regulated by phosphorylation during hemidesmosome disassembly. In summary, our study provides insights into the molecular mechanisms of hemidesmosome architecture and regulation.