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 α6β4, bullous pemphigoid antigen 180 (BP180 or BPAG2), and tetraspanin CD151, and the cytoplasmic proteins plectin, and BP230 (BPAG1e). Integrin α6β4 is connected to intermediate filaments via plectin and BP230, which bind to the cyto-domain of the β4 subunit. To unravel the molecular basis of the BP230-β4 interaction, we first mapped their mutual binding sites and subsequently solved the crystal structure of a human BP230-β4 complex, which was refined to 2.05 Ǻ 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 β4. 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 β4. Disruption of the BP230-β4 interface prevents the recruitment of BP230 to hemidesmosomes in keratinocytes in culture, revealing a key role of the BP230-β4 interaction for the assembly of hemidesmosomes. Phosphomimetic substitutions of T1663 of β4, and T39 and S46 of BP230, disrupt binding, suggesting that the BP230-β4 interaction might be regulated by phosphorylation during hemidesmosome disassembly. In summary, our study provides insights into the molecular mechanisms of hemidesmosome architecture and regulation.

We thank ALBA-CELLS, Diamond Light Source, and EMBL-DESY for access to synchrotron-radiation facilities. This work was funded by the Spanish Ministry of Science, Innovation and Universities (grants BFU2009-08389, BFU2015-69499-P, CTQ2015-64486-R), Aragon Government, and Castilla-León Government. Grants were co-funded by the European Regional Development Fund. This work received funding from the European Community’s Seventh Framework Programme (FP7/2007-2013) under BioStruct-X (grant n0. 283570)

This poster was presented at the Biennial Structural Biology Conference "Structural Biology: Deeper into the Cell", organized by Instruct-ERIC (https://instruct-eric.eu/biennial2019), which was held in Alcalá de Henares (Spain) from May 22 till May 24, 2019.

No