Molecular basis for recognition and surface export of integrin α3β1 by the tetraspanin CD151

Abstract

Integrin α3β1 (ITGα3β1) is a member of an integrin subfamily that binds to laminin proteins and promotes attachment of epithelial cells to the basement membrane. ITGα3β1 forms a complex with the tetraspanin CD151, and loss-of-function mutations in both ITGα3 and CD151 cause epidermolysis bullosa, a severe skin blistering disease thought to result from a defect in cellular attachment to the basement membrane. Though functionally linked, the molecular details of how CD151 regulates integrin signaling are unclear. The work of this dissertation identifies a previously unappreciated role for CD151. In Chapter 2, we report the cryo-EM structure of an ITGα3β1 complex with CD151, identifying a conserved interface between the ITGα3 calf-2 domain and CD151 predicted to remain accessible independent of integrin conformation. Cell based assays of wild-type and mutant CD151 show that this interface is required for complex formation. Strikingly, CRISPR-mediated CD151 knockout cells have a defect in ITGα3β1 surface export that is variably penetrant among knockout clones, and surface delivery of ITGα3β1 is restored in CD151 knockout cells upon re-expression of wild-type CD151 but remains reduced upon expression of interface-disruptive CD151 mutant proteins. Together, these studies identify CD151 as a chaperone that promotes ITGα3β1 surface export and define the molecular basis for binding of CD151 to ITGα3β1, providing a biochemical rationale for their related loss-of-function phenotypes in epidermolysis bullosa. In Chapter 3, I report an AlphaFold optimized and modular laminin E8 fragment expression and purification method for studying laminin binding integrin biology. Importantly, these studies also identify an understudied tetraspanin, Tspan11, which can compensate for loss of CD151. Collectively, this work determines that tetraspanins CD151 and Tspan11 chaperone ITGα3β1 surface export and provides key insights for future tetraspanin-integrin research.