Structure and Function of Tetraspanin Complexes

Project Summary Tetraspanins are an ancient and exceptionally well-conserved family of four pass transmembrane proteins – numbering 33 unique members in humans – that have essential but poorly understood functions in many different cellular contexts. Among tetraspanin proteins, CD81 and the C8 subfamily stand out as particularly important in human biology. CD81 is a binding partner of CD19 in the B cell co-receptor complex, a key regulator of B cell receptor (BCR) signaling. Relatively little is known, however, about how the dynamic association of CD81 with CD19 regulates its association with the B cell receptor and ability to induce a signaling response, critical knowledge gaps that impede therapeutic efforts to modulate or target this pathway. The subset of tetraspanins with eight extracellular cysteine residues (the “C8” tetraspanins) facilitate the maturation and function of the transmembrane metalloprotease ADAM10, which has numerous important roles in physiology and disease. In the immune system, ADAM10 promotes T cell proliferation and cytokine production as the primary sheddase of the low-affinity IgE receptor CD23, and it cleaves Lag-3 in response to stimulation of the T cell receptor. ADAM10 also catalyzes ligand-dependent physiologic Notch cleavage, an essential step in Notch signal transduction, and is the protease responsible for constitutive alpha secretase processing of the Alzheimer’s precursor protein APP, resulting in production of a non-toxic product instead of the toxic A-beta (Aβ) fragment produced by beta- secretase. The objective of this work is to provide a deep and comprehensive understanding of the molecular basis for the function of CD81 and the C8 tetraspanins using structural, biochemical, and dynamic approaches, building from our recent progress in visualizing the structure of CD81 in its free and CD19-bound states. To achieve this goal, we plan to elucidate the dynamics of the CD19-CD81 co-receptor complex in response to B cell activation, determine the molecular basis for recognition of ADAM10 by C8 tetraspanins, and uncover how C8-tetraspanin binding to ADAM10 modulates substrate selection and proteolytic activity. Together, successful completion of these aims will provide a deep understanding of how CD81 modulates signal transduction by the B cell co-receptor, and how the C8 tetraspanins influence ADAM10 metalloprotease function. These findings will inform future therapeutic efforts targeting these important biological processes.