A Structural Change in Butyrophilin upon Phosphoantigen Binding Underlies Phosphoantigen-Mediated Vγ9Vδ2 T Cell Activation

Human V gamma 9V delta 2 T cells respond to microbial infections and malignancy by sensing diphosphate-containing metabolites called phosphoantigens, which bind to the intracellular domain of butyrophilin 3A1, triggering extracellular interactions with the V gamma 9V delta 2 T cell receptor (TCR). Here, we examined the molecular basis of this inside-out triggering mechanism. Crystal structures of intracellular butyrophilin 3A proteins alone or in complex with the potent microbial phosphoantigen HMBPP or a synthetic analog revealed key features of phosphoantigens and butyrophilins required for gamma delta T cell activation. Analyses with chemical probes and molecular dynamic simulations demonstrated that dimerized intracellular proteins cooperate in sensing HMBPP to enhance the efficiency of gamma delta T cell activation. HMBPP binding to butyrophilin doubled the binding force between a gamma delta T cell and a target cell during outside signaling, as measured by single-cell force microscopy. Our findings provide insight into the inside-out triggering of V gamma 9V delta 2 T cell activation by phosphoantigen-bound butyrophilin, facilitating immunotherapeutic drug design.