Sensor function for butyrophilin 3A1 in prenyl pyrophosphate stimulation of human Vγ2Vδ2 T cells (APP5P.100)

Abstract

Abstract Vγ2Vδ2 T cells play important roles in human immunity to pathogens and in cancer immunotherapy by responding to isoprenoid metabolites such as (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate and isopentenyl pyrophosphate. The immunoglobulin superfamily protein, butyrophilin 3A1 (BTN3A1), has been shown to be required for prenyl pyrophosphate stimulation. We proposed that the intracellular B30.2 domain of BTN3A1 binds prenyl pyrophosphates resulting in a change in the extracellular BTN3A1 dimer that is detected by Vγ2Vδ2 TCRs. Such B30.2 binding has been recently demonstrated. However, others have reported that the extracellular BTN3A1 IgV domain binds prenyl pyrophosphates leading to the proposal that the Vγ2Vδ2 TCR recognizes the complex. To distinguish between these mechanisms, we mutagenized residues in the binding sites to alanine and tested the mutant BTN3A1 proteins for their ability to mediate prenyl pyrophosphate stimulation of Vγ2Vδ2 T cells to proliferate and secrete TNF-α. Mutagenesis of residues in the IgV site had no effect whereas mutagenesis of residues within the basic pocket and surrounding V regions of the B30.2 domain abrogated prenyl pyrophosphate-induced proliferation and reduced or abolished TNF-α production. Some mutations further from the B30.2 site also diminished stimulation suggesting that the B30.2 domain may interact with a second protein. These findings support intracellular sensing of prenyl pyrophosphates rather than extracellular presentation.