The butyrophilin 3A1 intracellular domain undergoes a conformational change involving the juxtamembrane region

Abstract

Small isoprenoid diphosphates, such as ()-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP), are ligands of the internal domain of BTN3A1. Ligand binding in target cells promotes activation of Vγ9Vδ2 T cells. We demonstrate by small-angle X-ray scattering (SAXS) that HMBPP binding to the internal domain of BTN3A1 induces a conformational change in the position of the B30.2 domain relative to the juxtamembrane (JM) region. To better understand the molecular details of this conformational rearrangement, NMR spectroscopy was used to discover that the JM region interacts with HMBPP, specifically at the diphosphate. The spectral location of the affected amide peaks, partial NMR assignments, and JM mutants (STAA or TA) investigated, confirm that the backbone amide of at least one Thr (Thr), adjacent to conserved Ser, comes close to the HMBPP diphosphate, whereas double mutation of nonconserved residues (Ser/Thr) may perturb the local fold. Cellular mutation of either of the identified Thr residues reduces the activation of Vγ9Vδ2 T cells by HMBPP, zoledronate, and POM-C-HMBP, but not by a partial agonist BTN3 antibody. Taken together, our results show that ligand binding to BTN3A1 induces a conformational change within the intracellular domain that involves the JM region and is required for full activation.-Nguyen, K., Li, J., Puthenveetil, R., Lin, X., Poe, M. M., Hsiao, C.-H. C., Vinogradova, O., Wiemer, A. J. The butyrophilin 3A1 intracellular domain undergoes a conformational change involving the juxtamembrane region.

The Armadillo (Dasypus novemcinctus): A Witness but Not a Functional Example for the Emergence of the Butyrophilin 3/Vγ9Vδ2 System in Placental Mammals

“…The human BTN3A3 gene is comprised of nine protein-coding exons with exon 2 encoding the BTN3-V region, exon 3 encoding BTN3-C, exon 4 representing part of the transmembrane region, followed by four relatively small exons (5–8) and the B30.2 exon (9) (Figure 1 A) ( 42 ). The alpaca BTN3 -like genomic sequence is organized in a locus strikingly homologous to human BTN3A3 (Figure 1 B), showing exons with nucleotide sequence identities to human BTN3A3 ranging from 73 to 93% and conserved intron lengths.…”

Towards Deciphering the Hidden Mechanisms That Contribute to the Antigenic Activation Process of Human Vγ9Vδ2 T Cells

“…Interestingly, HMBPP could interact with residues within both the B30.2 and the JTM region at different contact points. Furthermore, this report also indicates that both key residues Ser/Thr 296/297 and Thr 304 fall within a critical functional BTN3A1 JTM region ( 65 ).…”

“…Based on both length and flexibility characteristics, the authors propose that the B30.2 domain of BTN3A1 is moved toward the JTM region and closer to the membrane upon ligand binding. Such intracellular changes would be sensed by γδ T cells through modifications of either the extracellular domain or interactions with other molecular partners ( 65 ). Non-BTN3A proteins composed of an intracellular B30.2 domain have been shown to naturally multimerize and this status is important to fulfill their functions ( 74 , 75 ).…”

Improving the Efficiency of Vγ9Vδ2 T-Cell Immunotherapy in Cancer

“…The molecular details of signal transduction are a current research topic and matter of debate, especially regarding two different models: originally, the “antigen presenting model” by Vavassori et al ( 145 ) assuming that CD277 and the TCR interact directly following PAg binding to an extracellular CD277 domain. Recent experimental evidence rather supports a second, so called “allosteric model” by Harly et al, postulating that PAgs interact with the intracellular B30.2 domain of CD277 ( 147 ) either directly ( 148 ) or indirectly ( 149 , 150 ) and induce a conformational change that is transferred to the extracellular parts of the CD277 molecule ( 147 , 151 ). PAg sensing may additionally involve molecules like Rho-GTPase ( 152 ) or Periplakin and is modulated by mechanisms enabling transmembranous PAg transport or via hydrolyzation of PAgs by ecto-ATPase CD39 ( 106 , 153 ).…”