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 Ig superfamily protein butyrophilin (BTN)3A1 was 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 was demonstrated recently. However, other investigators 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 two binding sites 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 on Vγ2Vδ2 T cell proliferation or secretion of TNF-α. In contrast, mutagenesis of residues within the basic pocket and surrounding V regions of the B30.2 domain abrogated prenyl pyrophosphate-induced proliferation. Mutations of residues making hydrogen bonds to the pyrophosphate moiety also abrogated TNF-α secretion, as did mutation of aromatic residues making contact with the alkenyl chain. Some mutations further from the B30.2 binding site also diminished stimulation, suggesting that the B30.2 domain may interact with a second protein. These findings support intracellular sensing of prenyl pyrophosphates by BTN3A1 rather than extracellular presentation.
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http://dx.doi.org/10.4049/jimmunol.1500314 | DOI Listing |
PLoS One
January 2025
Department of Biochemistry, Memorial University of Newfoundland, St. John's, Newfoundland and Labrador, Canada.
Geranylgeranyl pyrophosphate synthase (GGPPS), a key enzyme in protein prenylation, plays a critical role in cellular signal transduction and is a promising target for cancer therapy. However, the enzyme's native hexameric quaternary structure presents challenges for crystallographic studies. The primary objective of this study was to engineer dimeric forms of human GGPPS to facilitate high-resolution crystallographic analysis of its ligand binding interactions.
View Article and Find Full Text PDFMethods Mol Biol
January 2025
Department of Molecular and Cellular Biology, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan.
Functionally derivatized analogs of prenyl lipids are valuable tools for the detection and analysis of prenylated proteins. Using a biotinylated analog of geranylgeranyl, we previously identified Ykt6 as a substrate for a novel protein prenyltransferase, termed geranylgeranyltransferase type III (GGTase-III). Ykt6 is an evolutionarily highly conserved SNARE protein that regulates multiple intracellular trafficking pathways, including intra-Golgi trafficking and autophagosome-lysosome fusion.
View Article and Find Full Text PDFJ Cell Mol Med
December 2024
Cardiovascular Center, Medical College of Wisconsin, Milwaukee, Wisconsin, USA.
Fibrosis, characterised by excessive extracellular matrix deposition, contributes to both organ failure and significant mortality worldwide. Whereas fibroblasts are activated into myofibroblasts, marked by phenotypic factors such as α-smooth muscle actin (α-SMA), periostin, fibroblast activation protein (FAP) and heat shock protein 47 (HSP47), the cellular processes of trans-differentiation for fibrosis development remain poorly understood. Herein, we hypothesised that the molecular signalling of geranylgeranyl pyrophosphate (GGPP), a crucial biochemical molecule for protein prenylation, is essential in the regulation of profibrotic mechanisms for fibroblast-to-myofibroblast activation.
View Article and Find Full Text PDFBiochemistry
January 2025
Department of Chemistry, University of Minnesota-Twin Cities, Minneapolis, Minnesota 55455, United States.
Prenylation consists of the modification of proteins with either farnesyl diphosphate (FPP) or geranylgeranyl diphosphate (GGPP) at a cysteine near the C-terminus of target proteins to generate thioether-linked lipidated proteins. In recent work, metabolic labeling with alkyne-containing isoprenoid analogues including C15AlkOPP has been used to identify prenylated proteins and track their levels in different diseases. Here, a systematic study of the impact of isoprenoid length on proteins labeled with these probes was performed.
View Article and Find Full Text PDFPlant Cell Physiol
November 2024
Laboratory of Plant Gene Expression, Research Institute for Sustainable Humanosphere, Kyoto University, Gokasho, Uji, Kyoto 611-0011, Japan.
Plant membrane-bound prenyltransferases (PTs) catalyse the transfer of prenyl groups to acceptor substrates, phenols, using prenyl diphosphates as the donor substrate. The presence of prenyl residues in the reaction products, prenylated phenols, is key to the expression of a variety of physiological activities. Plant PTs generally exhibit high specificities for both substrate recognition and prenylation sites, while the molecular mechanism involved in these enzymatic properties is largely unknown.
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