# CELL BIOLOGY AND MOLECULAR MECHANISMS OF HUMAN GAMMA/DELTA T CELL ACTIVATION

> **NIH NIH R01** · UNIVERSITY OF MICHIGAN AT ANN ARBOR · 2021 · $383,280

## Abstract

PROJECT SUMMARY/ABSTRACT
Gamma/delta (γδ) T cells are innate-like lymphocytes that express unconventional antigen receptors (γδ TCR)
on their surface. Unlike conventional αβ T cells, they do not recognize peptide-MHC antigens but are instead
activated by invariant stress-inducible receptors and host/pathogen-associated metabolites. Most γδ T cells in
human peripheral blood express the Vγ9Vδ2 TCR and are activated by phosphoisoprenoid (PiP) metabolites
produced by medically important microbes, viruses and cancer cells. Vγ9Vδ2 T cell activation by PiPs is TCR-
dependent, and requires contact with PiP-producing target cells that express the B7-family protein butyrophilin
3A1 (BTN3A1). However, the molecular mechanisms by which PiPs and BTN3A1 initiate signaling and activate
Vγ9Vδ2 T cells remain unresolved. Our overall objective in this proposal is to identify the cell biological and
molecular basis of PiP-dependent Vγ9Vδ2 T cell activation. In αβ T cells, antigen-induced signaling occurs at
the cell-cell contact interface between T cells and antigen-bearing target cells, known as the immunological
synapse. We propose to employ cutting edge approaches that include super-resolution fluorescence imaging,
electron microscopy/tomography (EM), CRISPR/Cas9-based genomic editing, and proximity-based chemical
tagging, to test our central hypothesis: that PiP-mediated Vγ9Vδ2 T cell activation occurs through a uniquely
configured immunological synapse, in which TCR, accessory receptors (including BTN3A1), cytoskeletal
elements, and lipid domains act in cis and across the synapse to initiate signaling. Our substantial preliminary
investigations establish the feasibility of our approach and point to a need for `close contacts' and synaptic
TCR/phosphatase segregation for effective PiP-dependent T cell activation. Surprisingly, we find that
expression of BTN3A1 is required in both target cells and T cells, pointing to the possibility of an unsuspected
trans-synaptic interaction of BTN3A1. To test our central hypothesis, we propose two Specific Aims: 1.) Using
super-resolution imaging and EM, in conjunction with biochemical, signaling and functional assays, establish
the PiP-dependent molecular organization, membrane topography, and key signaling events at the Vγ9Vδ2 T
cell synapse; and 2.) Using genome-editing and RNAi approaches, combined with BTN3A1 structure/function
mutagenesis, establish the role of PiP-induced changes in cell-surface organization of BTN3A1 in Vγ9Vδ2 T
cell activation. Our approach is innovative as it uses cutting-edge imaging and biochemical methods to test a
novel model for PiP-mediated Vγ9Vδ2T cell activation. The proposed research is significant as it will identify
the fundamental biochemical, biophysical and cell biological requirements for Vγ9Vδ2 T cell activation, and
provide a mechanistically-based framework to harness γδ T cell immunity for cellular immunotherapy.

## Key facts

- **NIH application ID:** 10166762
- **Project number:** 5R01AI134999-03
- **Recipient organization:** UNIVERSITY OF MICHIGAN AT ANN ARBOR
- **Principal Investigator:** Kaushik Choudhuri
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $383,280
- **Award type:** 5
- **Project period:** 2019-06-11 → 2024-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10166762

## Citation

> US National Institutes of Health, RePORTER application 10166762, CELL BIOLOGY AND MOLECULAR MECHANISMS OF HUMAN GAMMA/DELTA T CELL ACTIVATION (5R01AI134999-03). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/10166762. Licensed CC0.

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