# A ligand-centered framework for γδ T cell activation in colorectal cancer revealed by single-cell and transformer-based perturbation

## TL;DR

This study identifies key ligands and transcription factors involved in γδ T cell activation in colorectal cancer, offering insights for improved immunotherapies.

## Contribution

A novel ligand inference pipeline combining single-cell data and in silico perturbation to reveal γδ T cell activation mechanisms in CRC.

## Key findings

- IL-15 and TNFSF9 (4-1BBL) promote γδ T cell effector function.
- NCR2 and KLRC3 (NKG2E) are linked to γδ T cell activation when overexpressed.
- Monocytes and dendritic cells are key contributors to γδ T cell activation in the tumor microenvironment.

## Abstract

Understanding the activation mechanisms of γδ T cells in colorectal cancer (CRC) is critical for harnessing their therapeutic potential. Here, using an atlas of human CRC-infiltrating γδ T cells that we built by integrating multiple single-cell RNA-seq datasets, we developed a γδ T cell-refined ligand inference pipeline by combining differential gene expression, gene regulatory network prediction, ligand inference, and in silico perturbation analysis. This approach identified ligands, including IL-15 and TNFSF9 (4-1BBL), as candidates promoting γδ T cell effector function and highlighted NCR2 and KLRC3 (NKG2E), whose in silico overexpression was associated with γδ T cell activation. Ligand enrichment analyses further indicated that monocytes and dendritic cells are key contributors to γδ T cell activation in the tumor microenvironment. Our results also highlighted transcription factors IKZF1, FOSL2, and FOXO1 in the less activated γδ T cells and IRF1, KLF2, and BHLHE40 in the effector γδ T cells that plausibly regulated the differential activation state. Together, our results offer a systems-level view of the signaling and transcriptional programs governing γδ T cell phenotypes in CRC and provide a foundation for γδ T cell-based immunotherapies with enhanced antitumor functions.