In this project, we will further investigate how γδ TCR repertoires are shaped and which cognate ligands can trigger γδ TCR signaling. We hypothesize that the peripheral γδ TCR repertoire is shaped through recognition of specific antigen via the γδ TCR and that antigen-specific triggering of the γδ TCR induces intracellular signaling events leading to proliferation and survival of the respective γδ T cells. The principal method to test our hypotheses is already established in our group, namely the analysis of human γδ TCR repertoires by next generation sequencing of rearranged TRG and TRD gene pools. We will use it to explore how exposure to microbial, viral, or neoplastic stimuli will modulate the peripheral γδ TCR repertoire in cord blood, pediatric, and adult cohorts. Furthermore, we will establish new methods to perform efficient high throughput analysis of paired TRG and TRD TCR chains. We will share and advance this know-how with the entire DFG Research Unit. In return, new collaborations will enable us to identify how individual γδ TCR recombinations respond to neoplasm and microbial exposure and warrant access to γδ T cells from clinically relevant cancer patient cohorts. In addition, we will combine γδ TCR repertoire analyses and protein biochemistry to identify cognate ligands of responsive γδ TCR clones. We expect that cognate antigens of γδ TCRs may either be directly of microbial or viral origin or danger-indicating self-structures that were induced by tumor formation as well as by microbial or viral infection. Defining how γδ TCRs recognize microbial antigen and/or danger-associated self-ligands will pave the way for the therapeutic manipulation of immune surveillance by γδ T cells.This project proposal is part of the DFG Research Unit FOR 2799 „Receiving and Translating Signals via the γδ T Cell Receptor“ and designed to collaborate with all other projects within the research group.