The role of T cell receptor affinity in CD4+ T cell differentiation

Abstract

Naïve helper T cells become activated when their T cell receptor (TCR) recognizes a microbial peptide presented on MHCII (p:MHCII) by dendritic cells (DCs). During this interaction, DCs provide polarizing cues to guide T cell differentiation towards specific fates, like T follicular helper cells which help B cells. The affinity of TCR for its cognate p:MHCII is known to influence the fate adopted by CD4+ T cells, but the mechanisms responsible for this effect are not completely understood. The mechanism could be T cell extrinsic, by affecting the DC subset a T cell contacts thereby influencing the polarizing factors received, or T cell intrinsic, by modulating TCR signal strength thus regulating genes involved in T cell differentiation. The T cell extrinsic hypothesis was tested with a quantitative, imaging approach called histo-cytometry to analyze the T cell-DC interactions. This technique involves time-consuming manual image analysis. Therefore, we wrote software to automate histo-cytometry analysis, reducing hands-on analysis time by up to 90%. With histo-cytometry, we determined that TCR affinity did not affect the DC subset contacted by CD4+ T cells, rendering the cell extrinsic hypothesis unlikely. We therefore focused on the T cell intrinsic hypothesis by performing a CRISPR/Cas9 screen to identify TCR regulated genes that influence T cell differentiation. This screen revealed two novel regulators of helper T cell differentiation, Eef1e1 and Gbp2, which are expressed at a higher level in high affinity than low affinity T cells. These results suggest that TCR affinity influences helper T cell differentiation intrinsically by regulating genes that control the differentiation process. We next explored the relationship between CD4+ T cell differentiation and cytotoxic activity. This study determined that cytotoxic function of helper T cells was dependent on the Fas pathway and Fas was upregulated on target cells proportional to the TCR affinity of the responding T cells. Additionally, we identified that many different helper T cell subsets express FasL and have cytotoxic potential. Thus, my work could benefit future vaccines by providing greater understanding of how helper T cell fate decisions occur and how these decisions influence cytotoxic function.