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PD1 is an inhibitory protein which interacts with its ligands to restrict T cell function. This interaction may prevent immune clearance of cancers and is a target for cancer immunotherapy. Using Nr4a3-Tocky technology, which tracks T cell receptor (TCR) signalling, we investigated PD1 expression and regulation of T cell activation.
Splenocytes from OTI Nr4a3-Tocky mice were used as a source of T cells. These cells use a fluorescent Timer protein to trace TCR signalling over time. Splenocytes were stimulated with ova peptides of varying affinity and incubated to investigate PD1 expression over time. Murine PDL1-Fc-IgG was used to ligate PD1. PD1 and T cell activation markers were measured by flow cytometry.
In activated T cells, PD1 expression was correlated closely with T cell activation. Increasing the incubation time, stimulant peptide dose and affinity increased the percentage PD1 expression, and the PD1 mean fluorescence intensity. When ligated by PDL1, the proportion of immature TCR-induced Timer protein was reduced, suggesting attenuation in TCR signalling.
Nr4a3-Tocky proved to be effective for investigating the interaction between PD1 and PDL1 in TCR signalling. The close coupling of PD1 expression to T cell activation suggests PD1 functions as a T cell activation marker. Ligation by PDL1 attenuates TCR signalling.
Future work will investigate whether these findings are supported in murine tumour models, to further understand the role of PD1 and PDL1 in cancer development and the impact of immunotherapy, and to understand the role of PD1 on other T cell effector functions.