The role of TCF-1 in early fate decisions of memory T cells

Adoptive T cell immunotherapies entail the ex vivo genetic engineering of autologous T cells to be able to effectively identify and eliminate cancer cells upon reinfusion into cancer patients. These therapies have shown significant clinical success in haematological malignancies and two chimeric antigen receptor (CAR) T cell therapies have thus far been approved by the European Medicines Agency. However, relapse remains a significant issue, partly as a result of poor long-term CAR T cell persistence. Enriching adoptive immunotherapies with memory T cells has been demonstrated to significantly improve T cell persistence and long-term tumour control in both mouse models and clinical trials.
The progressive differentiation model of T cell fate holds that T cell differentiation proceeds via a hierarchical tree beginning with naïve T cells, followed by potentially long-lived but weakly cytotoxic memory T cells and finally short-lived but highly cytotoxic effector T cells. T cell factor 1 (TCF1) is a key transcription factor in the establishment and maintenance of memory T cell character and has gained significant scientific interest in recent years as a factor which could be modulated to increase memory character in CAR T cell infusions. In naïve T cells TCF1 promotes the accessibility of chromatin sites associated with memory fate, but during the course of effector T cell differentiation TCF1 expression is irreversibly lost and many of these chromatin sites become inaccessible. Only those cells which maintain TCF1 expression in the divisions following T cell activation become long-lived memory T cells.
The precise functions of TCF1 in the initial fate determination, maintenance and function of memory T cells remain unclear. Elucidating these functions is of great clinical interest as current CAR T cell manufacturing practices are known to bias cell fate towards effectors, and understanding the mechanisms which can counteract this process could allow the generation of cell products with enhanced therapeutic potential.
Therefore, our aim is to clarify the chromatin structures and transcriptional programs directly regulated by TCF1. We will conditionally knock down TCF1 at the protein level by engineering a modified TCF1 tagged with an artificial degron which directs TCF1 towards rapid degradation by the E3 ubiquitin ligase system upon the administration of an inducing ligand. This will allow us to characterise the effects of acute TCF1 depletion on chromatin accessibility by ATAC-seq, and on nascent mRNA expression by nascent RNA-seq. Unlike DNA-level knockout and mRNA-level knockdown models reported in prior literature, degron-based protein-level knockdowns operate as rapidly as the <24h timescales of nuclear remodelling. This eliminates noise from the longer-term secondary effects of TCF1 knockdown, allowing us for the first time to precisely characterise the primary functions of TCF1 at chosen stages in T cell differentiation. We may also conduct these experiments at the single cell level so as to be able to order cells according to their position in the cell cycle, as this is likely to impact chromatin architectures and transcriptomes. We will conduct these experiments in the human T lymphocyte cell line Jurkat.