Abstract
T lymphocytes are critical components of an effective adaptive immune response, with different subsets performing proinflammatory and immuno-suppressive roles. In response to foreign antigens, naïve T cells activate expand and differentiate into effector T cells which mediate pathogen elimination. During the aftermath of pathogen clearance, a subset of effector T cells revert back to quiescence and differentiate into memory T cells. The Forkhead box O transcription factors FOXO1 and FOXO3 (to lesser extent) are essential for naïve and memory T cell differentiation. However, FOXO1 and FOXO3 function is not restricted to quiescent T cells; there is increasing evidence for a role in differentiation of effector T cells and in innate lymphocytes. The molecular mechanisms of how FOXO1 and FOXO3 control these functions is still poorly defined.In this thesis, we examine the relative expression of FOXO1 and FOXO3 in different lymphocyte populations and investigate exactly how FOXO1 and FOXO3 control lymphocyte functions. We use two novel knockin mouse lines developed by the Cantrell laboratory that contain LoxP flanked Foxo1-GFP and Foxo3-GFP reporter genes in the endogenous loci. This gave us the unique opportunity to accurately quantify and compare FOXO1 and FOXO3 expression and subcellular localisation in immune cells at a single cell resolution, as well as to functionally test the consequence of FOXO1 or FOXO3 deletion in T cells.
Using the FOXO1-GFP and FOXO3-GFP knockin mice we quantified and compared FOXO1 and FOXO3 expression during T cell development and in peripheral T cells ex vivo. We show that the expression of FOXO1 increases with thymocyte development and is abundantly expressed in conventional naïve and effector T lymphocytes. Conversely, we found that FOXO3 is highly expressed in thymic progenitors and it is preferentially expressed in ‘innate’ like lymphocytes; invariant NKT (iNKT) cells and intraepithelial lymphocytes (IEL).
Using the FOXO1-GFP knockin mouse, we studied T cell antigen receptor (TCR) control of FOXO1 nuclear localisation at a single cell resolution in vitro. We revealed that FOXO1 nuclear export is digital and is highly dependent on the quality and quantity of the TCR ligand. We show that FOXO1 is indeed expressed in CD8+ cytotoxic T lymphocytes (CTL) but repressed by the P1(3,4,5)P3/PDK1/AKT signalling pathway. Interestingly FOXO1 is not regulated by glucose or amino acid availability in CTL, which are essential regulators of CTL growth, proliferation and effector function.
Using FOXO1GFPflx/GFPflx Granzyme B cre mice, we investigated whether deletion of FOXO1 expression in activated T cells would modify the in vivo anti-tumour T cell response. We found that loss of FOXO1 expression in effector T cells reduced the growth rate of tumours and that FOXO1-deficient tumour infiltrating CD8+ T cells were bigger in cell size and more granular than FOXO1 expressing CD8+ T cells. Thus, we propose a potential therapeutic strategy where targeting FOXO1 activity may enhance T cell responses to tumour antigens, improving the clearance of malignant tissues.
Although FOXO3 is highly expressed during T cell development, loss of FOXO3 expression in early thymic progenitors did not impair the development of conventional T or iNKT cells. Global transcriptome analyses in Foxo3 null hepatic CD4+ T and iNKT cells revealed a very small percentage of transcripts to be deregulated in these T cell subsets, suggesting that FOXO3 is dispensable for iNKT development and function.
Overall, these results suggest that FOXO1 and FOXO3 are not co-expressed equally in different T cell subsets and have distinct and nonredundant roles in controlling T cell function.
| Date of Award | 2019 |
|---|---|
| Original language | English |
| Awarding Institution |
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| Sponsors | Medical Research Council |
| Supervisor | Doreen Cantrell (Supervisor) |