T-Cell Receptor Signaling: A Delicate Orchestration

T-cell receptors (TCRs) are crucial components of T cell immunity, enabling the detection and subsequent elimination of abnormal, cancerous and infected cells.

Dysfunctional T cell receptor signaling can lead to various immune deficiency or autoimmune diseases. In cancer treatment the study and modification of T cell receptor signaling is important for CAR-T cell therapy as well as modulation of the tumor microenvironment (TME). Modulating T cell activation can be important to avoid rejection of organ or stem cell transplants by the immune response.

The TCR is composed of two subunits, which are non-covalently associated with dimers of cluster of differentiation 3 (CD3) chains ¹. Most T cell receptors are composed of an alpha and beta subunit, which are responsible for recognising antigens presented by major histocompatibility complex (MHC) molecules. The CD3 molecules are important for transducing the activation signals to the T cell following stimulation of the alpha/beta T cell receptor by MHC interaction. A small population of T cells express an alternative gamma/delta T cell receptor. γ/δ T cells differ in their function and activation from α/β T cells. γ/δ T cells normally comprise about 1-5% of circulating lymphocytes but their fraction expands in response to tumors, inflammation, and infection. They recognize antigens independently from MHC and play a role in innate and adaptive immune responses ².

In this blog we focus on signaling of the α/β T cell receptor. α/β T cell receptor signaling is activated following recognition of self or foreign antigen-derived peptides presented by MHC molecules ³. These peptides may be presented either by MHC class I molecules (present on all nucleated cells) or MHC class II molecules (on antigen-presenting cells). In addition to the T cell receptor, CD4 (expressed by T helper cells) or CD8 (expressed on cytotoxic T cells) co-receptors interact with an invariant chain of the MHC molecules. The CD4 co-receptor interacts with MHC class II molecules and the CD8 co-receptor interacts with MHC class I molecules ⁴. These interactions are important to direct cytotoxic T cells to infected or cancerous cells, or direct T helper cells to antigen-presenting cells. Some cancers and viruses have evolved to downregulate MHC class I expression to avoid T cell immunity ⁵.

The interaction of the TCR with antigens presented by MHC leads to the activation of several pathways like Ca2+-Calcineurin-NFAT, PKCθ-IKK-NKκβ, RASGRP1-RAS-ERK1/2 and TSC1/2 – mTOR ⁶. In combination with co-stimulatory receptors these pathways regulate cytokine production, cell proliferation and differentiation.

Initial interaction of the TCR with antigens presented by MHC molecules leads to the activation of several protein tyrosine kinases (PTK), including Src family members LCK and FYN ⁶. LCK and FYN phosphorylate immunoreceptor tyrosine-based activation motifs (ITAMs) on the cytoplasmic side of the CD3 complex ⁷. This leads to recruitment and activation of the Syk family kinase Zap-70 which in turn phosphorylates the transmembrane adaptor protein linker for activation of T cells (LAT) and other effectors, resulting in the activation of phospholipase C-γ1 (PLCγ1) ⁷. PLCγ hydrolyzes phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2), generating second messengers like diacylglycerol (DAG) and inositol trisphosphate (IP3) ⁶. IP3, through a signaling cascade involving Ca2+ release from the endoplasmatic reticulum, leads to the nuclear translocation and activation of nuclear factor of activated T cells (NFAT). ⁷ DAG leads to the activation of Ras proteins, a family of small GTPases, subsequent MAPK and ERK1/2 activation and expression of transcription factors JUN and FOS, which, in a complex with NFAT, contribute to T cell activation by inducing IL-2 and other effectors ⁶. Additionally, DAG activates proteinase PKCθ resulting in phosphorylation of CARMA1 and a formation of a complex with B cell lymphoma/leukemia 10 (BCL10) and mucosa-associated lymphoid tissue translocation protein-1 (MALT1) which ultimately leads to NF-κβ entering the nucleus and activating transcription ⁸.

Effective T cell receptor signaling involves not only the activation of the T cell receptor but co-stimulation of additional cell surface receptors such as CD28, which is activated by binding to its ligands CD80 and CD86 present on antigen-presenting cells. CD28 stimulation amplifies TCR receptor signaling, for example by recruiting phosphatidylinositol-3-kinase (PI3K) which leads to the recruitment of AKT, activation of the mTOR pathway and in turn to prolonged nuclear localization of NFAT ⁶. The mTOR pathway also promotes differentiation of naïve CD4 T cells to T helper 1, 2 and 17 lineages, while inhibiting the differentiation to regulatory T cells (Treg), which have immune suppressive function ⁹.

Defects in, or disruption of, any of the components of the T cell signalling pathway can result in immune deficiencies and diseases, or T cell exhaustion. Various mutations and gene fusions within the TCR signaling pathway have been detected in peripheral T cell lymphoma (PTCL) and angioimmunoblastic T cell lymphoma (AITL), for example mutations involving FYN, PLCγ1, PI3K, ITK or CD28. ¹⁰ Patients with ZAP70 mutations lack circulating CD8+ T cells and have defective CD4+ T cells and mutations in LCK lead to serious immune dysregulation. ¹¹. Interrupted NFAT activation can cause severe combined immunodeficiency ⁶. Furthermore, when NFAT acts on its own, without the JUN/FOS complex, it activates several genes contributing to T cell exhaustion. (1) For example, it induces transcription of CBL-B, an ubiquitin ligase upregulated in exhausted T cells. ¹² CBL-B deficiency has been shown to inhibit T cell exhaustion and CBL-1 depletion has been suggested to improve CAR-T cell function. ¹² Other genetic modifications of the TCR signaling pathways in engineered CAR T cells are being tested to prevent exhaustion, for example the overexpression of c-JUN. ¹³

In addition to receptors that activate TCR signalling, there are also inhibitory receptors such as CTLA-4 and PD-1, which modulate TCR signaling to prevent overactivation of T cells. The dysregulation of this negative feedback plays a role in the pathogenesis of autoimmune diseases like Diabetes type 1, rheumatoid arthritis, Systemic lupus erythematosus, and inflammatory bowel disease ¹⁴ In the tumor microenvironment, on the other hand, this negative feedback is often enhanced. Monoclonal antibodies that inhibit PD-1 or CTLA-4, such as nivolumab, pembrolizumab, and ipilimumab, have been approved for treating several types of cancers, including metastatic melanoma, renal cell carcinoma, and non-small cell lung cancer. ¹⁵

Here at Revvity Cambridge, we have a large collection of cell models across haploid and cancer lines. Using Kegg Pathway Assessment, we compared known genes implicated in T-cell receptor signaling within our catalogue of HAP1 and Cancer cell lines. Check out the list below for gene coverage and links to product pages.

Gene knockouts available in HAP1 Cell Lines
Gene knockouts available in HAP1 and Cancer Cell Lines
Gene knockouts available in Cancer Cell Lines

References

1. R. A. Mariuzza, P. Agnihotri, and J. Orban, “The structural basis of T-cell receptor (TCR) activation: An enduring enigma,” Journal of Biological Chemistry, vol. 295, no. 4. American Society for Biochemistry and Molecular Biology Inc., pp. 914–925, Jan. 24, 2020. doi: 10.1074/jbc.REV119.009411.
2. M. Yazdanifar, G. Barbarito, A. Bertaina, and I. Airoldi, “γδ T Cells: The Ideal Tool for Cancer Immunotherapy,” Cells, vol. 9, no. 5. NLM (Medline), May 24, 2020. doi: 10.3390/cells9051305.
3. T. Thaventhiran, “T Cell Co-inhibitory Receptors-Functions and Signalling Mechanisms,” J Clin Cell Immunol, vol. 01, no. S12, 2013, doi: 10.4172/2155-9899.s12-005.
4. J. A. L. J. et al. Alberts B, “T Cells and MHC Proteins.,” in Molecular Biology of the Cell., 4th ed., New York: Garland Science , 2002. Accessed: Jun. 19, 2024. [Online]. Available: https://www.ncbi.nlm.nih.gov/books/NBK26926/
5. B. C. Taylor and J. M. Balko, “Mechanisms of MHC-I Downregulation and Role in Immunotherapy Response,” Frontiers in Immunology, vol. 13. Frontiers Media S.A., Feb. 28, 2022. doi: 10.3389/fimmu.2022.844866.
6. K. Shah, A. Al-Haidari, J. Sun, and J. U. Kazi, “T cell receptor (TCR) signaling in health and disease,” Signal Transduction and Targeted Therapy, vol. 6, no. 1. Springer Nature, Dec. 01, 2021. doi: 10.1038/s41392-021-00823-w.
7. J. R. Hwang, Y. Byeon, D. Kim, and S. G. Park, “Recent insights of T cell receptor-mediated signaling pathways for T cell activation and development,” Experimental and Molecular Medicine, vol. 52, no. 5. Springer Nature, pp. 750–761, May 01, 2020. doi: 10.1038/s12276-020-0435-8.
8. J. Schulze-Luehrmann and S. Ghosh, “Antigen-Receptor Signaling to Nuclear Factor κB,” Immunity, vol. 25, no. 5. pp. 701–715, Nov. 2006. doi: 10.1016/j.immuni.2006.10.010.
9. H. Chi, “Regulation and function of mTOR signalling in T cell fate decisions,” Nature Reviews Immunology, vol. 12, no. 5. pp. 325–338, May 2012. doi: 10.1038/nri3198.
10. X. Liu, J. Ning, X. Liu, and W. C. Chan, “Mutations Affecting Genes in the Proximal T-Cell Receptor Signaling Pathway in Peripheral T-Cell Lymphoma,” Cancers, vol. 14, no. 15. MDPI, Aug. 01, 2022. doi: 10.3390/cancers14153716.
11. L. D. Notarangelo, “Immunodeficiency and immune dysregulation associated with proximal defects of T cell receptor signaling,” Current Opinion in Immunology, vol. 31. Elsevier Ltd, pp. 97–101, Dec. 01, 2014. doi: 10.1016/j.coi.2014.10.003.
12. J. Kumar et al., “Deletion of Cbl-b inhibits CD8 + T-cell exhaustion and promotes CAR T-cell function,” J Immunother Cancer, vol. 9, no. 1, Jan. 2021, doi: 10.1136/jitc-2020-001688.
13. R. C. Lynn et al., “c-Jun overexpression in CAR T cells induces exhaustion resistance,” Nature, vol. 576, no. 7786, pp. 293–300, Dec. 2019, doi: 10.1038/s41586-019-1805-z.
14. J. H. Buckner, “Mechanisms of impaired regulation by CD4+ CD25+ FOXP3+ regulatory T cells in human autoimmune diseases,” Nature Reviews Immunology, vol. 10, no. 12. pp. 849–859, Dec. 2010. doi: 10.1038/nri2889.
15. E. I. Buchbinder and A. Desai, “CTLA-4 and PD-1 pathways similarities, differences, and implications of their inhibition,” American Journal of Clinical Oncology: Cancer Clinical Trials, vol. 39, no. 1. Lippincott Williams and Wilkins, pp. 98–106, 2016. doi: 10.1097/COC.0000000000000239.