Thymocyte selection-associated high mobility group box protein TOX is a protein that in humans is encoded by the TOXgene.[5][6][7] TOX drives T-cell exhaustion[8][9] and plays a role in innate lymphoid cell development.[10][11]
Structure
The TOX gene encodes a protein that belongs to a large superfamily of chromatin associated proteins that share an approximately 75 amino acid DNA binding motif, the HMG (high mobility group)-box (named after that found in the canonical member of the family, high mobility group protein 1). Some high mobility group (HMG) box proteins (e.g., LEF1) contain a single HMG box motif and bind DNA in a sequence-specific manner, while other members of this family (e.g., HMGB1) have multiple HMG boxes and bind DNA in a sequence-independent but structure-dependent manner. While TOX has a single HMG-box motif,[7] it is predicted to bind DNA in a sequence-independent manner.[12]
TOX subfamily
TOX is a member of a small subfamily of proteins (TOX2, TOX3, and TOX4) that share almost identical HMG-box sequences.[12] TOX2 has been identified to play a role in the differentiation of T follicular helper cell.[13] TOX2 is thought to be a downstream signal of BCL-6.[13] TOX3 has been identified as a breast cancer susceptibility locus.[14][15] TOX is highly expressed in the thymus, the site of development of T lymphocytes.[10]Knockout mice that lack TOX have a severe defect in development of certain subsets of T lymphocytes.[16]
Function
T cell exhaustion
TOX is necessary for T cell persistence but also drives T cell exhaustion.[17][18][19] An increase in TOX expression is characterized by a weakening of the effector functions of the cytotoxic T cell and upregulation of inhibitory receptors on the cytotoxic T cells.[20][21] TOX promotes the exhausted T cell phenotype through epigenetic remodeling.[20][22]PD-1 is an inhibitory marker on T cells that increases when TOX is unregulated.[20][23][22] This allows for cancerous cells to evade the cytotoxic T cells through upregulated expression of PD-L1.[24]
Effector function
Markers of effector functions that are decreased when TOX is overexpressed are KLRG1, TNF, and IFN-gamma.[8] IFN-gamma and TNF-alpha production are also increased when the Tox and Tox2 genes are deleted.[9] Upregulation of effector function in cells lacking TOX is not always seen and it has been proposed that inhibitory receptor function is separated from effector CD8+ cytotoxic T cell function.[8] T-cell exhaustion does not occur when TOX is deleted from CD8+ T cells, but the cells instead adopt the KLRG1+ terminal effector state and undergo apoptosis, or programmed cell death.[9] It was therefore proposed that TOX prevents this terminal differentiation and instead promotes exhaustion so that the T-cell has a slightly more sustained response.[9]
Cancer & chronic infection
In cancer or during chronic viral infection, T-cell exhaustion occurs when cytotoxic T-cells are constantly stimulated.[8][25] TOX is upregulated in CD8+ T cells from chronic infection when compared to acute infection.[8] Patients with cancer typically have high levels of TOX in their tumor-infiltrating lymphocytes,[8] and anti-tumor immunity is heightened when Tox and Tox2 are deleted.[9] TOX and TOX2-deficient tumor-specific CAR T cells additionally have increased antitumor effector cell function as well as decreased levels of inhibitory receptors.[8]
Activation
NFAT transcription factors are essential for activating TOX in CD8+ T-cells,[8] and it has been suggested that TOX is a downstream target of NFAT.[9] The expression and function of NR4a (a target of NFAT) and TOX are strongly linked with reduced NR4a expression in Tox double knockout T cells and minimized Tox expression in NR4a triple knockout T cells.[9]
T-cell development
TOX is necessary for positive selection in developing thymocytes.[26] Knock out TOX mice shows a requirement of TOX for the CD4 T cell lineage,[26] however CD8 single positive T-cells were still able to develop.[26]
Notch signaling can aid in the development of all innate lymphoid cells, but in TOX-deficient cells, Notch target genes are expressed at low levels, so it is possible that TOX is required for downstream activation of these Notch target genes.[10] TOX was also found to bind Hes1, a Notch target gene, in embryonic kidney cells.[10]
Several ILC3 populations are reduced in the absence of TOX, implicating TOX’s role in their development.[10] In the small intestine, major ILC3 populations are normal in TOX-deficient cells, suggesting that gut ILC3 development may occur independently of TOX.[10] Some ILC3 populations in the gut expand in the absence of TOX.[10]
It has been proposed that NFIL3 and TOX regulate the transition of common lymphoid progenitor to early innate lymphoid progenitor.[11] In NFIL3-deficient mice, the expression of TOX is downregulated, indicating that NFIL3 is directly affecting the expression of TOX which is then acting downstream in ILC development.[11] TOX-deficient mice and NFIL3-deficient mice both lack mature ILCs and ILC progenitors.[11]
^Wilkinson B, Chen JY, Han P, Rufner KM, Goularte OD, Kaye J (March 2002). "TOX: an HMG box protein implicated in the regulation of thymocyte selection". Nature Immunology. 3 (3): 272–280. doi:10.1038/ni767. PMID11850626. S2CID19716719.
^ abCheng Y, Shao Z, Chen L, Zheng Q, Zhang Q, Ding W, et al. (January 2021). "Role, function and regulation of the thymocyte selection-associated high mobility group box protein in CD8+ T cell exhaustion". Immunology Letters. 229: 1–7. doi:10.1016/j.imlet.2020.11.004. PMID33186634. S2CID226948315.
