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An age-dependent increase in interferon (IFN)-γ expression by intestinal intraepithelial lymphocytes (IELs) contributes to the acquisition of resistance to infection by pathogens. However, how IELs acquire the ability to produce IFN-γ remains to be elucidated. Here, we report that IELs in the small intestine acquire the ability to rapidly produce IFN-γ at two distinct life stages. TCRαβ+ IELs (αβIELs) started producing IFN-γ at 4 weeks of age, within 1 week after weaning. In contrast, TCRγδ+ IELs (γδIELs) started producing IFN-γ at 7 weeks of age. In mice lacking Eγ4, an enhancer of the TCRγ locus (Eγ4−/− mice), Thy-1+ Vγ5+ γδIELs, a major subpopulation of γδIELs, were specifically reduced and their ability to produce IFN-γ was severely impaired, whereas Vγ2+ γδIELs normally produced IFN-γ. In Eγ4−/− mice, TCR expression levels were reduced in Vγ5+ γδIEL precursors in the thymus but unchanged in the Vγ5+ IELs. Nevertheless, TCR responsiveness in Vγ5+ γδIELs was impaired in Eγ4−/− mice, suggesting that the TCR signal received in the thymus may determine TCR responsiveness and the ability to produce IFN-γ in the gut. These results suggest that αβIELs and γδIELs start producing IFN-γ at different life stages and that the ability of Vγ5+ γδIELs to produce IFN-γ in the gut may be predetermined by TCR signalling in IEL precursors in the thymus. [ABSTRACT FROM AUTHOR]

γδ T cells expressing Vγ5Vδ1 TCR originally develop in the embryonic thymus and migrate to the epidermis, forming dendritic epidermal T cells (DETCs) throughout life. It is thought that a TCR signal is essential for their development; e.g. lack of TCR signal-transducer ZAP70 significantly decreases DETC numbers. On the other hand, lack of ZAP70 does not affect Vγ5Vδ1+ T cells in the embryonic thymus; thus, the involvement of TCR signaling remains elusive. Here, we used SKG mice with attenuated TCR signaling rather than gene-knockout mice. In SKG mice, Vγ5+ T cells showed a marked decrease [10% of wild-type (WT)] in adult epidermis; however, there was just a moderate decrease (50% of WT) in the embryonic thymus. In early postnatal epidermis in SKG mice, substantial numbers of Vγ5+ T cells were observed (50% of WT). Their activation markers including CD122, a component of the IL-15 receptor indispensable for DETC proliferation, were comparable to those of WT. However, the Vγ5+ T cells in SKG mice did not proliferate and form DETCs thereafter. Furthermore, in SKG/+ mice, the number of thymic Vγ5Vδ1+ T cells increased, compared to SKG mice; however, the number of DETCs remained significantly lower than in WT, similar to SKG mice. Our results suggest that signaling via Vγ5Vδ1 TCR is indispensable for DETC development, with distinct contributions to embryonic development and postnatal proliferation. [ABSTRACT FROM AUTHOR]

The signal of the IL-7R and signal transducers and activators of transcription (STAT) 5 plays an essential role in γδ T-cell development by inducing V–J recombination in the TCRγ locus. Previously, we have shown that STAT5 binds to the Jγ promoters and controls chromatin accessibility by histone acetylation. However, little is known on control mechanism of Vγ region by the IL-7R. To elucidate the regulation by STAT5, we first analyzed the chromatin status of Vγ region in primary thymocytes. The levels of histone H3 acetylation are high at Vγ5, HsA element and Vγ2 in Rag2−/− thymocytes but low in IL-7R α-chain (IL-7Rα)-deficient early thymocytes, suggesting that IL-7R signaling controls the accessibility of the Vγ region. In addition, high levels of histone H3 acetylation and germ line transcription were induced at Vγ5 and HsA by cytokine and STAT5 in cytokine-dependent Ba/F3 and other hematopoietic cell lines. Importantly, the chromatin accessibility of Vγ5 gene is increased by cytokine signal. Furthermore, STAT5 was not recruited to a non-canonical STAT-binding motif in the endogenous chromatin of the Vγ5 promoter by cytokine stimulation, while STAT5 binds to a consensus motif in the HsA element. In accordance with this result, STAT5 does not directly activate the Vγ5 promoter by reporter assay. These results suggested that while STAT5 directly binds to HsA element and induces its histone acetylation, STAT5 indirectly activates the Vγ5 promoter. Thus, this study implies a potential role of STAT5 in accessibility control of Vγ region, especially at Vγ5 and HsA. [ABSTRACT FROM AUTHOR]

The IL-7R controls local accessibility of joining (J) γ gene segments in the mouse TCRγ locus by recruiting signal transducers and activators of transcription (STAT) 5 and transcriptional coactivators to the Jγ germ line promoters and inducing histone acetylation and germ line transcription. Because STAT consensus motifs are conserved not only in the Jγ promoters but also in the TCRγ 3′ enhancer (Eγ) elements, it is possible that STAT5 interacts with and activates Eγ. To address this question, we first showed that the lysine 4 residue of histone H3 is substantially methylated at Eγ1 and Eγ4 elements in wild-type early thymocytes and that the levels of the methylation are reduced in IL-7R α chain-deficient mice. We also showed that STAT5 has potential to elevate histone acetylation of the Eγ elements in a cytokine-dependent cell line by cytokine stimulation. Next, we demonstrated that STAT5 is recruited to the STAT consensus motifs in the Eγ elements after cytokine stimulation and that transcription factors Runt-related (Runx) and c-Myb are constitutively recruited to Eγ. Furthermore, we showed that STAT5 augments basal Eγ activity controlled by Runx and c-Myb. These results suggest that STAT5 is recruited to the consensus motifs in the Eγ elements by cytokine stimulation and augments basal Eγ activity independent of Runx and c-Myb. Therefore, this study implies that the Eγ elements might be activated in two successive steps, first by Runx and c-Myb and next by STAT5. [ABSTRACT FROM PUBLISHER]

γδ T cells are implicated in autoimmune diseases but their precise function remains unclear. In multiple sclerosis (MS) brain tissue, γδ T cells co-localize with heat shock protein (hsp)65+ oligodendrocytes and are oligoclonally restricted in the Vδ2Jδ3 lineage. To investigate the homing properties and the degree of heterogeneity of Vδ2Jδ3+ γδ T cells in MS, peripheral blood lymphocytes (PBL) from 34 MS patients, 42 controls (14 autoimmune control patients, 28 healthy volunteers), and 11 lymphatic tissues of MS patients and controls were studied by RT-PCR and sequencing. Vδ2Jδ3 TCR rearrangement was detected in 27 out of 28 healthy controls, and was significantly less frequent in MS patients (24 out of 34) and autoimmune control patients (seven out of 14). It was present only in five out of 11 tissue specimens, none of them from MS patients. Direct sequencing and cloning/automated sequencing of the Vδ2Jδ3 PCR products indicated heterogeneity in healthy controls and oligoclonality in MS patients, but also in autoimmune control patients. Differences between MS patients and healthy controls were more accentuated in exacerbating hospitalized patients than in clinically stable outpatients participating in a clinical trial. Only one Vδ2Jδ3 sequence of a total of 85 different sequences obtained was shared between two MS patients. Taken together, evidence for clonal expansion of Vδ2Jδ3+ γδ T cells was found in PBL of MS patients. This T cell subset, previously shown to be present in 100% of chronic-active MS plaques, might home to the CNS in MS, resulting in its under-representation in the blood. [ABSTRACT FROM AUTHOR]

In the mouse thymus, pre-T cells are defined by their CD3−CD4−CD8− triple-negative, CD4410/− CD25+ phenotype. We made a rat mAb IF-7, that, among all Tcell subsets analyzed, reacted exclusively with pre-T cells. Molecular cloning revealed that the antigen recognized by IF-7 was identical to BP-3/BST-1, a glycosyl-phosphatidylinositol-linked, CD38-related molecule previously described asa possible co-activation molecule of pre-B cells. We found that IF-7 cross-linking enhances the proliferative response ofsorted pre-T cells to anti-CD3 stimulation. In addition, IF-7 enhances and accelerates the development of fetal thymic organ culture (FTOC), although the γδ lineage is unaffected by the treatment. In addition, sorted IF-7+ pre-T cells give preferentially rise to αβ TCR+ thymocytes in FTOC. Our observations strongly suggest that BP-3/BST-1 is implicated in both early B and T cell growth and development, and is an early marker for the αβ lineage. [ABSTRACT FROM AUTHOR]