Immune Regulatory Roles of Suppressor Of Cytokine Signaling (SOCS) Molecules

在SOCS家族成员中，SOCS1和SOCS3的作用已经通过基因缺失的小鼠模型进行了广泛的评估。SOCS1和SOCS3之间是否存在冗余尚不清楚，主要是因为目前还没有SOCS1/SOCS3双重缺陷的小鼠模型。因此，我们的目标是利用SOCS1和SOCS3基因缺失的小鼠来培育SOCS1、SOCS3双缺陷小鼠，并用T细胞特异性Cre重组酶转基因小鼠来培育它们。我们已经培育了这些小鼠一年多了，但仍然没有达到预期的基因类型。但是，通过扩大育种，我们希望在短期内获得T细胞特异性SOCS1/SOCS3缺陷小鼠，用于深入分析。除了SOCS1和SOCS3，我们的研究也集中在SOCS4，它仍然是一个特征不佳的SOCS家族，在未成熟的胸腺细胞中大量表达，但在成熟的T细胞中表达下调。我们最近利用基因陷阱ES细胞系统建立了SOCS4缺陷小鼠，并通过实时逆转录聚合酶链式反应证实了这些小鼠中SOCS4基因的表达缺失。SOCS4缺陷小鼠的大体表型分析没有发现发育异常，也没有观察到SOCS4缺陷对其免疫系统的任何不良影响。此外，在SOCS4缺乏的T细胞中，IL-7信号仍然没有受到影响，这表明SOCS4与其他SOCS家族成员存在潜在的冗余。因此，我们也在产生SOCS4，SOCS1双缺乏和SOCS4，SOCS3双缺乏小鼠的过程中来检查这些点。相反，为了检验强制过表达SOCS4是否会影响细胞因子信号，我们产生了T细胞特异性的SOCS4转基因小鼠。在这里，我们发现SOCS4的过度表达抑制了T细胞的发育和分化。具体地说，我们发现转基因SOCS4的表达干扰了T细胞的存活和动态平衡，因此外周淋巴组织中的初始T细胞数量显著减少。然而，SOCS4对T细胞的有害作用的分子基础尚不清楚，因为我们发现在SOCS4转基因的T细胞中细胞因子信号转导没有受到影响。我们之前报道过核因子ThPOK上调T细胞中SOCS1和SOCS3的转录[Luckey MA等人，2014，NAT。免疫。]。由于ThPOK只在CD4T细胞中表达，而在CD8T细胞和未成熟胸腺细胞中不表达，目前尚不清楚SOCS1是如何在CD4、CD8双阳性胸腺细胞或CD8T细胞中诱导表达的。为此，我们最近将兴趣转移到小肠中的CD4+CD8aa+上皮内T细胞，这些T细胞来源于CD4T细胞，然后在肠道中分化为CD4+CD8aa+T细胞。重要的是，CD4T细胞向CD4+CD8aa+T细胞的转化是由ThPOK的丢失和CD8谱系特异性因子Runx3d的获得所介导的。SOCS家族分子在这一过程中的表达变化尚不清楚。但是，我们认为这是一个有趣的新场所，可以深入了解ThPOK/Runx3d表达通路上SOCS家族分子表达的调控过程。因此，我们进行了一系列研究，以了解是什么控制了CD4+CD8aa+上皮内T细胞的产生，我们发现趋化因子受体CCR9是这一过程的新调节因子(Li C.et al.，2022，Mucosal Immunol.)，我们还发现动态平衡的IL-7和IL-15信号是这种T细胞转换的抑制因子(Li C.et al.，2022，Cell Mol Immunol.)。有了这些新的见解，我们目前正在评估SOCS家族成员在肠道T细胞分化和维持中的作用，以及在其他非淋巴组织中的作用。

Among SOCS family members, the roles of SOCS1 and SOCS3 have been extensively assessed using mouse models of genetic deletion. Whether there is a redundancy between SOCS1 and SOCS3 is unclear, mostly because mouse models of SOCS1/SOCS3 double deficiency have not been available. Thus, we aimed to generate SOCS1, SOCS3-double deficient mice by using SOCS1-floxed and SOCS3-floxed mice, and by breeding them with T cell specific Cre recombinase transgenic mice. We have been breeding these mice for more than a year, but still did not achieve the desired genotypes. But, by expanding the breeding, we hope to obtain T cell specific SOCS1/SOCS3-deficient mice anytime soon for their in-depth analyses. In parallel to SOCS1 and SOCS3, our study also has been focused on SOCS4 which remains a poorly characterized SOCS family that is abundantly expressed in immature thymocytes but whose expression is downregulated on mature T cells. We recently generated SOCS4-deficient mice utilizing a gene-trap ES cell system, and we verified the lack of SOCS4 mRNA expression in these mice by real-time reverse transcription PCR. Gross phenotypic analysis of SOCS4-deficient mice did not show abnormalities in their development, and we also did not observe any adverse effect of SOCS4-deficiency in their immune system. Moreover, IL-7 signaling remained unaffected in SOCS4-deficient T cells, suggesting a potential redundancy of SOCS4 with other SOCS family members. Thus, we are also in the process of generating SOCS4, SOCS1 double deficient and SOCS4, SOCS3 double deficient mice to examine these points. Conversely, to examine if the forced overexpression of SOCS4 would affect cytokine signaling, we generated T cell-specific SOCS4-transgenic mice. Here, we found that SOCS4 overexpression suppressed the development and differentiation of T cells. Specifically, we found that transgenic SOCS4 expression interfered with T cell survival and homeostasis so that naive T cell numbers in peripheral lymphoid tissues were significantly reduced. The molecular basis of the detrimental effect of SOCS4 on T cells, however, remains unclear because we found that cytokine signaling was unaffected in SOCS4 transgenic T cells. We previously reported that the nuclear factor ThPOK upregulates the transcription of SOCS1 and SOCS3 in T cells [Luckey MA et al., 2014, Nat. Immunol.]. Because ThPOK is only expressed in CD4 T cells and absent in CD8 T cells and immature thymocytes, it has been unclear how SOCS1 is induced in CD4, CD8 double positive thymocytes or in CD8 T cells. To this end, we have recently shifted our interest to CD4+CD8aa+ intraepithelial T cells in the small intestine, which are derived from CD4 T cells but then differentiate into CD4+CD8aa+ T cells in the gut. Importantly, the conversion of CD4 T cells into CD4+CD8aa+ T cells is mediated by the loss of ThPOK and the acquisition of the CD8 lineage specifying factor Runx3d. How the expression of SOCS family molecules changes during this process is not known. But, we considered this an interesting new venue to gain insights into the regulatory process of SOCS family molecule expression along the ThPOK/Runx3d expression pathway. Thus, we performed a series of studies to understand what controls the generation of CD4+CD8aa+ intraepithelial T cells, and we identified the chemokine receptor CCR9 as a new regulator of this process (Li C. et al., 2022, Mucosal Immunol.), and we also found homeostatic IL-7 and IL-15 signaling as suppressors of this T cell conversion (Li C. et al., 2022, Cell Mol Immunol.). Equipped with these new insights, we are currently assessing the role of SOCS family members in the differentiation and maintenance of T cells in the gut, but also in other non-lymphoid tissues.