TOX, A Novel Regulator of Thymocyte Selection

描述(由申请人提供)：越来越清楚的是，胸腺的工作包括比最初意识到的更复杂的情况。这个器官不仅是CD4和CD8T细胞的生成器，也是自然杀伤T(NKT)细胞、上皮内T淋巴细胞、调节性T(Treg)细胞和先天类CD8T细胞高度专门化谱系的发育场所。控制这些不同的T细胞亚系与普通前体细胞分化的详细分子机制仍有待确定。在这里，我们重点研究TOX(胸腺细胞选择相关HMG盒蛋白)在这方面的作用，TOX是一种核DNA结合蛋白，在我们的实验室中首次被确定为胸腺选择事件的调节因子。我们培育了TOX缺陷小鼠，证明了TOX对于规范的CD4T细胞谱系的发育以及NKT和Treg细胞的发育都是必需的。在这些突变动物中，对T细胞发育的特定阶段的抑制以前没有报道过，这使得对这一过程有了新的见解。此外，我们还发现，TOX在胸腺之外发挥作用，包括在NK细胞发育和淋巴器官形成中起着不可或缺的作用。我们在这里提出了三个具体的目标来描述TOX在免疫系统发育中的作用和作用机制。首先，我们将研究TOX与调节胸腺中CD4和ThPOK表达的关系，后者是一种主要的CD4谱系承诺因子。此外，我们提出了一种新的方法来识别参与建立CD4谱系基因计划的TOX依赖基因，并将比较TOX在不同T细胞谱系发育中的作用。其次，我们将分析TOX在淋巴器官发生发展中的作用，重点分析TOX调节淋巴组织诱导细胞发育的潜力。我们还复制了在培养系统中没有TOX的情况下对NK细胞发育的抑制。我们建议对TOX在这一背景下的作用进行详细的分析，包括TOX是否可能调节另一个与TOX具有某些功能特性的核因子。第三，我们提出了在体内CD4T细胞发育的背景下，识别TOX的直接基因靶点和蛋白结合伙伴的最先进的方法。毒素在啮齿动物和人类之间高度保守，其表达模式也是如此。因此，这些研究也应该为人类免疫系统的工作机制提供洞察力。TOX定义了一个小的蛋白质亚家族，包括另外三个成员，其中一个被认为是乳腺癌的易感基因，而TOX本身在一些癌细胞和肿瘤中高度表达。因此，了解TOX的作用机制可能会影响人类健康和疾病。

DESCRIPTION (provided by applicant): It has become increasingly clear that the workings of the thymus include greater complexity than first appreciated. Not only is this organ a generator of CD4 and CD8 T cells, but it is also a site for development of highly specialized lineages of natural killer T (NKT) cells, intraepithelial T lymphocytes, regulatory T (Treg) cells, and innate-like CD8 T cells. The detailed molecular mechanisms that control the differentiation of these distinct T cell sublineages from common precursors remain to be determined. Here we focus on the role of TOX (thymocyte selection-associated HMG box protein) in this context, a nuclear DNA-binding protein first identified in our laboratory as a regulator of thymic selection events. We produced TOX-deficient mice, demonstrating that TOX is required for development of the canonical CD4 T cell lineage as well as NKT and Treg cell development. Inhibition of the specific stage of T cell development that is affected in these mutant animals has not been reported previously, allowing novel insights into the process. In addition, we have found that TOX plays roles outside the thymus, including an obligatory role in NK cell development and lymph node organogenesis. We propose here three specific aims to delineate the role and mechanism of action of TOX in development of the immune system. First, we will investigate the relationship between TOX and regulation of CD4 and ThPOK expression in the thymus, the latter a dominant CD4 lineage commitment factor. In addition, we propose a novel approach to identify TOX-dependent genes involved in establishment of the CD4 lineage gene program, and will compare the role of TOX in development of different T cell lineages. Second, we will analyze the role of TOX in development of lymph node organogenesis, focusing on the potential for TOX to regulate lymphoid tissue inducer cell development. We also have replicated the inhibition of NK cell development in the absence of TOX in a culture system. We propose detailed analysis of the role for TOX in this context, including whether TOX might function to regulate another nuclear factor that shares some functional properties with TOX. Third, we propose state-of-the-art approaches to identify direct gene targets and protein binding partners of TOX, in an in vivo context of CD4 T cell development. TOX is highly conserved between rodents and humans, as is its expression pattern. Thus, these studies should provide insight into workings of the human immune system as well. TOX defines a small protein subfamily that includes three other members, one of which has been implicated as a breast cancer susceptibility locus, while TOX itself is highly expressed in some cancer cell lines and tumors. Understanding the mechanism of action of TOX is therefore likely to impact both human health and disease.