Role of Nuclear Factor TOX in Germinal Center Reactions

描述（由申请人提供）：产生高亲和力保护性抗体反应需要T细胞和B细胞在次要淋巴组织的特定区域（称为生发中心）协同工作。了解这些细胞相互作用的调控对于疫苗开发和理解生发中心淋巴细胞在肿瘤发展和自身免疫中的作用是重要的。T滤泡辅助细胞（TFH）是一种特化的CD4 T细胞，能够迁移到滤泡中并向B细胞传递辅助信号，因此在体液免疫反应的功能成熟中起关键作用。基因表达的全局分析表明，生发中心B细胞和TFH都具有独特的基因表达模式，将它们与其他原始和活化淋巴细胞亚群区分开来。据报道，在生发中心B细胞和TFH中特异性表达的一个基因编码TOX，这是一种在啮齿动物和人类之间高度保守的蛋白质，在我们的实验室中首次被确定为T细胞发育的调节剂。我们已经生产了TOX转基因小鼠和全球和有条件的TOX缺陷小鼠，并表明TOX是CD4 T细胞谱系发育所必需的。使用这些转基因动物的组合，我们在这里提出确定TOX是否也在生发中心反应中发挥作用。我们的方法使我们能够区分tox缺乏对B细胞和T细胞的影响。因此，将条件性tox缺陷小鼠培育为表达Cre重组酶的B细胞谱系小鼠，将用于寻找免疫后生发中心B细胞形成和功能的缺陷。提出了两种互补的方法来研究TOX在TFH发育和功能中的类似作用；用胸腺表达的TOX转基因对全球TOX缺陷小鼠进行互补，并在体外通过cre介导的CD4+ T细胞中TOX位点的缺失，随后在体内测试TFH的发育和功能。为了补充这些研究，我们还建议在小鼠中产生TOX报告敲入菌株，这将使我们能够在免疫反应期间跟踪活细胞和体内TOX的表达。

DESCRIPTION (provided by applicant): The generation of a high affinity protective antibody response requires T and B cell collaboration in specialized areas of secondary lymphoid tissue known as germinal centers. Understanding the regulation of these cellular interactions is important for vaccine development and for understanding the role of germinal center lymphocytes in tumor development and autoimmunity. T follicular helper cells (TFH) are specialized CD4 T cells that are able to migrate into follicles and deliver helper signals to B cells, and thus play a critical role in functional maturation of a humoral immune response. Global analysis of gene expression has shown that both germinal center B cells and TFH have unique patterns of gene expression that distinguish them from other subsets of naive and activated lymphocytes. One of the genes reported to be expressed specifically in both germinal center B cells and TFH encodes TOX, a protein highly conserved between rodents and humans and first identified in our laboratory as a regulator of T cell development. We have produced TOX transgenic mice and globally and conditionally TOX-deficient mice and have shown that TOX is required for development of the CD4 T cell lineage. Using combinations of these genetically modified animals, we propose here to determine whether TOX also plays a role in germinal center reactions. Our approaches allow us to distinguish the effects of TOX-deficiency on B and T cells. Thus, breeding conditionally TOX-deficient mice to mice that express Cre recombinase in the B cell lineage will be used to look for defects in germinal center B cell formation and function upon immunization. Two complementary approaches are proposed to study similarly the role of TOX in TFH development and function; complementation of globally TOX-deficient mice with a thymically expressed TOX transgene and in vitro Cre-mediated deletion of the Tox locus in CD4+ T cells that are subsequently tested for TFH development and function in vivo. To complement these studies we also propose to generate a TOX reporter knock-in strain of mice that will allow us to track expression of TOX in viable cells and in vivo during an immune response. PUBLIC HEALTH RELEVANCE: A protective antibody-mediated immune response requires T and B lymphocyte interactions in specialized areas of secondary lymphoid tissue known as germinal centers. Understanding the regulation of these interactions is important for vaccine development and for understanding the role of germinal center lymphocytes in tumor development and autoimmunity. This research program is directed at understanding the role of a specific nuclear protein, highly conserved in rodents and humans, in regulating these germinal center reactions.