AKNA, germinal center reaction and immunity.

超出提供的空间。体液免疫反应是由次级淋巴器官富含T细胞区域的B细胞激活，导致生发中心(GC)前体募集到次级毛囊，在那里分化为记忆细胞而启动的。B细胞的激活需要同源受体/配体的相互作用，其中T细胞CD40配体(CD154)与Bcetl-CD40的连接为细胞的增殖、存活和B细胞记忆的诱导提供了必不可少的刺激。本研究主要研究生发中心B淋巴细胞表达的AT-Hook蛋白AKNA，它与富含A/T的CD40和CD154启动子调控元件结合并调控其转录。与此一致，小鼠AKNA(MoAKNA)也是一种核蛋白，由B和T淋巴细胞表达，具有AT-钩状DNA结合基序，上调CD154，诱导CD40表达。总体而言，这些发现表明AKNA在调节GC内的Ag依赖反应中发挥作用。其具体目的是：1.体外研究AKNA在基因转录中的作用。利用小鼠模型，我们将确定moAKNA在CD40、CD154和其他基因靶点表达中的作用。我们将研究moAKNA介导的基因调控机制，并评估其DNA结合要求和特异性。2.为了检测树突状细胞(DC)是否表达moAKNA，以及它是否在DC的成熟和功能中发挥作用，我们将在依赖Fit3配体(FItL)的体内DC动员和不同阶段特异性DC亚群的分选后分析moAKNA的表达。我们还将检查AKNA的表达是否随着抗原提呈细胞(APC)表型的获得以及CD40和CD154.3的表达而达到峰值。在体内评估AKNA功能的意义。将产生AKNA缺陷小鼠，以阐明其在免疫反应中的功能。我们将检查免疫后二次免疫反应的发展，检查共刺激分子的表达，生发中心的形成和抗体的产生。淋巴结构、细胞增殖、表型、CD40和CD154表达的组织学分析以及血清免疫球蛋白的测定将评估功能成熟B淋巴细胞库的存在或缺失。将检测DC的成熟度及其以抗原和CD40/CD154依赖的方式刺激T细胞的能力。将测试胸腺内T细胞的分化、选择及其归巢到次级卵泡并提供同源帮助的能力。AKNA缺陷细胞的免疫功能将在体内和体外检测其对Ag刺激的反应、增殖、产生抗体和/或细胞因子、对生存/死亡信号的反应以及形成免疫记忆的能力。表演网站========================================Section End===========================================

EXCEED THE SPACE PROVIDED. Humoral immune responses are initiated by the activation of B cells in the T cell-rich areas of secondary lymphoid organs that lead to the recruitment of germinal center (GC) precursors into secondary follicles, where they differentiate into memory cells. B cell activation requires cognate receptor/ligand interactions, among which, the ligation of B cetl- CD40 by the T celI-CD40 ligand (CD154) provides the essential stimuli for cell proliferation, survival, and induction of B cell memory. The present study focuses on the AT-hook protein AKNA that is expressed by germinal center B- lymphocytes, binds to A/T-rich regulatory elements of CD40 and CD154 promoters and regulates their transcription. In keeping with this notion, mouse AKNA (moAKNA) is also a nuclear protein that is expressed by B and T lymphocytes, exhibits the AT-hook DNA-binding motifs, upregulates CD154 and induces the expression of CD40. Collectively these findings suggest that AKNA plays a role in the regulation of Ag-dependent responses within GCs. The specific aims are: 1. To assess in vitro the role of AKNA in gene transcription. Using the murine model, we shall determine moAKNA's role in the expression of CD40, CD154 and other gene targets. We shall study the mechanisms of moAKNA-mediated gene regulation and assess its DNA-binding requirements and specificity. 2. To examine whether moAKNA is expressed by dendritic cells (DC) and whether it plays any role in DC maturation and function, moAKNA expression will be analyzed following in vivo Fit3 ligand(FItL)-dependent DC mobilization, and sorting of distinct stage-specific DC subsets. We shall also examine whether AKNA expression peaks with the acquisition of the antigen presenting cell (APC) phenotype and the expression of CD40 and CD154.3. To assess in vivo the significance of AKNA's function. AKNA-deficient mice will be generated to elucidate its function in the immune response. We shall examine the development of secondary immune responses after immunization, examine the expression of costimulatory molecules, formation of germinal centers and antibody production. The histological analysis of lymphoid architecture, cell proliferation, phenotype, CD40 and CD154 expression, and determination of serum Ig will assess the presence or absence of functional mature B lymphocyte repertoires. DC maturation and their ability to stimulate T cells in an Ag and CD40/CD154-dependent manner will be examined. Intrathymic T cell differentiation, selection, and their capacity to home to the secondary follicles and provide cognate help will be tested. Immune functions of AKNA-deficient cells will be examined in vivo and in vitro for their capacity to respond to Ag stimuli, proliferate, produce antibodies and/or cytokines, respond to survival/death signaling, and develop immunological memory. PERFORMANCE SITE ========================================Section End===========================================