BCR MEDIATED SIGNALING AND CYCLING B CELL BIOLOGY

抗原驱动的B细胞反应发生在次级淋巴组织中。 活跃的周期B细胞迁移到生发中心的毛囊 (GC)反应发展。由于循环GC B细胞被编程为 经历细胞凋亡，它们必须获得生存和/或增殖 来自GC环境的信号才能生存。目前的研究 利用体外模型系统探索循环B的生物学 细胞。为此，从小的静息B细胞中分离出 小鼠的脾在批量培养中被激活2或4天 脂多糖，3步恢复活跃循环的B细胞 Percoll渐变。保护自行车运动所需的第一个生存信号 B细胞持续进入凋亡是通过占用 Bcr抗原。这是这一进程的第一步 亲和力成熟，因为只有最适合的克隆才能竞争或 限制性抗原。在第一个具体目标中，概述了实验 为了检验CD19/CD21共同受体在 在将这一最初的生存信号传递给循环中的B细胞的过程中发挥作用。 为此，将重新刺激循环中的HEL转基因B细胞 用可溶的或固定化的HEL或HEL-C3d融合蛋白。自C3d以来 是CD21的配体，这种融合蛋白有效地招募了 CD19/CD21共同受体定位于BCR。我们之前的研究 已经通过循环B表示的BCR显示了信号的强度 细胞决定了Bc l-2家族的表达模式。骑自行车B 细胞表达促凋亡蛋白Bax和Bclxs。交联性 可溶性F(ab‘)2抗MU对B细胞BCR的诱导作用 抗细胞凋亡蛋白BclxL的出现，同时优化杂交. 固定化抗Mu抗体连接bcr同时诱导Bclxl和Bcl2 抗细胞凋亡蛋白。第二个具体说明中描述的实验 AIM将研究这些不同BCR的生物学后果- 生存/死亡蛋白的诱导模式。我们之前的研究已经 还表明通过循环B表示的通过BCR的信号强度 细胞决定了它们对Th细胞衍生信号的反应能力。 第三个特定目标的实验将检验这样的假设 BCR派生的信号在一定程度上通过调节 CD40信号复合体的组成。总而言之，这些实验 将扩大我们对外部信号如何影响命运的理解 循环B细胞。

Antigen driven B cell responses occur within secondary lymphoid tissues. Actively cycling B cells migrate into follicles where germinal center (GC) responses develop. Since cycling GC B cells are programmed to undergo apoptosis, they must receive viability and/or proliferative signals from the GC environment in order to survival. The present study utilizes an in vitro model system to explore the biology of cycling B cells. For this purpose, small resting B cells isolated from the spleens of mice are activated in bulk culture for 2 or 4 days with lipopolysaccharide, and the actively cycling B cells recovered on 3-step Percoll gradients. The first survival signal required to protect cycling B cells from continuing into apoptosis is delivered by occupancy of the BCR by antigen. This represents the first step in the process of affinity maturation, since only the best fit clones can compete or limiting antigen. In the first specific aim, experiments are outlined to test the hypothesis that the CD19/CD21 co-receptor plays an important role during delivery of this initial survival signal to cycling B cells. For this purpose, cycling HEL transgenic B cells will be restimulated with soluble or immobilized HEL or a HEL-C3d fusion protein. Since C3d is the ligand for CD21, this fusion protein effectively recruits the CD19/CD21 co-receptor to sites of occupied BCR. Our previous studies have shown that strength of signal through BCR expressed by cycling B cells determines the pattern of Bcl-2 family expression. Cycling B cells express the pro-apoptotic proteins Bax and Bcl-XS. Cross-linking BCR on cycling B cells with soluble F(ab')2 anti-mu induces the appearance of the anti-apoptitic protein Bcl-XL, while optimal cross- linking of BCR by immobilized anti-mu induces both Bcl-XL and Bcl-2 anti-apoptotic proteins. Experiments described in the second specific aim will examine the biological consequences of these different BCR- induced patterns of survival/death proteins. Our previous studies have also shown that strength of signal through BCR expressed by cycling B cells determines their ability to respond to Th cell-derived signals. Experiments in the third specific aim will test the hypothesis that the BCR-derived signals accomplish this, in part, by regulating the composition of the CD40 signaling complex. In sum, these experiments will expand our understanding of how external signals impact on the fate of cycling B cells.