Regulation of cyclin D3 in B lymphocyte development

描述（由申请方提供）：B细胞发育是一个高度有序和受调控的过程，在此过程中，谱系定型前B细胞产生不同的外周克隆B淋巴细胞群，显示出广泛的抗原特异性。在B淋巴细胞生成的早期，Igm与替代轻链和伊加/IgB组装形成前BCR，其首先扩增携带一个框内重链重排的前B细胞群，然后启动轻链重排。在以前的研究中，我们证明了细胞周期蛋白D3是前B细胞克隆扩增所唯一需要的（Nat Immunol 7：489）。细胞周期蛋白D3的积累和细胞周期的启动受GC依赖性信号的协调调节，GC依赖性信号增强Ccnd 3 mRNA和通过前BCR的信号增加核细胞周期蛋白D3的可用性。在初步结果中，以及在目前出版的工作中（Nat Immunol），我们证明Ras/MEK/ERK信号传导途径的前BCR依赖性激活协调Ccnd 3转录的终止与Igk重组的起始。ERK下游，特定的信号传导效应物介导每个离散的发育事件。通过ERK介导的Id蛋白的抑制和E2 A的去抑制诱导Igk转录，而Aiolos的诱导沉默Ccnd 3，导致细胞周期退出。除了确定协调B细胞发育的中心途径外，这些发现还为理解前BCR和IL-7依赖性信号之间的相互作用提供了框架。具体来说，我们证明，Igk转录E2 A和STAT 5之间的竞争控制的内含子Igk增强子（Eki）。相比之下，细胞周期蛋白D3蛋白由PI-3激酶调节，PI-3激酶通过一种新的淋巴特异性机制控制细胞周期蛋白D3相对于抑制剂p27 kip 1和CDK 4/Rb效应复合物的亚核位置。这些观察结果引起了一个模型，其中初始前B细胞增殖是由STAT 5和PI-3 k的协调激活介导的。STAT 5增强Ccnd 3转录并抑制Igk重组，而PI-3 k驱动生产性细胞周期蛋白D3/CDK 4/Rb复合物的形成。随后的Igk重组需要从STAT 5逃逸和通过前BCR激活ERK。该模型的核心预测将在以下具体目标中进行检验：目标1。研究E2 A和STAT 5如何整合调节Igk基因转录。目标二。研究PI-3激酶如何调节细胞周期蛋白D3的核定位以及与CDK 4、Rb和p27 kip 1的组装。目的3：确定，在体内，为什么细胞周期蛋白D3作出独特的贡献淋巴细胞的发展。 公共卫生相关性：骨髓中B细胞的发育由骨髓中存在的细胞因子和注定成为B细胞的淋巴细胞前体上的前B细胞抗原受体（前BCR）的表达决定。在这项拨款申请中，我们将确定细胞内信号如何通过前BCR和细胞因子受体传递，整合到直接复杂的发育过程中。这些研究将导致对导致白血病的免疫系统过程的产生有更好的理解。

DESCRIPTION (provided by applicant): B cell development is a highly ordered and regulated process during which lineage-committed pro-B cells give rise to diverse peripheral populations of clonal B lymphocytes displaying a broad repertoire of antigen specificities. Early in B lymphopoiesis, Igm assembles with surrogate light chain and Iga/Igb to form a pre-BCR that first expands pre-B cell populations bearing one in-frame heavy chain rearrangements and then initiates light chain rearrangement. In previous studies, we demonstrated that cyclin D3 is uniquely required for clonal expansion of pre-B cells (Nat Immunol 7:489). The accumulation of cyclin D3, and the initiation of cell cycle, is coordinately regulated by gc-dependent signals that enhance Ccnd3 mRNA and signals through the pre-BCR that increase the availability of nuclear cyclin D3. In Preliminary Results, and in work now in press (Nat Immunol), we demonstrate that pre-BCR dependent activation of the Ras/MEK/ERK signaling pathway coordinates the termination of Ccnd3 transcription with the initiation Igk recombination. Downstream of ERK, specific signaling effectors mediate each discrete developmental event. Igk transcription is induced by ERK- mediated suppression of the Id proteins, and derepression of E2A, while induction of Aiolos silences Ccnd3 leading to cell cycle exit. In addition to identifying a central pathway coordinating B cell development, these findings provide a framework for understanding the interplay between pre-BCR and IL-7 dependent signals. Specifically, we demonstrate that Igk transcription is controlled by competition between E2A and STAT 5 at the intronic Igk enhancer (Eki). In contrast, cyclin D3 protein is regulated by PI-3 kinase that, through a novel, lymphoid specific mechanism, controls the subnuclear location of cyclin D3 relative to the inhibitor p27kip1 and the CDK4/Rb effector complex. These observations evoke a model in which initial pre-B cell proliferation is mediated by the coordinated activation of STAT 5 and PI-3k. STAT 5 enhances Ccnd3 transcription and suppresses Igk recombination while PI-3k drives the formation of productive cyclin D3/CDK4/Rb complexes. Subsequent Igk recombination requires both escape from STAT 5 and activation of ERK by the pre-BCR. The central predictions of this model will be tested in the following Specific Aims: Aim 1. To determine how E2A and STAT 5 integrate to regulate Igk transcription. Aim 2. To determine how PI-3 kinase regulates cyclin D3 nuclear localization and assembly with CDK4, Rb and p27kip1. Aim 3: To determine, in vivo, why cyclin D3 makes a unique contribution to lymphocyte development. PUBLIC HEALTH RELEVANCE: The development of B cells in the bone marrow is determined by both cytokines present in the bone marrow and by expression of the pre-B cell antigen receptor (pre-BCR) on lymphocyte precursors destined to become B cells. In this grant application, we will determine how intracellular signals, delivered through the pre-BCR and cytokine receptors, integrate to direct complex developmental processes. These studies will lead to a greater understanding of both the generation of the immune system of those processes that lead to leukemia.