Intracellular Signaling by the Precursor B Cell Receptor

描述（由申请方提供）：前体B细胞受体（preBCR）转导的信号是B细胞发育所必需的。前BCR组分遗传缺陷的人和小鼠患有前B和更成熟的B细胞的缺乏或缺乏，这是导致无丙种球蛋白血症状态和由此产生的严重免疫缺陷的原因。下游前BCR信号转导分子的失调或功能丧失突变与B细胞白血病和淋巴瘤相关。前BCR信号传导的一个神秘方面是受体信号传导被激活的生理手段。配体依赖性和配体非依赖性机制的证据已在细胞培养系统中得到证实，并被认为是生理性的。这两种模型的共同点是，受体定位于细胞表面是至关重要的，无论是参与配体还是驻留在那里的关键信号转导分子，因为表面表达与受体活性直接相关。与此模型相反，我们最近已经证明，前BCR复合物直接远离细胞膜，并重新分布在细胞内，后ER室是一样或几乎一样的活性野生型前BCR。这是通过将内膜定位序列附加到形成前BCR的免疫球蛋白μ重链（IG HC）的胞质尾区，并在前BCR缺陷型转化细胞和原代前B细胞中检测所得的“重定向”前BCR活性来实现的。这些数据支持了前BCR不需要细胞表面的细胞外配体来进行信号传导的模型，并且前BCR在ER后区室（如高尔基体）中具有活性。然而，与preBCR信号传导相关的几个发育重要过程，如等位基因排斥，无法进行评价，这些过程仍然可能需要依赖于preBCR表面表达的活动。因此，本小R03提案中的实验目的是通过评价这些重定向的preBCR复合物的体内生物活性来建立这些发现。通过建立表达重定向IG HC的转基因小鼠，并评价其支持B细胞发育的能力，以及在遗传学上preBCR缺陷和preBCR充足的小鼠中实现等位基因排除，将在一个目标中实现这一目标。这些信息对于更好地理解在正常免疫发育和B细胞病理中决定受体活性的分子机制将是重要的。这项提议中的实验旨在了解一种被称为前BCR的关键分子如何控制B细胞的发育，B细胞是制造免疫反应所必需的抗体的细胞。遗传上不能制造前BCR的人是严重的免疫缺陷。所获得的信息是需要更好地了解一些B细胞白血病的起源，可以追溯到影响前BCR功能的突变。

DESCRIPTION (provided by applicant): Signals transduced by the precursor B cell receptor (preBCR) are necessary for B cell development. Humans and mice genetically deficient in preBCR components suffer from an absence or paucity of pre-B and more mature B cells that is responsible for a state of agammaglobulinemia and resultant severe immune deficiency. Deregulating or loss-of-function mutations of downstream preBCR signal transduction molecules are associated with B cell leukemia and lymphoma. An enigmatic aspect of preBCR signaling is the physiological means by which receptor signaling is activated. Evidence for both ligand-dependent and ligand-independent mechanisms has been demonstrated in cell culture systems and suggested to be physiological. Common to both models has been the idea that receptor localization to the cell surface is critical, either to engage ligands or key signal transduction molecules that reside there, because surface expression has directly correlated with receptor activity. Contrary to this model, we have recently demonstrated that preBCR complexes directed away from the cell membrane and redistributed among intracellular, post-ER compartments were as or nearly as active as wild-type preBCRs. This was accomplished by appending endomembrane localization sequences to the cytoplasmic tail of a preBCR-forming immunoglobulin mu heavy chain (Ig HC), and testing the resulting "redirected" preBCR activity in preBCR-deficient transformed and primary pro-B cells. These data support the model that preBCRs do not require extracellular ligands at the cell surface for signaling, and that the preBCR is active in post-ER compartments like the Golgi. However, several developmentally important processes linked to preBCR signaling, such as allelic exclusion, could not be evaluated, and it remains possible that these require activities dependent on preBCR surface expression. Therefore, the purpose of the experiments in this small R03 proposal is to build on these findings by evaluating the biological activity of these redirected preBCR complexes in vivo. This will be accomplished in one Aim by establishing transgenic mice that express the redirected Ig HCs and evaluating their ability to support B cell development and effect allelic exclusion in genetically preBCR-deficient and preBCR-sufficient mice. This information will be important to better understand the molecular mechanisms that determine receptor activity in normal immune development and in B cell pathologies. The experiments in this proposal aim to understand how a critical molecule known as the preBCR controls the development of B cells, the cells that make antibodies essential for an immune response. People that are genetically unable to make a preBCR are severely immune deficient. The information obtained is needed to better understand the origin of some B cell leukemias that can be traced to mutations that affect the functioning of the preBCR.