In vivo functions of Ig-beta ubiquitinylation

描述（由申请方提供）：需要两种协调信号来从幼稚卵泡B细胞中引发有效的体液免疫应答。当B细胞抗原受体（BCR）识别多价或膜限制性抗原并启动相互关联的信号级联网络时，在细胞表面立即提供第一种。相反，必要的第二信号出现在专门的晚期内体（包含隔室或MIIC的MHC II）专有的过程之后。这些包括用肽装载MHC II类，随后募集T细胞辅助，以及通过toll样受体（TLR）7和9的配体结合。这些信号2激活受体的激活依赖于BCR在细胞表面捕获配体并将它们沿着胞吞转运的能力。 通向受体的通路。然而，BCR靶向晚期内体并由此将信号1与信号2偶联的分子机制知之甚少。我们先前已经证明，IG泛素化是细胞系中将内化的BCR复合物靶向晚期内体所必需的。我们还证明了BCR内吞靶向MIIC不是一个组成性过程，而是在外周无反应性细胞中受调节以防止异常B细胞活化的过程。我们现在已经推导出了IG泛素化（IG？）这证实了它在引导BCR复合物进入晚期内体中的作用。此外，我们现在证明了IG泛素化是选择IgM高细胞进入未成熟B细胞库以及Td和Ti体液免疫应答所必需的。对IG的要求泛素化不依赖于抗原特异性，并与基础和BCR诱导的信号传导中的特异性缺陷相关。这些信号传导缺陷的体外互补恢复了BCR靶向晚期内体。基于这些和其他数据在初步结果中，我们假设，BCR泛素化，很像酪氨酸磷酸化，在耦合的BCR信号传导过程中发挥了关键作用，B细胞发育和外周活化所必需的。此外，我们提出，泛素化在BCR稳态和选择和维持B细胞中起着额外的作用，而与抗原特异性无关。这一中心假设将在以下具体目标中得到检验：目标1。确定IG泛素化对未成熟B细胞选择的作用。目标二。确定IG泛素化在外周免疫应答中的作用。目标3：确定IG调节BCR信号启动和表面稳态的机制。!

DESCRIPTION (provided by applicant): Two coordinated signals are required to elicit productive humoral immune responses from na¿ve follicular B cells. The first is provided immediately at the cell surface when the B cell antigen receptor (BCR) recognizes polyvalent or membrane-restricted antigens and initiates an interrelated web of signaling cascades. In contrast, the necessary second signals arise later from processes exclusive to specialized late endosomes (the MHC II containing compartment or MIIC). These include the loading of MHC class II with peptides, and subsequent recruitment of T cell help, and ligand binding by toll-like receptors (TLR) 7 and 9. Activation of these signal 2 enabling receptors is dependent upon the abilities of the BCR to capture ligands at the cell surface and to deliver them along the endocytic pathway to their receptors. However, the molecular mechanisms by which the BCR target late endosomes, and thereby couples signal 1 with signal 2, are poorly understood. We have previously demonstrated that Ig¿ ubiquitination is necessary in cell lines for targeting internalized BCR complexes to late endosomes. We have also demonstrated that BCR endocytic targeting to the MIIC is not a constitutive process but one that is regulated in peripheral anergic cells to prevent aberrant B cell activation. We have now gone on to derive an in vivo model of Ig¿ ubiquitination (Ig¿???) that confirms its role in ushering BCR complexes into late endosomes. In addition, we now demonstrate that Ig¿ ubiquitination is required for selection of IgMhigh cells into the immature B cell pool and for both Td and Ti humoral immune responses. The requirement for Ig¿ ubiquitination was independent of antigen specificity and was associated with specific defects in basal and BCR-induced signaling. Complementation of these signaling defects in vitro restored BCR targeting to late endosomes. Based on these and other data presented in Preliminary Results, we hypothesize that BCR ubiquitination, much like tyrosine phosphorylation, plays a critical role in coupling the BCR to signaling processes necessary for B cell development and peripheral activation. Furthermore, we propose that ubiquitination plays additional roles in BCR homeostasis and in selecting and maintaining B cells irrespective of antigenic specificity. This central hypothesis will be tested in the followin Specific Aims: Aim 1. To determine how Ig¿ ubiquitination contributes to immature B cell selection. Aim 2. To determine the role of Ig¿ ubiquitination in peripheral immune responses. Aim 3. Determine the mechanisms by which Ig¿ regulates BCR signal initiation and surface homeostasis. !