Regulation of B cell differentiation by eIF4E

 描述(申请人提供)：雷帕霉素(MTOR)的哺乳动物靶标在抗原刺激后促进淋巴细胞的生长、分裂和分化。MTOR是一种蛋白激酶，在mTORC1和mTORC2两个复合体中发挥作用。雷帕霉素是一种变构的mTORC1抑制剂，能深刻抑制B细胞的增殖和分化。相反，在包括关节炎和狼疮在内的自身抗体驱动的疾病患者的淋巴细胞中，mTORC1活性升高。然而，mTORC1信号促进B细胞分化的机制尚不清楚。我们已经报道，mTORC1的遗传或药物抑制抑制了激活的B细胞中抗体类别的转换。随后，我们确定了真核细胞起始因子4E(EIF4E)在促进B细胞增殖和抗体类别转换方面的新作用。EIF4E以细胞类型特异性的方式调节mRNAs亚集的翻译，并受到作为mTORC1直接底物的eIF4E结合蛋白(4E-Bps)的反对。利用新的化学工具和遗传小鼠模型，我们已经确定mTORC1、4E-BPS和eIF4E在类别转换中的作用可以与它们在增殖中的作用分开。这项提案的总体目标是确定mTORC1和eIF4E促进抗体类别转换的机制。第一个目标是测试这样的假设，即有效的类别切换需要eIF4E，而4E-BPS与之相反。我们将测量在存在或不存在RAP的情况下，eIF4E或4E-BPS表达增加或减少的小鼠激活的B细胞的同型转换。还将使用通过ATP竞争方式抑制mTORC1的新型化合物。4E-BP/eIF4E信号轴的作用也将在体内通过测量免疫小鼠的抗体反应和生发中心形成来测试。第二个目标将定义eIF4E调节参与B细胞分化的mRNAs翻译的机制。越来越多的证据表明，翻译效率在不同的mRNAs之间差异很大，受调控的翻译对蛋白质组的调节程度与转录和表观遗传调节相似。因此，全基因组翻译组研究揭示了肿瘤发生和其他过程中的新调控机制。我们将使用核糖体图谱对eIF4E靶标进行全球、无偏见的鉴定，这些靶标在经历抗体类别转换的激活B细胞中进行差异翻译。该项目具有重要意义，因为对由eIF4E控制并被mTORC1抑制的翻译组的更好理解可能揭示抑制致病自身抗体反应的新策略，具有比雷帕霉素更好的治疗窗口。这项工作还将阐明正在开发的eIF4E抑制剂对癌症的潜在免疫抑制作用。

 DESCRIPTION (provided by applicant): The mammalian target of rapamycin (mTOR) drives lymphocyte growth, division, and differentiation following antigen stimulation. mTOR is a protein kinase that functions in two complexes, mTORC1 and mTORC2. Rapamycin is an allosteric mTORC1 inhibitor that profoundly suppresses B cell proliferation and differentiation. Conversely, elevated mTORC1 activity is seen in lymphocytes from patients with autoantibody-driven disorders including arthritis and lupus. However, the mechanisms by which mTORC1 signaling promotes B cell differentiation remain poorly defined. We have reported that genetic or pharmacological inhibition of mTORC1 suppresses antibody class switching in activated B cells. Subsequently we identified novel roles for eukaryotic initiation factor 4E (eIF4E), a downstream mTORC1 effector, in promoting B cell proliferation and antibody class switching. eIF4E regulates the translation of subsets of mRNAs in a cell type-specific manner, and is opposed by eIF4E-binding proteins (4E-BPs) that are direct substrates of mTORC1. Using novel chemical tools and genetic mouse models we have determined that the roles of mTORC1, 4E-BPs and eIF4E in class switching can be separated from their roles in proliferation. The general goal of this proposal is to define the mechanisms by which mTORC1 and eIF4E promote antibody class switching. The first Aim is to test the hypothesis that efficient class switching requires eIF4E an is opposed by 4E-BPs. We will measure isotype switching in activated B cells from mice with increased or decreased expression of eIF4E or 4E-BPs, in the presence or absence of RAP. Novel compounds that inhibit mTORC1 through an ATP-competitive manner will also be used. The role of the 4E-BP/eIF4E signaling axis will also be tested in vivo by measuring antibody responses and germinal center formation in immunized mice. The second Aim will define the mechanisms by which eIF4E regulates translation of mRNAs involved in B cell differentiation. Increasing evidence indicates that translation efficiency varies widely among individual mRNAs, and that regulated translation modulates the proteome to a similar degree as transcriptional and epigenetic regulation. In accord, genome-wide translatome studies have uncovered novel regulatory mechanisms in tumorigenesis and other processes. We will use ribosome profiling for global, unbiased identification of eIF4E targets that are differentially translated in activated B cells undergoing antibody class switching. This project is significant because a greater understanding of the translatome controlled by eIF4E and suppressed by mTORC1 inhibition might reveal novel strategies for suppressing pathogenic autoantibody responses, with a better therapeutic window than rapamycin. This work will also shed light on potential immunosuppressive effects of eIF4E inhibitors under development for cancer.