Dissecting the Role Ubiquitin E3 Ligase UBR5 in Lymphomagenesis

摘要 套细胞淋巴瘤（MCL）是一种罕见的侵袭性非霍奇金淋巴瘤。可惜有限 目前已有MCL的治疗方法，这表明需要进一步阐明其分子机制 调节疾病的转化和进展。大多数MCL患者的t（11; 14） 易位导致CyclinD1过度表达，导致细胞广泛增殖， 淋巴结的分化起源于淋巴结的套区，然而，额外的突变是必要的， 转型下一代测序已经在MCL患者中发现了许多新的突变 包括泛素E3连接酶UBR 5。E3泛素连接酶作为底物识别组分， 泛素蛋白酶体系统的蛋白质降解。在196名MCL患者的队列中，UBR 5是第3位 最常见的突变基因，约60%的突变在UBR5的HECT结构域内发现， 可以接受泛素分子并将其转移到底物上。为了了解UBR 5 HECT的作用， 我们使用新的CRISPR/Cas 9技术产生了条件小鼠。 HECT结构域的缺失导致脾脏中的前生殖中心B细胞的阻滞， B1和边缘B细胞亚群。此外，脾脏中的滤泡B细胞表型异常， 最终分化为分泌抗体的浆细胞。蛋白质组学研究揭示蛋白质上调 与缺乏UBR 5的HECT结构域的B细胞中通过剪接体的mRNA剪接相关。这些 研究提示：1）UBR 5突变沿着cyclinD1的表达可能导致疾病 套细胞淋巴瘤的进展（目的1），2）了解UBR5突变的分子机制 U5剪接体的异常表达可为MCL提供潜在的治疗靶点（目的2） 蛋白质阻断B细胞成熟并促进淋巴瘤发生（Aim 3）。在本申请中，我们提出研究 了解UBR 5及其相互作用蛋白在B细胞淋巴瘤发生中的作用，并确定其分子机制。 在B细胞中由UBR5调节的途径。我们的研究目的是为套细胞的研究提供新的视角 淋巴瘤转化、进展和潜在的未来治疗靶点。

ABSTRACT Mantle cell lymphoma (MCL) is a rare and aggressive non-Hodgkin’s lymphoma. Unfortunately limited therapies for MCL are currently available suggesting a need to further unravel molecular mechanisms regulating transformation and progression of the disease. The majority of MCL patients have a t(11;14) translocation leading to overexpression of CyclinD1 resulting in extensive proliferation and block in differentiation originating in the mantle zone of the lymph node, however additional mutations are necessary for transformation. Next generation sequencing has identified a number of novel mutations in MCL patients including the ubiquitin E3 ligase UBR5. E3 ubiquitin ligases serve as the substrate-recognizing component for protein degradation by the ubiquitin proteasome system. In a cohort of 196 MCL patients UBR5 was the 3rd most frequently mutated gene and ~60% of the mutations were found within the HECT domain of UBR5, which can accept and transfer ubiquitin molecules to the substrate. In order to understand the role of UBR5 HECT domain in B-lymphoid development we generated a conditional mouse using novel CRISPR/Cas 9 technology. Loss of the HECT domain leads to a block in pre-germinal center B cells in the spleen with a reduction of both B1 and marginal B cell subsets. In addition, follicular B cells in the spleen are phenotypically abnormal and fail to terminally differentiate to anti-body secreting plasma cells. Proteomic studies reveal up-regulation of proteins associated with mRNA splicing via the spliceosome in B cells lacking the HECT domain of UBR5. These studies suggest that 1) cooperation of UBR5 mutations along with expression of cyclinD1 may led to disease progression of mantle cell lymphoma (Aim 1), 2) understanding molecular mechanism of UBR5 mutations could provide potential therapeutic targets in MCL (Aim 2), and 3) aberrant expression of U5 spliceosome proteins block B cell maturation and promote lymphomagenesis (Aim 3). In this application we propose studies to understand the role of UBR5 and its interacting proteins in B-cell lymphomagenesis and define molecular pathways regulated by UBR5 in B-cells. Our goal of the proposed studies is to provide insights to mantle cell lymphoma transformation, progression and potential future therapeutics targets.