Molecular mechanisms of B cell malignant transformation

B细胞恶性转化的分子机制

• 批准号: 8372542
• 负责人: PING XIE
• 金额: $ 33.62万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-04 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8372542
• 关键词: Address; Affect; Apoptosis; Apoptotic; B-Cell Lymphomas; B-Cell Neoplasm; B-Lymphocytes; B-Lymphoma Development; Biochemical; Biological Assay; Biological Models; Cell Death; Cell Line; Cell Survival; Cells; Cessation of life; Chronic Lymphocytic Leukemia; Co-Immunoprecipitations; Communication; Deletion Mutation; Development; Endoplasmic Reticulum; Equilibrium; Family; Fractionation; Gene Expression; Genes; Grant; Human; Image; Knowledge; Luciferases; Lymphoma; MCL1 protein; Malignant - descriptor; Malignant Neoplasms; Mantle Cell Lymphoma; Mature B-Lymphocyte; Mediating; Mediator of activation protein; Microarray Analysis; Microscopic; Mitochondria; Modeling; Molecular; Molecular Profiling; Multiple Myeloma; Mus; Mutagenesis; Mutation; Non-Hodgkin' s Lymphoma; Oncogenic; Organelles; Pathway interactions; Patients; Physiological; Premalignant; Protein Family; Proteins; Reporter; Role; Scheme; Signal Transduction; Stimulus; Structure-Activity Relationship; T-Lymphocyte; TNF receptor-associated factor 3; TNFRSF5 gene; Testing; Therapeutic Intervention; Tumor Suppressor Genes; Tumor Suppressor Proteins; United States; Up-Regulation; Vesicle; bak protein; base; biological adaptation to stress; chromatin immunoprecipitation; combat; design; endoplasmic reticulum stress; human RTN4 protein; information gathering; insight; interest; macrophage; mitochondrial membrane; mouse model; mutant; novel; overexpression; protein transport; small hairpin RNA; therapeutic target; trafficking; transcription factor; treatment strategy; tumor; tumorigenesis; vector

DESCRIPTION (provided by applicant): TRAF3 is a novel tumor suppressor gene identified in a variety of human B lymphoma, including splenic marginal zone lymphoma, B cell chronic lymphocytic leukemia and mantle cell lymphoma, as well as multiple myeloma (MM). To explore the role of TRAF3, we recently generated a new genetically modified mouse model that has the TRAF3 gene specifically deleted in B cells (B-TRAF3-/- mice). We found that TRAF3 deletion results in prolonged survival of mature B cells, which eventually leads to spontaneous development of B lymphomas in mice. This proposal aims to understand how TRAF3 inactivation promotes oncogenic B cell survival. We first found that the mechanism of aberrant B cell survival mediated by TRAF3 inactivation is fundamentally different from that of normal B cell survival induced by physiological stimuli. To identify novel targets of TRAF3 inactivation, we performed microarray analysis to compare the global gene expression profiles of splenic B cells purified from young, tumor-free B-TRAF3-/- mice and littermate control mice. Strikingly, a number of genes identified in our microarray analysis are implicated in subcellular organelle or vesicle trafficking, including Rhbdf1, Rasgrp3, Ehd1, and Kif11, etc. Current evidence indicates that many survival proteins are trafficking between two important organelles, endoplasmic reticulum (ER) and mitochondria, and that the balance between mitochondrial levels of pro-survival proteins and anti-survival proteins controls the threshold of cell death. Interestingly, or preliminary results demonstrated that mitochondrial levels of the anti-survival protein Bak were decreased, and mitochondrial levels of the pro- survival protein Mcl-1 were increased in premalignant TRAF3-/- B cells. However, total cellular levels of Bak and Mcl-1 were not altered in these cells. Based on our new findings, we will test the central hypothesis that TRAF3 inactivation modulates the trafficking of Bak and Mcl-1 between ER and mitochondria to increase the threshold of cell death in B cells. To address this, we propose complementary studies using B-TRAF3-/- mice and human patient-derived multiple myeloma cell lines with TRAF3 deletions or mutations as model systems. In Aim 1, using microarray analysis and a stringent prioritization scheme, our preliminary results led us to focus on a novel target of TRAF3, Rhbdf1, which is implicated in ER protein trafficking. To delineate the causal role of Rhbdf1 in oncogenic B cell survival, we will employ lentiviral shRNA vector-mediated knockdown, ectopic overexpression, and mutagenesis of Rhbdf1. In Aim 2, we will perform extensive biochemical fractionation, confocal microscopic imaging, and co-immunoprecipitation studies to understand how TRAF3 inactivation and Rhbdf1 modulate the trafficking of Bak and Mcl-1 between ER and mitochondria. Furthermore, we will interrogate how TRAF3 signaling interplays and cross-talks with ER stress responses. In summary, this study will provide new insights into the exact mechanisms of TRAF3 inactivation-mediated oncogenic B cell survival, and will open up new avenues for the treatment of B lymphoma and MM. PUBLIC HEALTH RELEVANCE: Lymphoma is the 5th most common human cancer with approximately 21,000 annual deaths in the United States. TRAF3 is a novel tumor suppressor gene identified in a variety of human B lymphoma, including splenic marginal zone lymphoma, B cell chronic lymphocytic leukemia and mantle cell lymphoma, as well as multiple myeloma. This proposal aims to investigate a novel target of TRAF3, Rhbdf1, and a new signaling mechanism that connect TRAF3 inactivation to oncogenic B cell survival. To address the gaps in knowledge, we propose complementary studies using a new mouse B lymphoma model with the TRAF3 gene specifically deleted in B lymphocytes and human patient-derived multiple myeloma cell lines with TRAF3 deletions or mutations. Information gathered from the proposed studies will advance the understanding of the pathological mechanisms underlying B cell oncogenesis, and will open up new avenues for the development of treatment strategies to combat B lymphoma and multiple myeloma.

描述(申请人提供)：TRAF3是一种新的肿瘤抑制基因，在多种人类B淋巴瘤中发现，包括脾边缘带淋巴瘤、B细胞慢性淋巴细胞白血病和套细胞淋巴瘤以及多发性骨髓瘤(MM)。为了探索TRAF3的作用，我们最近建立了一种新的转基因小鼠模型，该模型具有B细胞中TRAF3基因的特异性缺失(B-TRAF3-/-小鼠)。我们发现，TRAF3缺失会导致成熟B细胞存活时间延长，最终导致小鼠B淋巴瘤的自发发展。这项建议旨在了解TRAF3失活如何促进致癌B细胞存活。我们首次发现，TRAF3失活介导的异常B细胞存活机制与生理刺激诱导的正常B细胞存活机制是根本不同的。为了确定TRAF3失活的新靶点，我们进行了微阵列分析，比较了从年轻的、无肿瘤的B-TRAF3-/-小鼠和产仔对照小鼠中纯化的脾B细胞的整体基因表达谱。值得注意的是，我们的芯片分析中发现了一些与亚细胞细胞器或囊泡运输有关的基因，包括Rhbdf1、Rasgrp3、Ehd1和Kif11等。目前的证据表明，许多生存蛋白在内质网(ER)和线粒体这两个重要细胞器之间运输，线粒体促进生存蛋白和抗生存蛋白水平之间的平衡控制着细胞死亡的阈值。有趣的是，或初步结果表明，在癌前病变的TRAF3-/-B细胞中，抗生存蛋白Bak的线粒体水平下降，而促生存蛋白Mcl-1的线粒体水平上升。然而，在这些细胞中，Bak和Mcl-1的总细胞水平没有变化。基于我们的新发现，我们将检验核心假设，即TRAF3失活调控Bak和Mcl-1在内质网和线粒体之间的运输，以提高B细胞中细胞死亡的阈值。为了解决这一问题，我们建议使用B-TRAF3-/-小鼠和带有TRAF3缺失或突变的人多发性骨髓瘤细胞系作为模型系统进行互补研究。在目标1中，使用微阵列分析和严格的优先方案，我们的初步结果使我们专注于TRAF3的一个新靶点，Rhbdf1，它与内质网蛋白运输有关。为了阐明Rhbdf1在致癌B细胞存活中的因果作用，我们将采用慢病毒shRNA载体介导的Rhbdf1的敲除、异位过表达和突变。在目标2中，我们将进行广泛的生化分级、共聚焦显微镜成像和免疫共沉淀研究，以了解TRAF3失活和Rhbdf1如何调节Bak和Mcl-1在内质网和线粒体之间的运输。此外，我们将询问TRAF3信号如何与内质网应激反应相互作用和相互作用。综上所述，本研究将为TRAF3失活介导的致癌B细胞存活的确切机制提供新的见解，并将为B淋巴瘤和MM的治疗开辟新的途径。 与公共卫生相关：淋巴瘤是第五大最常见的人类癌症，在美国每年约有2.1万人死亡。TRAF3是一种新的肿瘤抑制基因，在多种人类B淋巴瘤中发现，包括脾边缘带淋巴瘤、B细胞慢性淋巴细胞白血病和套细胞淋巴瘤以及多发性骨髓瘤。该建议旨在研究TRAF3的新靶点Rhbdf1，以及将TRAF3失活与致癌B细胞存活联系起来的新的信号机制。为了解决知识上的空白，我们提出了一项补充研究，使用了一种新的小鼠B淋巴瘤模型，该模型的TRAF3基因在B淋巴细胞和人类患者来源的多发性骨髓瘤细胞系中特异性缺失，TRAF3缺失或突变。从拟议的研究中收集的信息将促进对B细胞肿瘤发生的病理机制的理解，并将为开发抗击B淋巴瘤和多发性骨髓瘤的治疗策略开辟新的途径。