Defining the partner interaction network of the tetraspanin CD53 in regulating B cell trafficking

定义四跨膜蛋白 CD53 在调节 B 细胞运输中的伙伴相互作用网络

• 批准号: 10660927
• 负责人: Weikai Li
• 金额: $ 56.53万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-06 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10660927
• 关键词: Adhesions; Affect; Annexins; Antibody Formation; Antigens; B-Cell Development; B-Cell Leukemia; B-Lymphocytes; Binding; Biochemical; Biological Assay; Bone Marrow; Cell Adhesion; Cell Communication; Cell physiology; Cell surface; Cells; Chemotactic Factors; Coculture Techniques; Collaborations; Complex; Cytoskeleton; Data; Electrons; Endothelium; Family; Family member; Goals; Growth; Hematopoiesis; Homing; Human; Immune response; Immunoglobulins; Immunohistochemistry; Impairment; In Vitro; Infection; Integral Membrane Protein; Label; Link; Malignant - descriptor; Malignant Neoplasms; Marrow; Mass Spectrum Analysis; Mediating; Methods; Microscopic; Modeling; Molecular; Molecular Conformation; Mus; Population; Predisposition; Process; Proliferating; Proteins; Public Health; Reporting; Research; Research Personnel; Role; Serum; Signal Transduction; Signaling Protein; Spleen; Structure; Techniques; Testing; cell motility; chemokine; cohort; cytokine; experimental study; immune system function; in vivo; insight; knock-down; leukemia/lymphoma; lymphoid organ; member; migration; monolayer; novel; novel therapeutic intervention; response; secondary lymphoid organ; stathmin; therapeutic target; tool; trafficking

Project Summary/Abstract The goal of this project is to understand how CD53 regulates B cell trafficking, and to determine the CD53- partner network underlying this process. CD53 is a member of the tetraspanin family of transmembrane proteins that organize multi-protein networks on the cell surface to regulate a wide variety of cellular processes such as proliferation, homing and survival. Loss of CD53 is associated with impaired immune system function. CD53 is highly expressed on both normal and malignant B cells, however its role in these cells is not clear. We previously reported that CD53 is required for normal B cell development in the bone marrow. In preliminary data, we now find that CD53 is essential to both normal and malignant B cell trafficking, with the loss of CD53 causing significant impairment in B cell adhesion, migration, bone marrow homing and antibody production. To understand the underlying molecular mechanisms, we have determined the crystal structure of CD53; in the entire tetraspanin family, this the first structure captured in an active conformation. We reveal how conformational changes influence CD53 partner interactions by mass spectrometry (MS) based footprinting. We have used proximity labeling to identify several candidate CD53 partners, whose functions link chemokine signaling to cell motility. These data support our hypothesis that CD53 coordinates a complex of adhesion, signaling and cell motility proteins that facilitate B cell trafficking. Both normal and malignant B cells rely on accurate trafficking to their niches in the bone marrow and secondary lymphoid organs to optimize their maturation and function. Thus, an understanding of the interactions that guide B cell trafficking are important not only for optimizing normal B cell function, but will also reveal potential therapeutic targets of malignant B cells. Using a combination of in vitro and vivo adhesion and migration studies in combination with proximity labeling, quantitative MS, live-cell MS- based footprinting, biochemical and electron microscopic analyses, this dual-PI proposal presents a 5-year plan to: 1) elucidate the CD53-partner interaction network regulating B cell trafficking, and 2) determine the functional consequences of disrupting the CD53-partner interactions. Armed with our newly developed tools, we will reveal a novel network of protein interactions coordinating cell signaling and motility to regulate B cell adhesion, migration, chemokine signaling and niche localization. We will uncover how this CD53-mediated network responds to chemokine signaling during B cell migration and how cross-cell interactions are established during B cell homing. Thus, the proposed studies will significantly advance our understanding of how B cell trafficking is coordinated and regulated. Given the conserved structure and functional redundancy of tetraspanin family members, this will lead to our long-term goal of elucidating tetraspanin/partner relationships that apply to other roles of CD53 in immune system function and malignancy as well as the functions of other tetraspanin family members.

项目总结/摘要 本项目的目标是了解CD53如何调节B细胞的运输，并确定CD53- 这一进程背后的伙伴网络。CD53是跨膜蛋白的四跨膜蛋白家族的成员 组织细胞表面的多蛋白质网络以调节多种细胞过程，例如 增殖归巢和生存CD53的丢失与免疫系统功能受损有关。CD53是 在正常和恶性B细胞上高度表达，然而其在这些细胞中的作用尚不清楚。我们之前 报道CD 53是骨髓中正常B细胞发育所必需的。在初步数据中，我们现在 发现CD53对于正常和恶性B细胞运输都是必需的， B细胞粘附、迁移、骨髓归巢和抗体产生的显著损害。到 了解潜在的分子机制，我们已经确定了CD 53的晶体结构; 在整个四跨膜蛋白家族中，这是第一个以活性构象捕获的结构。我们揭示了构象 通过基于质谱法（MS）的足迹法，变化影响CD53伴侣相互作用。我们已经使用 邻近标记以鉴定几个候选的CD53配偶体，其功能将趋化因子信号传导与细胞增殖相关联。 能动性这些数据支持了我们的假设，即CD53协调粘附、信号传导和细胞增殖的复合体。 促进B细胞运输的运动蛋白。正常和恶性B细胞都依赖于准确的运输， 它们在骨髓和次级淋巴器官中的小生境，以优化它们的成熟和功能。因此，在本发明中， 理解引导B细胞运输的相互作用不仅对于优化正常的B 细胞功能，而且还将揭示恶性B细胞的潜在治疗靶点。使用体外结合 以及结合邻近标记、定量MS、活细胞MS的体内粘附和迁移研究。 基于足迹法、生物化学和电子显微镜分析，该双PI提案提出了一个5年计划 目的：1）阐明调节B细胞运输的CD 53-配偶体相互作用网络，和2）确定功能性 破坏CD53-伴侣相互作用的后果。借助我们新开发的工具，我们将揭示 一种新的蛋白质相互作用网络，协调细胞信号传导和运动以调节B细胞粘附， 迁移、趋化因子信号传导和小生境定位。我们将揭示这个CD53介导的网络 在B细胞迁移过程中对趋化因子信号传导的反应以及在迁移过程中如何建立跨细胞相互作用 B细胞归巢。因此，拟议中的研究将大大推进我们对B细胞贩运的理解 是协调和规范的。鉴于四跨膜蛋白家族的保守结构和功能冗余， 成员们，这将导致我们的长期目标，阐明四跨膜蛋白/伙伴关系，适用于其他 CD53在免疫系统功能和恶性肿瘤中的作用以及其他四跨膜蛋白家族的功能 成员