Control of ribosomal function in cells of the hematopoietic system

造血系统细胞核糖体功能的控制

• 批准号: 10509989
• 负责人: Andrew Levine
• 金额: $ 11.11万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10509989
• 关键词: Affect; Area; Award; Biochemical; Biological Assay; Biology; Bone Marrow; Bone marrow failure; Cell physiology; Cells; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Craniofacial Abnormalities; Data Science; Defect; Development; Development Plans; Diagnostic; Disease; Doctor of Philosophy; Elements; Environment; FMRP; Foundations; Functional disorder; Future; Genes; Genetic Translation; Goals; Grant; Hematopoiesis; Hematopoietic; Hematopoietic System; Housekeeping; Human; Immune; Immunologist; Immunology; In Vitro; Infection; Inflammation; Inflammatory; Inflammatory Response; Innate Immune System; Lipopolysaccharides; Macrophage Activation; Mediating; Mentors; Messenger RNA; Methods; Modification; Molecular; Mouse Protein; Mutagenesis; Mutation; Nature; Nutrient; Pathologist; Phase; Physicians; Poly I-C; Process; Production; Protein Biosynthesis; Proteins; Proteomics; Protocols documentation; RNA; RNA Binding; RNA Sequences; Regulation; Regulatory Element; Research; Research Ethics; Research Personnel; Ribosomal Interaction; Ribosomal Proteins; Ribosomal RNA; Ribosomes; Role; Scientist; Signal Pathway; Signal Transduction; Specificity; Stimulus; Structure; Testing; Tissues; Training; Transcript; Translating; Translation Initiation; Translational Regulation; Translations; United States National Institutes of Health; Work; career development; cell type; cytokine; experimental study; extracellular; immune activation; immune function; immunoregulation; interest; macrophage; monocyte; neutrophil; novel; programs; proteostasis; response; ribosome profiling

PROJECT SUMMARY/ABSTRACT Translation is a dynamic process with a high degree of specificity conferred by multiple factors including mRNA transcript sequence, composition of the translation machinery, and cellular milieu. A striking demonstration of this specificity is the tissue-specific (e.g. craniofacial anomalies, bone marrow failure) rather than global nature of the developmental defects seen in human ribosomopathies caused by congenital mutations in ribosomal factors. Moreover, ribosomopathy-driven bone marrow failure is characterized by selective, rather than global, defects in mRNA translation. Thus, even the composition of the ribosome can affect mRNA translational efficiency in a tissue-specific manner. These observations point to a particularly nuanced regulation of translation in the hematopoietic system; elucidating the mechanisms of this regulation is therefore essential for understanding hematopoietic cell development, function, and dysfunction in disease states. As such, my long-term research goal is to elucidate mechanisms of translational control in hematolymphoid cells during normal immune function and in disease states, focusing on ribosome-associated proteins (RAPs)—a poorly studied class of proteins which physically interact with the core ribosomal proteins to tune the translational efficiency of specific mRNAs. My central hypothesis is that, in immune cells poised for rapid responses to infection, inflammatory stimuli may induce dynamic interactions of RAPs with ribosomes, where they may effect rapid changes in the translational efficiency of specific mRNAs with important immunoregulatory consequences. In previous work, I have defined the repertoire of ribosome-bound RAPs in in vitro bone marrow-derived macrophages. In Aim 1 of this proposal, I will use global, unbiased proteomics to temporally profile RAP-ribosome interactions in activated macrophages following TLR stimulation. In Aim 2, I will use CRISPR gene editing, ribosome profiling, and cytokine secretion assays to 1) identify mRNA targets that are specifically regulated by two RAPs I have already identified, Rnf213 and Helz2, as well as other RAPs identified in Aim 1, and 2) determine the function of these RAPs in the macrophage inflammatory response. Having completed MD-PhD training and clinical training as a clinical pathologist and hematopathologist, I am applying for the K38 Award to support my goal of becoming an independent physician investigator. UCSF’s exceptional training environment, especially in the areas of immunology and proteostasis, make it an ideal place for me to pursue my efforts. Critical elements of my career development plan include leveraging the expertise of my primary mentor Dr. Davide Ruggero, a pioneer in the field of ribosome biology, and my co-mentors Dr. Jason Cyster, an expert immunologist, and Dr. Karthik Ganapathi, an expert diagnostic hematopathologist; guidance by physician-scientist advisors; coursework in data science and research ethics; and other professional development activities. All together, I wholeheartedly believe this career development plan will provide a strong foundation on which to ultimately build an independent research program.

项目摘要/摘要 翻译是一个动态的过程，具有高度的特异性，受多种因素的影响，包括信使 转录序列，翻译机器的组成，以及细胞环境。一个引人注目的示范 这种特异性是特定于组织的(例如，颅面畸形、骨髓衰竭)，而不是全球性的 由核糖体先天突变引起的人类核糖体疾病的发育缺陷 各种因素。此外，核糖体病导致的骨髓衰竭的特点是选择性的，而不是全局性的， 信使核糖核酸翻译的缺陷。因此，即使是核糖体的组成也能影响mRNA的翻译 以特定于组织的方式提高效率。这些观察指向了一种特别微妙的监管 在造血系统中的翻译；因此，阐明这种调节的机制对于 了解疾病状态下造血细胞的发育、功能和功能障碍。 因此，我的长期研究目标是阐明翻译控制的机制 免疫功能正常和疾病状态下的血淋巴细胞，主要集中在核糖体相关 蛋白质(RAP)-一类研究较少的蛋白质，它与核心核糖体蛋白物理上相互作用以 调整特定mRNAs的翻译效率。我的中心假设是，在免疫细胞中， 对感染的快速反应，炎症刺激可诱导RAP与核糖体的动态相互作用， 它们可能会影响特定mRNAs的翻译效率的快速变化，具有重要的 免疫调节后果。在以前的工作中，我已经定义了与核糖体结合的RAP谱系 在体外，骨髓来源的巨噬细胞。在这项提案的目标1中，我将使用全球、公正的蛋白质组学来 TLR刺激后活化巨噬细胞中RAP-核糖体相互作用的时间分布。在目标2中，我 将使用CRISPR基因编辑、核糖体分析和细胞因子分泌分析来1)识别 由我已经确定的两个RAP(Rnf213和Helz2)以及其他RAP具体管理 目的1和2)确定这些RAP在巨噬细胞炎症反应中的功能。 作为一名临床病理学家和血液病理学家，完成了医学博士培训和临床培训， 我正在申请K38奖，以支持我成为一名独立内科研究员的目标。加州大学旧金山分校 特殊的训练环境，特别是在免疫学和蛋白质平衡方面，使它成为一个理想的地方 让我继续努力。我职业发展计划的关键要素包括利用专业知识 我的主要导师大卫·鲁杰罗博士，核糖体生物学领域的先驱，以及我的共同导师大卫·鲁格罗博士。 免疫学家Jason Cyster和诊断血液病理学家Karthik Ganapathi博士； 医生-科学家顾问的指导；数据科学和研究道德方面的课程；以及其他专业人员 发展活动。综上所述，我衷心相信这份职业发展计划将提供一个强有力的 最终建立一个独立研究计划的基础。