Regulation of protein ubiquitination in hematopoietic cytokine signaling

造血细胞因子信号传导中蛋白质泛素化的调节

基本信息

批准号：
9310835
负责人：
Wei Tong
金额：
$ 56.7万
依托单位：
CHILDREN'S HOSP OF PHILADELPHIA
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-04-18 至 2021-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9310835
关键词：
Acute Myelocytic Leukemia Adaptor Signaling Protein Address Alleles Attenuated Automobile Driving Biochemical Blood Cells Bone Marrow Transplantation CBL Protein CBL gene Cell Count Cell Line Cell Proliferation Cells Chronic Clonal Hematopoietic Stem Cell Collaborations Cytokine Receptors Cytokine Signaling Data Development Disease Disease remission Engineering Exhibits FDA approved Family Frequencies Genetic Goals Granulocyte-Macrophage Colony-Stimulating Factor Receptors Growth Half-Life Hematopoiesis Hematopoietic Hematopoietic Neoplasms Hematopoietic stem cells Homeostasis Human Hypersensitivity Interleukin-3 Janus kinase Janus kinase 2 Juvenile Myelomonocytic Leukemia Kinetics Knock-in Mouse Knock-out Knockout Mice MPL gene Malignant - descriptor Mediating Modeling Molecular Molecular Conformation Mus Mutation Myelogenous Myeloproliferative disease Patients Pharmacology Phenocopy Phosphorylation Phosphotransferases Play Protein Tyrosine Kinase Proteins Proteomics Regulation Role Series Signal Pathway Signal Transduction Site Stem cell transplant Stem cells Testing Therapeutic Tyrosine Ubiquitin Ubiquitination Ursidae Family Viral actionable mutation cell growth cytokine gain of function human stem cells in vivo induced pluripotent stem cell insight kinase inhibitor loss of function mutation mouse model mutant novel novel therapeutics outcome forecast overexpression patient population progenitor receptor therapeutic target treatment strategy ubiquitin-protein ligase

项目摘要

Abstract Hematopoietic stem and progenitor cell (HSPC) homeostasis is regulated by intricate cytokine receptor and tyrosine kinase signaling pathways. Janus Kinase 2 (JAK2) is the key tyrosine kinase in the signaling pathway of an array of hematopoietic receptors, including thrombopoietin (TPO) receptor (MPL) in hematopoietic stem cells (HSCs) and granulocyte macrophage colony-stimulating factor receptor (GM-CSFR) in myeloid progenitors. While JAK2 plays an essential role in hematopoietic development, uncontrolled JAK2 signaling results in hematopoietic malignancies. Gain-of-function JAK2 mutations such as V617F are found in large populations of patients with myeloproliferative neoplasms (MPNs), a clonal HSC disease. Genetically targeting JAK2 abrogates MPN in mice; the JAK2V617F mutation is the driving mutation found in the HSCs of human MPN patients. However, current FDA-approved JAK kinase inhibitors have low curative potential, indicating the need for a better understanding of the regulation of JAK2 for efficient targeting. A number of E3 ubiquitin ligases for JAK2 have been proposed, but none of them, when lost in vivo, increases JAK2 protein level in HSPCs, expands HSC pool, or enhances multiple lineage hematopoiesis. Thus, the E3 ligase(s) regulating JAK2 turnover remain elusive. Through a series of biochemical and functional studies, we found that JAK2 stability is regulated by the CBL family E3 ubiquitin ligases, c-CBL and CBL-b, via the adaptor protein LNK (also called SH2B3). C-Cbl-/- mice phenocopy Lnk-/- mice, exhibiting an augmented HSPC pool with superior transplantability and hypersensitivity to cytokines. Importantly, CBL loss-of-function mutations have been found in a wide range of myeloid malignancies with the most frequent occurrence in chronic and juvenile myelomonocytic leukemia (CMML and JMML), both of which bear poor prognosis. Here, we propose to define the molecular basis underlying the regulation of JAK2 stability and signaling through this novel signaling axis, CBL/LNK/JAK2, and exploit it for therapeutic strategies in treating myeloid neoplasms. The following specific aims will be addressed: 1. Investigate mechanisms by which CBL regulates JAK2 ubiquitination, stability and signaling in hematopoietic cell lines and HSPCs from Cbl deficient and Cbl E3 ligase inactive mouse models. 2. Determine the influence of CBL on the stability of constitutively active JAK2 mutants and mutant JAK2- mediated MPN development. 3. Determine the role of CBL in regulating JAK2 level and signaling in primary human progenitors and explore therapeutic potential of JAK inhibition in treating murine and human myeloid malignancies with CBL mutations. Collectively, our data point to a novel direct role of CBL-mediated JAK2 ubiquitination and degradation in cytokine signaling and hematopoiesis. The proposed studies will likely provide mechanistic insights into the regulation of wild type JAK2 in normal HSPCs as well as JAK2 mutants in MPNs. In addition, our studies may reveal novel therapeutic strategies for the treatment of CBLmut myeloid malignancies with poor prognosis.

抽象的造血干细胞和祖细胞 (HSPC) 的稳态受复杂的细胞因子受体和酪氨酸激酶信号通路。 Janus 激酶 2 (JAK2) 是信号通路中关键的酪氨酸激酶一系列造血受体，包括造血干中的血小板生成素 (TPO) 受体 (MPL) 骨髓细胞 (HSC) 和粒细胞巨噬细胞集落刺激因子受体 (GM-CSFR) 祖先。虽然 JAK2 在造血发育中发挥着重要作用，但不受控制的 JAK2 信号传导导致造血系统恶性肿瘤。功能获得性 JAK2 突变（例如 V617F）发现于大骨髓增生性肿瘤 (MPN)（一种克隆性 HSC 疾病）患者群体。基因靶向 JAK2 消除小鼠 MPN； JAK2V617F 突变是在人类 MPN 的 HSC 中发现的驱动突变患者。然而，目前 FDA 批准的 JAK 激酶抑制剂的治疗潜力较低，表明需要更好地了解 JAK2 的调控以实现有效靶向。多种 E3 泛素连接酶 JAK2 已被提出，但它们在体内丢失时都不会增加 HSPC 中的 JAK2 蛋白水平，扩大 HSC 库，或增强多谱系造血作用。因此，调节 JAK2 的 E3 连接酶营业额仍然难以捉摸。通过一系列的生化和功能研究，我们发现JAK2的稳定性由 CBL 家族 E3 泛素连接酶 c-CBL 和 CBL-b 通过接头蛋白 LNK（也称为 SH2B3）。 C-Cbl-/- 小鼠表型 Lnk-/- 小鼠，表现出增强的 HSPC 库，具有优越的性能可移植性和对细胞因子的过敏。重要的是，已发现 CBL 功能丧失突变在多种骨髓恶性肿瘤中，最常见于慢性和青少年粒单核细胞白血病（CMML 和 JMML），两者预后均较差。在这里，我们建议定义通过这种新型信号轴调节 JAK2 稳定性和信号传导的分子基础， CBL/LNK/JAK2，并将其用于治疗骨髓肿瘤的治疗策略。具体如下将解决以下目标： 1. 研究 CBL 调节 JAK2 泛素化、稳定性和来自 Cbl 缺陷和 Cbl E3 连接酶失活小鼠模型的造血细胞系和 HSPC 中的信号传导。 2. 确定CBL对组成型活性JAK2突变体和突变体JAK2-稳定性的影响介导 MPN 的发展。 3.确定CBL在原发性中调节JAK2水平和信号传导的作用人类祖细胞并探索 JAK 抑制在治疗小鼠和人类骨髓细胞中的治疗潜力具有 CBL 突变的恶性肿瘤。总的来说，我们的数据表明 CBL 介导的 JAK2 具有新颖的直接作用细胞因子信号转导和造血作用中的泛素化和降解。拟议的研究可能会提供对正常 HSPC 中野生型 JAK2 以及 JAK2 突变体中 JAK2 调节的机制见解 MPN。此外，我们的研究可能揭示治疗 CBLmut 骨髓细胞的新治疗策略预后不良的恶性肿瘤。