PDGF Receptor Regulation by CBL

CBL 对 PDGF 受体的调节

• 批准号: 7178463
• 负责人: Hamid Band
• 金额: $ 15.44万
• 依托单位: NORTHSHORE UNIVERSITY HEALTHSYSTEM
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-03-01 至 2007-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7178463
• 关键词: Accounting; Antineoplastic Agents; Apoptosis; Back; Binding; Biochemical; CBL Protein; CBL gene; Cancer Biology; Cell Proliferation; Cell physiology; Cell surface; Cells; Chimeric Proteins; Clathrin-Coated Vesicles; Complex; Cytoplasmic Tail; Defect; Development; Dimerization; Dominant-Negative Mutation; Down-Regulation; Embryo; Endocytosis; Epidermal Growth Factor Receptor; Equilibrium; Event; Family; Fibroblasts; Glioma; Goals; Homeostasis; Human; Hyperactive behavior; Lead; Ligand Binding; Ligands; Link; Lysosomes; Malignant Neoplasms; Mammals; Mediating; Modeling; Molecular; Mus; Myeloid Leukemia; Numbers; Oncogene Proteins; Oncogenes; PDGF receptor tyrosine kinase; PDGFRB gene; Phosphorylation; Phosphotransferases; Physiological; Platelet-Derived Growth Factor Receptor; Play; Preventive; Process; Protein Tyrosine Kinase; Proteins; Radiation; Radio; Receptor Activation; Receptor Down-Regulation; Receptor Protein-Tyrosine Kinases; Receptor Signaling; Recycling; Regulation; Research; Role; Route; Signal Transduction; Signaling Protein; Sorting - Cell Movement; System; TSG101 gene; Testing; Therapeutic Agents; Tissues; Tumor Suppressor Proteins; Ubiquitin; Ubiquitination; Vacuole; Work; Wound Healing; Yeasts; angiogenesis; base; cell killing; chemotherapeutic agent; chemotherapy; inhibitor/antagonist; insight; migration; mutant; neoplastic cell; novel; novel therapeutics; receptor; trafficking; tumorigenesis; ubiquitin ligase; yeast genetics

DESCRIPTION (provided by applicant): PDGF receptor tyrosine kinases play crucial physiological roles in cell proliferation, migration and differentiation during development, wound healing and tissue remodeling. Activation of these receptors is linked to cell proliferation; invasiveness and angiogenesis in human cancers, in particular glial tumors, and PDGFR fusion oncogenes are responsible for a subset of myelogenous leukemia. Indeed, inhibitors of PDGFR kinase activity are being evaluated as anti-cancer agents. Understanding the mechanisms that regulate the level of signaling downstream of PDGF receptors is therefore a major goal of research in cell and cancer biology. Ligand-induced downregulation, representing a balance between lysosomal degradation and recycling to cell surface, constitutes a major determinant of signaling potency of receptor tyrosine kinases. Our recent work has established the Cbl proto-oncoprotein as a crucial regulator of PDGF receptor down-regulation. Cbl, an ubiquitin ligase, targets activated PDGF receptors for ubiquitination that in turn facilitates their lysosomal sorting. How Cbl-dependent ubiquitination functions as a lysosomal sorting signal for PDGF receptors is unknown. Based on recent yeast genetic studies, we hypothesize the role of a novel endosomal sorting complex, ESCRT-1, which incorporates the TSG101 tumor suppressor protein, in Cbl- and ubiquitin-dependent lysosomal sorting of PDGF receptor. Here, we will test this hypothesis using a number of complementary approaches. We will use Cbl-deficient mouse embryonic fibroblasts, Cbl-insensitive PDGFR mutants expressed in PDGFR-null fibroblasts, cells with a conditional defect in ubiquitination, and PDGFR-ubiquitin fusion proteins to establish the essential role of Cbl-dependent ubiquitination in PDGFR down-regulation. We will use dual immunolabeling studies to identify the compartment(s) where Cbl-dependent lysosomal sorting of PDGFR occurs. We will use biochemical and co-localization analyses to establish that PDGFR and ESCRT-1 interact, and use over-expression and dominant-negative approaches to establish the requirement of this novel protein complex in lysosomal sorting of PDGFR. Functional analyses will assess if ESCRT-1 complex is critical in Cbl-mediated downregulation of PDGFR signaling. Insights gained through these studies should enhance the molecular understanding of a basic cellular process, downregulation of activated receptor tyrosine kinases. Given the role of PDGF receptors in maligant cell proliferation, invasiveness and angiogenesis, our studies are particularly relevant to cancer. Identification of novel regulatory mechanisms of receptor tyrosine kinase function may provide avenues to develop newer therapeutic agents and/or chemo- and radio-sensitizers.

描述（由申请人提供）：PDGF受体酪氨酸激酶在发育过程中细胞增殖、迁移和分化、伤口愈合和组织重塑中起着至关重要的生理作用。这些受体的激活与细胞增殖有关；人类癌症，特别是神经胶质肿瘤的侵袭性和血管生成，以及PDGFR融合癌基因是骨髓性白血病的一个亚群的原因。事实上，PDGFR激酶活性抑制剂正在被评估为抗癌药物。因此，了解调节PDGF受体下游信号水平的机制是细胞和癌症生物学研究的主要目标。配体诱导的下调，代表了溶酶体降解和细胞表面再循环之间的平衡，构成了受体酪氨酸激酶信号效力的主要决定因素。我们最近的工作已经确定了Cbl原癌蛋白是PDGF受体下调的关键调节因子。Cbl是一种泛素连接酶，靶向激活的PDGF受体进行泛素化，从而促进它们的溶酶体分选。ccl依赖性泛素化如何作为PDGF受体的溶酶体分选信号起作用尚不清楚。基于最近的酵母遗传学研究，我们假设了一种新的内体分选复合体ESCRT-1在PDGF受体的Cbl和泛素依赖性溶酶体分选中的作用，ESCRT-1包含TSG101肿瘤抑制蛋白。在这里，我们将使用一些互补的方法来检验这一假设。我们将使用缺乏ccl的小鼠胚胎成纤维细胞、在PDGFR缺失的成纤维细胞中表达的ccl不敏感的PDGFR突变体、泛素化有条件缺陷的细胞以及PDGFR-泛素融合蛋白来确定ccl依赖的泛素化在PDGFR下调中的重要作用。我们将使用双重免疫标记研究来确定发生ccl依赖性PDGFR溶酶体分选的隔室。我们将使用生化和共定位分析来确定PDGFR和ESCRT-1相互作用，并使用过表达和显性阴性方法来确定PDGFR溶酶体分选对这种新型蛋白复合物的需求。功能分析将评估ESCRT-1复合物是否在ccl介导的PDGFR信号下调中起关键作用。通过这些研究获得的见解应该增强对基本细胞过程的分子理解，即激活受体酪氨酸激酶的下调。鉴于PDGF受体在恶性细胞增殖、侵袭性和血管生成中的作用，我们的研究与癌症特别相关。受体酪氨酸激酶功能的新调控机制的鉴定可能为开发新的治疗剂和/或化学和放射增敏剂提供途径。