Calreticulin-mediated protein folding in health and disease

健康和疾病中钙网蛋白介导的蛋白质折叠

• 批准号: 10599361
• 负责人: MALINI RAGHAVAN
• 金额: $ 45.13万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-03-10 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10599361
• 关键词: Abnormal Cell; Address; Affect; Affinity; Alleles; Antigen Presentation; Antigen-Presenting Cells; Antigens; Apoptotic; Base Pairing; Binding; Binding Proteins; Binding Sites; Blood Platelets; Bone Marrow; Bone Marrow Cells; C-terminal; CD8-Positive T-Lymphocytes; CD8B1 gene; Calcium; Calcium Binding; Calcium Signaling; Cell Maturation; Cell Proliferation; Cell secretion; Cell surface; Cells; Charge; Chronic; Complex; Dendritic Cells; Dependence; Disease; Disulfides; Endoplasmic Reticulum; Environment; Fibrosis; Frameshift Mutation; Genes; Globulins; Glycoproteins; Growth Factor Receptors; Health; Hemorrhagic Thrombocythemia; Homeostasis; Human; Hyperplasia; Immune Evasion; Immune response; Immune system; Immunity; Immunologics; Immunotherapeutic agent; Impairment; Induced Mutation; Infection; Inflammation; Knowledge; Light; Link; MPL gene; Major Histocompatibility Complex; Malignant Neoplasms; Measures; Mediating; Mediator; Megakaryocytes; Mission; Molecular; Molecular Chaperones; Mutagenesis; Mutate; Mutation; Myelofibrosis; Myeloproliferative disease; Nature; Oncogenic; Pathway interactions; Patients; Peptides; Phagocytosis; Phagocytosis Induction; Phosphatidylserines; Primary Myelofibrosis; Process; Production; Proteins; Recurrence; Research; Sampling; Site; Somatic Mutation; Specificity; Splenomegaly; T cell response; United States National Institutes of Health; calreticulin; cell transformation; combat; dimer; disability; disulfide bond; driver mutation; human disease; insertion/deletion mutation; insight; leukemia; lysyl-aspartyl-glutamyl-leucine; monocyte; mutant; neoantigens; neoplasm immunotherapy; novel; peptide I; precursor cell; protein activation; protein folding; response; tool

Abstract Calreticulin (CRT) is a calcium-binding endoplasmic reticulum (ER) chaperone that is important for the folding of many N-linked glycoproteins. It is best known for its function in the assembly of major histocompatibility complex (MHC) class I (MHC-I) molecules, which present peptide antigens to CD8+ T cells for protective immunity against cancers and infections. Recent studies show that somatic frameshift mutations that alter the C-terminus of CRT are one class of driver mutations in myeloproliferative neoplasms (MPN). A 52 base-pair deletion mutant CRTDel52 and a 5 base-pair insertion mutant CRTIns5 are recurrent mutations in essential thrombocythemia (ET), primary myelofibrosis (PMF) and post-ET myelofibrosis (MF). It is our overall hypothesis that the changes to the sequence and charge of the C-terminal domains of CRT induced by the mutations enable both megakaryocyte transformation and immune system evasion in MPN. Although oncogenic transformation in megakaryocytes requires both mutated CRT and the thrombopoietin receptor/myeloproliferative leukemia protein (Mpl), the molecular mechanisms of mutant CRT-mediated constitutive activation of Mpl are unknown. CRT is known to be important for ER calcium storage and cellular calcium signaling, but precisely how MPN mutations alter these processes is not understood. The human MHC-I locus is highly polymorphic. While it is known that MPN-linked mutant CRTs are ineffectively incorporated into the MHC-I peptide-loading complex (PLC), it is unknown whether MPN CRT mutants have differential influences on the assembly of various MHC-I allotypes. Studies are proposed here to address these gaps in knowledge. We present evidence that disulfide-linked dimers of CRTDel52 are important for Mpl activation. The nature of CRTDel52 dimers and their complexes with Mpl will be elucidated using structural and mutagenesis studies. The effects of CRT deficiency, haploinsufficiency and mutation upon cellular calcium signaling will be examined. Based on the knowledge of variable dependencies on CRT for cell-surface expression of MHC-I allotypes, we will examine whether MPN CRT mutants impair MHC class I assembly for some allotypes, but alter assembly and peptide display for some other MHC-I allotypes. Such alterations could be exploited for immunotherapeutic targeting of MPN. Finally, mutant CRT expression is also detectable in blood monocytes of MPN patients, leading to predictions of alterations to antigen presenting cell (APC) calcium signaling and phagocytosis, which will be examined. Taken together, these studies will further our understanding of how mutations of a ubiquitous ER chaperone drive cell transformation, and influence the fundamental immunological processes of antigen presentation and phagocytosis.

摘要 钙网蛋白（CRT）是一种钙结合的内质网（ER）伴侣，在折叠 许多N-连接的糖蛋白。它最为人所知的功能是在组装主要组织相容性 MHC复合物I类（MHC-I）分子，其将肽抗原呈递给CD 8 + T细胞以提供保护性 对癌症和感染的免疫力。最近的研究表明，体细胞移码突变，改变了 CRT的C端是骨髓增生性肿瘤（MPN）中的一类驱动突变。52个碱基对 缺失突变体CRTDel 52和5个碱基对插入突变体CRTIns 5是必需的CRTIns 5的复发突变。 血小板增多症（ET）、原发性骨髓纤维化（PMF）和ET后骨髓纤维化（MF）。这是我们的整体 假设CRT的C-末端结构域的序列和电荷的变化是由 突变使MPN中的巨核细胞转化和免疫系统逃避成为可能。虽然 巨核细胞的致癌转化需要突变的CRT和血小板生成素 受体/骨髓增生性白血病蛋白（Mpl），突变CRT介导的分子机制 Mpl的组成性激活是未知的。已知CRT对于ER钙储存和细胞增殖是重要的。 钙信号，但确切地说，MPN突变如何改变这些过程还不清楚。人类 MHC-I位点具有高度多态性。虽然已知MPN连锁的突变型CRT不能有效地抑制 由于MPN CRT突变体被整合到MHC-I肽加载复合物（PLC）中，目前尚不清楚MPN CRT突变体是否具有 对各种MHC-I同种异型组装的差异影响。在此提出研究来解决这些问题 知识的差距。我们提出的证据表明，CRTDel 52的二硫键连接的二聚体对于Mpl activation. CRTDel 52二聚体和它们与Mpl的复合物的性质将使用结构和分子生物学方法来阐明。 诱变研究。CRT缺陷、单倍不足和突变对细胞钙的影响 将检查信令。基于细胞表面CRT的变量依赖性知识， MHC-I同种异型表达，我们将研究MPN CRT突变体是否损害MHC-I类组装， 一些同种异型，但改变一些其他MHC-I同种异型的组装和肽展示。这种改变可以 用于MPN的免疫靶向。最后，突变CRT表达也可在 MPN患者的血液单核细胞，导致预测抗原呈递细胞（APC）钙 信号传导和吞噬作用，这将被检查。综合起来，这些研究将进一步促进我们的 了解普遍存在的ER伴侣蛋白突变如何驱动细胞转化，并影响细胞增殖。 抗原呈递和吞噬作用的基本免疫过程。