Cellular Interactions with Thrombospondin

细胞与血小板反应蛋白的相互作用

基本信息

批准号：
10487187
负责人：
david d roberts
金额：
$ 156.25万
依托单位：
DIVISION OF CLINICAL SCIENCES - NCI
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10487187
关键词：
7-methylguanosine Acute Alkylation Angiogenesis Inhibition Anthracycline Autophagocytosis Blocking Antibodies Blood Platelets Blood Pressure Blood flow Bone Marrow CD47 gene CD8-Positive T-Lymphocytes Cell Surface Receptors Cell Survival Cells Co-Immunoprecipitations Code Complex Cyclic GMP Cyclic GMP-Dependent Protein Kinases Cysteine DNA Repair Data Databases Extracellular Matrix Family suidae Frequencies Gene Family Gene Proteins Genes Genome Glycoproteins Growth Hemostatic function Hereditary Disease Human Individual Integrin Binding Ionizing radiation Irradiated tumor Ischemia KDR gene Knockout Mice Ligand Binding Ligands Link Maintenance Malignant Neoplasms Mass Spectrum Analysis Mediating Membrane Messenger RNA Metabolism MicroRNAs Missense Mutation Modification Mus Myocardium NOS3 gene Natural Killer Cells Nitric Oxide Non-Malignant Nuclear Export Null Lymphocytes Play Probability Proteins RNA RNA Caps Radiation Injuries Rattus Receptor Cell Receptor Signaling Recovery Regulation Reperfusion Injury Reporting Reproduction Role Signal Pathway Signal Transduction Signal Transduction Pathway Smooth Muscle Myocytes Soluble Guanylate Cyclase Stromal Cells T-Lymphocyte THBS1 gene THBS2 gene Tenascin Therapeutic Thrombospondin 1 Thrombospondins Tissue Survival Tissues Transgenic Mice Tumor Immunity UBQLN1 gene Untranslated RNA Vascular Smooth Muscle adaptive immune response angiogenesis anti-cancer anti-tumor immune response base c-myc Genes cancer cell cancer immunotherapy cancer stem cell cell type connective tissue growth factor cytotoxic environmental stressor exome sequencing exportin 1 protein extracellular vesicles gene discovery immune checkpoint insight ischemic injury knock-down leptomycin B loss of function loss of function mutation macrophage malignant breast neoplasm member mouse model neoplastic cell novel radiation resistance receptor receptor binding response self-renewal soft tissue stem cells therapeutic target tissue culture tissue stress trafficking transcription factor transcriptomics tumor tumor progression tumor xenograft vesicular release

项目摘要

CD47 is a marker of self and a signaling receptor for thrombospondin-1 that is also an enriched membrane component of extracellular vesicles (EVs) released by various cell types. Previous studies identified CD47-dependent functional effects of T cell EVs on target cells, mediated by delivery of their RNA contents, and enrichment of specific subsets of coding and noncoding RNAs in CD47+ EVs. Here, transcriptomic analyses of EVs released by human and murine T cells revealed a global CD47-dependent enrichment of 5'-7-methylguanosine-capped microRNAs and mRNAs in EVs relative to levels in the cells. Knockdown or loss of CD47 in wild type Jurkat T cells or treatment with thrombospondin-1 enhanced levels of specific capped-RNAs released in EVs, and re-expressing CD47 in null cells decreased their levels. Mass spectrometry and co-immunoprecipitation identified specific interactions of CD47 with components of the exportin-1/Ran nuclear export complex and its known cargo proteins and between the CD47 cytoplasmic adapter ubiquilin-1 and the exportin-1/Ran complex. Interaction with CD47 was inhibited following alkylation of exportin-1 at Cys528 by leptomycin B. Leptomycin B modification of exportin-1 increased levels of cap-dependent RNAs and their association with exportin-1 in EVs released from wild type but not CD47-deficient cells. Therefore, CD47 regulates the trafficking of cap-dependent RNAs to EVs through physical interactions with the exportin-1/Ran transport complex. Targeted gene disruption in mice has provided valuable insights into the functions of matricellular proteins. Apart from missense and loss of function mutations that have been associated with inherited diseases, however, their functions in humans remain unclear. The availability of deep exome sequencing data from over 140,000 individuals in the Genome Aggregation Database provided an opportunity to examine intolerance to loss of function and missense mutations in human matricellular genes. The probability of loss-of-function intolerance (pLI) differed widely within members of the thrombospondin, CYR61/CTGF/NOV (CCN), tenascin, small integrin-binding ligand N-linked glycoproteins (SIBLING), and secreted protein, acidic and rich in cysteine (SPARC) gene families. Notably, pLI values in humans had limited correlation with viability of the corresponding homozygous null mice. Among the thrombospondins, only THBS1 was highly loss-intolerant (pLI = 1). In contrast, Thbs1 is not essential for viability in mice. Several known thrombospondin-1 receptors were similarly loss-intolerant, although thrombospondin-1 is not the exclusive ligand for some of these receptors. The frequencies of missense mutations in THBS1 and the gene encoding its signaling receptor CD47 indicated conservation of some residues implicated in specific receptor binding. Deficits in missense mutations were also observed for other thrombospondin genes and for SPARC, SPOCK1, SPOCK2, TNR, and DSPP. The intolerance of THBS1 to loss of function in humans and elevated pLI values for THBS2, SPARC, SPOCK1, TNR, and CCN1 support important functions for these matricellular protein genes in humans, some of which may relate to functions in reproduction or responding to environmental stresses.

CD47是自我的标记和血小板传播1的信号受体，它也是各种细胞类型释放的细胞外囊泡（EV）的富集膜成分。先前的研究确定了T细胞EVS对靶细胞的CD47依赖性功能效应，通过递送其RNA含量介导，以及在CD47+ EV中富集编码和非编码RNA的特定子集。在这里，人和鼠T细胞释放的电动汽车的转录组分析显示，与细胞水平相对于电动汽车中的5'-7-7-甲基鸟苷笼型microRNA和mRNA的全球CD47依赖性富集。野生型Jurkat T细胞中CD47的敲低或丢失或用血小板蛋白1的处理增强了EV中释放的特定限制RNA水平，并在零细胞中重新表达CD47降低了其水平。质谱和共免疫沉淀确定了CD47与Exportin-1/RAN核出口复合物及其已知的货物蛋白的特定相互作用，以及CD47细胞质适配器Ubiquilin-1和Exportin-1/RAN复合物之间的特定相互作用。嗜血霉素B的烷基化在Cys528上的Exportin-1烷基化烷基化B.嗜经霉素B的EXTORTIN-1的相互作用在Cys528上的相互作用增加了cap依赖性RNA的水平及其与野生型释放的EVS中的Evelin-1的相关性，但不是CD47缺乏的细胞。因此，CD47通过与Exportin-1/RAN运输复合物的物理相互作用来调节CAP依赖性RNA对EV的运输。小鼠的靶向基因破坏为矩阵蛋白的功能提供了宝贵的见解。但是，除了与遗传疾病相关的功能突变的丧失外，它们在人类中的功能仍然不清楚。来自基因组聚集数据库中140,000多名个体的深层外显子组测序数据提供了一个机会，可以检查人类母细胞基因中功能丧失和功能丧失和错义突变的不耐受。功能丧失不宽容（PLI）的可能性在血小板蛋白，CYR61/CTGF/NOV（CCN），Tenascin，小内整合素结合配体N-链接糖蛋白（sibling）（sibling），酸性蛋白质，酸性蛋白质，酸性蛋白质，酸性蛋白质（Sparcarce）（Sparc）的Gene formals（Sparc）Gene中的可能性差异很大。值得注意的是，人类中的PLI值与相应纯合子无效小鼠的生存力的相关性有限。在血小板传播中，只有THBS1是高度损失智力（PLI = 1）。相比之下，THBS1对于小鼠的生存能力并不是必需的。尽管某些受体中的几种已知的血小板传播1受体同样具有损失智能剂，但并不是其中一些受体的独家配体。 THBS1中错义突变的频率和编码其信号受体CD47的基因表明保存了与特定受体结合有关的某些残基。对于其他血小板传播基因以及SPARC，SPOCK1，SPOCK2，TNR和DSPP，还观察到错义突变的缺陷。 THBS1对人类功能丧失的不耐受和THBS2，SPARC，SPOCK1，TNR和CCN1的PLI值的升高支持人类这些基质蛋白基因的重要功能，其中一些可能与生殖或对环境应力的反应有关。