Programmed Cell Removal (PrCR) by Macrophages: recognition and phagocytosis of target cells

巨噬细胞的程序性细胞去除（PrCR）：靶细胞的识别和吞噬作用

基本信息

批准号：
10576906
负责人：
IRVING L. WEISSMAN
金额：
$ 40.47万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-01 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10576906
关键词：
Adhesions Affect Aging Agonist American Society of Clinical Oncology Apoptotic Asialoglycoproteins Atherosclerosis BCL2 gene Binding Binding Sites Biochemical Biochemical Pathway Blocking Antibodies Blood Bone Marrow C-terminal CD47 gene Cell Communication Cell Membrane Proteins Cell secretion Cell surface Cells Cellular biology Clinical Clinical Trials Degenerative Disorder Development Disease Disease remission Eating Enzymes Event Excision Exposure to Extracellular Space Fibroblasts Generations Glycoproteins Hematopoietic stem cells Heterogeneity Homeostasis Human In Vitro Individual Inflammation Inflammatory Invaded Investigation KDEL Motif Knowledge Label Lesion Life Ligands Longevity Macrophage Macrophage Activation Malignant Neoplasms Mediating Membrane Molecular Chaperones Multipotent Stem Cells Mus Neoplasms Neuraminidase Non-Malignant Normal tissue morphology Pathogenicity Pathologic Pathway interactions Peritoneal Phagocytes Phagocytosis Phase Process Proteins Proteolytic Processing Proteomics Publishing Reporting Research Rest Role Sialic Acids Sialoglycoproteins Signal Transduction Smooth Muscle Smooth Muscle Myocytes Sterility Surface Systemic Scleroderma TLR3 gene Testing Therapeutic Tissues Toll-like receptors Translating calreticulin cancer cell cancer type disorder prevention in vivo leukemic stem cell lysyl-aspartyl-glutamyl-leucine neoplastic cell neutrophil new therapeutic target nonalcoholic steatohepatitis novel overexpression prevent receptor therapeutic development

项目摘要

Project Summary Macrophage-mediated programmed cell removal (PrCR) allows clearance of living cells. We have shown that this phagocytic process can eliminate cancer cells that present an ‘eat me’ signal and have their dominant 'don't eat me' molecules blocked. We then further extended this to the clearance of other pathogenic or ‘expired’ cells. The key novel findings of our recent studies are: 1. Activated macrophages produce and secrete calreticulin (CRT); 2. Secreted CRT binds to surface asialoglycans on target cells to create ‘eat me signals' for macrophages. 3. Cancer cells and neutrophiles modulate their surface to expose asialoglycan binding sites for CRT that acts as an ‘eat me’ signal for macrophages. We propose that by binding both to asialoglycans and to pro-phagocytic receptors on macrophages (CD91/LPR1), CRT can bridge target cells to macrophages for clearance via PrCR. CRT is normally a resident ER protein containing a C-terminal KDEL retention signal, but it’s been shown that in dying cells CRT can be translocated to the cell surface. We found that upon macrophage activation via toll-like receptors (TLR), CRT can both translocate to the cell surface and be secreted, leading to increased PrCR of either WT (dying) or Bcl-2+ (viable) peritoneal neutrophils and cancer cells on which the ‘don’t eat me’ signal CD47 is either absent or blocked. Based on these findings we proporse: (1) to elucidate the signals affecting the macrophage that result in CRT translocation to the cell surface and secretion of soluble forms of CRT; (2) to elucidate the mechanisms regulating the availability of asialoglycan-containing binding sites for CRT on target cells. Elucidating the individual mechanisms in macrophages and target cells required for PrCR and understanding the cross-talk within macrophage:target-cell interaction can have broad therapeutic implications. In Aim 1 we will investigate which signals stimulate macrophages to increase cell surface expression and secretion of CRT for PrCR and the heterogeneity of macrophages that can carry out PrCR in vitro and sterile inflammation in vivo. In Aim 2 we will employ proteomic analysis to define and characterize the different proteoforms of CRT originating in macrophages before or after stimulation: the ER form vs. cell-surface- bound, vs. soluble secreted CRT, vs. CRT that is bound to asialoglycans on target cells. We have preliminary evidence that the secreted form of CRT does not contain the KDEL motif and that potential proteolytic processing leads to the formation of the different CRT forms. Lastly in Aim 3 we will study the initiating signaling events and enzymes that affect the addition or removal of sialic acid, the activity of which determines the level of exposed asialoglycans and thus the binding of CRT to the surface of cells destined for elimination. These studies will shed light on a novel mechanism by which macrophages detect cells that are to be removed from the body. Our findings will have a broad relevance to cancer, degenerative disease, and inflammatory lesions and will likely reveal new therapeutic targets for life altering disease conditions.

项目摘要巨噬细胞介导的编程细胞去除（PRCR）允许清除活细胞。我们已经表明这种吞噬过程可以消除发出“我的饮食”信号并具有主导性的癌细胞“不要吃我的分子被阻塞了。然后，我们将其扩展到清除其他病原或“过期”细胞的清除率。我们最近研究的主要新颖发现是：1。活化的巨噬细胞产生和秘密钙网蛋白（CRT）; 2。分泌的CRT与靶细胞上的表面Asialogllycan结合，以创建“我的信号” 巨噬细胞。 3。癌细胞和中性粒细胞调节其表面以暴露于Asialoglycan结合位点 CRT充当巨噬细胞的“吃我”信号。我们提出，通过将其绑定到Asialogllycans和巨噬细胞（CD91/LPR1）的促脑细胞受体，CRT可以将靶细胞桥接到巨噬细胞上通过PRCR清除。 CRT通常是含有C末端KDEL保留信号的常驻ER蛋白，但已经表明，在垂死的细胞中，CRT可以转移到细胞表面。我们发现巨噬细胞通过Toll样受体（TLR）激活，CRT既可以转移到细胞表面并分泌，从而导致增加了WT（垂死）或Bcl-2+（可行）腹膜中性粒细胞和癌细胞的PRCR增加。吃我的信号CD47是不存在的或阻塞的。基于这些发现，我们主张：（1）阐明影响巨噬细胞的信号，导致CRT转移到细胞表面和固体的分泌 CRT的形式；（2）阐明了计算含有含有asialogly的结合位点的机制对于目标细胞上的CRT。阐明PRCR所需的巨噬细胞和目标细胞中的各个机制并了解巨噬细胞中的串扰：目标细胞相互作用可以进行广泛的治疗含义。在AIM 1中，我们将研究哪些信号刺激巨噬细胞增加细胞表面 CRT对PRCR的表达和分泌以及巨噬细胞的异质性，可以在体内体外和无菌炎症。在AIM 2中，我们将采用蛋白质组学分析来定义和表征 CRT的不同蛋白质类型源自巨噬细胞之前或之后的巨噬细胞：ER形式与细胞表面 - 绑定，与固体分泌的CRT，与靶细胞上的Asialoglycans结合的CRT。我们有初步 CRT的分泌形式不包含KDEL基序和潜在蛋白水解处理的证据导致形成不同的CRT形式。最后，在AIM 3中，我们将研究启动信号事件和影响唾液酸添加或去除的酶，其活性决定了暴露的水平 Asialoglycans，因此CRT与注定要消除的细胞表面结合。这些研究将丢弃对巨噬细胞检测要从体内去除的细胞进行的新机制进行了启示。我们的调查结果将与癌症，退化性疾病和炎症性病变具有广泛的相关性，并且很可能揭示了改变疾病疾病的新治疗靶标。