Human Cytomegalovirus Entry into Cells Mediated by Pentamer and Trimer Complexes

五聚体和三聚体复合物介导的人巨细胞病毒进入细胞

• 批准号: 10468251
• 负责人: Theodore S Jardetzky
• 金额: $ 75.42万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-17 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10468251
• 关键词: Address; Affect; Antibodies; Antibody Response; Benign; Binding; Biochemical; Blood Vessels; Capsid; Cell Surface Receptors; Cell surface; Cells; Cellular Membrane; Complex; Cytomegalovirus; Cytomegalovirus Vaccines; Cytoplasm; Data; Defect; Development; Disease; Endosomes; Endothelial Cells; Endothelium; Epithelial; Epithelial Cells; Epitopes; Fetal Development; Fibroblasts; Follow-Up Studies; Foundations; Glycoproteins; Grant; Hepatitis; Hepatocyte; Herpesviridae; Human; Image; Immune Targeting; Immune system; Immunity; Immunize; Infection; Maps; Mediating; Membrane Fusion; Modeling; Molecular Conformation; Mutagenesis; Nervous system structure; Neuroglia; Newborn Infant; Oryctolagus cuniculus; PDGFRB gene; Pathology; Pathway interactions; Peptides; Platelet-Derived Growth Factor alpha Receptor; Pneumonia; Prevalence; Production; Property; Proteins; Receptor Cell; Role; Site; Structure; Structure-Activity Relationship; Systemic disease; Testing; Transplant Recipients; Tropism; Vaccines; Viral; Viral Fusion Proteins; Virion; Virus; Virus Diseases; base; cell type; design; experimental study; graft failure; human imaging; immunosuppressed; in vivo; interest; macrophage; monocyte; mutant; neonate; organ transplant rejection; particle; receptor; trafficking

Human cytomegalovirus (HCMV) is a ubiquitous, usually benign virus. Nevertheless, HCMV frequently contributes to rejection of organs in transplant patients and causes systemic disease and defects in the development of the CNS in neonates. HCMV infects many different cell types including epithelial and endothelial, glial cells, fibroblasts and monocyte-macrophages. This broad tropism is facilitated by a capacity to enter different cell types via distinct entry pathways involving different viral glycoproteins including: gH/gL/UL128-131, denoted the pentamer, gH/gL/gO, the trimer. Our model for how HCMV enters epithelial and endothelial cells suggests that HCMV trimers bind to cell surface receptors, e.g. PDGFRα viruses are internalized and pentamer acts in endosomes to promote gB-mediated fusion in fibroblasts, then of the virion envelope with cellular membranes. There is a third form of gH/gL, a complex of gH/gL with gB, gB-gH/gL and we do not know whether gB-gH/gL promotes in virus entry. Given their importance in virus entry, trimer and pentamer are also important targets of antibodies (Abs) and are considered key players in the design of HCMV vaccines. Four aims are proposed: Aim 1. To characterize pathways of HCMV entry into fibroblasts and epithelial and endothelial cells and determine where trimer and pentamer function. This aim will test the hypothesis that trimer binding to cellular receptors leads to cell surface traffic followed by internalization of virus particles into cells and downstream pentamer-mediated effects promoting virus exit from endosomes into the cytoplasm. Aim 2. To determine the structures of trimer and trimer:PDGFRα, define trimer structure/function relationships and identify other trimer receptors. We have a preliminary structure of trimer with its receptor PDGFR We will extend these structural studies and use site directed mutants to test function. Other studies will identify trimer receptors important for entry into epithelial and endothelial cells. Aim 3. To investigate how HCMV gB-gH/gL complexes function. We will test the hypothesis that gB- gH/gL is important for HCMV entry by using mutant forms of gB and gH/gL to block assembly of gB-gH/gL. Aim 4. To characterize trimer- specific Abs in human sera and compare to pentamer Abs. We made striking observations that trimer- and pentamer-specific Abs in human sera synergize to neutralize HCMV. We will extend these studies characterize the prevalence and potency of trimer-specific Abs and identify the epitopes in trimer recognized by these Ab.

人巨细胞病毒（HCMV）是一种普遍存在的，通常是良性的病毒。然而，HCMV经常导致移植患者的器官排斥，并导致新生儿CNS发育的全身性疾病和缺陷。HCMV感染许多不同的细胞类型，包括上皮细胞和内皮细胞、神经胶质细胞、成纤维细胞和单核细胞-巨噬细胞。这种广泛的嗜性通过经由涉及不同病毒糖蛋白的不同进入途径进入不同细胞类型的能力来促进，所述不同病毒糖蛋白包括：gH/gL/UL 128 -131，表示为五聚体，gH/gL/gO，三聚体。我们关于HCMV如何进入上皮和内皮细胞的模型表明，HCMV三聚体与细胞表面受体结合，例如PDGFRα病毒被内化，五聚体在核内体中起作用，促进成纤维细胞中gB介导的融合，然后是病毒体包膜与细胞膜的融合。还有第三种形式的gH/gL，gH/gL与gB的复合物，gB-gH/gL，我们不知道gB-gH/gL是否促进病毒进入。鉴于它们在病毒进入中的重要性，三聚体和五聚体也是抗体（Ab）的重要靶标，并且被认为是HCMV疫苗设计中的关键参与者。提出了四个目标：目标1。表征HCMV进入成纤维细胞、上皮细胞和内皮细胞的途径，并确定三聚体和五聚体的功能。该目的将检验以下假设：三聚体与细胞受体结合导致细胞表面运输，随后是病毒颗粒内化到细胞中，以及下游五聚体介导的促进病毒从内体进入细胞质的作用。目标二。确定三聚体和三聚体：PDGFRα的结构，确定三聚体结构/功能关系并鉴定其他三聚体受体。我们有一个三聚体与其受体PDGFR的初步结构，我们将扩展这些结构研究，并使用定点突变体来测试功能。其他研究将确定三聚体受体进入上皮和内皮细胞的重要性。目标3.探讨HCMV gB-gH/gL复合物的功能。我们将通过使用gB和gH/gL的突变形式来阻断gB-gH/gL的组装来检验gB-gH/gL对于HCMV进入是重要的假设。目标4。表征人血清中的三聚体特异性Ab并与五聚体Ab进行比较。我们做了惊人的观察，三聚体和五聚体特异性抗体在人血清中协同作用，以中和HCMV。我们将扩展这些研究，表征三聚体特异性抗体的流行和效力，并确定这些抗体识别的三聚体中的表位。