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: 10120270
• 负责人: Theodore S Jardetzky
• 金额: $ 76.77万
• 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/10120270
• 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; Epithelium; 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; Organ; 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; 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.

人类巨细胞病毒（HCMV）是一种普遍存在的良性病毒。然而，HCMV经常导致移植患者器官排斥，并导致新生儿中枢神经系统发育的全身性疾病和缺陷。HCMV感染许多不同类型的细胞，包括上皮细胞和内皮细胞、胶质细胞、成纤维细胞和单核巨噬细胞。这种广泛的趋向性是通过不同的病毒糖蛋白途径进入不同细胞类型的能力促进的，包括：gH/gL/UL128-131，表示五聚体，gH/gL/gO，三聚体。我们关于HCMV如何进入上皮细胞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，五聚体在核内体中作用，促进gb介导的成纤维细胞融合，然后是病毒粒子包膜与细胞膜的融合。还有第三种形式的gH/gL， gH/gL与gB， gB-gH/gL的复合体，我们不知道gB-gH/gL是否促进病毒进入。鉴于三聚体和五聚体在病毒进入中的重要性，它们也是抗体（Abs）的重要靶点，被认为是HCMV疫苗设计中的关键角色。提出四个目标：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。这一目的将验证三聚体与细胞受体结合导致细胞表面交通，随后病毒颗粒内化进入细胞和下游五聚体介导的效应，促进病毒从内体进入细胞质。目标2。为了确定三聚体和三聚体PDGFRα的结构，确定三聚体的结构/功能关系，并鉴定其他三聚体受体。我们已经有了三聚体及其受体PDGFR的初步结构，我们将扩展这些结构研究并使用位点定向突变来测试功能。其他研究将确定三聚体受体对进入上皮细胞和内皮细胞很重要。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，来验证gB-gH/gL对HCMV进入很重要的假设。目标4。为了表征人血清中的三聚体特异性抗体，并与五聚体抗体进行比较。我们发现人血清中的三聚体和五聚体特异性抗体能够协同中和HCMV。我们将扩展这些研究，表征三聚体特异性抗体的患病率和效力，并确定这些抗体识别的三聚体中的表位。