Mechanisms and Consequences of Human Cytomegalovirus-Induced Cell Fusion

先天性巨细胞病毒感染是世界上最常见的先天性感染，约占所有新生儿的1%。高达15%的先天感染儿童患有长期的神经后遗症，如感觉神经性听力丧失或智力低下。关于决定病毒经胎盘传播的可能性和子代致病性的病毒和宿主因素知之甚少。巨细胞病毒感染其人类宿主中的许多不同类型的细胞和组织。上皮细胞和髓样细胞在病毒传播中起着重要作用。部分编码蛋白表现出很大的变异性，表明存在不同致病性的HCMV毒株。病毒包膜糖蛋白与进入受体相互作用，并介导病毒包膜与细胞膜的融合，由于它们是中和抗体的靶标，已被认为是病毒基因型和基因相关致病性的决定因素。然而，病毒糖蛋白在病毒附着、融合和进入细胞中的功能差异还没有被评估为致病的基础。我们最近发现，HCMV包膜糖蛋白B(GB)的毒株特异性变异可以导致病毒快速进入细胞，细胞融合(合胞体形成)和细胞基因组不稳定。此外，其他人的结果表明，在先天性HCMV分离株中存在形成合胞体的HCMV毒株。这些结果引发了这样一个问题：病毒糖蛋白的功能差异是否可能是不同毒株在传播和致病方面存在差异的基础。在拟议的项目中，我们希望确定在成纤维细胞、上皮细胞和巨噬细胞中介导细胞融合的HCMV包膜糖蛋白的变异体。我们将使用特征的HCMV毒株和先天性HCMV分离株来确定哪些包膜糖蛋白对合体表型有贡献，以及合体形成变异体是否经常出现在先天性HCMV分离株中。最后，我们将在体外感染人类胎盘组织样本，以确定相关细胞类型感染的变异体特异性差异。此外，我们的目标是建立一种系统，使我们能够在小动物模型中评估合胞变异体的致病性。

Congenital HCMV infection is the most common congenital infection worldwide, affecting approximately 1% of all newborns. Up to 15% of congenitally infected children suffer from long-term neurological sequelae such as sensineural hearing loss or mental retardation. Little is known about the viral and host factors determining the likelihood of transplacentar virus transmission and pathogenicity in the offspring.HCMV infects many different cell types and tissues in its human host. Epithelial and myeloid cells play important roles in viral dissemination. Some of the HCMV-encoded protein show substantial variability, suggesting that HCMV strains with different pathogenicity exist. Viral envelope glycoproteins, which interact with entry receptors and mediate fusion of the viral envelope with cellular membranes, have been proposed as determinants of viral genotypes and genotype-associated pathogenicity because they are targets of neutralizing antibodies. However, functional differences of viral glycoproteins in viral attachment, fusion, and entry have not been evaluated as the basis of pathogenicity.We have recently shown that strain-specific variants in the HCMV envelope glycoprotein B (gB) can cause rapid viral entry into cells, cell fusion (syncytium formation), and cellular genome instability. Moreover, results of others have shown that syncytium-forming HCMV strains are present in congenital HCMV isolates. These results give rise to the question whether functional differences of viral glycoproteins might be the basis of strain-specific differences in transmission and pathogenicity.In the proposed project we want to identify variants of HCMV envelope glycoproteins that mediate cell fusion in fibroblasts, epithelial cells, and macrophages. We will use characterized HCMV strains as well as congenital HCMV isolates to determine which envelope glycoproteins contribute to a syncytial phenotype and whether syncytium-forming variants are a frequent occurrence in congenital HCMV isolates. Finally, we will infect human placental tissue samples ex vivo to determine variant-specific differences in the infection of relevant cell types. Moreover, we aim to establish a system that allows us to evaluate the pathogenicity of syncytial variants in a small animal model.