Cell type-specific KSHV entry mediators for infection of epithelia and fibroblasts

用于感染上皮细胞和成纤维细胞的细胞类型特异性 KSHV 进入介质

• 批准号: 512328936
• 负责人: Dr. Alexander Hahn
• 金额: --
• 依托单位: Abteilung Infektionsbiologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/512328936?language=en
• 关键词: Cell; type; specific; KSHV; entry

Kaposi’s sarcoma-associated herpesvirus (KSHV) is a major cause of morbidity and mortality in Sub-Saharan Africa and poses a threat to immunocompromised individuals worldwide. KSHV enters host cells through mechanisms involving a variety of interactions of viral glycoproteins with cellular receptors, among those the interaction with integrins through glycoprotein B and the interaction with molecules of the Eph family of receptor tyrosine kinases through the gH/gL glycoprotein complex. Previous studies from our group and others have shown that, while important, these known receptor interactions do not fully explain KSHV entry into host cells, also depending on the target cell. We therefore focused on the less well-studied glycoprotein K8.1, a highly glycosylated and immunogenic protein of the KSHV envelope. We observed that deletion of the gene coding for K8.1 leads to strongly reduced infectivity on epithelial cells. We speculated that this is due to the interaction of K8.1 with a so-far unknown receptor on these cells and therefore have identified a number of K8.1-interacting cellular proteins by affinity purification and mass spectrometry that we will now test for function in the KSHV entry process through gene knockout. We will further pinpoint the exact function of K8.1 and its interaction with cellular receptors with regard to the attachment step, endocytosis, and membrane fusion. In a second approach, we performed preliminary studies to identify cellular proteins that interact with the known KSHV entry receptor EphA2. We identified both known and novel interactors and analyzed these candidates through gene knockout. Knockout of one of these factors, a small GTPase-binding protein, caused a significant reduction in the susceptibility to KSHV infection of a number of different cell lines and primary cells, but not of all cells that were tested. The reduction in infection upon knockout was most pronounced in fibroblasts but not observable e.g. in an epithelial cell line. We now want to study the exact function of this GTPase-binding protein for both endocytosis of KSHV and of the EphA2 receptor.

卡波西肉瘤相关疱疹病毒（KSHV）是撒哈拉以南非洲地区发病和死亡的主要原因，并对全球免疫功能低下的个体构成威胁。KSHV通过涉及病毒糖蛋白与细胞受体的各种相互作用的机制进入宿主细胞，其中包括通过糖蛋白B与整联蛋白的相互作用和通过gH/gL糖蛋白复合物与受体酪氨酸激酶的Eph家族分子的相互作用。我们小组和其他人以前的研究表明，虽然重要，但这些已知的受体相互作用并不能完全解释KSHV进入宿主细胞，这也取决于靶细胞。因此，我们专注于研究较少的糖蛋白K8.1，高度糖基化和免疫原性蛋白的KSHV包膜。我们观察到编码K8.1的基因的缺失导致对上皮细胞的感染性大大降低。我们推测这是由于K8.1与这些细胞上迄今未知的受体相互作用，因此已经通过亲和纯化和质谱鉴定了许多K8.1相互作用的细胞蛋白，我们现在将通过基因敲除测试KSHV进入过程中的功能。我们将进一步确定K8.1的确切功能及其与细胞受体的相互作用，包括附着步骤、内吞作用和膜融合。在第二种方法中，我们进行了初步研究，以鉴定与已知的KSHV进入受体EphA 2相互作用的细胞蛋白。我们确定了已知和新型相互作用因子，并通过基因敲除分析了这些候选因子。敲除这些因子之一，一种小的GTP酶结合蛋白，导致许多不同细胞系和原代细胞对KSHV感染的易感性显著降低，但不是所有测试的细胞。敲除后感染的减少在成纤维细胞中最明显，但在例如上皮细胞系中不可观察到。我们现在想研究这种GTP酶结合蛋白对KSHV和EphA 2受体的内吞作用的确切功能。