SLC26A9介导EB病毒感染鼻咽上皮细胞的作用及分子机制

Epstein-Barr virus infection is closely related to the pathogenesis of various tumors, and the incidence of nasopharyngeal carcinoma has been studied for nearly half a century, but the receptors and infection mechanisms of nasopharyngeal epithelial cells are still unclear. Using the expression profile chip technology, our group found that SLC26A9 was highly expressed in the nasopharyngeal epithelial cell model with high Epstein-Barr virus infection. In addition, we found that interference with SLC26A9 significantly inhibited Epstein-Barr virus infection of nasopharyngeal epithelial cells, while overexpression of SLC26A9 promoted viral infection. Therefore, we speculate that SLC26A9 can be used as a key receptor for Epstein-Barr virus to mediate viral infection of nasopharyngeal epithelial cells. This project intends to further use CRISPR/Cas9, antibody blocking, protein blocking, immunofluorescence and other techniques to confirm that SLC26A9 is a key receptor for Epstein-Barr virus infection of nasopharyngeal epithelial cells. Specific pathways of SLC26A9-mediated Epstein-Barr virus infection in epithelial cells were revealed by inhibitor blocking experiments, virus binding and entry experiments. The binding proteins interacting with SLC26A9 were screened by Pull-down and mass spectrometry, and the relationship between these binding proteins and viral infection was investigated. To clarify the specific molecular mechanism of SLC26A9 as a key receptor-mediated Epstein-Barr virus infection, it provides a new target for nasopharyngeal carcinoma intervention.

EB病毒感染与多种肿瘤发病密切相关，与鼻咽癌的发病研究已近半个世纪，但其感染鼻咽上皮细胞的受体及感染机制仍不清楚。本课题组通过表达谱芯片发现EB病毒高效感染的鼻咽上皮细胞模型中SLC26A9高表达。同时我们发现干扰SLC26A9显著抑制EB病毒感染鼻咽上皮细胞，而过表达SLC26A9能促进病毒的感染。因此，我们推测SLC26A9作为EB病毒的受体介导病毒感染鼻咽上皮细胞。本项目拟进一步利用CRISPR/Cas9、抗体阻断、蛋白阻断、免疫荧光等技术证实SLC26A9是EB病毒感染鼻咽上皮细胞的关键受体；利用抑制剂阻断和病毒结合进入等实验揭示SLC26A9介导EB病毒感染上皮细胞的具体路径；利用Pull-down和质谱技术筛选与SLC26A9相互作用的结合蛋白，并研究这些结合蛋白与病毒感染的关系，进一步明确SLC26A9作为关键受体介导EB病毒感染的具体分子机制，为鼻咽癌干预提供新靶标。

EB病毒（EBV）被认为是一种与上皮和淋巴肿瘤相关的病原体。EB病毒进入靶细胞是一个复杂的过程，由病毒包膜糖蛋白与细胞受体的结合和相互作用启动。近期，我们课题组报道了Neuropilin 1（NRP1）和 Ephrin receptor A2（EphA2）显著增强EB病毒进入鼻咽上皮细胞。然而，EB病毒感染上皮细胞的机制尚不清楚。近期，我们通过芯片技术和RNA干扰筛选分析发现，SLC26A9对EB病毒进入上皮细胞很重要。SLC26A9过表达显著增加了EB病毒感染上皮细胞，而敲低SLC26A9则降低了EB病毒的感染。SLC26A9抗体能有效阻断EB病毒感染上皮细胞。EB病毒糖蛋白gH和gB是B细胞和上皮细胞感染的关键融合蛋白。我们发现SLC26A9直接与EB病毒 gH和gB相互作用，以促进EB病毒內吞和融合。利用激光共焦显微镜技术，我们观察到SLC26A9与EEA1（内体标记物）和LAMP1（溶酶体标记物）共定位，表明SLC26A6介导的EBV内化可能依赖于内吞作用。此外，敲低SLC26A9能抑制溶酶体酸化。缺失STAS结构域和酸化抑制剂氯喹都能有效阻断EB病毒感染上皮细胞。总之，我们的结果表明SLC26A9可以通过介导内吞酸化以促进EB病毒感染。