PIN1与EB病毒核抗原-1（EBNA1)特异性结合调控EBNA1功能促进鼻咽癌发生

Epstein–Barr Virus (EBV) maintenance is essential for tumorigenesis of nasopharyngeal carcinoma, so it is important to find out the key mechanism of long-term EBV infection in nasopharyngeal carcinoma. We have reported that prolyl isomerase PIN1 can activate the oncogenic signaling pathways and confer tumorigenic properties to nasopharyngeal carcinoma. In addition, the consistent over-expression of PIN1 in EBV-associated NPC suggested that it plays a role in EBV infection. However, the contributions of PIN1 over-expression to EBV-associated nasopharyngeal carcinoma development is still unclear. Our recent data has shown that PIN1 was highly expressed in NPC tumor lines and primary tumors that also showed concomitant EBNA1 over-expression by Western blot and immunohistochemistry .PIN1 was also found to co-localize and interact with EBNA1 in the EBV-positive NPC cells by immunofluorescence staining and co-immunoprecipitation assay respectively. In this study, we aimed to investigate the effect of PIN1 on EBNA1 functions and its role in the tumorigenesis of EBV-associated NPC. We hypothesis the WW domain of PIN1 interacts with EBNA1 at the specific serine-proline motif (Ser383 and Ser393). The binding may contribute to EBV long-term infection by modulating the EBV transcription activity and EBV DNA replication via OriP. In addition, the binding contributes to the transformation of nasopharynx epithelia cells, correlates with increased cancer signal pathway activity, regulates the downstream target oncogene and promotes the cell oncogenic properties. Our investigation indicates that PIN1 may serve as a potential therapeutic target for this EBV-associated nasopharyngeal carcinoma and may provide new perspective of tumorigenesis in EBV-associated nasopharyngeal carcinoma .

EBV的持续感染是鼻咽癌发生发展的必要条件，因此识别维持鼻咽癌中EBV持续感染的关键分子机制尤为重要。前期我们已报道鼻咽癌中肽基脯氨酰异构酶（PIN1）可激活肿瘤信号通路具有癌基因功能，且与EBV病毒感染密切相关，但PIN1与鼻咽癌发生的机制，尚不清楚。我们的近期预实发现：鼻咽癌石蜡切片中PIN1与EBV病毒共表达；鼻咽癌细胞系中EB病毒核抗原-1（EBNA1）和PIN1可发生直接相互作用。综合前期已发表结果及近期预实验数据，我们提出科学假说：①PIN1的WW区域与EBNA1Ser383、Ser393位点结合，增强EBV转录活性和EBV病毒DNA复制，维持EBV持续感染；② EBNA1和PIN1结合，促进上皮细胞转化，放大激活肿瘤信号通路，调控下游癌基因，增强细胞的致瘤性。本课题的完成有助于揭示鼻咽癌早期发病的新机制，为PIN1成为防治鼻咽癌的新靶点提供理论和数据支持。

鼻咽癌(Nasopharyngeal carcinoma)是一种与EBV病毒(Epstein-Barr virus, EBV)密切相关的上皮性恶性肿瘤。EB 病毒编码的EB病毒核抗原1（EBNA1）在鼻咽癌中持续表达且与病毒潜伏及病毒DNA复制密切相关。作为一种磷酸化蛋白质，EBNA1 的功能会受蛋白磷酸化和去磷酸化的影响。PIN1是一种新的肽脯氨酰异构酶 ,特异性地催化蛋白质中磷酸化的Ser/Thr-Pro酰胺键的顺反异构,改变蛋白质的构象 ,调控蛋白质的活性，磷酸化状态，蛋白与蛋白的相互作用,蛋白的稳定及亚细胞定位。肽脯氨酰异构酶PIN1 是否可以通过改变EBNA1的功能，从而影响其在鼻咽癌中的作用, 尽而成为治疗鼻咽癌的潜在靶基因, 是一项有意义的研究。.课题组按研究计划执行。从人体组织标本、细胞及动物实验3个层次对PIN1促进EBV相关鼻咽癌发生发展的机制进行了深入的研究。1完成了人体组织标本中鼻咽癌中内源性PIN1与EBNA1共定位及其与肿瘤关系的研究； 2完成PIN1功能的相关研究，PIN1过表达促进上皮细胞转化，放大激活肿瘤信号通路，增强细胞的致瘤性的研究，PIN1抑制降低细胞增殖，抑制肿瘤的生长。3完成EBNA1和PIN1相互结合对EBNA1功能的影响的研究，包括鉴别EBNA1与PIN1结合的位点及该结合对病毒转录和复制的影响。免疫荧光染色技术和免疫沉淀技术分别证实在C666-1中EBNA1和PIN1发生直接相互作用。EBNA1上Ser383和Ser393 区域，PIN1的 WW 区域是EBNA1和PIN1发生直接相互作用的位点。BNA1和PIN1的相互作用可以调节的 EBV 转录活性和 EBV DNA 复制, 从而影响鼻咽癌的发生发展。 4课题组在进行信号通路研究时，意外的发现Hippo信号通路的核心蛋白YAP1在EBV高表达的鼻咽癌细胞系中低表达，过表达EBNA1可以促进YAP1的磷酸化，而减少YAP1的整体表达。YAP1基因在鼻咽癌的发生发展中起到的作用，是我们后续研究的兴趣点。.我们的研究显示PIN1的过表达对鼻咽癌的发生起到重要的作用。EBNA1和PIN1的相互作用可以调节EBV的功能从而影响鼻咽癌的发生发展。这些都为PIN1成为治疗EBV相关肿瘤的靶向治疗目标奠定了坚实的基础。