Pioche-Durieu Catherine, Keryer Cécile, Souquère Sylvie, Bosq Jacques, Faigle Wolfgang, Loew Damarys, Hirashima Mitsuomi, Nishi Nozomu, Middeldorp Jaap, Busson Pierre
Institut Gustave Roussy, CNRS UMR 8126, Villejuif Cedex 94805, France.
J Virol. 2005 Nov;79(21):13326-37. doi: 10.1128/JVI.79.21.13326-13337.2005.
Nasopharyngeal carcinomas (NPC) are etiologically related to the Epstein-Barr virus (EBV), and malignant NPC cells have consistent although heterogeneous expression of the EBV latent membrane protein 1 (LMP1). LMP1 trafficking and signaling require its incorporation into membrane rafts. Conversely, raft environment is likely to modulate LMP1 activity. In order to investigate NPC-specific raft partners of LMP1, rafts derived from the C15 NPC xenograft were submitted to preparative immunoprecipitation of LMP1 combined with mass spectrometry analysis of coimmunoprecipitated proteins. Through this procedure, galectin 9, a beta-galactoside binding lectin and Hodgkin tumor antigen, was identified as a novel LMP1 partner. LMP1 interaction with galectin 9 was confirmed by coimmunoprecipitation and Western blotting in whole-cell extracts of NPC and EBV-transformed B cells (lymphoblastoid cell lines [LCLs]). Using mutant proteins expressed in HeLa cells, LMP1 was shown to bind galectin 9 in a TRAF3-independent manner. Galectin 9 is abundant in NPC biopsies as well as in LCLs, whereas it is absent in Burkitt lymphoma cells. In subsequent experiments, NPC cells were treated with Simvastatin, a drug reported to dissociate LMP1 from membrane rafts in EBV-transformed B cells. We found no significant effects of Simvastatin on the distribution of LMP1 and galectin 9 in NPC cell rafts. However, Simvastatin was highly cytotoxic for NPC cells, regardless of the presence or absence of LMP1. This suggests that Simvastatin is a potentially useful agent for the treatment of NPCs although it has distinct mechanisms of action in NPC and LCL cells.
鼻咽癌（NPC）在病因上与爱泼斯坦-巴尔病毒（EBV）相关，恶性NPC细胞对EBV潜伏膜蛋白1（LMP1）具有一致但异质性的表达。LMP1的运输和信号传导需要其整合到膜筏中。相反，筏环境可能调节LMP1的活性。为了研究LMP1的NPC特异性筏伴侣，将源自C15 NPC异种移植的筏进行LMP1的制备性免疫沉淀，并结合对共免疫沉淀蛋白的质谱分析。通过该程序，半乳糖凝集素9（一种β-半乳糖苷结合凝集素和霍奇金肿瘤抗原）被鉴定为一种新的LMP1伴侣。在NPC和EBV转化的B细胞（淋巴母细胞系[LCLs]）的全细胞提取物中，通过共免疫沉淀和蛋白质印迹证实了LMP1与半乳糖凝集素9的相互作用。使用在HeLa细胞中表达的突变蛋白，显示LMP1以不依赖TRAF3的方式结合半乳糖凝集素9。半乳糖凝集素9在NPC活检组织以及LCLs中丰富，而在伯基特淋巴瘤细胞中不存在。在随后的实验中，用辛伐他汀处理NPC细胞，辛伐他汀是一种据报道可使EBV转化的B细胞中的LMP1与膜筏解离的药物。我们发现辛伐他汀对NPC细胞筏中LMP1和半乳糖凝集素9的分布没有显著影响。然而，无论是否存在LMP1，辛伐他汀对NPC细胞都具有高度细胞毒性。这表明辛伐他汀虽然在NPC和LCL细胞中具有不同的作用机制，但它是一种潜在的治疗NPC的有用药物。
