EHD2与IRF-1相互作用在非霍奇金淋巴瘤CAM-DR中的作用及机制

The activation of NF-κB is considered as one of the major reasons for cell adhesion-mediated drug resistance (CAM-DR). It has been reported that interferon regulatory factor-1 (IRF-1) plays an important role in NF-κB activation. Our research group have recently found that IRF-1 and EHD2 are involved in the process of CAM-DR of non-Hodgkin's lymphoma (NHL), and they can interact with each other. Furthermore, our preliminary study showed that EHD2 could promote the degradation of IRF-1. Based on these findings, we speculate that EHD2 may influence NF-κB activity by regulating IRF-1 expression，which may be critically involved in the process of CAM-DR. In this project, we aim to first establish NHL cell adhesion model, then analyse the interaction between EHD2 and IRF-1, as well as their regulatory mechanism. Moreover, the influence of the molecular mechanism on NF-κB activity and CAM-DR will be analyzed. Finally, we will clarify the correlation between the expression of EHD2 and IRF-1 in NHL clinical specimens, and investigate the role of EHD2-IRF-1 interaction in the process of CAM-DR utilizing nude mouse model. Our research may help to gain a better insight into the mechanism underlying CAM-DR and may be a promising therapeutic target.

NF-κB通路的活化是细胞粘附介导的耐药（CAM-DR）形成的重要原因之一。干扰素调节因子1（IRF-1）的表达在NF-κB活化中起重要作用。课题组前期研究发现IRF-1、EHD2与非霍奇金淋巴瘤（NHL）CAM-DR有关，并发现两者相互作用。预实验表明EHD2可调节IRF-1的降解。基于此，提出EHD2可能通过调节IRF-1的表达影响NF-κB通路参与CAM-DR进程。本课题拟首先构建NHL细胞粘附模型，分析EHD2与IRF-1在粘附过程中的相互作用及调节机制。其次，探讨其相互作用如何影响NF-κB通路进而影响CAM-DR进程。最后验证并分析EHD2、IRF-1与NHL患者的临床病理关系，并在耐药裸鼠模型中研究这种相互作用对NHL细胞CAM-DR的影响。课题的研究将进一步揭示NHL细胞CAM-DR发生发展的分子机制。

NF-κB 通路活化与细胞粘附介导的耐药（CAM-DR） 密切相关。根据前期工作基础我们提出假说：在淋巴瘤中EHD2与IRF-1相互作用可促进IRF-1泛素化降解，其相互作用通过激活NF-κB信号通路促进CAM-DR进程。为验证上述假说，我们首先在OCI-Ly8以及OCI-Ly10细胞中验证了EHD2与IRF-1的相互作用。接下来分析了EHD2对IRF-1蛋白表达水平的影响；通过放线菌酮追踪分析了EHD2对IRF-1半衰期的影响，同时分析了EHD2对IRF-1泛素化修饰的影响。通过CCK-8和台盼蓝拒染实验分析了EHD2、IRF-1对细胞增殖和死亡的影响。然后利用纤粘蛋白（FN）包被构建细胞粘附模型，免疫印记检测EHD2、IRF-1在悬浮及粘附培养过程中的表达变化。构建粘附模型，药物刺激实验分析EHD2、IRF-1对CAM-DR的影响。最后分析了EHD2与IRF-1相互作用对NF-κB通路活性的影响，并分析了其对NF-κB下游凋亡相关蛋白表达的影响。通过本项目的研究，已明确在淋巴瘤中EHD2与IRF-1相互作用可促进IRF-1泛素化降解，其相互作用通过激活NF-κB信号通路促进CAM-DR进程。课题的研究结果将进一步揭示CAM-DR发生发展的分子机制。