p38/JNK MAPKs 通路在博卡病毒MVC感染致宿主细胞病变与病毒复制 的作用机制探讨

Bocavirus subfamily is a member of the parvovirus family, and the infection of bocavirus is closely related to respiratory and intestinal infectious diseases. Some studies have reported that the pathogenesis of some virus infection is associated with MAPKs signaling pathway,but there is rare report in the parvovirus infection. In the preliminary study, we successful obtained the bocavirus MVC infectious clone vector, analyzed the characteristics of virus genome and related genetic features, and revealed the apoptosis process in target cells by MVC infection. Recently, our results showed that p53 protein plays a key role in the apoptosis of MVC infectious cells, which is associated with the activity of p38/JNK proteins. So we speculate p38/JNK MAPKs-p53 pathway plays an important role in cytopathy induced by MVC virus infection. This study aims to reveal the expression and phosphorylation levels of p38, JNK and its upstream- and downstream-related genes in cells infected with MVC, and to reveal cell apoptosis and the expression and phospholation levels of their downstream regulatory proteins in MVC infetious cells by using the inhibitors of p38 and JNK proteins blocking p38/JNK MAPKs pathway. Furthermore, the study the effets of viral genomic replication ability change of MVC on p38/JNK MAPKs signaling pathway is also carry out in WRD cells transfected with MVC transfectious clone and its different viral genes mutants, respectively. Moreover, the viral genome DNA replication, transcription and virus particle formation are also carried out by blocking p38/JNK MAPKs pathway with inhibitors of p38 and JNK proteins. The completion of this research will deepen the understanding of the pathogenesis of infection of bocavirus, but also will provide an opportunity for the study of the pathogenesis of Bocavirus and other parvovirus members infection.

博卡病毒（属细小病毒家族）感染与呼吸道及肠道感染性疾病密切相关。研究报道，有些病毒感染致病与MAPKs信号通路变化有关，但在细小病毒方面鲜有报道。课题组前期获得博卡病毒MVC感染性克隆，分析病毒基因组特性，揭示了MVC感染致细胞凋亡性病变。近期发现宿主细胞p53蛋白在病毒感染致细胞凋亡中发挥关键作用，且与p38/JNK蛋白活性相关，推测p38/JNK MAPKs-p53通路在MVC感染致病中发挥作用。本研究拟揭示p38/JNK MAPKs 通路在MVC感染中相关调控因子表达及活性变化；研究p38/JNK通路阻滞对MVC感染致细胞病变及相关因子表达及活性影响；利用MVC感染性克隆和突变体，研究病毒基因组复制能力改变对p38/JNK通路的影响；研究p38/JNK通路阻滞对病毒复制、基因表达及病毒子形成的影响。研究成果将深化博卡病毒致病机制的认识，并对其他细小病毒家族感染致病机制研究提供借鉴。

通过博卡病毒MVC感染WRD靶细胞后，揭示了博卡病毒MVC感染WRD靶细胞后，引起靶细胞内p38/JNK MAPKs信号通路的激活，该通路的激活与MVC感染引起靶细胞的凋亡和病毒基因组的复制和病毒基因的表达存在密切关系。完成了VC感染WRD靶细胞后，能够诱导p38/JNK MAPKs 通路中相关调控因子表达升高和磷酸化水平增强，p38/JNK抑制剂的干预，p53、ATF2、Mnk1、eIF4E基因的蛋白表达水平降低，MVC感染的细胞活力上升，细胞凋亡水平下降低。p38/JNK MAPKs信号通路的激活，与博卡病毒基因组复制、病毒基因的表达存在密切关系，p38/JNK抑制剂的干预后，MVC病毒基因组复制能力下降，病毒蛋白表达水平降低。增加了：制备了特异性靶向博卡病毒VP1结构蛋白的多克隆抗体，构建并筛选了分别靶向p38、JNK基因的RNA干扰载体，并对病毒基因表达及病毒颗粒形成开展了初步研究。

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