J Virol：浙江大学周继勇等阐明伴侣蛋白CCTγ如何促进狂犬病毒的复制

Journal of Virology（《病毒学杂志》）在2013年3月1日在线发表了我国杭州浙江大学农业部动物疫病病原学重点实验室周继勇教授等人的一项最新研究结果。研究人员研究了嗜神经型狂犬病毒（RABV）感染过程中神经元CCTγ的表达情况、细胞定位和作用，该研究阐明了狂犬病毒为了自身的存活如何利用宿主细胞的分子伴侣及其区室化功能。

作为已知的最古老感染性疾病，狂犬病仍然对世界公共卫生造成严重威胁。而位于真核生物细胞基质的伴侣蛋白TRiC/CCT复合物通过水解ATP来促进蛋白质折叠。在本研究中，研究人员以小鼠N2a细胞作为模型，来研究嗜神经型狂犬病毒感染过程中神经元伴侣蛋白CCTγ的表达情况、细胞定位和作用。

在小鼠N2a细胞感染RABV之后，研究人员通过双向电泳和质谱分析法发现，有24种蛋白的水平发生变化，其中20种蛋白水平上调，另外4种下调。在感染RABV的小鼠N2a细胞或共转染编码RABV核蛋白（nucleoprotein (N)）和磷蛋白（phosphoprotein (P)）基因的N2a细胞，共聚焦显微镜（confocal microscopy）结果显示，表达水平上调的CCTγ与病毒蛋白N和P共定位，它们在细胞核周围区域形成空心的环状内含物（即狂犬病包涵体(NBs)）。当小鼠N2a细胞只表达病毒蛋白N或P时，则不会形成这些包含物。通过慢病毒介导的RNA干扰技术降低CCTγ的表达水平，可显著抑制RABV的复制。

这些结果表明，由病毒蛋白N和P组成的复合物可将CCTγ招募至NBs；而且将伴侣蛋白CCTγ识别为宿主因子，CCTγ具有促进细胞内RABV复制的作用。这项工作阐明了狂犬病毒为了自身的存活是如何利用细胞分子伴侣及其区室化功能。

Cellular Chaperonin CCTγ Contributes to Rabies Virus Replication during Infection
ABSTRACT
Rabies, as the oldest known infectious disease, remains a serious threat to public health worldwide. The eukaryotic cytosolic chaperonin TRiC/CCT complex facilitates the folding of proteins through ATP hydrolysis. Here we investigated expression, cellular localization and function of neuronal CCTγ during neurotropic rabies virus (RABV) infection using mouse N2a cells as a model. Following RABV infection, 24 altered proteins were identified using two-dimensional electrophoresis and mass spectrometry, including 20 up-regulated proteins and 4 down-regulated proteins. In mouse N2a cells infected with RABV or co-transfected with RABV genes encoding nucleoprotein (N) and phosphoprotein (P), confocal microscopy demonstrated that up-regulated cellular CCTγ was co-localized with viral proteins N and P, which formed a hollow cricoid inclusion within the region around the nucleus. These inclusions, which correspond to Negri Bodies (NBs), did not form in mouse N2a cells only expressing viral proteins N or P. Knockdown of CCTγ by lentivirus-mediated RNA interference led to a significant inhibition of RABV replication. These results demonstrate that the complex consisting of viral proteins N and P recruits CCTγ to NBs and identify the chaperonin CCTγ as a host factor facilitates intracellular RABV replication. This work illustrates how viruses can utilize cellular chaperonins and compartmentalization for their own benefit.