Division of Hematology, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland 20892, USA.
J Neurovirol. 2009 May;15(3):229-37. doi: 10.1080/13550280902913636.
Neurovirulence is one of the pathological complications associated with vaccinia virus (VV) infection/vaccination. Although the viral N1L protein has been identified as the neurovirulence factor, none of the host N1L-interacting factors have been identified so far. In the present study, we identified N1L-interacting proteins by screening a human brain cDNA expression library with N1L as a bait protein in a yeast two-hybrid analysis. The analysis revealed that N1L interacts with human brain-originated cellular basement membrane-associated chondroitin sulfate proteoglycan (bamacan). The N1L-binding domain of bamacan was mapped to its C-terminal 227 amino acids. The N1L-bamacan interaction was further confirmed in both VV-infected and N1L-transfected mammalian cells. Following the confirmation of the protein interactions by coimmunoprecipitation experiments, confocal microscopic analysis revealed that N1L colocalizes with bamacan both in VV-infected B-SC-1 cells as well as in mice neuronal tissue. Furthermore, a human neural cell line, which expresses bamacan to moderately elevated levels relative to a non-neural cell line, supported enhanced viral growth. Overall, these studies clearly suggest that bamacan interacts with the VV-N1L and such interactions seem to play a positive role in promoting the viral growth and perhaps contribute to the virulence of VV in neural cells.
神经毒力是与牛痘病毒（VV）感染/接种相关的病理并发症之一。虽然已经确定病毒的 N1L 蛋白是神经毒力因子，但迄今为止尚未鉴定出任何宿主 N1L 相互作用因子。在本研究中，我们通过酵母双杂交分析用 N1L 作为诱饵蛋白筛选人脑组织 cDNA 表达文库，鉴定出与 N1L 相互作用的蛋白。分析表明，N1L 与人脑源性细胞基底膜相关软骨素蛋白聚糖（bamacan）相互作用。bamacan 的 N1L 结合域被映射到其 C 末端的 227 个氨基酸。在 VV 感染和 N1L 转染的哺乳动物细胞中进一步证实了 N1L-bamacan 相互作用。通过共免疫沉淀实验证实了蛋白质相互作用后，共聚焦显微镜分析显示，N1L 与 bamacan 在 VV 感染的 B-SC-1 细胞以及小鼠神经元组织中共定位。此外，与非神经细胞系相比，表达 bamacan 水平适度升高的人神经细胞系支持增强的病毒生长。总的来说，这些研究清楚地表明，bamacan 与 VV-N1L 相互作用，这种相互作用似乎在促进病毒生长中起积极作用，并且可能有助于 VV 在神经细胞中的毒力。
