Arita Minetaro, Shimizu Hiroyuki, Miyamura Tatsuo
Department of Virology II, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama-shi, Tokyo 208-0011, Japan.
J Gen Virol. 2004 Jul;85(Pt 7):1933-1944. doi: 10.1099/vir.0.19768-0.
Sabin vaccine strains of poliovirus (PV) contain major attenuation determinants in the internal ribosomal entry site (IRES), an area that directs viral protein synthesis. To examine the effect of reduced viral protein synthesis on PV neurovirulence, spacer sequences, consisting of short open reading frames of different lengths, were introduced between the IRES and the initiation codon of viral polyprotein, resulting in PV mutants with reduced viral protein synthesis. These PV mutants had a viral protein synthesis activity 8.8-55 % of that of the parental Mahoney strain as measured in HeLa S3 cells. Only viruses with more than 28 % of the wild-type activity had intact spacer sequences following plaque purification. Mutants with 17 % or 21 % of the wild-type activity were unstable and a mutant with 8.8 % was lethal. The neurovirulence of PV mutants was evaluated in transgenic mice carrying the human PV receptor gene. In this test, mutants with more than 28 % of the wild-type activity remained neurovirulent, while a mutant with 17 % of wild-type activity exhibited a partially attenuated phenotype. This mutant stably replicated in the spinal cord; however, the stability was severely affected during the course of virus infection from the cerebrum to the spinal cord. These results suggest that reduced viral protein synthesis activity as measured in cultured cells (17-55 % of the wild-type activity) is not the main determinant of PV attenuation.
脊髓灰质炎病毒（PV）的萨宾疫苗株在内部核糖体进入位点（IRES）中含有主要的减毒决定因素，该区域指导病毒蛋白的合成。为了研究病毒蛋白合成减少对PV神经毒力的影响，在IRES和病毒多聚蛋白的起始密码子之间引入了由不同长度的短开放阅读框组成的间隔序列，从而产生了病毒蛋白合成减少的PV突变体。在HeLa S3细胞中检测发现，这些PV突变体的病毒蛋白合成活性是亲本马奥尼株的8.8%-55%。在空斑纯化后，只有病毒蛋白合成活性超过野生型活性28%的病毒具有完整的间隔序列。病毒蛋白合成活性为野生型活性17%或21%的突变体不稳定，而活性为8.8%的突变体是致死性的。在携带人PV受体基因的转基因小鼠中评估了PV突变体的神经毒力。在该试验中，病毒蛋白合成活性超过野生型活性28%的突变体仍然具有神经毒力，而病毒蛋白合成活性为野生型活性17%的突变体表现出部分减毒的表型。该突变体在脊髓中稳定复制；然而，在病毒从大脑到脊髓的感染过程中，其稳定性受到严重影响。这些结果表明，在培养细胞中测得的病毒蛋白合成活性降低（为野生型活性的17%-55%）不是PV减毒的主要决定因素。
