Moffatt S, Yaegashi N, Tada K, Tanaka N, Sugamura K
Department of Microbiology and Immunology, Tohoku University School of Medicine, Sendai, Japan.
J Virol. 1998 Apr;72(4):3018-28. doi: 10.1128/JVI.72.4.3018-3028.1998.
Infection of erythroid-lineage cells by human parvovirus B19 is characterized by a gradual cytocidal effect. Accumulating evidence now implicates the nonstructural (NS1) protein of the virus in cytotoxicity, but the mechanism underlying the NS1-induced cell death is not known. Using a stringent regulatory system, we demonstrate that NS1 cytotoxicity is closely related to apoptosis, as evidenced by cell morphology, genomic DNA fragmentation, and cell cycle analysis with the human erythroleukemia cell line K562 and the erythropoietin-dependent megakaryocytic cell line UT-7/Epo. Apoptosis was significantly inhibited by an interleukin-1beta (IL-1beta)-converting enzyme (ICE)/CED-3 family protease inhibitor, Ac-DEVD-CHO (CPP32; caspase 3), whereas a similar inhibitor of ICE (caspase 1), Ac-YVAD-CHO, had no effect. Furthermore, stable expression of the human Bcl-2 proto-oncogene resulted in near-total protection from cell death in response to NS1 induction. Mutations engineered into the nucleoside triphosphate-binding domain of NS1 significantly rescued cells from NS1-induced apoptosis without having any effect on NS1-induced activation of the IL-6 gene expression which is mediated by NF-kappaB. Furthermore, using pentoxifylline, an inhibitor of NF-kappaB activation, we demonstrate that the NF-kappaB-mediated IL-6 activation by NS1 is uncoupled from the apoptotic pathway. This functional dissection indicates a complexity underlying the biochemical function of human parvovirus NS1 in transcriptional activation and induction of apoptosis. Our findings indicate that NS1 of parvovirus B19 induces cell death by apoptosis in at least erythroid-lineage cells by a pathway that involves caspase 3, whose activation may be a key event during NS1-induced cell death.
人细小病毒B19对红系细胞的感染具有逐渐的杀细胞效应。越来越多的证据表明，该病毒的非结构（NS1）蛋白具有细胞毒性，但其诱导细胞死亡的机制尚不清楚。利用一个严格的调控系统，我们证明NS1的细胞毒性与凋亡密切相关，这在人红白血病细胞系K562和促红细胞生成素依赖性巨核细胞系UT-7/Epo的细胞形态、基因组DNA片段化及细胞周期分析中得到了证实。白细胞介素-1β（IL-1β）转化酶（ICE）/CED-3家族蛋白酶抑制剂Ac-DEVD-CHO（CPP32；半胱天冬酶3）可显著抑制凋亡，而类似的ICE（半胱天冬酶1）抑制剂Ac-YVAD-CHO则无此作用。此外，人Bcl-2原癌基因的稳定表达可使细胞在NS1诱导下几乎完全免受细胞死亡。对NS1核苷三磷酸结合结构域进行的工程突变可显著使细胞从NS1诱导的凋亡中获救，且对NS1诱导的由核因子κB介导的IL-6基因表达激活没有任何影响。此外，使用核因子κB激活抑制剂己酮可可碱，我们证明NS1介导的核因子κB对IL-6的激活与凋亡途径无关。这种功能剖析表明人细小病毒NS1在转录激活和凋亡诱导中的生化功能具有复杂性。我们的研究结果表明，细小病毒B19的NS1至少在红系细胞中通过一条涉及半胱天冬酶3的途径诱导细胞凋亡，半胱天冬酶3的激活可能是NS1诱导细胞死亡过程中的关键事件。
