Muramatsu S, Ishido S, Fujita T, Itoh M, Hotta H
Department of Microbiology, Kobe University School of Medicine, Hyogo, Japan.
J Virol. 1997 Jul;71(7):4954-61. doi: 10.1128/JVI.71.7.4954-4961.1997.
Subcellular localization of the NS2 and NS3 proteins of hepatitis C virus was analyzed. In stable Ltk transfectants inducibly expressing an NS2-NS3 polyprotein (amino acids [aa] 810 to 1463), processed full-size NS2 (aa 810 to 1026) was detected exclusively in a cytoplasmic membrane fraction. On the other hand, the other processed product, carboxy-truncated NS3 (NS3 deltaC1463; aa 1027 to 1463), was present in both cytoplasmic and nuclear fractions. To further analyze subcellular localization of NS3, NS3 deltaC1459 (aa 1027 to 1459), full-size NS3 (NS3F; aa 1027 to 1657), and both amino- and carboxy-truncated NS3 (NS3 deltaNdeltaC; aa 1201 to 1459) were expressed in HeLa cells by using a vaccinia virus-T7 hybrid expression system. NS3 deltaC1459 and NS3F accumulated in the nucleus as well as in the cytoplasm, exhibiting a dot-like staining pattern. On the other hand, NS3 deltaNdeltaC was localized predominantly in the cytoplasm, suggesting the presence of a nuclear localization signal(s) in the amino-terminal sequence of NS3. NS4A, a viral cofactor for the NS3 protease, inhibited nuclear transport of NS3 deltaC1459 and NS3F, with the latter inhibited to a lesser extent than was the former. Interestingly, wild-type p53 tumor suppressor augmented nuclear localization of NS3 deltaC1459 and NS3F, whereas mutant-type p53 inhibited nuclear localization and augmented cytoplasmic localization of NS3 deltaC1459. However, subcellular localization of NS3 deltaNdeltaC was not affected by either type of p53. Wild-type p53-mediated nuclear accumulation of NS3 deltaC1459 and NS3F was inhibited partially, but not completely, by coexpressed NS4A, with NS3F again affected less prominently than was NS3 deltaC1459.
对丙型肝炎病毒NS2和NS3蛋白的亚细胞定位进行了分析。在可诱导表达NS2-NS3多聚蛋白（氨基酸[aa]810至1463）的稳定Ltk转染细胞中，仅在细胞质膜组分中检测到加工后的全长NS2（aa 810至1026）。另一方面，另一种加工产物，羧基截短的NS3（NS3 deltaC1463；aa 1027至1463），存在于细胞质和细胞核组分中。为了进一步分析NS3的亚细胞定位，通过痘苗病毒-T7杂交表达系统在HeLa细胞中表达了NS3 deltaC1459（aa 1027至1459）、全长NS3（NS3F；aa 1027至1657）以及氨基和羧基均截短的NS3（NS3 deltaNdeltaC；aa 1201至1459）。NS3 deltaC1459和NS3F在细胞核和细胞质中均有积累，呈现点状染色模式。另一方面，NS3 deltaNdeltaC主要定位于细胞质，这表明NS3的氨基末端序列中存在核定位信号。NS4A是NS3蛋白酶的病毒辅助因子，它抑制了NS3 deltaC1459和NS3F的核转运，其中NS3F受到的抑制程度小于NS3 deltaC1459。有趣的是，野生型p53肿瘤抑制因子增强了NS3 deltaC1459和NS3F的核定位，而突变型p53则抑制了NS3 deltaC1459的核定位并增强了其细胞质定位。然而，两种类型的p53均未影响NS3 deltaNdeltaC的亚细胞定位。共表达的NS4A部分但未完全抑制野生型p53介导的NS3 deltaC1459和NS3F的核积累，其中NS3F受到的影响再次不如NS3 deltaC1459显著。
