Identification of a functional, CRM-1-dependent nuclear export signal in hepatitis C virus core protein.
Identification of a functional, CRM-1-dependent nuclear export signal in hepatitis C virus core protein.
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DOI：
10.1371/journal.pone.0025854
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发表时间：
2011
期刊：
影响因子：
3.7
通讯作者：
Budkowska A
中科院分区：
文献类型：
--
作者：
Cerutti A;Maillard P;Minisini R;Vidalain PO;Roohvand F;Pecheur EI;Pirisi M;Budkowska A
丙型肝炎病毒（HCV）感染是世界范围内慢性肝病的主要原因。HCV核心蛋白参与核衣壳的形成，但它也与多种细胞质和核分子相互作用，并在肝脏疾病和肝癌的发生发展中发挥关键作用。核心蛋白在HCV感染期间主要存在于细胞质中，但在肝癌患者和核心转基因小鼠中也存在于细胞核中。HCV核心区含有核定位信号（NLS），但尚未发现核输出信号（内斯）。我们在这里表明，aa（109-133）区域指导的核心易位从细胞核到细胞质的CRM-1介导的核输出途径。在所鉴定的内斯或编码成熟核心aa（1-173）的序列中的三个疏水残基（L119、I123和L126）的突变显著增强了转染的Huh 7细胞中相应蛋白质的核定位。核心结构域II中的内斯和相邻的疏水序列都是维持核心蛋白或其片段在胞质区室中所必需的。JFH 1复制模型的电子显微镜研究表明，在病毒进入细胞后几分钟，核心易位到细胞核中。在感染早期，通过链霉素B治疗阻断核质出口，导致通过共聚焦显微镜检测到核中的核心蛋白，并且与病毒复制的减少相一致。我们的数据表明，功能性NLS和内斯直接HCV核心蛋白穿梭于细胞质和细胞核之间，至少有一些核心蛋白转运到细胞核。HCV核心区的这些新特性可能是病毒增殖和与核分子相互作用所必需的，影响细胞信号传导和HCV感染的发病机制。
Hepatitis C virus (HCV) infection is a major cause of chronic liver disease worldwide. HCV core protein is involved in nucleocapsid formation, but it also interacts with multiple cytoplasmic and nuclear molecules and plays a crucial role in the development of liver disease and hepatocarcinogenesis. The core protein is found mostly in the cytoplasm during HCV infection, but also in the nucleus in patients with hepatocarcinoma and in core-transgenic mice. HCV core contains nuclear localization signals (NLS), but no nuclear export signal (NES) has yet been identified. We show here that the aa(109–133) region directs the translocation of core from the nucleus to the cytoplasm by the CRM-1-mediated nuclear export pathway. Mutagenesis of the three hydrophobic residues (L119, I123 and L126) in the identified NES or in the sequence encoding the mature core aa(1–173) significantly enhanced the nuclear localisation of the corresponding proteins in transfected Huh7 cells. Both the NES and the adjacent hydrophobic sequence in domain II of core were required to maintain the core protein or its fragments in the cytoplasmic compartment. Electron microscopy studies of the JFH1 replication model demonstrated that core was translocated into the nucleus a few minutes after the virus entered the cell. The blockade of nucleocytoplasmic export by leptomycin B treatment early in infection led to the detection of core protein in the nucleus by confocal microscopy and coincided with a decrease in virus replication. Our data suggest that the functional NLS and NES direct HCV core protein shuttling between the cytoplasmic and nuclear compartments, with at least some core protein transported to the nucleus. These new properties of HCV core may be essential for virus multiplication and interaction with nuclear molecules, influence cell signaling and the pathogenesis of HCV infection.
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