Identification of cellular and viral factors related to anti-hepatitis C virus activity of cyclophilin inhibitor
Identification of cellular and viral factors related to anti-hepatitis C virus activity of cyclophilin inhibitor
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DOI：
10.1111/j.1349-7006.2009.01263.x
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发表时间：
2009-10-01
期刊：
影响因子：
5.7
通讯作者：
Shimotohno, Kunitada
中科院分区：
文献类型：
--
作者：
Goto, Kaku;Watashi, Koichi;Shimotohno, Kunitada
迄今为止，我们已经报道了一种免疫抑制剂环孢素A（CsA），一种众所周知的亲环素（CyP）抑制剂（CPI），强烈抑制丙型肝炎病毒（HCV）在细胞培养中的复制，和CyPB是病毒复制的细胞辅助因子。为了进一步研究CPI的抗病毒机制，我们在此通过在CsA与G418存在下培养HCV亚基因组复制子细胞4周来开发携带CsA抗性HCV复制子的细胞。从分离的CsA抗性细胞的总RNA转染到幼稚Huh 7细胞赋予CsA抗性，表明复制子RNA本身负责抗性表型。在已鉴定的氨基酸突变中，NS 5A中的D320 E赋予CsA抗性。携带D320 E突变的复制子对干扰素-α敏感，但对CsA和其他CPI（包括NIM 811和sanglifehrin A）具有抗性。敲除个别CyP亚型揭示CyP 40，除了CyPA和CyPB，有助于病毒复制，CsA抗性复制子获得独立于CyPA的有效复制。这些数据为CyP调控HCV复制的机制以及设计新的和特异性的CPI抗HCV策略提供了重要证据。（Cancer Sci 2009; 100：1943-1950）。
We have so far reported that an immunosuppressant cyclosporin A (CsA), a well-known cyclophilin (CyP) inhibitor (CPI), strongly suppressed hepatitis C virus (HCV) replication in cell culture, and that CyPB was a cellular cofactor for viral replication. To further investigate antiviral mechanisms of CPI, we here developed cells carrying CsA-resistant HCV replicons, by culturing the HCV subgenomic replicon cells for 4 weeks in the presence of CsA with G418. Transfection of total RNA from the isolated CsA-resistant cells to naive Huh7 cells conferred CsA resistance, suggesting that the replicon RNA itself was responsible for the resistant phenotype. Of the identified amino acid mutations, D320E in NS5A conferred the CsA resistance. The replicon carrying the D320E mutation was sensitive to interferon-alpha, but was resistant to CsA and other CPIs including NIM811 and sanglifehrin A. Knockdown of individual CyP subtypes revealed CyP40, in addition to CyPA and CyPB, contributed to viral replication, and CsA-resistant replicons acquired independence from CyPA for efficient replication. These data provide important evidence on the mechanisms underlying the regulation of HCV replication by CyP and for designing novel and specific anti-HCV strategies with CPIs. (Cancer Sci 2009; 100: 1943-1950).