Li You, Zheng Xin, Xie Ling, Kapustina Maryna, Shirasaki Takayoshi, Yonish Bryan, Chen Xian, Hirai-Yuki Asuka, Nagata Noriyo, Suzuki Ryosuke, Isogawa Masanori, Vogt Matthew R, Muramatsu Masamichi, Lemon Stanley M
Department of Pediatrics, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
Department of Virology II, National Institute of Infectious Diseases, Japan Institute for Health Security, Tokyo, Japan.
J Virol. 2025 Jul 22;99(7):e0050225. doi: 10.1128/jvi.00502-25. Epub 2025 Jun 5.
HSP90 heat shock chaperones are essential for maintaining cellular proteostasis, as well as the ATP-dependent folding and functional maturation of many viral proteins. As a result, inhibitors of HSP90 have broad antiviral activity, disrupting replication of many viruses at concentrations below those causing cytotoxicity. Among the , HSP90 inhibitors block replication of multiple , and species, in some cases, by preventing post-translational processing and assembly of P1 capsid proteins. Hepatitis A virus (HAV), classified within the genus , has been suggested to be an exception among picornaviruses and to replicate independently of HSP90, possibly because its slow translational kinetics could facilitate co-translational folding and assembly of its capsid proteins. However, we show here that HAV replication is highly dependent upon HSP90, both in human hepatocyte-derived cell lines, in which the 50% inhibitory concentration of geldanamycin was 8.7-11.8 nM, and in mice. Label-free proteomics experiments suggested that HSP90 interacts with capsid proteins or their precursors and may thus facilitate the folding and assembly of capsid proteins, as it does for enteroviruses and aphthoviruses. By contrast, there was no evidence for HSP90 interacting with any nonstructural protein, and HSP90 inhibitors did not impair 3C proteolytic activity. Despite this, and in contrast to previous studies of enteroviruses and aphthoviruses, geldanamycin potently inhibited replication of a subgenomic HAV replicon. We conclude that HAV is no exception from the HSP90-dependent nature of other picornaviruses and indeed is more dependent on HSP90 than other picornaviruses for amplification of its genome.IMPORTANCEHepatitis A virus (HAV), a common cause of acute infectious hepatitis, has been reported to differ from other picornaviruses in not requiring heat shock protein HSP90 for efficient replication. However, we show here that productive HAV infection is highly dependent on HSP90 and that HAV replication is potently blocked both in cell culture and in the murine liver by chemical inhibitors of HSP90. Such inhibitors also disrupt the replication of a subgenomic HAV RNA replicon, indicating that HSP90 is required for the assembly of functional replication organelles. This highlights a key difference from other picornaviruses for which HSP90 is required primarily, if not exclusively, for the maturation of the P1 capsid proteins.
热休克蛋白90（HSP90）热休克伴侣蛋白对于维持细胞蛋白质稳态以及许多病毒蛋白的ATP依赖性折叠和功能成熟至关重要。因此，HSP90抑制剂具有广泛的抗病毒活性，能在低于细胞毒性浓度的情况下破坏多种病毒的复制。在这些病毒中，HSP90抑制剂可阻断多种肠道病毒、口蹄疫病毒和甲型肝炎病毒的复制，在某些情况下，是通过阻止P1衣壳蛋白的翻译后加工和组装来实现的。甲型肝炎病毒（HAV）属于嗜肝病毒属，有人认为它是小RNA病毒中的一个例外，其复制不依赖于HSP90，可能是因为其缓慢的翻译动力学有助于衣壳蛋白的共翻译折叠和组装。然而，我们在此表明，HAV复制高度依赖于HSP90，无论是在人肝细胞衍生的细胞系中（在该细胞系中，格尔德霉素的50%抑制浓度为8.7 - 11.8 nM），还是在小鼠体内。无标记蛋白质组学实验表明，HSP90与衣壳蛋白或其前体相互作用，因此可能像对肠道病毒和口蹄疫病毒那样促进衣壳蛋白的折叠和组装。相比之下，没有证据表明HSP90与任何非结构蛋白相互作用，并且HSP90抑制剂不会损害3C蛋白酶活性。尽管如此，与之前对肠道病毒和口蹄疫病毒的研究不同，格尔德霉素能有效抑制亚基因组HAV复制子的复制。我们得出结论，HAV在依赖HSP90这一特性上与其他小RNA病毒并无不同，实际上在其基因组扩增方面比其他小RNA病毒更依赖于HSP90。
重要性
甲型肝炎病毒（HAV）是急性传染性肝炎的常见病因，据报道它与其他小RNA病毒不同，高效复制不需要热休克蛋白HSP90。然而，我们在此表明，HAV的有效感染高度依赖于HSP90，并且HSP90的化学抑制剂在细胞培养和小鼠肝脏中均能有效阻断HAV复制。此类抑制剂还会破坏亚基因组HAV RNA复制子的复制，表明HSP90是功能性复制细胞器组装所必需的。这突出了与其他小RNA病毒的一个关键区别，对于其他小RNA病毒，HSP90主要（如果不是唯一）是P1衣壳蛋白成熟所必需的。
