Localization determines function: N-terminally truncated NS5A fragments accumulate in the nucleus and impair HCV replication

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• 作者：Daniel Sauter ; Kiyoshi Himmelsbach ; Malte Kriegs ; Monica Carvajal Yepes ; Eberhard Hildt
• 关键词：Hepatitis C virus ; Regulatory protein ; NS5A ; Signal transduction
• 刊名：Journal of Hepatology
• 出版年：2009
• 出版时间：May 2009
• 年：2009
• 卷：50
• 期：5
• 页码：861-871
• 全文大小：613 K

文摘

Background/Aims

The Hepatitis C Virus (HCV) nonstructural protein 5A (NS5A) is an essential part of the ER-localized HCV–replicon complex. Although NS5A harbours a conserved NLS in its C-terminal domain, NS5A is associated with the cytoplasmic face of the ER by an amphipathic helix close to its N-terminus.

Methods

Intracellular distribution of NS5A in HCV replicating cells was analyzed by confocal microscopy and subcellular fractionation. The effect on HCV replication was analyzed using the JFH-1-based infection/replication system.

Results

During viral life cycle N-terminally truncated NS5A fragments are caspase-dependent formed that lack the ER-attachment signal and are localized within the nucleus. These N-terminally truncated fragments inhibit HCV replication. If their formation is blocked by inhibition of caspases HCV replication is increased.

The C-terminal domain of NS5A binds to c-Raf and thereby localizes it to the replicon complex. This interaction is essential for HCV replication. The N-terminally truncated NS5A fragments are still able to bind c-Raf. However, due to their nuclear localization they withdraw c-Raf from the replicon complex into the nucleus resulting in an impaired HCV replication.

Conclusions

Formation of N-terminally truncated NS5A fragments could represent a mechanism to regulate HCV replication by withdrawal of essential factors from the replicon complex.