HCV NS5A dimer interface residues regulate HCV replication by controlling its self-interaction, hyperphosphorylation, subcellular localization and interaction with cyclophilin A.
HCV NS5A dimer interface residues regulate HCV replication by controlling its self-interaction, hyperphosphorylation, subcellular localization and interaction with cyclophilin A.
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DOI：
10.1371/journal.ppat.1007177
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发表时间：
2018-07
期刊：
影响因子：
6.7
通讯作者：
Yi M
中科院分区：
文献类型：
--
作者：
Shanmugam S;Nichols AK;Saravanabalaji D;Welsch C;Yi M
The HCV NS5A protein plays multiple roles during viral replication, including viral genome replication and virus particle assembly. The crystal structures of the NS5A N-terminal domain indicated the potential existence of the NS5A dimers formed via at least two or more distinct dimeric interfaces. However, it is unknown whether these different forms of NS5A dimers are involved in its numerous functions. To address this question, we mutated the residues lining the two different NS5A dimer interfaces and determined their effects on NS5A self-interaction, NS5A-cyclophilin A (CypA) interaction, HCV RNA replication and infectious virus production. We found that the mutations targeting either of two dimeric interfaces disrupted the NS5A self-interaction in cells. The NS5A dimer-interrupting mutations also inhibited both viral RNA replication and infectious virus production with some genotypic differences. We also determined that reduced NS5A self-interaction was associated with altered NS5A-CypA interaction, NS5A hyperphosphorylation and NS5A subcellular localization, providing the mechanistic bases for the role of NS5A self-interaction in multiple steps of HCV replication. The NS5A oligomers formed via different interfaces are likely its functional form, since the residues at two different dimeric interfaces played similar roles in different aspects of NS5A functions and, consequently, HCV replication. In conclusion, this study provides novel insight into the functional significance of NS5A self-interaction in different steps of the HCV replication, potentially, in the form of oligomers formed via multiple dimeric interfaces. HCV NS5A is a multifunctional protein involved in both viral RNA replication and infectious virus production, and is a target of one of the most potent antivirals available to date. However, the mode of action of NS5A inhibitors is still unclear due to the lack of mechanistic detail regarding NS5A functions during HCV life cycles. In this study, we have provided evidence that surface-exposed NS5A residues involved in two different dimeric interactions in crystal structures are indeed involved in NS5A self-interactions in cells. We also showed that these NS5A residues play critical role in HCV RNA replication and infectious virus production by regulating NS5A hyperphosphorylation, its subcellular localization and its interaction with host protein CypA. Overall, our data support the functional significance of “NS5A oligomers” formed via multiple interfaces in HCV replication. We speculate that the NS5A inhibitors exploited the NS5A oligomer-dependent functions during HCV replication, rather than targeting individual NS5A, which consequently resulted in their high potency.
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DOI：
10.3390/v6114227
发表时间：
2014-11-06
期刊：
Viruses
影响因子：
--
作者：
Issur M;Götte M
通讯作者：
Götte M
影响因子：
3.7
作者：
Coelmont L;Hanoulle X;Chatterji U;Berger C;Snoeck J;Bobardt M;Lim P;Vliegen I;Paeshuyse J;Vuagniaux G;Vandamme AM;Bartenschlager R;Gallay P;Lippens G;Neyts J
通讯作者：
Neyts J
影响因子：
5.4
作者：
Hwang, Jungwook;Huang, Luyun;Cameron, Craig E.
通讯作者：
Cameron, Craig E.
影响因子：
3.7
作者：
Keum, Sun Ju;Park, Sung Mi;Jang, Sung Key
通讯作者：
Jang, Sung Key
影响因子：
5.4
作者：
Blight, KJ;McKeating, JA;Rice, CM
通讯作者：
Rice, CM