A temperature sensitive mutant of heat shock protein 70 reveals an essential role during the early steps of tombusvirus replication.
A temperature sensitive mutant of heat shock protein 70 reveals an essential role during the early steps of tombusvirus replication.
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DOI：
10.1016/j.virol.2009.08.003
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发表时间：
2009-11-10
期刊：
影响因子：
3.7
通讯作者：
Nagy, Peter D.
中科院分区：
文献类型：
--
作者：
Wang, Robert Yung-Liang;Stork, Jozsef;Pogany, Judit;Nagy, Peter D.
正链RNA病毒通过选择宿主蛋白进行复制，可以支持稳健的复制并抑制宿主的抗病毒应答。番茄丛矮病毒（Tomato bushy stunt tombusvirus，TBSV）将细胞内大量的分子伴侣热休克蛋白70（Hsp 70）募集到复制酶复合体中。利用酵母模型宿主，我们证明了热休克蛋白70基因的四个成员SSA亚家族是TBSV复制所必需的。组成型表达的SSA 1和SSA 2是病毒复制酶中的驻留蛋白，可以通过热诱导的SSA 3和/或SSA 4进行补充以用于TBSV复制。使用酵母菌株携带的温度敏感ssa 1 ts，但缺乏功能SSA 2/3/4，我们表明，Ssa 1 pts的失活导致的缺陷，在膜定位的病毒复制蛋白，导致在胞质分布的病毒蛋白和缺乏复制酶活性。Ssa 1 pts的体外复制酶组装试验表明，在复制酶组装过程中需要功能性Ssa 1 p，但在负链或正链合成过程中不需要。在ssa 1 ts酵母中进行的从非允许到允许的温度转换实验表明，在没有其他SSA基因的情况下，重新激活的Ssa 1 pts可以促进有效的TBSV复制。我们还证明，纯化的重组Ssa 3 p可以促进TBSV复制酶在酵母膜上的体外组装，表明Ssa 3 p可以完全补充Ssa 1 p的功能。两者合计，热休克蛋白70蛋白质的胞质SSA亚家族在TBSV复制中发挥重要的和多重的作用。
By co-opting host proteins for their replication, plus-stranded RNA viruses can support robust replication and suppress host anti-viral responses. Tomato bushy stunt tombusvirus (TBSV) recruit the cellular heat shock protein 70 (Hsp70), an abundant cytosolic chaperone, into the replicase complex. By taking advantage of yeast model host, we demonstrate that the four-member SSA subfamily of HSP70 genes is essential for TBSV replication. The constitutively-expressed SSA1 and SSA2, which are resident proteins in the viral replicase, can be complemented by the heat-inducible SSA3 and/or SSA4 for TBSV replication. Using a yeast strain carrying a temperature sensitive ssa1ts, but lacking functional SSA2/3/4, we show that inactivation of Ssa1pts led to a defect in membrane localization of the viral replication proteins, resulting in cytosolic distribution of the viral proteins and lack of replicase activity. An in vitro replicase assembly assay with Ssa1pts revealed that functional Ssa1p is required during the replicase assembly process, but not during minus- or plus-strand synthesis. Temperature shift experiments from nonpermissive to permissive in ssa1ts yeast revealed that the re-activated Ssa1pts could promote efficient TBSV replication in the absence of other SSA genes. We also demonstrate that the purified recombinant Ssa3p can facilitate the in vitro assembly of the TBSV replicase on yeast membranes, demonstrating that Ssa3p can fully complement the function of Ssa1p. Taken together, the cytosolic SSA subfamily of Hsp70 proteins play essential and multiple roles in TBSV replication.
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影响因子：
3.7
作者：
Li, Zhenghe;Pogany, Judit;Panavas, Tadas;Xu, Kai;Esposito, Anthony M.;Kinzy, Terri Goss;Nagy, Peter D.
通讯作者：
Nagy, Peter D.
影响因子：
3.7
作者：
Nagy, PD;Pogany, J
通讯作者：
Pogany, J
影响因子：
3.7
作者：
Dufresne, Philippe J.;Thivierge, Karine;Fortin, Marc G.
通讯作者：
Fortin, Marc G.
影响因子：
64.8
作者：
Krishnan, Manoj N.;Ng, Aylwin;Sukumaran, Bindu;Gilfoy, Felicia D.;Uchil, Pradeep D.;Sultana, Hameeda;Brass, Abraham L.;Adametz, Rachel;Tsui, Melody;Qian, Feng;Montgomery, Ruth R.;Lev, Sima;Mason, Peter W.;Koski, Raymond A.;Elledge, Stephen J.;Xavier, Ramnik J.;Agaisse, Herve;Fikrig, Erol
通讯作者：
Fikrig, Erol
DOI：
10.1073/pnas.93.26.15289
发表时间：
1996-12-24
影响因子：
11.1
作者：
Aranda, MA;Escaler, M;Maule, AJ
通讯作者：
Maule, AJ