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Amino Acid Sequence Motifs Essential for P0-Mediated Suppression of RNA Silencing in an Isolate of Potato leafroll virus from Inner Mongolia

机译:内蒙古马铃薯卷叶病毒分离物中P0介导的RNA沉默抑制所必需的氨基酸序列基序。

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Polerovirus P0 suppressors of host gene silencing contain a consensus F-box-like motif with Leu/Pro (L/P) requirements for suppressor activity. The Inner Mongolian Potato leafroll virus (PLRV) P0 protein (P0(PL-IM)) has an unusual F-box-like motif that contains a Trp/Gly (W/G) sequence and an additional GW/WG-like motif (G139/W140/G141) that is lacking in other P0 proteins. We used Agrobacterium infiltration-mediated RNA silencing assays to establish that P0(PL-IM) has a strong suppressor activity. Mutagenesis experiments demonstrated that the P0(PL-IM) F-box-like motif encompasses amino acids 76-LPRHLHYECLEWGLLCG THP-95, and that the suppressor activity is abolished by L76A, W87A, or G88A substitution. The suppressor activity is also weakened substantially by mutations within the G139/W140/G141 region and is eliminated by a mutation (F220R) in a C-terminal conserved sequence of P0(PL-IM). As has been observed with other P0 proteins, P0(PL-IM) suppression is correlated with reduced accumulation of the host AGO1-silencing complex protein. However, P0(PL-IM) fails to bind SKP1, which functions in a proteasome pathway that may be involved in AGO1 degradation. These results suggest that P0(PL-IM) may suppress RNA silencing by using an alternative pathway to target AGO1 for degradation. Our results help improve our understanding of the molecular mechanisms involved in PLRV infection.
机译:宿主基因沉默的脊髓灰质炎病毒P0抑制子包含一个共有的F盒样基序,具有抑制活性的Leu / Pro(L / P)要求。内蒙古马铃薯卷叶病毒(PLRV)P0蛋白(P0(PL-IM))具有不寻常的F-box-like基序,其中包含Trp / Gly(W / G)序列和一个额外的GW / WG-like基序( G139 / W140 / G141)在其他P0蛋白中缺乏。我们使用农杆菌浸润介导的RNA沉默测定法来确定P0(PL-IM)具有很强的抑制活性。诱变实验表明,P0(PL-IM)F-box-like基序包含氨基酸76-LPRHLHYECLEWGLLCG THP-95,并且抑制活性被L76A,W87A或G88A取代所废除。 G139 / W140 / G141区域内的突变也显着减弱了抑制活性,P0(PL-IM)的C端保守序列中的突变(F220R)消除了抑制活性。正如其他P0蛋白所观察到的,P0(PL-IM)抑制与宿主AGO1沉默复合蛋白的积累减少有关。但是,P0(PL-IM)无法结合SKP1,后者在可能参与AGO1降解的蛋白酶体途径中起作用。这些结果表明,P0(PL-IM)可能通过使用替代途径靶向降解的AGO1来抑制RNA沉默。我们的结果有助于增进我们对参与PLRV感染的分子机制的了解。

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