Laboratory of Virology, Wageningen University, Wageningen, The Netherlands.
J Virol. 2012 May;86(9):4981-8. doi: 10.1128/JVI.06801-11. Epub 2012 Feb 29.
Baculovirus occlusion-derived virus (ODV) infects insect midgut cells under alkaline conditions, a process mediated by highly conserved per os infectivity factors (PIFs), P74 (PIF0), PIF1, PIF2, PIF3, PIF4, and PIF5 (ODV-E56). Previously, a multimolecular complex composed of PIF1, PIF2, PIF3, and P74 was identified which was proposed to play an essential role during ODV entry. Recently, more proteins have been identified that play important roles in ODV oral infectivity, including PIF4, PIF5, and SF58, which might work in concert with previously known PIFs to facilitate ODV infection. In order to understand the ODV entry mechanism, the identification of all components of the PIF complex is crucial. Hence, the aim of this study was to identify additional components of the PIF complex. Coimmunoprecipitation (CoIP) combined with proteomic analysis was used to identify the components of the Autographa californica multiple nucleopolyhedrovirus (AcMNPV) PIF complex. PIF4 and P95 (AC83) were identified as components of the PIF complex while PIF5 was not, and this was confirmed with blue native PAGE and a second CoIP. Deletion of the pif4 gene impaired complex formation, but deletion of pif5 did not. Differentially denaturing SDS-PAGE further revealed that PIF4 forms a stable complex with PIF1, PIF2, and PIF3. P95 and P74 are more loosely associated with this complex. Three other proteins, AC5, AC68, and AC108 (homologue of SF58), were also found by the proteomic analysis to be associated with the PIF complex. Finally the functional significance of the PIF protein interactions is discussed.
杆状病毒出芽型病毒 (ODV) 在碱性条件下感染昆虫中肠细胞，这一过程由高度保守的经口感染因子 (PIFs) 介导，包括 P74 (PIF0)、PIF1、PIF2、PIF3、PIF4 和 PIF5 (ODV-E56)。先前，已鉴定出一种由 PIF1、PIF2、PIF3 和 P74 组成的多分子复合物，该复合物被认为在 ODV 进入过程中发挥重要作用。最近，更多的蛋白质已被鉴定出来，它们在 ODV 的经口感染中发挥重要作用，包括 PIF4、PIF5 和 SF58，它们可能与先前已知的 PIFs 协同作用，促进 ODV 感染。为了了解 ODV 进入机制，鉴定 PIF 复合物的所有组成部分至关重要。因此，本研究的目的是鉴定 PIF 复合物的其他组成部分。使用免疫共沉淀 (CoIP) 结合蛋白质组学分析来鉴定苜蓿银纹夜蛾多核型多角体病毒 (AcMNPV) PIF 复合物的组成部分。鉴定出 PIF4 和 P95 (AC83) 是 PIF 复合物的组成部分，而 PIF5 则不是，这一点通过蓝色非变性 PAGE 和第二次 CoIP 得到了证实。pif4 基因缺失会损害复合物的形成，但 pif5 基因缺失则不会。差异变性 SDS-PAGE 进一步表明，PIF4 与 PIF1、PIF2 和 PIF3 形成稳定的复合物。P95 和 P74 与该复合物的结合则较为松散。蛋白质组学分析还发现另外 3 种蛋白质 AC5、AC68 和 AC108（SF58 的同源物）与 PIF 复合物相关。最后，讨论了 PIF 蛋白相互作用的功能意义。
