Abstract
The production of plant helical virus-like particles (VLPs) via plant-based expression has been problematic with previous studies suggesting that an RNA scaffold may be necessary for their efficient production. To examine this, we compared the accumulation of VLPs from two potexviruses, papaya mosaic virus and alternanthera mosaic virus (AltMV), when the coat proteins were expressed from a replicating potato virus X- based vector (pEff) and a non-replicating vector (pEAQ-HT). Significantly greater quantities of VLPs could be purified when pEff was used. The pEff system was also very efficient at producing VLPs of helical viruses from different virus families. Examination of the RNA content of AltMV and tobacco mosaic virus VLPs produced from pEff revealed the presence of vector-derived RNA sequences, suggesting that the replicating RNA acts as a scaffold for VLP assembly. Cryo-EM analysis of the AltMV VLPs showed they had a structure very similar to that of authentic potexvirus particles. Thus, we conclude that vectors generating replicating forms of RNA, such as pEff, are very efficient for producing helical VLPs.
Highlights
Half of known RNA plant viruses have helical structures [1]
We showed that expression of the coat proteins of unrelated plant virus genera (TMV, cucumber vein yellowing virus (CVYV, genus Ipomovirus, family Potyviridae) and sweet potato feathery mottle virus (SPFMV, genus Potyvirus, family Potyviridae)) from the pEff vector leads to the efficient formation of helical virus-like particles (VLPs)
To investigate whether the use of a replicating vector could enhance the production of filamentous VLPs, we compared expression of two potexvirus coat proteins from alternanthera mosaic virus (AltMV) (AltMV-CP) and papaya mosaic virus (PapMV) (PapMV-CP), using pEAQ-HT or pEff. (Figure 1)
Summary
Half of known RNA plant viruses have helical structures [1]. Such viruses can be spread by vectors, mechanical damage or propagation and can cause severe symptoms in their host plant species with consequential yield losses impacting farmers and food security. The only exception to this was found when a replicating PVX vector was used to express the coat protein of the related potexvirus, alternanthera mosaic virus (AltMV; [16]) These results suggested that co-expression of the coat protein with an appropriate, replicating RNA might be the key to producing VLPs of helical plant viruses. The possibility that replicating RNA may be necessary for the efficient generation of helical VLPs would be consistent with our recent studies on the requirements for RNA packaging in the isometric particles of cowpea mosaic virus (CPMV; [17]) and satellite tobacco necrosis virus (STNV; [18]) To test this hypothesis, we have compared the level of coat protein and VLP accumulation of two potexviruses, AltMV and papaya mosaic virus (PapMV) when expressed from the non-replicating pEAQ-HT vector and the replicating PVX-based vector pEff [19]. We conclude that the deployment of a replicating vector, such as pEff, is an efficient approach to producing structurally relevant VLPs of helical viruses
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