Structural Insights into the Molecular Mechanisms of Cauliflower Mosaic Virus Transmission by Its Insect Vector

Abstract

Cauliflower mosaic virus (CaMV) is transmitted from plant to plant through a seemingly simple interaction with insect vectors. This process involves an aphid receptor and two viral proteins, P2 and P3. P2 binds to both the aphid receptor and P3, itself tightly associated with the virus particle, with the ensemble forming a transmissible viral complex. Here, we describe the conformations of both unliganded CaMV P3 protein and its virion-associated form. X-ray crystallography revealed that the N-terminal domain of unliganded P3 is a tetrameric parallel coiled coil with a unique organization showing two successive four-stranded subdomains with opposite supercoiling handedness stabilized by a ring of interchain disulfide bridges. A structural model of virus-liganded P3 proteins, folding as an antiparallel coiled-coil network coating the virus surface, was derived from molecular modeling. Our results highlight the structural and biological versatility of this coiled-coil structure and provide new insights into the molecular mechanisms involved in CaMV acquisition and transmission by the insect vector.Cauliflower mosaic virus (CaMV) is the type member of the plant virus family Caulimoviridae. This family is grouped together with hepadnaviruses into the pararetrovirus group due to its mode of replication via reverse transcription of a pregenomic RNA intermediate (17). The CaMV genome is a double-stranded circular DNA of approximately 8,000 bp, comprising seven major open reading frames (ORFs), only six of which have clearly identified biological functions. The product of ORF VI (P6) is expressed early in infection from the monocistronic 19S RNA. In addition to its recently reported role in the suppression of RNA silencing (14), P6 rapidly self-aggregates into the so-called electron-dense inclusion bodies, demonstrated to be the viral factory (17). Within these structures, P6 then trans-activates the translation of ORFs I to V from the polycistronic pregenomic 35S RNA according to a complex reinitiation process (39). ORFs I to V encode the major capsid protein (P4, encoded by ORF IV), reverse transcriptase (P5, encoded by ORF V), and three auxiliary proteins: P1 (ORF I) is involved in cell-to-cell and long-distance within-plant movement, P2 (ORF II) is involved in aphid transmission, and P3 (ORF III) is tightly associated with the virus particles and has a complex regulatory function in the virus infection cycle, particularly during plant-to-plant vector transmission.The CaMV viral particle is roughly spherical, 520 Å in diameter, with an icosahedral T7 symmetry. It is constituted of three concentric shells, built from 420 capsid protein subunits, surrounding a large inner cavity with a diameter of approximately 250 Å (4, 37). The physical association between P3 and viral particles has been consistently demonstrated in mature CaMV virions by copurification, immunolabeling experiments, and in vitro interactions (7,(23)(24)(25). Recently, cryoelectron microscopy (cryo-EM) image reconstruction further showed that P3 molecu...

The helper-component protease transmission factor of tobacco etch potyvirus binds specifically to an aphid ribosomal protein homologous to the laminin receptor precursor

“…Interestingly, an equivalent RPS2 product was identified in cast cuticles from Anopheles gambiae after moulting (He et al, 2007), a fact that provides at least preliminary evidence for its presence in insect cuticles. Further work will be needed to confirm whether this is also the case for aphids, and particularly for the structures considered essential for virus transmission that have been described recently in aphid stylets (Uzest et al, 2010). Our results also raise an interesting question from an evolutionary perspective, that of how an insect ribosomal protein could become a specific receptor for a plant virus.…”

“…In this virus, particles form a stable complex with the viral P3 protein, which is then retained in the aphid vector stylets via an interaction with the aphid transmission factor P2 (Leh et al, 1999;Plisson et al, 2005;Hoh et al, 2010 …”

“…In this virus, particles form a stable complex with the viral P3 protein, which is then retained in the aphid vector stylets via an interaction with the aphid transmission factor P2 (Leh et al, 1999;Plisson et al, 2005;Hoh et al, 2010). Taking advantage of the ability to label the P2 protein in vivo with GFP, retention of P3-virion complexes was shown to take place by protein-protein interactions near the tip of the stylet (Uzest et al, 2007), a site where a new anatomical structure named the 'acrostyle' was recently described (Uzest et al, 2010).…”

Virus Factories of Cauliflower Mosaic Virus Are Virion Reservoirs That Engage Actively in Vector Transmission

“…Another signal pattern might trigger release of virions from VFs without simultaneous mixed-network formation, allowing virions to interact with P1 and leading to cell-to-cell movement. The nature of the signals and whether they overlap remains unknown; we propose that the previously hypothesized redox switch involving the cysteine ring of P3 plays a role as a signal receptor (40). P3 is indeed a good candidate, because it is strategically well placed on the virus capsid surface (10), interacting with P1 and P2, and it is found in both VFs and TBs.…”

Host cell processes to accomplish mechanical and non-circulative virus transmission

“…e During aphid vector probing punctures, the stylets visit a small number of epidermis and mesophyll cells. f-g According to the speculation by Hoh et al (2010), a change in the redox state induced by the aphid puncture and/or the saliva could result in disintegration of the TB and the release of P2 and P3 (orange). h Subsequently, P2 can be acquired by the aphid and attach to a receptor protein in the stylet cuticle.…”