Structure-guided mutagenesis of the capsid protein indicates that a nanovirus requires assembled viral particles for systemic infection

2023

Article référencé dans le Web of Science WOS:000935338800001

Trapani S., Bhat E. A., Yvon M., Lai-Kee-Him J., Hoh F., Vernerey M. S., Pirolles E., Bonnamy M., Schoehn G., Zeddam Jean-Louis, Blanc S., Bron P.

PLoS Pathogens, 2023, 19 (1), p. e1011086 [ 25 p.] ISSN 1553-7366

The genome of multipartite viruses is divided in two or more segments, each encapsidated separately in an individual viral particle. An unresolved question about these viral systems is the maintenance of the genome integrity. Two opposing hypotheses exist. One proposes that, somehow, the distinct segments can be sorted and assembled into a macromolecular complex containing at least one copy of each. This imposes either a propagation as non-encapsidated nucleic acids that could interact to form such "sorted" complexes, or structural differences between viral particles containing distinct segments also allowing assembly of sorted complexes. The other hypothesis postulates that the different segments spread independently, individually packaged in structurally similar particles, which eventually come together at random. We first determined the atomic structure of viral particles of a nanovirus (FBNSV), where no major structural differences were detected depending on the encapsidated segment. We then carried out structure-guided mutagenesis to prevent particle assembly, and thereby compromised systemic infection of host plants. These results strongly suggest that the viral genome moves long distance as assembled viral particles. Such a situation, where all particles of the viral population are alike, does not support the existence of a large macromolecular complex which would form by specifically sorting and assembling particles each containing a distinct segment. Instead, our results, together with other evidence from totally unrelated approaches, further support the independent and random propagation of the distinct segments of the FBNSV genome. Nanoviruses are plant multipartite viruses with a genome composed of six to eight circular single-stranded DNA segments. The distinct genome segments are encapsidated individually in icosahedral particles that measure approximate to 18 nm in diameter. Recent studies on the model species Faba bean necrotic stunt virus (FBNSV) revealed that complete sets of genomic segments rarely occur in infected plant cells and that the function encoded by a given viral segment can complement the others across neighbouring cells, presumably by translocation of the gene products through unknown molecular processes. This allows the viral genome to replicate, assemble into viral particles and infect anew, even with the distinct genome segments scattered in different cells. Here, we question the form under which the FBNSV genetic material propagates long distance within the vasculature of host plants and, in particular, whether viral particle assembly is required. Using structure-guided mutagenesis based on a 3.2 angstrom resolution cryogenic-electron-microscopy reconstruction of the FBNSV particles, we demonstrate that specific site-directed mutations preventing capsid formation systematically suppress FBNSV long-distance movement, and thus systemic infection of host plants, despite positive detection of the mutated coat protein when the corresponding segment is agroinfiltrated into plant leaves. These results strongly suggest that the viral genome does not propagate within the plant vascular system under the form of uncoated DNA molecules or DNA:coat-protein complexes, but rather moves long distance as assembled viral particles.

Sciences fondamentales / Techniques d'analyse et de recherche [020] ; Sciences du monde végétal [076]

Fonds IRD [F B010087460]

fdi:010087460