Structural analysis of the spiroplasma virus, SpV4: implications for evolutionary variation to obtain host diversity among the Microviridae

AbstractBackground: Spiroplasma virus, SpV4, is a small, non-enveloped virus that infects the helical mollicute Spiroplasma melliferum. SpV4 exhibits several similarities to the Chlamydia phage, Chp1, and the Coliphages α3, φK, G4 and φX174. All of these viruses are members of the Microviridae. These viruses have isometric capsids with T = 1 icosahedral symmetry, cause lytic infections and are the only icosahedral phages that contain single-stranded circular DNA genomes. The aim of this comparative study on these phages was to understand the role of their capsid proteins during host receptor recognition.Results: The three-dimensional structure of SpV4 was determined to 27 å resolution from images of frozen-hydrated particles. Cryo-electron microscopy (cryo-EM) revealed 20, ∼54 å long, ‘mushroom-like’ protrusions on the surface of the capsid. Each protrusion comprises a trimeric structure that extends radially along the threefold icosahedral axes of the capsid. A 71 amino acid portion of VP1 (the SpV4 capsid protein) was shown, by structural alignment with the atomic structure of the F capsid protein of φX174, to represent an insertion sequence between the E and F strands of the eight-stranded antiparallel β-barrel. Secondary structure prediction of this insertion sequence provided the basis for a probable structural motif, consisting of a six-stranded antiparallel β sheet connected by small turns. Three such motifs form the rigid stable trimeric structures (mushroom-like protrusions) at the threefold axes, with hydrophobic depressions at their distal surface.Conclusions: Sequence alignment and structural analysis indicate that distinct genera of the Microviridae might have evolved from a common primordial ancestor, with capsid surface variations, such as the SpV4 protrusions, resulting from gene fusion events that have enabled diverse host ranges. The hydrophobic nature of the cavity at the distal surface of the SpV4 protrusions suggests that this region may function as the receptor-recognition site during host infection

The abundant extrachromosomal DNA content of the Spiroplasma citri GII3-3X genome

<p>Abstract</p> <p>Background</p> <p><it>Spiroplama citri</it>, the causal agent of citrus stubborn disease, is a bacterium of the class <it>Mollicutes </it>and is transmitted by phloem-feeding leafhopper vectors. In order to characterize candidate genes potentially involved in spiroplasma transmission and pathogenicity, the genome of <it>S. citri </it>strain GII3-3X is currently being deciphered.</p> <p>Results</p> <p>Assembling 20,000 sequencing reads generated seven circular contigs, none of which fit the 1.8 Mb chromosome map or carried chromosomal markers. These contigs correspond to seven plasmids: pSci1 to pSci6, with sizes ranging from 12.9 to 35.3 kbp and pSciA of 7.8 kbp. Plasmids pSci were detected as multiple copies in strain GII3-3X. Plasmid copy numbers of pSci1-6, as deduced from sequencing coverage, were estimated at 10 to 14 copies per spiroplasma cell, representing 1.6 Mb of extrachromosomal DNA. Genes encoding proteins of the TrsE-TraE, Mob, TraD-TraG, and Soj-ParA protein families were predicted in most of the pSci sequences, in addition to members of 14 protein families of unknown function. Plasmid pSci6 encodes protein P32, a marker of insect transmissibility. Plasmids pSci1-5 code for eight different <it>S. citri </it>adhesion-related proteins (ScARPs) that are homologous to the previously described protein P89 and the <it>S. kunkelii </it>SkARP1. Conserved signal peptides and C-terminal transmembrane alpha helices were predicted in all ScARPs. The predicted surface-exposed N-terminal region possesses the following elements: (i) 6 to 8 repeats of 39 to 42 amino acids each (sarpin repeats), (ii) a central conserved region of 330 amino acids followed by (iii) a more variable domain of about 110 amino acids. The C-terminus, predicted to be cytoplasmic, consists of a 27 amino acid stretch enriched in arginine and lysine (KR) and an optional 23 amino acid stretch enriched in lysine, aspartate and glutamate (KDE). Plasmids pSci mainly present a linear increase of cumulative GC skew except in regions presenting conserved hairpin structures.</p> <p>Conclusion</p> <p>The genome of <it>S. citri </it>GII3-3X is characterized by abundant extrachromosomal elements. The pSci plasmids could not only be vertically inherited but also horizontally transmitted, as they encode proteins usually involved in DNA element partitioning and cell to cell DNA transfer. Because plasmids pSci1-5 encode surface proteins of the ScARP family and pSci6 was recently shown to confer insect transmissibility, diversity and abundance of <it>S. citri </it>plasmids may essentially aid the rapid adaptation of <it>S. citri </it>to more efficient transmission by different insect vectors and to various plant hosts.</p

Plasmid and viral vectors for gene cloning and expression in Spiroplasma citri

International audienc

SpV1 and SpV4, spiroplasma viruses with circular, single-stranded DNA genomes, and their contribution to the molecular biology of spiroplasmas

International audienc

Mutagenesis of a tryptophan codon from TGG to TGA in the CAT gene does not prevent its expression in the helical mollicute Spiroplasma citri

International audienc

Interaction entre le phytoplasme du stolbur et la tomate : recherche de lignées résistantes et effet de l'activation des voies de signalisation dépendantes du salicylate et du jasmonate

National audienc