Dataset Information

Nucleotide sequence of the Spiroplasma citri fibril protein gene.

ABSTRACT: Electron microscopic observation of spiroplasmas lysed by detergent (sodium deoxycholate) revealed the release of bundles of fibrils from the cells. Individual fibrils are 4 nm in diameter and possess a 9-nm periodicity along their length. These fibrils are thought to function as cytoskeletal structures involved in the shape and motility of spiroplasmas. Polyacrylamide gel electrophoresis of density gradient-purified fibrils showed a protein of approximately 55 kDa. Oligonucleotide probes were constructed from the N-terminal amino acid sequence of two peptides obtained after V8 protease hydrolysis of the fibril protein. The probes were used to identify the clones in a genomic DNA library of Spiroplasma citri that contained inserts carrying the probe sequence. Sequencing of a 3.3-kbp fragment yielded the full open reading frame of the fibril protein gene and the start of a second open reading frame of an unknown protein. The fibril protein is composed of 515 amino acids, which have a computed molecular mass of 59 kDa. Northern (RNA) blot hybridization and primer extension experiments showed that transcription of the fibril protein gene starts from a promoter located 100 nucleotides upstream of the initiation codon and stops at a rho-independent type terminator, leading to a 1.7-kbp transcript. Southern blot hybridization of genomic DNA using the fibril protein gene as the probe showed that a single copy of the gene is present in the chromosomes of both S. citri and Spiroplasma melliferum. The genotypic symbol fib is proposed for the spiroplasma fibril protein gene.

SUBMITTER: Williamson DL

PROVIDER: S-EPMC208096 | biostudies-other | 1991 Jul

REPOSITORIES: biostudies-other

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Nucleotide sequence of the Spiroplasma citri fibril protein gene.

Williamson D L DL Renaudin J J Bové J M JM

Journal of bacteriology 19910701 14

Electron microscopic observation of spiroplasmas lysed by detergent (sodium deoxycholate) revealed the release of bundles of fibrils from the cells. Individual fibrils are 4 nm in diameter and possess a 9-nm periodicity along their length. These fibrils are thought to function as cytoskeletal structures involved in the shape and motility of spiroplasmas. Polyacrylamide gel electrophoresis of density gradient-purified fibrils showed a protein of approximately 55 kDa. Oligonucleotide probes were c ...[more]

PMID: 1712358

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Project description:BackgroundSpiroplasma citri is a cell wall-less, plant pathogenic bacteria that colonizes two distinct hosts, the leafhopper vector and the host plant. Given the absence of a cell wall, surface proteins including lipoproteins and transmembrane polypeptides are expected to play key roles in spiroplasma/host interactions. Important functions in spiroplasma/insect interactions have been shown for a few surface proteins such as the major lipoprotein spiralin, the transmembrane S. citri adhesion-related proteins (ScARPs) and the sugar transporter subunit Sc76. S. citri efficient transmission from the insect to the plant is expected to rely on its ability to adapt to the different environments and more specifically to regulate the expression of genes encoding surface-exposed proteins.ResultsGenes encoding S. citri lipoproteins and ScARPs were investigated for their expression level in axenic medium, in the leafhopper vector Circulifer haematoceps and in the host plant (periwinkle Catharanthus roseus) either insect-infected or graft-inoculated. The vast majority of the lipoprotein genes tested (25/28) differentially responded to the various host environments. Considering their relative expression levels in the different environments, the possible involvement of the targeted genes in spiroplasma host adaptation was discussed. In addition, two S. citri strains differing notably in their ability to express adhesin ScARP2b and pyruvate dehydrogenase E1 component differed in their capacity to multiply in the two hosts, the plant and the leafhopper vector.ConclusionsThis study provided us with a list of genes differentially expressed in the different hosts, leading to the identification of factors that are thought to be involved in the process of S. citri host adaptation. The identification of such factors is a key step for further understanding of S. citri pathogenesis. Moreover the present work highlights the high capacity of S. citri in tightly regulating the expression level of a large set of surface protein genes, despite the small size of its genome.

Project description:BackgroundSpiroplasma citri BR3-3X and S. kunkelii CR2-3X cause serious diseases worldwide on citrus and maize species, respectively. S. citri BR3-3X harbors a plasmid, pBJS-Original (pBJS-O), that encodes the spiroplasma adhesion related protein 1 (SARP1), a protein implicated in binding of the pathogen to cells of its leafhopper vector, Circulifer tenellus. The S. kunkelii CR2-3X plasmid, pSKU146, encodes a homolog of SARP1, Sk-ARP1. Due to the close phylogenetic relationship of the two pathogens, we hypothesized that the two plasmids are closely related as well.ResultsThe nucleotide sequence of pBJS-O was determined and compared to the sequences of a plasmid from BR3-T (pBJS-T), which is a multiply passaged leafhopper transmissible derivative of BR3-3X, and to known plasmid sequences including that of pSKU146. In addition to arp1, the 13,374 bp pBJS-O sequence putatively contains nine genes, recognized as open reading frames (ORFs). Several pBJS-O ORFs have homologs on pSKU146. However, the sequences flanking soj-like genes on both plasmids were found to be more distant from one another than sequences in any other region. Further, unlike pSKU146, pBJS-O lacks the conserved oriT region characteristic of the IncP group of bacterial plasmids. We were unable to identify a region in pBJS-O resembling a known plasmid origin of transfer. In regions where sequence was available for the plasmid from both BR3-3X and BR3-T, the pBJS-T sequence had a 0.4 kb deletion relative to its progenitor, pBJS-O. Southern blot hybridization of extrachromosomal DNA from various S. citri strains and spiroplasma species to an arp-specific probe and a probe made from the entire plasmid DNA of BR3-3X revealed limited conservation of both sequences in the genus Spiroplasma. Finally, we also report the presence on the BR3-3X chromosome of arp2, an S. citri homolog of arp1 that encodes the predicted protein SARP2. The C-terminal domain of SARP2 is homologous to that of SARP1, but its N-terminal domain is distinct.ConclusionOur data suggest that pBJS is a novel S. citri plasmid that does not belong to any known plasmid incompatibility group. The differences between pBJS-O and pSKU146 suggest that one or more events of recombination have contributed to the divergence of the plasmids of the two sister Spiroplasma species; the plasmid from S. citri itself has diverged slightly during the derivation of S. citri BR3-T from BR3-3X. Our data also show that pBJS-O encodes the putative adhesin SARP1. The presence of traE and mob on pBJS-O suggests a role for the plasmid in spiroplasmal conjugation.

Dataset Information

Nucleotide sequence of the Spiroplasma citri fibril protein gene.

Publications

Nucleotide sequence of the Spiroplasma citri fibril protein gene.

Similar Datasets

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