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Molecular characterization of an endopolygalacturonase from Fusarium oxysporum expressed during early stages of infection.

ABSTRACT: The tomato vascular wilt pathogen Fusarium oxysporum f. sp. lycopersici produces an array of pectinolytic enzymes that may contribute to penetration and colonization of the host plant. Here we report the isolation of pg5, encoding a novel extracellular endopolygalacturonase (endoPG) that is highly conserved among different formae speciales of F. oxysporum. The putative mature pg5 product has a calculated molecular mass of 35 kDa and a pI of 8.3 and is more closely related to endoPGs from other fungal plant pathogens than to PG1, the major endoPG of F. oxysporum. Overexpression of pg5 in a bacterial heterologous system produced a 35-kDa protein with endoPG activity. Accumulation of pg5 transcript is induced by citrus pectin and D-galacturonic acid and repressed by glucose. As shown by reverse transcription-PCR, pg5 is expressed by F. oxysporum in tomato roots during the initial stages of infection. Targeted inactivation of pg5 has no detectable effect on virulence toward tomato plants.

SUBMITTER: Garcia-Maceira FI

PROVIDER: S-EPMC92854 | biostudies-literature | 2001 May

REPOSITORIES: biostudies-literature

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Molecular characterization of an endopolygalacturonase from Fusarium oxysporum expressed during early stages of infection.

García-Maceira F I FI Di Pietro A A Huertas-González M D MD Ruiz-Roldán M C MC Roncero M I MI

Applied and environmental microbiology 20010501 5

The tomato vascular wilt pathogen Fusarium oxysporum f. sp. lycopersici produces an array of pectinolytic enzymes that may contribute to penetration and colonization of the host plant. Here we report the isolation of pg5, encoding a novel extracellular endopolygalacturonase (endoPG) that is highly conserved among different formae speciales of F. oxysporum. The putative mature pg5 product has a calculated molecular mass of 35 kDa and a pI of 8.3 and is more closely related to endoPGs from other f ...[more]

PMID: 11319099

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Project description:BackgroundChrysanthemum Fusarium wilt is a common fungal disease caused by Fusarium oxysporum, which causes continuous cropping obstacles and huge losses to the chrysanthemum industry. The defense mechanism of chrysanthemum against F. oxysporum remains unclear, especially during the early stages of the disease. Therefore, in the present study, we analyzed chrysanthemum 'Jinba' samples inoculated with F. oxysporum at 0, 3, and 72 h using RNA-seq.ResultsThe results revealed that 7985 differentially expressed genes (DEGs) were co-expressed at 3 and 72 h after F. oxysporum infection. We analyzed the identified DEGs using Kyoto Encyclopedia of Genes and Genomes and Gene Ontology. The DEGs were primarily enriched in "Plant pathogen interaction", "MAPK signaling pathway", "Starch and sucrose metabolism", and "Biosynthesis of secondary metabolites". Genes related to the synthesis of secondary metabolites were upregulated in chrysanthemum early during the inoculation period. Furthermore, peroxidase, polyphenol oxidase, and phenylalanine ammonia-lyase enzymes were consistently produced to accumulate large amounts of phenolic compounds to resist F. oxysporum infection. Additionally, genes related to the proline metabolic pathway were upregulated, and proline levels accumulated within 72 h, regulating osmotic balance in chrysanthemum. Notably, the soluble sugar content in chrysanthemum decreased early during the inoculation period; we speculate that this is a self-protective mechanism of chrysanthemums for inhibiting fungal reproduction by reducing the sugar content in vivo. In the meantime, we screened for transcription factors that respond to F. oxysporum at an early stage and analyzed the relationship between WRKY and DEGs in the "Plant-pathogen interaction" pathway. We screened a key WRKY as a research target for subsequent experiments.ConclusionThis study revealed the relevant physiological responses and gene expression changes in chrysanthemum in response to F. oxysporum infection, and provided a relevant candidate gene pool for subsequent studies on chrysanthemum Fusarium wilt.

Project description:BACKGROUND:Upon exposure to unfavorable environmental conditions, plants need to respond quickly to maintain their homeostasis. For instance, physiological, biochemical and transcriptional changes occur during plant-pathogen interaction. In the case of Vanilla planifolia Jacks., a worldwide economically important crop, it is susceptible to Fusarium oxysporum f. sp. vanillae (Fov). This pathogen causes root and stem rot (RSR) in vanilla plants that lead to plant death. To investigate how vanilla plants, respond at the transcriptional level upon infection with Fov, here we employed the RNA-Seq approach to analyze the dynamics of whole-transcriptome changes during two-time frames of the infection. RESULTS:Analysis of global gene expression profiles upon infection by Fov indicated that the major transcriptional change occurred at 2?days post-inoculation (dpi), in comparison to 10 dpi. Briefly, the RNA-Seq analysis carried out in roots found that 3420 and 839 differentially expressed genes (DEGs) were detected at 2 and 10 dpi, respectively, as compared to the control. In the case of DEGs at 2 dpi, 1563 genes were found to be up-regulated, whereas 1857 genes were down-regulated. Moreover, functional categorization of DEGs at 2 dpi indicated that up-regulated genes are mainly associated to translation, whereas down-regulated genes are involved in cell wall remodeling. Among the translational-related transcripts, ribosomal proteins (RPs) were found increased their expression exclusively at 2 dpi. CONCLUSIONS:The screening of transcriptional changes of V. planifolia Jacks upon infection by Fov provides insights into the plant molecular response, particularly at early stages of infection. The accumulation of translational-related transcripts at early stages of infection potentially points to a transcriptional reprogramming coupled with a translational regulation in vanilla plants upon infection by Fov. Altogether, the results presented here highlight potential molecular players that might be further studied to improve Fov-induced resistance in vanilla plants.

Project description:Monitoring of pathogenic strains of Fusarium oxysporum (Fox), which cause wilt and rots on agricultural and ornamental plants, is important for predicting disease outbreaks. Since both pathogenic and non-pathogenic strains of Fox are ubiquitous and are able to colonize plant roots, detection of Fox DNA in plant material is not the ultimate proof of an ongoing infection which would cause damage to the plant. We followed the colonization of tomato plants by strains Fox f. sp. radicis-lycopersici ZUM2407 (a tomato foot and root rot pathogen), Fox f. sp. radiciscucumerinum V03-2g (a cucumber root rot pathogen) and Fox Fo47 (a well-known non-pathogenic biocontrol strain). We determined fungal DNA concentrations in tomato plantlets by quantitative PCR (qPCR) with primers complementary to the intergenic spacer region (IGS) of these three Fox strains. Two weeks after inoculation of tomato seedlings with these Fox strains, the DNA concentration of Forl ZUM2407 was five times higher than that of the non-compatible pathogen Forc V03-2g and 10 times higher than that of Fo47. In 3-week-old plantlets the concentration of Forl ZUM2407 DNA was at least 10 times higher than those of the other strains. The fungal DNA concentration, as determined by qPCR, appeared to be in good agreement with data of the score of visible symptoms of tomato foot and root rot obtained 3 weeks after inoculation of tomato with Forl ZUM2407. Our results show that targeting of the multicopy ribosomal operon results in a highly sensitive qPCR reaction for the detection of Fox DNA. Since formae speciales of Fox cannot be distinguished by comparison of ribosomal operons, detection of Fox DNA is not evidence of plant infection by a compatible pathogen. Nevertheless, the observed difference in levels of plant colonization between pathogenic and non-pathogenic strains strongly suggests that a concentration of Fox DNA in plant material above the threshold level of 0.005% is due to proliferation of pathogenic Fox.

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Molecular characterization of an endopolygalacturonase from Fusarium oxysporum expressed during early stages of infection.

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Molecular characterization of an endopolygalacturonase from Fusarium oxysporum expressed during early stages of infection.

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