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Identification and characterization of Sclerotinia sclerotiorum NADPH oxidases.

ABSTRACT: Numerous studies have shown both the detrimental and beneficial effects of reactive oxygen species (ROS) in animals, plants, and fungi. These organisms utilize controlled generation of ROS for signaling, pathogenicity, and development. Here, we show that ROS are essential for the pathogenic development of Sclerotinia sclerotiorum, an economically important fungal pathogen with a broad host range. Based on the organism's completed genome sequence, we identified two S. sclerotiorum NADPH oxidases (SsNox1 and SsNox2), which presumably are involved in ROS generation. RNA interference (RNAi) was used to examine the function of SsNox1 and SsNox2. Silencing of SsNox1 expression indicated a central role for this enzyme in both virulence and pathogenic (sclerotial) development, while inactivation of the SsNox2 gene resulted in limited sclerotial development, but the organism remained fully pathogenic. ?Ssnox1 strains had reduced ROS levels, were unable to develop sclerotia, and unexpectedly correlated with significantly reduced oxalate production. These results are in accordance with previous observations indicating that fungal NADPH oxidases are required for pathogenic development and are consistent with the importance of ROS regulation in the successful pathogenesis of S. sclerotiorum.

SUBMITTER: Kim HJ

PROVIDER: S-EPMC3209176 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Identification and characterization of Sclerotinia sclerotiorum NADPH oxidases.

Kim Hyo-jin HJ Chen Changbin C Kabbage Mehdi M Dickman Martin B MB

Applied and environmental microbiology 20110902 21

Numerous studies have shown both the detrimental and beneficial effects of reactive oxygen species (ROS) in animals, plants, and fungi. These organisms utilize controlled generation of ROS for signaling, pathogenicity, and development. Here, we show that ROS are essential for the pathogenic development of Sclerotinia sclerotiorum, an economically important fungal pathogen with a broad host range. Based on the organism's completed genome sequence, we identified two S. sclerotiorum NADPH oxidases ...[more]

PMID: 21890677

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Project description:BackgroundSeveral phytopathogens produce small non-coding RNAs of approximately 18-30 nucleotides (nt) which post-transcriptionally regulate gene expression. Commonly called small RNAs (sRNAs), these small molecules were also reported to be present in the necrotrophic pathogen Sclerotinia sclerotiorum. S. sclerotiorum causes diseases in more than 400 plant species, including the important oilseed crop Brassica napus. sRNAs can further be classified as microRNAs (miRNAs) and short interfering RNAs (siRNAs). Certain miRNAs can activate loci that produce further sRNAs; these secondary sRNA-producing loci are called 'phased siRNA' (PHAS) loci and have only been described in plants. To date, very few studies have characterized sRNAs and their endogenous targets in S. sclerotiorum.ResultsWe used Illumina sequencing to characterize sRNAs from fungal mycelial mats of S. sclerotiorum spread over B. napus leaves. In total, eight sRNA libraries were prepared from in vitro, 12 h post-inoculation (HPI), and 24 HPI mycelial mat samples. Cluster analysis identified 354 abundant sRNA clusters with reads of more than 100 Reads Per Million (RPM). Differential expression analysis revealed upregulation of 34 and 57 loci at 12 and 24 HPI, respectively, in comparison to in vitro samples. Among these, 25 loci were commonly upregulated. Altogether, 343 endogenous targets were identified from the major RNAs of 25 loci. Almost 88% of these targets were annotated as repeat element genes, while the remaining targets were non-repeat element genes. Fungal degradome reads confirmed cleavage of two transposable elements by one upregulated sRNA. Altogether, 24 milRNA loci were predicted with both mature and milRNA* (star) sequences; these are both criteria associated previously with experimentally verified miRNAs. Degradome sequencing data confirmed the cleavage of 14 targets. These targets were related to repeat element genes, phosphate acetyltransferases, RNA-binding factor, and exchange factor. A PHAS gene prediction tool identified 26 possible phased interfering loci with 147 phasiRNAs from the S. sclerotiorum genome, suggesting this pathogen might produce sRNAs that function similarly to miRNAs in higher eukaryotes.ConclusionsOur results provide new insights into sRNA populations and add a new resource for the study of sRNAs in S. sclerotiorum.

Dataset Information

Identification and characterization of Sclerotinia sclerotiorum NADPH oxidases.

Publications

Identification and characterization of Sclerotinia sclerotiorum NADPH oxidases.

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