A comparative proteome analysis reveals flagellin, chemotaxis regulated proteins and amylovoran to be involved in virulence differences between Erwinia amylovora strains

Holtappels, Michelle; Vrancken, Kristof; Schoofs, H.; Deckers, T.; Remans, Tony; Noben, Jean-Paul; Valcke, Roland

doi:10.1016/j.jprot.2015.03.036

Cited by 26 publications

(40 citation statements)

References 57 publications

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“…Although the analysis of the gene expression of E. amylovora using real-time PCR was already studied 38, 39 , to the best of our knowledge, this is the first study of the validation of reference genes using a multi-algorithm analysis for use with the relative quantification of target gene expression in this bacterial species.…”

Section: Discussionmentioning

confidence: 99%

Validation of reference genes for the normalization of the RT-qPCR gene expression of virulence genes of Erwinia amylovora in apple shoots

Kałużna

Kuras

Puławska

2017

Sci Rep

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To study the expression of pathogenicity-related genes in Erwinia amylovora, seven candidate reference genes (ffh, glyA, gyrA, proC, pykA, recA, rpoB) were selected and validated with the following five different mathematic algorithms: geNorm, NormFinder, BestKeeper, the delta CT method and the RefFinder web-based tool. An overall comprehensive ranking output from each of the selected software programs revealed that proC and recA, followed by ffh and pykA, were the most stably expressed genes and can be recommended for the normalization of RT-qPCR data. A combination of the three reference genes, proC, recA and ffh, allowed for the accurate expression analysis of amsB and hrpN genes and the calculation of their fold change in E. amylovora after its infection of susceptible and resistant apple cultivars. To the best of our knowledge, this is the first study presenting a list of the most suitable reference genes for use in the relative quantification of target gene expression in E. amylovora in planta, selected on the basis of a multi-algorithm analysis.

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Section: Discussionmentioning

confidence: 99%

Validation of reference genes for the normalization of the RT-qPCR gene expression of virulence genes of Erwinia amylovora in apple shoots

Kałużna

Kuras

Puławska

2017

Sci Rep

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“…Two main classes of motility reported in E. amylovora include flagellar motility (Raymundo and Ries, ) and twitching motility through type IV pili (Berry et al ., ). Flagellar motility provides a means whereby the E. amylovora cells can swim through flower nectar to reach host tissues, and it has been suggested that flagella play an important role in virulence (Holtappels et al ., ; Puławska et al ., ). Like other members of the Enterobacteriaceae family, E. amylovora bears peritrichous flagella.…”

Section: Introductionmentioning

confidence: 99%

Three Hfq‐dependent small RNAs regulate flagellar motility in the fire blight pathogen Erwinia amylovora

Schachterle

Zeng

Sundin

2019

Molecular Microbiology

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Erwinia amylovora, the causative agent of fire blight disease of apple and pear trees, causes disease on flowers by invading natural openings at the base of the floral cup. To reach these openings, the bacteria use flagellar motility to swim from stigma tips to the hypanthium and through nectar. We have previously shown that the Hfq-dependent sRNAs ArcZ, OmrAB and RmaA regulate swimming motility in E. amylovora. Here, we tested these three sRNAs to determine at what regulatory level they exert their effects and to what extent they can complement each other. We found that ArcZ and OmrAB repress the flagellar master regulator flhD post-transcriptionally. We also found that ArcZ and RmaA positively regulate flhD at the transcriptional level. The role of ArcZ as an activator of flagellar motility appears to be unique to E. amylovora and may have recently evolved. Our results suggest that the Hfq-dependent sRNAs ArcZ, OmrAB and RmaA play an integral role in regulation of flagellar motility by acting primarily on the master regulator, FlhD, but also through additional factors.

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“…Proteomics comparisons of E. amylovora strains with high and low virulence have revealed numerous differences potentially related to virulence and acclimation to the host environment (Holtappels et al, 2016b), including differential post-translational lysine acetylation (Wu et al, 2013) and a higher abundance of outer membrane proteins, such as TolC and OmpA, in a higher virulence strain (Holtappels et al, 2016a). In contrast, higher levels of flagellin in culture conditions are correlated with lower virulence (Holtappels et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

An Erwinia amylovora yjeK mutant exhibits reduced virulence, increased chemical sensitivity and numerous environmentally dependent proteomic alterations

Klee

Mostafa

Chen

et al. 2018

Molecular Plant Pathology

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The Gram-negative bacterium Erwinia amylovora causes fire blight, an economically important disease of apples and pears. Elongation factor P (EF-P) is a highly conserved protein that stimulates the formation of the first peptide bond of certain proteins and facilitates the translation of certain proteins, including those with polyproline motifs. YjeK and YjeA are two enzymes involved in the essential post-translational β-lysylation of EF-P at a conserved lysine residue, K34. EF-P, YjeA and YjeK have been shown to be essential for the full virulence of Escherichia coli, Salmonella species and Agrobacterium tumefaciens, with efp, yjeA and yjeK mutants having highly similar phenotypes. Here, we identified an E. amylovora yjeK::Tn5 transposon mutant with decreased virulence in apple fruit and trees. The yjeK::Tn5 mutant also showed pleiotropic phenotypes, including reduced growth in rich medium, lower extracellular polysaccharide production, reduced swimming motility and increased chemical sensitivity compared with the wild-type, whilst maintaining wild-type level growth in minimal medium. All yjeK::Tn5 mutant phenotypes were complemented in trans with a plasmid bearing a wild-type copy of yjeK. Comprehensive, quantitative proteomics analyses revealed numerous, environmentally dependent changes in the prevalence of a wide range of proteins, in higher abundance and lower abundance, in yjeK::Tn5 compared with the wild-type, and many of these alterations could be linked to yjeK::Tn5 mutant phenotypes. The environmental dependence of the yjeK::Tn5 mutant proteomic alterations suggests that YjeK could be required for aspects of the environmentally dependent regulation of protein translation. YjeK activity may be critical to overcoming stress, including the challenging host environment faced by invading pathogenic bacteria.

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A comparative proteome analysis reveals flagellin, chemotaxis regulated proteins and amylovoran to be involved in virulence differences between Erwinia amylovora strains

Cited by 26 publications

References 57 publications

Validation of reference genes for the normalization of the RT-qPCR gene expression of virulence genes of Erwinia amylovora in apple shoots

Validation of reference genes for the normalization of the RT-qPCR gene expression of virulence genes of Erwinia amylovora in apple shoots

Three Hfq‐dependent small RNAs regulate flagellar motility in the fire blight pathogen Erwinia amylovora

An Erwinia amylovora yjeK mutant exhibits reduced virulence, increased chemical sensitivity and numerous environmentally dependent proteomic alterations

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