Abstract:Over-expression of the potato Gibberellin Stimulated-Like 2 ( GSL2 ) gene in transgenic potato confers resistance to blackleg disease incited by Pectobacterium atrosepticum and confirms a role for GSL2 in plant defence. The Gibberellin Stimulated-Like 2 (GSL2) gene (also known as Snakin 2) encodes a cysteine-rich, low-molecular weight antimicrobial peptide produced in potato plants. This protein is thought to play important roles in the innate defence against invading microbes. Over-expression of the GSL2 gene… Show more
“…RGD residues) and functional (e.g. toxic activity) data supporting its relationship with cysteine-rich peptides from snakes venoms, St SN1 and St SN2 have been recently renamed as GSL1 and GSL2, respectively [8,9]. …”
Section: Introductionmentioning
confidence: 99%
“…Overexpression of SN1 in potato and wheat ( Triticum aestivum ), SN2 in potato and tomato ( Solanum lycopersicum ), and snakin-defensin hybrid protein in tobacco ( Nicotiana tabacum ) and potato restricts pathogen invasiveness and enhances tolerance to bacterial and fungal diseases, without altering the agronomic phenotype of these crops [6,9-12]. Furthermore, disease sensitivity is enhanced by silencing SN2 in wild tobacco ( Nicotiana benthamiana ), supporting the central role of snakin peptides in plant defense [13].…”
BackgroundThe production of antimicrobial peptides is a common defense strategy of living cells against a wide range of pathogens. Plant snakin peptides inhibit bacterial and fungal growth at extremely low concentrations. However, little is known of their molecular and ecological characteristics, including origin, evolutionary equivalence, specific functions and activity against beneficial microbes. The aim of this study was to identify and characterize snakin-1 from alfalfa (MsSN1).ResultsPhylogenetic analysis showed complete congruence between snakin-1 and plant trees. The antimicrobial activity of MsSN1 against bacterial and fungal pathogens of alfalfa was demonstrated in vitro and in vivo. Transgenic alfalfa overexpressing MsSN1 showed increased antimicrobial activity against virulent fungal strains. However, MsSN1 did not affect nitrogen-fixing bacterial strains only when these had an alfalfa origin.ConclusionsThe results reported here suggest that snakin peptides have important and ancestral roles in land plant innate immunity. Our data indicate a coevolutionary process, in which alfalfa exerts a selection pressure for resistance to MsSN1 on rhizobial bacteria. The increased antimicrobial activity against virulent fungal strains without altering the nitrogen-fixing symbiosis observed in MsSN1-overexpressing alfalfa transgenic plants opens the way to the production of effective legume transgenic cultivars for biotic stress resistance.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-014-0248-9) contains supplementary material, which is available to authorized users.
“…RGD residues) and functional (e.g. toxic activity) data supporting its relationship with cysteine-rich peptides from snakes venoms, St SN1 and St SN2 have been recently renamed as GSL1 and GSL2, respectively [8,9]. …”
Section: Introductionmentioning
confidence: 99%
“…Overexpression of SN1 in potato and wheat ( Triticum aestivum ), SN2 in potato and tomato ( Solanum lycopersicum ), and snakin-defensin hybrid protein in tobacco ( Nicotiana tabacum ) and potato restricts pathogen invasiveness and enhances tolerance to bacterial and fungal diseases, without altering the agronomic phenotype of these crops [6,9-12]. Furthermore, disease sensitivity is enhanced by silencing SN2 in wild tobacco ( Nicotiana benthamiana ), supporting the central role of snakin peptides in plant defense [13].…”
BackgroundThe production of antimicrobial peptides is a common defense strategy of living cells against a wide range of pathogens. Plant snakin peptides inhibit bacterial and fungal growth at extremely low concentrations. However, little is known of their molecular and ecological characteristics, including origin, evolutionary equivalence, specific functions and activity against beneficial microbes. The aim of this study was to identify and characterize snakin-1 from alfalfa (MsSN1).ResultsPhylogenetic analysis showed complete congruence between snakin-1 and plant trees. The antimicrobial activity of MsSN1 against bacterial and fungal pathogens of alfalfa was demonstrated in vitro and in vivo. Transgenic alfalfa overexpressing MsSN1 showed increased antimicrobial activity against virulent fungal strains. However, MsSN1 did not affect nitrogen-fixing bacterial strains only when these had an alfalfa origin.ConclusionsThe results reported here suggest that snakin peptides have important and ancestral roles in land plant innate immunity. Our data indicate a coevolutionary process, in which alfalfa exerts a selection pressure for resistance to MsSN1 on rhizobial bacteria. The increased antimicrobial activity against virulent fungal strains without altering the nitrogen-fixing symbiosis observed in MsSN1-overexpressing alfalfa transgenic plants opens the way to the production of effective legume transgenic cultivars for biotic stress resistance.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-014-0248-9) contains supplementary material, which is available to authorized users.
“…This is supported by the failure to recover viable plants following potato transformation with antisense constructs of GSL genes [6]. In contrast, over expression of GSL genes in potato does not cause obvious changes in plant phenotype [15]. GSL peptides have a very similar spectrum of activity against microbes [8, 9].…”
Section: Introductionmentioning
confidence: 99%
“…They induce rapid aggregation of both Gram-negative and Gram-positive bacteria, and although this response does not correlate directly with antimicrobial activity, it may play an in vivo role in controlling pathogen migration [7, 9, 11]. Transgenic plants over-expressing GSL genes have been shown to have increased resistance to a range of microbial pathogens [3, 15–17]. Figure 1
The DNA and amino acid sequence of the mature GSL1 (Genbank accession FJ195646) from
Solanum tuberosum
, after cleavage of the N-terminal signal sequence.…”
BackgroundThe Gibberellin Stimulated-Like (GSL) or Snakin peptides from higher plants are cysteine-rich, with broad spectrum activity against a range of bacterial and fungal pathogens. To detect GSL peptides in applications such as western blot analysis and enzyme-linked immunosorbent assays (ELISA), specific antibodies that recognise GSL peptides are required. However, the intrinsic antimicrobial activity of these peptides is likely to prevent their expression alone in bacterial or yeast expression systems for subsequent antibody production in animal hosts.ResultsTo overcome this issue we developed an Escherichia coli expression strategy based on the expression of the GSL1 peptide as a His-tagged thioredoxin fusion protein. The DNA sequence for the mature GSL1 peptide from potato (Solanum tuberosum L.) was cloned into the pET-32a expression vector to produce a construct encoding N-terminally tagged his6-thioredoxin-GSL1. The fusion protein was overexpressed in E. coli to produce soluble non-toxic protein. The GSL1 fusion protein could be easily purified by using affinity chromatography to yield ~1.3 mg of his6-thioredoxin-GSL1 per L of culture. The fusion protein was then injected into rabbits for antibody production. Western blot analysis showed that the antibodies obtained from rabbit sera specifically recognised the GSL1 peptide that had been expressed in a wheat germ cell-free expression system.ConclusionWe present here the first report of a GSL1 peptide expressed as a fusion protein with thioredoxin that has resulted in milligram quantities of soluble protein to be produced. We have also demonstrated that a wheat germ system can be used to successfully express small quantities of GSL1 peptide useful as positive control in western blot analysis. To our knowledge this is the first report of antibodies being produced against GSL1 peptide. The antibodies will be useful for analysis of GSL1peptides in western blot, localization by immunohistochemistry (IHC) and quantitation by ELISA.
“…In recent years, plant antimicrobial peptides snakin-1 and snakin-2 have been the subject of interest as primary candidates for generating broad-spectrum biotic stress tolerance in crops (Kovalskaya et al, 2011;Balaji and Smart 2012;GuzmanRodriguez et al, 2013;Rong et al, 2013;García et al, 2014;Meiyalaghan et al, 2014;Mohan et al, 2014). Despite the importance of roles played by snakin peptides in plant-microbe interactions in nature and the potential of these peptides to improve crop production, the molecular mechanisms underlying its inhibition of microbial cells are poorly understood.…”
Snakin-1, a peptide produced by higher plants, has broad-spectrum antibiotic activity, inhibiting organisms ranging from Bacteria to Eukaryotes. However, the mode of action against target organisms is poorly understood. As a first step to elucidate the mechanism, we screened a mutation library of Pseudomonas fluorescens Pf-5 in LB and agar medium supplemented with alfalfa snakin-1 (MsSN1). We identified three biofilm formation-related Pseudomonas mutants that showed increased resistance to MsSN1. Genetic, physiological and bioinformatics analysis validated the results of the mutant screens, indicating that bacterial adhesion protein lapA is probably the target of MsSN1. Collectively, these findings suggest that snakin-1 acts on microbial adhesion properties.
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