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A Similar Secretome Disturbance as a Hallmark of Non-pathogenic Botrytis cinerea ATMT-Mutants?

ABSTRACT: The gray mold fungus Botrytis cinerea is a necrotrophic pathogen able to infect hundreds of host plants, including high-value crops such as grapevine, strawberry and tomato. In order to decipher its infectious strategy, a library of 2,144 mutants was generated by random insertional mutagenesis using Agrobacterium tumefaciens-mediated transformation (ATMT). Twelve mutants exhibiting total loss of virulence toward different host plants were chosen for detailed analyses. Their molecular characterization revealed a single T-DNA insertion in different loci. Using a proteomics approach, the secretome of four of these strains was compared to that of the parental strain and a common profile of reduced lytic enzymes was recorded. Significant variations in this profile, notably deficiencies in the secretion of proteases and hemicellulases, were observed and validated by biochemical tests. They were also a hallmark of the remaining eight non-pathogenic strains, suggesting the importance of these secreted proteins in the infection process. In the twelve non-pathogenic mutants, the differentiation of infection cushions was also impaired, suggesting a link between the penetration structures and the secretion of proteins involved in the virulence of the pathogen.

SUBMITTER: de Vallee A

PROVIDER: S-EPMC6908482 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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A Similar Secretome Disturbance as a Hallmark of Non-pathogenic Botrytis cinerea ATMT-Mutants?

de Vallée Amélie A Bally Pascal P Bruel Christophe C Chandat Lucie L Choquer Mathias M Dieryckx Cindy C Dupuy Jean William JW Kaiser Sophie S Latorse Marie-Pascale MP Loisel Elise E Mey Géraldine G Morgant Guillaume G Rascle Christine C Schumacher Julia J Simon Adeline A Souibgui Eytham E Viaud Muriel M Villalba François F Poussereau Nathalie N

Frontiers in microbiology 20191206

The gray mold fungus Botrytis cinerea is a necrotrophic pathogen able to infect hundreds of host plants, including high-value crops such as grapevine, strawberry and tomato. In order to decipher its infectious strategy, a library of 2,144 mutants was generated by random insertional mutagenesis using Agrobacterium tumefaciens-mediated transformation (ATMT). Twelve mutants exhibiting total loss of virulence toward different host plants were chosen for detailed analyses. Their molecul ...[more]

PMID: 31866989

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Project description:Botrytis cinerea is a major plant pathogen infecting more than 1400 plant species. During invasion, the fungus rapidly kills host cells, which is believed to be supported by induction of programmed plant cell death. To comprehensively evaluate the contributions of most of the currently known plant cell death inducing proteins (CDIPs) and metabolites for necrotrophic infection, an optimized CRISPR/Cas9 protocol was established which allowed to perform serial marker-free mutagenesis to generate multiple deletion mutants lacking up to 12 CDIPs. Whole genome sequencing of a 6x and 12x deletion mutant revealed a low number of off-target mutations which were unrelated to Cas9-mediated cleavage. Secretome analyses confirmed the loss of secreted proteins encoded by the deleted genes. Infection tests with the mutants revealed a successive decrease in virulence with increasing numbers of mutated genes, and varying effects of the knockouts on different host plants. Comparative analysis of mutants confirmed significant roles of two polygalacturonases (PG1, PG2) and the phytotoxic metabolites botrydial and botcinins for infection, but revealed no or only weak effects of deletion of the other CDIPs. Nicotiana benthamiana plants with mutated or silenced coreceptors of pattern recognition receptors, SOBIR1 and BAK1, showed similar susceptibility as control plants to infection by B. cinerea wild type and a 12x deletion mutant. These results raise doubts about a major role of manipulation of these plant defence regulators for B. cinerea infection. Despite the loss of most of the known phytotoxic compounds, the on planta secretomes of the multiple mutants retained substantial phytotoxic activity, proving that further, as yet unknown CDIPs contribute to necrosis and virulence. Our study has addressed for the first time systematically the functional redundancy of fungal virulence factors, and demonstrates that B. cinerea releases a highly redundant cocktail of proteins to achieve necrotrophic infection of a wide variety of host plants.

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A Similar Secretome Disturbance as a Hallmark of Non-pathogenic Botrytis cinerea ATMT-Mutants?

Publications

A Similar Secretome Disturbance as a Hallmark of Non-pathogenic <i>Botrytis cinerea</i> ATMT-Mutants?

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