Effector discovery and characterisation in the Fusarium graminearum-wheat floral interaction

Walker, Catherine Holly (2020) Effector discovery and characterisation in the Fusarium graminearum-wheat floral interaction. PhD thesis, University of Nottingham.

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Fusarium graminearum is the causal agent of Fusarium Head Blight (FHB) – a highly hazardous wheat disease which results in crop losses and mycotoxin contamination. A recent transcriptomic investigation of the F. graminearum-wheat interaction revealed an up-regulation in hundreds of genes encoding small secreted proteins (SSPs), or putative effectors, during infection which we hypothesise contribute towards pathogenicity. The aim of this study was therefore to bioinformatically identify and then functionally characterise effector candidates in wheat and Nicotiana benthamiana using a range of post-genomics techniques.

In this study, an effector discovery pipeline was established involving interrogation of the in vitro and in planta transcriptional profiles of genes belonging to the F. graminearum predicted secretome. This pipeline yielded twenty-four candidates for functional characterisation, seven of which were characterised using the Barley Stripe Mosaic Virus-mediated overexpression (BSMV-VOX) system. Transient expression of two FgSSPs, FgSSP32 and FgSSP33, in mature wheat ears led to a significant reduction in FHB disease symptoms. Single gene deletions of FgSSP32 and FgSSP33 were then generated using a split-marker transformation approach but found not to result in any changes in fungal pathogenicity on wheat.

Agrobacterium-mediated transient expression of FgSSP32 and FgSSP33 in the model species N. benthamiana led to the discovery that both proteins induce necrosis of the vascular tissue. Expression of proteins lacking a signal peptide did not result in necrosis suggesting that these proteins function apoplastically. Biochemical characterisation of N. benthamiana leaves expressing FgSSP32 and FgSSP33 revealed that both proteins induce the production of a compound tentatively identified as 12-oxo-phytodienoic acid (12-OPDA) – a precursor to the phytohormone jasmonic acid (JA). In addition, apoplastic occlusions were observed in N. benthamiana leaves expressing FgSSP33 indicating a host defence response towards this protein.

Transcriptome data exploring FgSSP32 and FgSSP33 expression during the F. graminearum-wheat floral interaction, revealed that both genes are expressed during the symptomatic phase of infection – a phase characterised by penetration and colonisation of wheat cells by fungal hyphae. We therefore hypothesise that FgSSP32 and FgSSP33 contribute towards the symptomatic phase of F. graminearum infection by activating cell death responses leading to the release of nutrients for fungal sequestration. Further work is required to identify host interacting proteins which may lead to the identification of wheat genes involved in F. graminearum resistance or susceptibility.

Item Type: Thesis (University of Nottingham only) (PhD)
Supervisors: Hammond-Kosack, Kim
Kanyuka, Kostya
Keywords: Fusarium graminearum, Wheat disease, Plant pathology
Subjects: Q Science > QK Botany
S Agriculture > SB Plant culture
Faculties/Schools: UK Campuses > Faculty of Science > School of Biosciences
Item ID: 59960
Depositing User: Walker, Catherine
Date Deposited: 31 Jul 2020 04:40
Last Modified: 24 Jul 2022 04:30
URI: https://eprints.nottingham.ac.uk/id/eprint/59960

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