Heterologous Expression of the Pyrenophora tritici-repentis Effector Proteins ToxA and ToxB, and the Prevalence of Effector Sensitivity in Australian Cereal Crops
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Here, we evaluate the expression of the proteinaceous effectors ToxA and ToxB, produced by the necrotrophic fungal pathogen Pyrenophora tritici-repentis, which confer tan spot disease susceptibility on wheat. These necrotrophic effectors were expressed in two heterologous systems: Escherichia coli and Pichia pastoris. The E. coli SHuﬄe system was demonstrated to be superior to P. pastoris in generating high-levels of recombinant proteins that were soluble and stable. In addition, protein extracts from P. pastoris induced non-specific chlorosis on wheat, postulated to be caused by co-purified glucanases secreted by the host. Up to 79.6 μg/ml of ToxB was obtained using the SHuﬄe system in the absence of the native signal peptide, whilst the ToxA yield was considerably lower at 3.2 μg/ml. Results indicated that a histidine tag at the ToxA C-terminus interfered with effector functionality. Heterologously expressed ToxA and ToxB were tested on a panel of Australian cereals, including 122 varieties of bread wheat, 16 durum, 20 triticale and 5 barley varieties, as well as common plant model species including tobacco and Arabidopsis thaliana. A varying degree of effector sensitivities was observed, with a higher ToxB sensitivity and prevalence in the durum and triticale varieties. ToxB-induced chlorosis was also detected on barley. The heterologous expression of effectors that are easily scalable, will facilitate effector-assisted selection of varieties in wheat breeding programs as well as the investigation of P. tritici-repentis effectors in host and non-host interactions.
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PacBio genome sequencing reveals new insights into the genomic organisation of the multi-copy ToxB gene of the wheat fungal pathogen Pyrenophora tritici-repentisMoolhuijzen, Paula ; See, Pao Theen ; Moffat, Caroline (2020)Background: Necrotrophic effector proteins secreted by fungal pathogens are important virulence factors that mediate the development of disease in wheat. Pyrenophora tritici-repentis (Ptr), the causal agent of wheat tan ...
A conserved hypothetical gene is required but not sufficient for Ptr ToxC production in Pyrenophora tritici-repentis.Shi, Gongjun State; Kariyawasam, Gayan; Liu, Sanzhen; Leng, Yueqiang; Zhong, Shaobin; Ali, Shaukat; Moolhuijzen, Paula ; Moffat, Caroline ; Rasmussen, Jack B; Friesen, Timothy L; Faris, Justin D; Liu, Zhaohui (2022)The fungus Pyrenophora tritici-repentis (Ptr) causes tan spot, an important foliar disease of wheat worldwide. The fungal pathogen produce three necrotrophic effectors, namely Ptr ToxA, Ptr ToxB and Ptr ToxC to induce ...
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