Heterologous Expression of the Effector Proteins ToxA and ToxB, and the Prevalence of Effector Sensitivity in Australian Cereal Crops.

Here, we evaluate the expression of the proteinaceous effectors ToxA and ToxB, produced by the necrotrophic fungal pathogen , which confer tan spot disease susceptibility on wheat. These necrotrophic effectors were expressed in two heterologous systems: and . The SHuffle system was demonstrated to be superior to in generating high-levels of recombinant proteins that were soluble and stable. In addition, protein extracts from 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 . 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 effectors in host and non-host interactions.