ORCID Profile
0000-0002-0188-9353
Current Organisation
Australian National University
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Publisher: Cold Spring Harbor Laboratory
Date: 21-03-2020
DOI: 10.1101/2020.03.20.000117
Abstract: Eukaryotic cells deploy autophagy to eliminate invading microbes. In turn, pathogens have evolved effector proteins to counteract antimicrobial autophagy. How and why adapted pathogens co-opt autophagy for their own benefit is poorly understood. The Irish famine pathogen Phythophthora infestans secretes the effector protein PexRD54 that selectively activates an unknown plant autophagy pathway, while antagonizing antimicrobial autophagy. Here we show that PexRD54 induces autophagosome formation by bridging small GTPase Rab8a-decorated vesicles with autophagic compartments labelled by the core autophagy protein ATG8CL. Rab8a is required for pathogen-triggered and starvation-induced but not antimicrobial autophagy, revealing that specific trafficking pathways underpin selective autophagy. We discovered that Rab8a contributes to basal immunity against P. infestans, but PexRD54 erts a sub-population of Rab8a vesicles to lipid droplets that associate with autophagosomes. These are then erted towards pathogen feeding structures that are accommodated within the host cells. We propose that PexRD54 mimics starvation-induced autophagy by channeling host endomembrane trafficking towards the pathogen interface possibly to acquire nutrients. This work reveals that effectors can interconnect independent host compartments to stimulate complex cellular processes that benefit the pathogen.
Publisher: Springer Science and Business Media LLC
Date: 25-04-2017
DOI: 10.1038/S41598-017-01100-Z
Abstract: Pathogens secrete effector proteins to suppress host immunity, mediate nutrient uptake and subsequently enable parasitism. However, on non-adapted hosts, effectors can be detected as non-self by host immune receptors and activate non-host immunity. Nevertheless, the molecular mechanisms of effector triggered non-host resistance remain unknown. Here, we report that a small cysteine-rich protein PstSCR1 from the wheat rust pathogen Puccinia striiformis f. sp. tritici ( Pst ) activates immunity in the non-host solanaceous model plant Nicotiana benthamiana . PstSCR1 homologs were found to be conserved in Pst , and in its closest relatives, Puccinia graminis f. sp. tritici and Puccinia triticina . When PstSCR1 was expressed in N . benthamiana with its signal peptide, it provoked the plant immune system, whereas no stimulation was observed when it was expressed without its signal peptide. PstSCR1 expression in N . benthamiana significantly reduced infection capacity of the oomycete pathogens. Moreover, apoplast-targeted PstSCR1 triggered plant cell death in a dose dependent manner. However, in Brassinosteroid insensitive 1-Associated Kinase 1 ( SERK3 / BAK1 ) silenced N . benthamiana , cell death was remarkably decreased. Finally, purified PstSCR1 protein activated defence related gene expression in N . benthamiana . Our results show that a Pst -secreted protein, PstSCR1 can activate surface mediated immunity in non-adapted hosts and contribute to non-host resistance.
Publisher: Portland Press Ltd.
Date: 12-2020
DOI: 10.1042/BSR20201188
Abstract: The importance of wheat yellow rust disease, caused by Puccinia striiformis f. sp. tritici (Pst), has increased substantially due to the emergence of aggressive new Pst races in the last couple of decades. In an era of escalating human populations and climate change, it is vital to understand the infection mechanism of Pst in order to develop better strategies to combat wheat yellow disease. The present study focuses on the identification of small secreted proteins (SSPs) and candidate-secreted effector proteins (CSEPs) that are used by the pathogen to support infection and control disease development. We generated de novo assembled transcriptomes of Pst collected from wheat fields in central Anatolia. We inoculated both susceptible and resistant seedlings with Pst and analyzed haustoria formation. At 10 days post-inoculation (dpi), we analyzed the transcriptomes and identified 10550 Differentially Expressed Unigenes (DEGs), of which 6072 were Pst-mapped. Among those Pst-related genes, 227 were predicted as PstSSPs. In silico characterization was performed using an approach combining the transcriptomic data and data mining results to provide a reliable list to narrow down the ever-expanding repertoire of predicted effectorome. The comprehensive analysis detected 14 Differentially Expressed Small-Secreted Proteins (DESSPs) that overlapped with the genes in available literature data to serve as the best CSEPs for experimental validation. One of the CSEPs was cloned and studied to test the reliability of the presented data. Biological assays show that the randomly selected CSEP, Unigene17495 (PSTG_10917), localizes in the chloroplast and is able to suppress cell death induced by INF1 in a Nicotiana benthamiana heterologous expression system.
Publisher: Cold Spring Harbor Laboratory
Date: 27-05-2020
DOI: 10.1101/2020.05.27.120113
Abstract: Plant pathogens cause disease through secreted effector proteins, which act to modulate host physiology and promote infection. Typically, the sequences of effectors provide little functional information and further targeted experimentation is required. Here, we utilised a structure/function approach to study SnTox3, an effector from the necrotrophic fungal pathogen Parastagonospora nodorum , which causes cell death in wheat-lines carrying the sensitivity gene Snn3 . We developed a workflow for the production of SnTox3 in a heterologous host that enabled crystal structure determination. We show this approach can be successfully applied to effectors from other pathogenic fungi. Complementing this, an in-silico study uncovered the prevalence of an expanded subclass of effectors from fungi. The β-barrel fold of SnTox3 is a novel fold among fungal effectors. We demonstrate that SnTox3 is a pre-pro-protein and that the protease Kex2 removes the pro-domain. Our in-silico studies suggest that Kex2-processed pro-domain (designated here as K2PP) effectors are common in fungi, and we demonstrate this experimentally for effectors from Fusarium oxysporum f sp. lycopersici . We propose that K2PP effectors are highly prevalent among fungal effectors. The identification and classification of K2PP effectors has broad implications for the approaches used to study their function in fungal virulence.
Publisher: Cold Spring Harbor Laboratory
Date: 13-01-2020
DOI: 10.1101/2020.01.11.897884
Abstract: The importance of wheat yellow rust disease, caused by Puccinia striiformis f. sp. tritici ( Pst ), has increased substantially due to the emergence of aggressive new Pst races in the last couple of decades. In an era of escalating human populations and climate change, it is vital to understand the infection mechanism of Pst in order to develop better strategies to combat wheat yellow disease. This study focuses on the identification of small secreted proteins (SSPs) and candidate-secreted effector proteins (CSEPs) that are used by the pathogen to support infection and control disease development. We generated de novo assembled transcriptomes of Pst collected from wheat fields in central Anatolia. We inoculated both susceptible and resistant seedlings with Pst and analyzed haustoria formation. At 10 days post-inoculation (dpi), we analyzed the transcriptomes and identified 10,550 Differentially Expressed Unigenes (DEGs), of which 6,220 were Pst -related. Among those Pst -related genes, 230 were predicted as PstSSPs. In silico characterization was performed using an approach combining the transcriptomic data and data mining results to provide a reliable list to narrow down the ever-expanding repertoire of predicted effectorome. The comprehensive analysis detected 14 Differentially Expressed Small-Secreted Proteins (DESSPs) that overlapped with the genes in available literature data to serve as the best CSEPs for experimental validation. One of the CSEPs was cloned and studied to test the reliability of the presented data. Biological assays show that the randomly selected CSEP, Unigene17495 (PSTG_10917), localizes in the chloroplast and is able to suppress cell death induced by INF1 in a Nicotiana benthamiana heterologous expression system.
Publisher: Wiley
Date: 20-02-2022
DOI: 10.1111/TPJ.15677
Abstract: The plant pathogen Parastagonospora nodorum secretes necrotrophic effectors to promote disease. These effectors induce cell death on wheat cultivars carrying dominant susceptibility genes in an inverse gene‐for‐gene manner. However, the molecular mechanisms underpinning these interactions and resulting cell death remain unclear. Here, we used a yeast two‐hybrid library approach to identify wheat proteins that interact with the necrotrophic effector ToxA. Using this strategy, we identified an interaction between ToxA and a wheat transmembrane NDR/HIN1‐like protein (TaNHL10) and confirmed the interaction using in planta co‐immunoprecipitation and confocal microscopy co‐localization analysis. We showed that the C‐terminus of TaNHL10 is extracellular whilst the N‐terminus is localized in the cytoplasm. Further analyses using yeast two‐hybrid and confocal microscopy co‐localization showed that ToxA interacts with the C‐terminal LEA2 extracellular domain of TaNHL10. Random mutagenesis was then used to identify a ToxA mutant, ToxA N109D , which was unable to interact with TaNHL10 in yeast two‐hybrid assays. Subsequent heterologous expression and purification of ToxA N109D in Nicotiania benthamiana revealed that the mutated protein was unable to induce necrosis on Tsn1 ‐dominant wheat cultivars, confirming that the interaction of ToxA with TaNHL10 is required to induce cell death. Collectively, these data advance our understanding on how ToxA induces cell death during infection and further highlight the importance of host cell surface interactions in necrotrophic pathosystems.
Publisher: eLife Sciences Publications, Ltd
Date: 23-08-2021
DOI: 10.7554/ELIFE.65285
Abstract: Eukaryotic cells deploy autophagy to eliminate invading microbes. In turn, pathogens have evolved effector proteins to counteract antimicrobial autophagy. How adapted pathogens co-opt autophagy for their own benefit is poorly understood. The Irish famine pathogen Phytophthora infestans secretes the effector protein PexRD54 that selectively activates an unknown plant autophagy pathway that antagonizes antimicrobial autophagy at the pathogen interface. Here, we show that PexRD54 induces autophagosome formation by bridging vesicles decorated by the small GTPase Rab8a with autophagic compartments labeled by the core autophagy protein ATG8CL. Rab8a is required for pathogen-triggered and starvation-induced but not antimicrobial autophagy, revealing specific trafficking pathways underpin selective autophagy. By subverting Rab8a-mediated vesicle trafficking, PexRD54 utilizes lipid droplets to facilitate biogenesis of autophagosomes erted to pathogen feeding sites. Altogether, we show that PexRD54 mimics starvation-induced autophagy to subvert endomembrane trafficking at the host-pathogen interface, revealing how effectors bridge distinct host compartments to expedite colonization.
Publisher: Wiley
Date: 08-2021
DOI: 10.1002/PLD3.341
Abstract: Plant fungal pathogens cause devastating diseases on cereal plants and threaten global food security. During infection, these pathogens secrete proteinaceous effectors that promote disease. Some of these effectors from necrotrophic plant pathogens induce a cell death response (necrosis), which facilitates pathogen growth in planta . Characterization of these effectors typically requires heterologous expression, and microbial expression systems such as bacteria and yeast are the predominantly used. However, microbial expression systems often require optimization for any given effector and are, in general, not suitable for effectors involving cysteine bridges and posttranslational modifications for activity. Here, we describe a simple and efficient method for expressing such effectors in the model plant Nicotiana benthamiana . Briefly, an effector protein is transiently expressed and secreted into the apoplast of N. benthamiana by Agrobacterium‐mediated infiltration. Two to three days subsequent to agroinfiltration, the apoplast from the infiltrated leaves is extracted and can be directly used for phenotyping on host plants. The efficacy of this approach was demonstrated by expressing the ToxA, Tox3, and Tox1 necrosis‐inducing effectors from Parastagonospora nodorum . All three effectors produced in N. benthamiana were capable of inducing necrosis in wheat lines, and two of three showed visible bands on Coomassie‐stained gel. These data suggest that N. benthamiana –agroinfiltration system is a feasible tool to obtain fungal effectors, especially those that require disulfide bonds and posttranslational modifications. Furthermore, due to the low number of proteins typically observed in the apoplast (compared with intracellular), this simple and high‐throughput approach circumvents the requirement to lyse cells and further purifies the target proteins that are required in other heterologous systems. Because of its simplicity and potential for high‐throughput, this method is highly amenable to the phenotyping of candidate protein effectors on host plants.
Publisher: Wiley
Date: 03-01-2021
DOI: 10.1111/NPH.17128
Publisher: Wiley
Date: 24-06-2021
DOI: 10.1111/NPH.17516
Abstract: Plant pathogens cause disease through secreted effector proteins, which act to promote infection. Typically, the sequences of effectors provide little functional information and further targeted experimentation is required. Here, we utilized a structure/function approach to study SnTox3, an effector from the necrotrophic fungal pathogen Parastagonospora nodorum , which causes cell death in wheat‐lines carrying the sensitivity gene Snn3 . We developed a workflow for the production of SnTox3 in a heterologous host that enabled crystal structure determination and functional studies. We show this approach can be successfully applied to study effectors from other pathogenic fungi. The β‐barrel fold of SnTox3 is a novel fold among fungal effectors. Structure‐guided mutagenesis enabled the identification of residues required for Snn3 recognition. SnTox3 is a pre‐pro‐protein, and the pro‐domain of SnTox3 can be cleaved in vitro by the protease Kex2. Complementing this, an in silico study uncovered the prevalence of a conserved motif (LxxR) in an expanded set of putative pro‐domain‐containing fungal effectors, some of which can be cleaved by Kex2 in vitro . Our in vitro and in silico study suggests that Kex2‐processed pro‐domain (designated here as K2PP) effectors are common in fungi and this may have broad implications for the approaches used to study their functions.
Publisher: eLife Sciences Publications, Ltd
Date: 15-06-2021
Publisher: Cold Spring Harbor Laboratory
Date: 23-09-2021
DOI: 10.1101/2021.09.22.461440
Abstract: The plant pathogen Parastagonospora nodorum secretes necrotrophic effectors to promote disease. These effectors induce cell death on wheat cultivars carrying dominant susceptibility genes in an inverse gene-for-gene manner. However, the molecular mechanisms underpinning these interactions and resulting cell death remain unclear. Here, we used a yeast-two-hybrid library approach to identify wheat proteins that interact with the necrotrophic effector ToxA. Using this strategy, we identified an interaction between ToxA and a wheat transmembrane NDR/HIN1-like protein (TaNHL10) and confirmed the interaction using in-planta co-immunoprecipitation and confocal microscopy co-localization analysis. We showed that the C-terminus of TaNHL10 is extracellular whilst the N-terminus was localized in the cytoplasm. Further analyses using yeast-two-hybrid and confocal microscopy co-localization showed that ToxA interacts with the C-terminal LEA2 extracellular domain of TaNHL10. Random mutagenesis was then used to identify a ToxA mutant, ToxA N109D , which was unable to interact with TaNHL10 in yeast-two-hybrid assays. Subsequent heterologous expression and purification of ToxA N109D in Nicotiania benthamiana revealed that the mutated protein was unable to induce necrosis on Tsn1 -dominant wheat cultivars confirming that the interaction of ToxA with TaNHL10 is required to induce cell death. Collectively, these data advance our understanding on how ToxA induces cell death during infection and further highlights the importance of host cell surface interactions in necrotrophic pathosystems.
Publisher: Cold Spring Harbor Laboratory
Date: 03-05-2021
DOI: 10.1101/2021.05.02.442377
Abstract: Plant fungal pathogens cause devastating diseases on cereal plants and threaten global food security. During infection, these pathogens secrete proteinaceous effectors that promote disease. Some of these effectors from necrotrophic plant pathogens induce a cell death response (necrosis), which facilitates pathogen growth in planta . Characterisation of these effectors typically requires heterologous expression and microbial expression systems such as bacteria and yeast are the predominantly used. However, microbial expression systems often require optimization for any given effector and are, in general, not suitable for effectors involving cysteine bridges and posttranslational modifications for activity. Here, we describe a simple and efficient method for expressing such effectors in the model plant Nicotiana benthamiana . Briefly, an effector protein is transiently expressed and secreted into the apoplast of N. benthamiana by Agrobacterium-mediated infiltration. Two-to-three days subsequent to agroinfiltration, the apoplast from the infiltrated leaves is extracted and can be directly used for phenotyping on host plants. The efficacy of this approach was demonstrated by expressing the ToxA, Tox3 and Tox1 necrosis-inducing effectors from Parastagonospora nodorum . All three effectors produced in N. benthamiana were capable of inducing necrosis in wheat lines, and two of three showed visible bands on Coomassie-stained gel. These data suggest that N. benthamiana- agroinfiltration system is a feasible tool to obtain fungal effectors, especially those that require disulfide bonds and posttranslational modifications. Furthermore, due to the low number of proteins typically observed in the apoplast (compared to intracellular), this simple and high-throughput approach circumvents the requirement to lyse cells and further purify the target proteins that is required in other heterologous systems. Because of its simplicity and potential for high-throughput, this method is highly amenable to the phenotyping of candidate protein effectors on host plants.
No related grants have been discovered for Bayantes Dagvadorj.