An Arabidopsis downy mildew non-RxLR effector suppresses induced plant cell death to promote biotroph infection

Abstract Our understanding of obligate biotrophic pathogens is limited by lack of knowledge concerning the molecular function of virulence factors. We established Arabidopsis host-induced gene silencing (HIGS) to explore gene functions of Hyaloperonospora arabidopsidis, including CYSTEINE-RICH PROTEIN (HaCR)1, a potential secreted effector gene of this obligate biotrophic pathogen. HaCR1 HIGS resulted in H. arabidopsidis-induced local plant cell death and reduced pathogen reproduction. We functionally characterized HaCR1 by ectopic expression in Nicotiana benthamiana. HaCR1 was capable of inhibiting effector-triggered plant cell death. Consistent with this, HaCR1 expression in N. benthamiana led to stronger disease symptoms caused by the hemibiotrophic oomycete pathogen Phytophthora capsici, but reduced disease symptoms caused by the necrotrophic fungal pathogen Botrytis cinerea. Expressing HaCR1 in transgenic Arabidopsis confirmed higher susceptibility to H. arabidopsidis and to the bacterial hemibiotrophic pathogen Pseudomonas syringae. Increased H. arabidopsidis infection was in accordance with reduced PATHOGENESIS RELATED (PR)1 induction. Expression of full-length HaCR1 was required for its function, which was lost if the signal peptide was deleted, suggesting its site of action in the plant apoplast. This study provides phytopathological and molecular evidence for the importance of this widespread, but largely unexplored class of non-RxLR effectors in biotrophic oomycetes.

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A Phytophthora capsici RXLR effector manipulates plant immunity by targeting RAB proteins and disturbing vesicle-mediated protein trafficking pathway

10.22541/au.163697880.04261552/v1 ◽

2021 ◽

Author(s):

Tianli Li ◽

Gan Ai ◽

Xiaowei Fu ◽

Jin Liu ◽

Hai Zhu ◽

...

Keyword(s):

Membrane Trafficking ◽

Plant Immunity ◽

Phytophthora Capsici ◽

Functional Characterization ◽

Ectopic Expression ◽

Defense Responses ◽

Infection Process ◽

Small Gtpase ◽

Rab Proteins ◽

Rxlr Effector

The oomycete pathogen Phytophthora capsici encodes hundreds of RXLR effectors to enter plant cells and suppress host defense responses. Only few of them are conserved across different strains and species. Such ‘core effectors’ may target hub immunity pathways that are essential during Phytophthora pathogens interacting with their hosts. However, the underlying mechanisms of core RXLRs-mediated host immunity manipulation are largely unknown. Here, we report the functional characterization of a P. capsici RXLR effector, RXLR242. RXLR242 expression is highly induced during the infection process. Its ectopic expression in Nicotiana benthamiana promotes Phytophthora infection. RXLR242 physically interacts with a group of RAB proteins, which belong to the small GTPase family and function in specifying transport pathways in the intracellular membrane trafficking system. RXLR242 impedes the secretion of PATHOGENESIS-RELATED 1 (PR1) protein to the apoplast by interfering the formation of RABE1-7-labeled vesicles. Further analysis indicated that such phenomenon is resulted from competitive binding of RXLR242 to RABE1-7. RXLR242 also interferes trafficking of the membrane-located receptor FLAGELLIN-SENSING 2 (FLS2) through competitively interacting with RABA4-3. Taken together, our work demonstrates that RXLR242 manipulates plant immunity by targeting RAB proteins and disturbing vesicle-mediated protein transporting pathway in plant hosts.

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Pectobacterium carotovorum Elicits Plant Cell Death with DspE/F but the P. carotovorum DspE Does Not Suppress Callose or Induce Expression of Plant Genes Early in Plant–Microbe Interactions

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-06-10-0143 ◽

2011 ◽

Vol 24 (7) ◽

pp. 773-786 ◽

Cited By ~ 29

Author(s):

Hye-Sook Kim ◽

Phanit Thammarat ◽

Steven A. Lommel ◽

Clifford S. Hogan ◽

Amy O. Charkowski

Keyword(s):

Gene Expression ◽

Cell Death ◽

Pseudomonas Syringae ◽

Plant Cell ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Secretion System ◽

Broad Host Range ◽

Pectobacterium Carotovorum ◽

Callose Deposition

The broad-host-range bacterial soft rot pathogen Pectobacterium carotovorum causes a DspE/F-dependent plant cell death on Nicotiana benthamiana within 24 h postinoculation (hpi) followed by leaf maceration within 48 hpi. P. carotovorum strains with mutations in type III secretion system (T3SS) regulatory and structural genes, including the dspE/F operon, did not cause hypersensitive response (HR)-like cell death and or leaf maceration. A strain with a mutation in the type II secretion system caused HR-like plant cell death but no maceration. P. carotovorum was unable to impede callose deposition in N. benthamiana leaves, suggesting that P. carotovorum does not suppress this basal immunity function. Within 24 hpi, there was callose deposition along leaf veins and examination showed that the pathogen cells were localized along the veins. To further examine HR-like plant cell death induced by P. carotovorum, gene expression profiles in N. benthamiana leaves inoculated with wild-type and mutant P. carotovorum and Pseudomonas syringae strains were compared. The N. benthamiana gene expression profile of leaves infiltrated with Pectobacterium carotovorum was similar to leaves infiltrated with a Pseudomonas syringae T3SS mutant. These data support a model where Pectobacterium carotovorum uses the T3SS to induce plant cell death in order to promote leaf maceration rather than to suppress plant immunity.

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Bacterial Growth Restriction During Host Resistance to Pseudomonas syringae Is Associated with Leaf Water Loss and Localized Cessation of Vascular Activity in Arabidopsis thaliana

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-22-7-0857 ◽

2009 ◽

Vol 22 (7) ◽

pp. 857-867 ◽

Cited By ~ 43

Author(s):

Brian C. Freeman ◽

Gwyn A. Beattie

Keyword(s):

Arabidopsis Thaliana ◽

Cell Death ◽

Pseudomonas Syringae ◽

Plant Cell ◽

Bacterial Growth ◽

Water Loss ◽

Leaf Water ◽

Infection Site ◽

Vascular Activity ◽

Leaf Water Loss

The physiological mechanisms by which plants limit the growth of bacterial pathogens during gene-for-gene resistance are poorly understood. We characterized early events in the Arabidopsis thaliana–Pseudomonas syringae pathosystem to identify physiological changes for which the kinetics are consistent with bacterial growth restriction. Using a safranine-O dye solution to detect vascular activity, we demonstrated that A. thaliana Col-0 resistance to P. syringae pv. tomato DC3000 cells expressing avrRpm1 involved virtually complete cessation of vascular water movement into the infection site within only 3 h postinoculation (hpi), under the conditions tested. This vascular restriction preceded or was simultaneous with precipitous decreases in photosynthesis, stomatal conductance, and leaf transpiration, with the latter two remaining at detectable levels. Microscopic plant cell death was detected as early as 2 hpi. Interestingly, suppression of bacterial growth during AvrRpm1-mediated resistance was eliminated by physically blocking leaf water loss through the stomata without altering plant cell death and was nearly eliminated by incubating plants at high relative humidity. The majority of the population growth benefit from blocking leaf water loss occurred early after inoculation, i.e., between 4 and 8 hpi. Collectively, these results support a model in which A. thaliana suppresses P. syringae growth during gene-for-gene resistance, at least in part, by coupling restricted vascular flow to the infection site with water loss through partially open stomata; that is, the plants effectively starve the invading bacteria for water.

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Use of a Secretion Trap Screen in Pepper Following Phytophthora capsici Infection Reveals Novel Functions of Secreted Plant Proteins in Modulating Cell Death

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-08-10-0183 ◽

2011 ◽

Vol 24 (6) ◽

pp. 671-684 ◽

Cited By ~ 23

Author(s):

Seon-In Yeom ◽

Hyang-Ku Baek ◽

Sang-Keun Oh ◽

Won-Hee Kang ◽

Sang Jik Lee ◽

...

Keyword(s):

Cell Death ◽

Capsicum Annuum ◽

Pseudomonas Syringae ◽

Signal Sequence ◽

Phytophthora Capsici ◽

Tobacco Rattle Virus ◽

Secreted Proteins ◽

In Planta ◽

Cell Wall Modification ◽

Cas Genes

In plants, the primary defense against pathogens is mostly inducible and associated with cell wall modification and defense-related gene expression, including many secreted proteins. To study the role of secreted proteins, a yeast-based signal-sequence trap screening was conducted with the RNA from Phytophthora capsici-inoculated root of Capsicum annuum ‘Criollo de Morelos 334’ (CM334). In total, 101 Capsicum annuum secretome (CaS) clones were isolated and identified, of which 92 were predicted to have a secretory signal sequence at their N-terminus. To identify differences in expressed CaS genes between resistant and susceptible cultivars of pepper, reverse Northern blots and real-time reverse-transcription polymerase chain reaction were performed with RNA samples isolated at different time points following P. capsici inoculation. In an attempt to assign biological functions to CaS genes, we performed in planta knock-down assays using the Tobacco rattle virus-based gene-silencing method. Silencing of eight CaS genes in pepper resulted in suppression of the cell death induced by the non-host bacterial pathogen (Pseudomonas syringae pv. tomato T1). Three CaS genes induced phenotypic abnormalities in silenced plants and one, CaS259 (PR4-l), caused both cell death suppression and perturbed phenotypes. These results provide evidence that the CaS genes may play important roles in pathogen defense as well as developmental processes.

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The RXLR Effector PcAvh1 Is Required for Full Virulence of Phytophthora capsici

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-09-18-0251-r ◽

2019 ◽

Vol 32 (8) ◽

pp. 986-1000 ◽

Cited By ~ 9

Author(s):

Xiao-Ren Chen ◽

Ye Zhang ◽

Hai-Yang Li ◽

Zi-Hui Zhang ◽

Gui-Lin Sheng ◽

...

Keyword(s):

Plant Pathogens ◽

Plant Immunity ◽

Phytophthora Capsici ◽

Ectopic Expression ◽

Plant Cells ◽

Effector Proteins ◽

Bell Pepper ◽

Broad Host Range ◽

Virus Induced Gene Silencing ◽

Rxlr Effector

Plant pathogens employ diverse secreted effector proteins to manipulate host physiology and defense in order to foster diseases. The destructive Phytophthora pathogens encode hundreds of cytoplasmic effectors, which are believed to function inside the plant cells. Many of these cytoplasmic effectors contain the conserved N-terminal RXLR motif. Understanding the virulence function of RXLR effectors will provide important knowledge of Phytophthora pathogenesis. Here, we report the characterization of RXLR effector PcAvh1 from the broad–host range pathogen Phytophthora capsici. Only expressed during infection, PcAvh1 is quickly induced at the early infection stages. CRISPR/Cas9-knockout of PcAvh1 in P. capsici severely impairs virulence while overexpression enhances disease development in Nicotiana benthamiana and bell pepper, demonstrating that PcAvh1 is an essential virulence factor. Ectopic expression of PcAvh1 induces cell death in N. benthamiana, tomato, and bell pepper. Using yeast two-hybrid screening, we found that PcAvh1 interacts with the scaffolding subunit of the protein phosphatase 2A (PP2Aa) in plant cells. Virus-induced gene silencing of PP2Aa in N. benthamiana attenuates resistance to P. capsici and results in dwarfism, suggesting that PP2Aa regulates plant immunity and growth. Collectively, these results suggest that PcAvh1 contributes to P. capsici infection, probably through its interaction with host PP2Aa.

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An RXLR effector PlAvh142 from Peronophythora litchii triggers plant cell death and contributes to virulence

Molecular Plant Pathology ◽

10.1111/mpp.12905 ◽

2020 ◽

Vol 21 (3) ◽

pp. 415-428 ◽

Cited By ~ 1

Author(s):

Junjian Situ ◽

Liqun Jiang ◽

Xiaoning Fan ◽

Wensheng Yang ◽

Wen Li ◽

...

Keyword(s):

Cell Death ◽

Plant Cell ◽

Rxlr Effector

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The RxLR effector Avh241 from Phytophthora sojae requires plasma membrane localization to induce plant cell death

New Phytologist ◽

10.1111/j.1469-8137.2012.04241.x ◽

2012 ◽

Vol 196 (1) ◽

pp. 247-260 ◽

Cited By ~ 68

Author(s):

Xiaoli Yu ◽

Junli Tang ◽

Qunqing Wang ◽

Wenwu Ye ◽

Kai Tao ◽

...

Keyword(s):

Cell Death ◽

Plasma Membrane ◽

Plant Cell ◽

Phytophthora Sojae ◽

Membrane Localization ◽

Plasma Membrane Localization ◽

Rxlr Effector

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The CaAP2/ERF064 Regulates Dual Functions in Pepper: Plant Cell Death and Resistance to Phytophthora capsici

Genes ◽

10.3390/genes10070541 ◽

2019 ◽

Vol 10 (7) ◽

pp. 541 ◽

Cited By ~ 5

Author(s):

Jing-Hao Jin ◽

Huai-Xia Zhang ◽

Muhammad Ali ◽

Ai-Min Wei ◽

De-Xu Luo ◽

...

Keyword(s):

Cell Death ◽

Phytophthora Capsici ◽

Ectopic Expression ◽

Transcript Level ◽

Pepper Plant ◽

Phytophthora Blight ◽

Pr Genes ◽

Biotic Stresses ◽

Pathogenesis Related ◽

Phytophthora Resistance

Phytophthora blight is one of the most destructive diseases of pepper (Capsicum annuum L.) globally. The APETALA2/Ethylene Responsive Factors (AP2/ERF) genes play a crucial role in plant response to biotic stresses but, to date, have not been studied in the context of Phytophthora resistance in pepper. Here, we documented potential roles for the pepper CaAP2/ERF064 gene in inducing cell death and conferring resistance to Phytophthora capsici (P. capsici) infection. Results revealed that the N-terminal, AP2 domain, and C-terminal of CaAP2/ERF064 protein is responsible for triggering cell death in Nicotiana benthamiana (N. benthamiana). Moreover, the transcription of CaAP2/ERF064 in plant is synergistically regulated by the Methyl-Jasmonate (MeJA) and ethephon (ET) signaling pathway. CaAP2/ERF064 was found to regulate the expression of CaBPR1, which is a pathogenesis-related (PR) gene of pepper. Furthermore, the silencing of CaAP2/ERF064 compromised the pepper plant resistance to P. capsici by reducing the transcript level of defense-related genes CaBPR1, CaPO2, and CaSAR82, while the ectopic expression of CaAP2/ERF064 in N. benthamiana plant elevated the expression level of NbPR1b and enhanced resistance to P. capsici. These results suggest that CaAP2/ERF064 could positively regulate the defense response against P. capsici by modulating the transcription of PR genes in the plant.

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Uncoupling Salicylic Acid-Dependent Cell Death and Defense-Related Responses From Disease Resistance in the Arabidopsis Mutant acd5

Genetics ◽

10.1093/genetics/156.1.341 ◽

2000 ◽

Vol 156 (1) ◽

pp. 341-350

Author(s):

Jean T Greenberg ◽

F Paul Silverman ◽

Hua Liang

Keyword(s):

Cell Death ◽

Salicylic Acid ◽

Disease Resistance ◽

Pseudomonas Syringae ◽

Higher Plants ◽

Ethylene Signaling ◽

Symptom Development ◽

Wild Type ◽

Dependent Cell ◽

Arabidopsis Mutant

Abstract Salicylic acid (SA) is required for resistance to many diseases in higher plants. SA-dependent cell death and defense-related responses have been correlated with disease resistance. The accelerated cell death 5 mutant of Arabidopsis provides additional genetic evidence that SA regulates cell death and defense-related responses. However, in acd5, these events are uncoupled from disease resistance. acd5 plants are more susceptible to Pseudomonas syringae early in development and show spontaneous SA accumulation, cell death, and defense-related markers later in development. In acd5 plants, cell death and defense-related responses are SA dependent but they do not confer disease resistance. Double mutants with acd5 and nonexpressor of PR1, in which SA signaling is partially blocked, show greatly attenuated cell death, indicating a role for NPR1 in controlling cell death. The hormone ethylene potentiates the effects of SA and is important for disease symptom development in Arabidopsis. Double mutants of acd5 and ethylene insensitive 2, in which ethylene signaling is blocked, show decreased cell death, supporting a role for ethylene in cell death control. We propose that acd5 plants mimic P. syringae-infected wild-type plants and that both SA and ethylene are normally involved in regulating cell death during some susceptible pathogen infections.

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BAK1 Is Not a Target of the Pseudomonas syringae Effector AvrPto

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-04-10-0096 ◽

2011 ◽

Vol 24 (1) ◽

pp. 100-107 ◽

Cited By ~ 49

Author(s):

Tingting Xiang ◽

Na Zong ◽

Jie Zhang ◽

Jinfeng Chen ◽

Mingsheng Chen ◽

...

Keyword(s):

Cell Surface ◽

Immune Responses ◽

Pseudomonas Syringae ◽

Plant Cell ◽

Crucial Role ◽

Pathogenic Bacteria ◽

Effector Proteins ◽

Receptor Like Kinase ◽

Receptor Kinases ◽

New Evidence

Plant cell surface-localized receptor kinases such as FLS2, EFR, and CERK1 play a crucial role in detecting invading pathogenic bacteria. Upon stimulation by bacterium-derived ligands, FLS2 and EFR interact with BAK1, a receptor-like kinase, to activate immune responses. A number of Pseudomonas syringae effector proteins are known to block immune responses mediated by these receptors. Previous reports suggested that both FLS2 and BAK1 could be targeted by the P. syringae effector AvrPto to inhibit plant defenses. Here, we provide new evidence further supporting that FLS2 but not BAK1 is targeted by AvrPto in plants. The AvrPto-FLS2 interaction prevented the phosphorylation of BIK1, a downstream component of the FLS2 pathway.

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