Proteasome-Dependent Degradation of RPM1 Desensitizes the RPM1-Mediated Hypersensitive Response

Mackey David,윤대진,Nam Jaesung 한국식물학회 2021 Journal of Plant Biology Vol.64 No.3

The intracellular plant resistance (R) proteins, nucleotide-binding and leucine-rich repeat (NLR) proteins, mediate resistance to pathogens by enabling recognition and rapid response. The response consists of the induction of a defensive suite that typically culminates in the hypersensitive response (HR), death of the plant cells at and around an infection site. The Arabidopsis intracellular innate immune receptor protein RESISTANCE TO PSEUDOMONAS MACULICOLA1 (RPM1) is a coiled-coil (CC) type of NLR protein that specifies resistance to strains of the bacterial pathogen Pseudomonas syringae expressing the type III effector proteins AvrRpm1 and AvrB. We previously demonstrated that RPM1-myc (an epitope-tagged version of RPM1) disappears coincident with the onset of HR induced by AvrRpm1. Infection with P. syringae expressing two other type III effector proteins, AvrRpt2 and AvrRps4, also initiated RPM1-myc disappearance at time points coincident with the HR they initiate through the NLR proteins RESISTANCE TO P. SYRINGAE2 (RPS2) and RESISTANCE TO P. SYRINGAE 4 (RPS4), respectively. Here, we use mutants impaired in NLR gene dependent signaling to demonstrate that disappearance of RPM1-myc requires normal NLR gene dependent signaling steps, but does not require HR. Inhibitors of the 26S proteasome block the disappearance of RPM1-myc and enhance RPM1-myc-dependent cell death. Our data are consistent with a model in which RPM1 is degraded by the 26S proteasome to limit the extent of RPM1-dependent signaling and/or cell death. Furthermore, AvrRpt2 induces disappearance of RPM1-myc in rps2 mutant plants without HR, suggesting that RPM1 is part of the host target of the virulence activity of AvrRpt2.

Responses of Arabidopsis thaliana to Challenge by Pseudomonas syringae

김민갑,Sun Young Kim,Woe Yeon Kim,David Mackey,이상열 한국분자세포생물학회 2008 Molecules and cells Vol.25 No.3

Plants are continually exposed to a variety of potentially pathogenic microbes, and the interactions between plants and pathogenic invaders determine the outcome, disease or disease resistance. To defend themselves, plants have developed a sophisticated immune system. Unlike animals, however, they do not have specialized immune cells and, thus all plant cells appear to have the innate ability to recognize pathogens and turn on an appropriate defense response. Using genetic, genomic and biochemical methods, tremendous advances have been made in understanding how plants recognize pathogens and mount effective defenses. The primary immune response is induced by microbe-associated molecular patterns (MAMPs). MAMP receptors recognize the presence of probable pathogens and evoke defense. In the co-evolution of plant-microbe interactions, pathogens gained the ability to make and deliver effector proteins to suppress MAMP-induced defense responses. In response to effector proteins, plants acquired R-proteins to directly or indirectly monitor the presence of effector proteins and activate an effective defense response. In this review we will describe and discuss the plant immune responses induced by two types of elicitors, PAMPs and effector proteins.

The Pseudomonas syringae type III effectors AvrRpm1 and AvrRpt2 promote virulence dependent on the F-box protein COI1

Geng, Xueqing,Shen, Mingzhe,Kim, Jin Hee,Mackey, David Springer-Verlag 2016 Plant cell reports Vol.35 No.4

<P>Type III effectors AvrRpm1 and AvrRpt2 promote bacterial growth dependent on a COI1-mediated pathway in the absence of the RPM1 and RPS2 resistance proteins. The type III effectors, AvrRpm1 and AvrRpt2, promote bacterial virulence by suppressing host defense responses. The defense suppressing activities of AvrRpm1 and AvrRpt2 are best studied in the absence of the resistance proteins RPM1 and RPS2, which induce defense responses to them. We tested whether the type III effectors could modulate a CORONATINE INSENSITIVE1 (COI1)-mediated hormone signaling pathway to promote virulence. COI1 has been demonstrated to contribute in the induction of chlorosis during Pseudomonas syringae infection. By comparing the activity of inducibly expressed AvrRpm1-HA or AvrRpt2-HA in rpm1rps2 and rpm1rps2coi1 backgrounds, we demonstrate that both effectors promote bacterial growth dependent on a COI1-mediated pathway and additively with the action of coronatine (COR) and that AvrRpt2-HA induces COI1-dependent chlorosis. Further, PATHOGENESIS RELATED1 (PR-1) expression resulting from inducible expression of AvrRpm1-HA or AvrRpt2-HA is elevated in coi1 plants consistent with the effectors activating JA-signaling to antagonize SA-signaling. In addition, we found that AvrRpm1-HA or AvrRpt2-HA requires COI1 to promote bacterial growth through suppression of both SA-dependent and SA-independent defense responses. Collectively, these results indicate that type III effectors AvrRpm1 and AvrRpt2 promote bacterial virulence by targeting a COI1-dependent signaling pathway.</P>

The <i>Pseudomonas syringae</i> type III effector AvrRpm1 induces significant defenses by activating the Arabidopsis nucleotide-binding leucine-rich repeat protein RPS2

Kim, Min Gab,Geng, Xueqing,Lee, Sang Yeol,Mackey, David Blackwell Publishing Ltd 2009 The Plant journal Vol.57 No.4

<P>Summary</P><P>Plant disease resistance (R) proteins recognize potential pathogens expressing corresponding avirulence (Avr) proteins through ‘gene-for-gene’ interactions. RPM1 is an Arabidopsis R-protein that triggers a robust defense response upon recognizing the <I>Pseudomonas syringae</I> effector AvrRpm1. Avr-proteins of phytopathogenic bacteria include type III effector proteins that are often capable of enhancing virulence when not recognized by an R-protein. In <I>rpm1</I> plants, AvrRpm1 suppresses basal defenses induced by microbe-associated molecular patterns. Here, we show that expression of AvrRpm1 in <I>rpm1</I> plants induced PR-1, a classical defense marker, and symptoms including chlorosis and necrosis. PR-1 expression and symptoms were reduced in plants with mutations in defense signaling genes (<I>pad4</I>, <I>sid2</I>, <I>npr1</I>, <I>rar1</I>, and <I>ndr1</I>) and were strongly reduced in <I>rpm1 rps2</I> plants, indicating that AvrRpm1 elicits defense signaling through the Arabidopsis R-protein, RPS2. Bacteria expressing AvrRpm1 grew more on <I>rpm1 rps2</I> than on <I>rpm1</I> plants. Thus, independent of its classical ‘gene-for-gene’ activation of RPM1, AvrRpm1 also induces functionally relevant defenses that are dependent on RPS2. Finally, AvrRpm1 suppressed host defenses and promoted the growth of type III secretion mutant bacteria equally well in <I>rps2</I> and <I>RPS2</I> plants, indicating that virulence activity of over-expressed AvrRpm1 predominates over defenses induced by weak activation of RPS2.</P>

The phytotoxin coronatine is a multifunctional component of the virulence armament of <i>Pseudomonas syringae</i>

Geng, Xueqing,Jin, Lin,Shimada, Mikiko,Kim, Min Gab,Mackey, David Springer Berlin Heidelberg 2014 Planta Vol.240 No.6

<P>Plant pathogens deploy an array of virulence factors to suppress host defense and promote pathogenicity. Numerous strains of <I>Pseudomonas syringae</I> produce the phytotoxin coronatine (COR). A major aspect of COR function is its ability to mimic a bioactive jasmonic acid (JA) conjugate and thus target the JA-receptor COR-insensitive 1 (COI1). Biological activities of COR include stimulation of JA-signaling and consequent suppression of SA-dependent defense through antagonistic crosstalk, antagonism of stomatal closure to allow bacterial entry into the interior of plant leaves, contribution to chlorotic symptoms in infected plants, and suppression of plant cell wall defense through perturbation of secondary metabolism. Here, we review the virulence function of COR, including updates on these established activities as well as more recent findings revealing COI1-independent activity of COR and shedding light on cooperative or redundant defense suppression between COR and type III effector proteins.</P>

Regulated Disorder: Posttranslational Modifications Control the RIN4 Plant Immune Signaling Hub

Toruñ,o, Tania Y.,Shen, Mingzhe,Coaker, Gitta,Mackey, David Scientific Societies 2019 Molecular plant-microbe interactions Vol.32 No.1

Responses of Arabidopsis thaliana to Challenge by Pseudomonas syringae

Kim, Min Gab,Kim, Sun Yong,Kim, Woe Yeon,Mackey, David,Lee, Sang Yeol Korean Society for Molecular Biology 2008 Molecules and cells Vol.25 No.3

Plants are continually exposed to a variety of potentially pathogenic microbes, and the interactions between plants and pathogenic invaders determine the outcome, disease or disease resistance. To defend themselves, plants have developed a sophisticated immune system. Unlike animals, however, they do not have specialized immune cells and, thus all plant cells appear to have the innate ability to recognize pathogens and turn on an appropriate defense response. Using genetic, genomic and biochemical methods, tremendous advances have been made in understanding how plants recognize pathogens and mount effective defenses. The primary immune response is induced by microbe-associated molecular patterns (MAMPs). MAMP receptors recognize the presence of probable pathogens and evoke defense. In the co-evolution of plant-microbe interactions, pathogens gained the ability to make and deliver effector proteins to suppress MAMP-induced defense responses. In response to effector proteins, plants acquired R-proteins to directly or indirectly monitor the presence of effector proteins and activate an effective defense response. In this review we will describe and discuss the plant immune responses induced by two types of elicitors, PAMPs and effector proteins.

Perturbation of Maize Phenylpropanoid Metabolism by an AvrE Family Type III Effector from <i>Pantoea stewartii</i>

Asselin, Jo Ann E.,Lin, Jinshan,Perez-Quintero, Alvaro L.,Gentzel, Irene,Majerczak, Doris,Opiyo, Stephen O.,Zhao, Wanying,Paek, Seung-Mann,Kim, Min Gab,Coplin, David L.,Blakeslee, Joshua J.,Mackey, Da American Society of Plant Biologists 2015 Plant Physiology Vol.167 No.3

<P><I>The virulence activity of an effector protein belonging to the widely conserved AvrE family is linked to its ability to cause system-wide reprogramming of phenylpropanoid metabolism in susceptible maize seedlings.</I></P><P>AvrE family type III effector proteins share the ability to suppress host defenses, induce disease-associated cell death, and promote bacterial growth. However, despite widespread contributions to numerous bacterial diseases in agriculturally important plants, the mode of action of these effectors remains largely unknown. WtsE is an AvrE family member required for the ability of <I>Pantoea stewartii</I> ssp. <I>stewartii</I> (<I>Pnss</I>) to proliferate efficiently and cause wilt and leaf blight symptoms in maize (<I>Zea mays</I>) plants. Notably, when WtsE is delivered by a heterologous system into the leaf cells of susceptible maize seedlings, it alone produces water-soaked disease symptoms reminiscent of those produced by <I>Pnss</I>. Thus, WtsE is a pathogenicity and virulence factor in maize, and an <I>Escherichia coli</I> heterologous delivery system can be used to study the activity of WtsE in isolation from other factors produced by <I>Pnss</I>. Transcriptional profiling of maize revealed the effects of WtsE, including induction of genes involved in secondary metabolism and suppression of genes involved in photosynthesis. Targeted metabolite quantification revealed that WtsE perturbs maize metabolism, including the induction of coumaroyl tyramine. The ability of mutant WtsE derivatives to elicit transcriptional and metabolic changes in susceptible maize seedlings correlated with their ability to promote disease. Furthermore, chemical inhibitors that block metabolic flux into the phenylpropanoid pathways targeted by WtsE also disrupted the pathogenicity and virulence activity of WtsE. While numerous metabolites produced downstream of the shikimate pathway are known to promote plant defense, our results indicate that misregulated induction of phenylpropanoid metabolism also can be used to promote pathogen virulence.</P>

Direct and Indirect Targeting of PP2A by Conserved Bacterial Type-III Effector Proteins

Jin, Lin,Ham, Jong Hyun,Hage, Rosemary,Zhao, Wanying,Soto-Herná,ndez, Jaricelis,Lee, Sang Yeol,Paek, Seung-Mann,Kim, Min Gab,Boone, Charles,Coplin, David L.,Mackey, David Public Library of Science 2016 PLoS pathogens Vol.12 No.5

<▼1><P>Bacterial AvrE-family Type-III effector proteins (T3Es) contribute significantly to the virulence of plant-pathogenic species of <I>Pseudomonas</I>, <I>Pantoea</I>, <I>Ralstonia</I>, <I>Erwinia</I>, <I>Dickeya</I> and <I>Pectobacterium</I>, with hosts ranging from monocots to dicots. However, the mode of action of AvrE-family T3Es remains enigmatic, due in large part to their toxicity when expressed in plant or yeast cells. To search for targets of WtsE, an AvrE-family T3E from the maize pathogen <I>Pantoea stewartii</I> subsp. <I>stewartii</I>, we employed a yeast-two-hybrid screen with non-lethal fragments of WtsE and a synthetic genetic array with full-length WtsE. Together these screens indicate that WtsE targets maize protein phosphatase 2A (PP2A) heterotrimeric enzyme complexes via direct interaction with B’ regulatory subunits. AvrE1, another AvrE-family T3E from <I>Pseudomonas syringae</I> pv. tomato strain DC3000 (<I>Pto</I> DC3000), associates with specific PP2A B’ subunit proteins from its susceptible host Arabidopsis that are homologous to the maize B’ subunits shown to interact with WtsE. Additionally, AvrE1 was observed to associate with the WtsE-interacting maize proteins, indicating that PP2A B’ subunits are likely conserved targets of AvrE-family T3Es. Notably, the ability of AvrE1 to promote bacterial growth and/or suppress callose deposition was compromised in Arabidopsis plants with mutations of PP2A genes. Also, chemical inhibition of PP2A activity blocked the virulence activity of both WtsE and AvrE1 <I>in planta</I>. The function of HopM1, a <I>Pto</I> DC3000 T3E that is functionally redundant to AvrE1, was also impaired in specific PP2A mutant lines, although no direct interaction with B’ subunits was observed. These results indicate that sub-component specific PP2A complexes are targeted by bacterial T3Es, including direct targeting by members of the widely conserved AvrE-family.</P></▼1><▼2><P><B>Author Summary</B></P><P>Gram-negative bacterial pathogens employ type-III effector (T3E) proteins to suppress host immunity and promote disease symptoms. AvrE-family T3Es, which are widely distributed among plant-pathogenic bacteria, suppress host defense responses and also contribute to water-soaking, which is perhaps the most common symptom of bacterial diseases and likely results in the release of nutrients from host cells to promote pathogen growth. Despite the central virulence functions of AvrE-family T3Es, their mode of action remains enigmatic largely due to their cell toxicity. We report here that two AvrE-family T3Es, WtsE from the maize pathogen <I>Pantoea stewartii</I> subsp. <I>stewartii</I> and AvrE1 from the tomato and Arabidopsis pathogen <I>Pseudomonas syringae</I> pv. tomato, each target protein phosphatase 2A (PP2A) complexes in susceptible hosts via direct interaction/association with specific B’ regulatory subunits. Chemical inhibitors were used to demonstrate that PP2A activity is required for the virulence functions of WtsE and AvrE1. PP2A isoform specificity was also tested using mutants of Arabidopsis. More generally, PP2A subunits regulate, both positively and negatively, rapid pattern-triggered immune responses in Arabidopsis. Thus, bacterial T3Es target sub-component specific PP2A complexes to manipulate host immunity and cause disease symptoms during infection.</P></▼2>

Effect of Hydroxycinnamic Acid Amides, Coumaroyl Tyramine and Coumaroyl Tryptamine on Biotic Stress Response in Arabidopsis

Macoy Donah Mary J.,Uddin Shahab,안경익,Peseth Son,Ryu Gyeong Ryul,차준영,Lee Jong-Yeol,Bae Dongryeoul,백승만,정혜진,Mackey David,이상열,김외연,김민갑 한국식물학회 2022 Journal of Plant Biology Vol.65 No.2

Coumaroyl tyramine (CT) and coumaroyl tryptamine (CTr) are neutral hydroxycinnamic acid amides (HCAAs) that accumulate in plants in response to pathogen infection. In this study, we showed that inoculation of Pseudomonas syringae pv. tomato DC3000 (Pto) and Erwinia carotovora carotovora (ECC) increased the accumulation of CT in Arabidopsis thaliana leaves at 24 h post-inoculation. Both CT and CTr increased the accumulation of PATHOGENESIS-RELATED 1 (PR1) protein. However, CT and CTr had no significant effect on resistance to biotrophic pathogens in both compatible and incompatible plant–pathogen interactions. Nonetheless, our results revealed that CTr plays a critical role in increasing plant susceptibility to the necrotrophic pathogen ECC. Exogenous application of CT and CTr increased the induction of callose deposition in both the absence and presence of ECC. Increased callose deposition was detected in salicylic acid induction-deficient Arabidopsis mutant sid2 but to a lesser extent in ethylene signaling mutants, ein2-1 and etr1. Overall, our results suggest that ethylene signaling is related to CT and CTr-induced callose deposition and contributes to plant defense against pathogens, whereas salicylic acid is not required for this response.