Recycling of Arabidopsis plasma membrane PEN1 syntaxin

Penetration resistance against powdery mildews is one of the best-studied processes of plant innate immunity. One vital component is the plant syntaxin, PEN1, which is required for timely deposition of callose and extracellular membrane material, as well as PEN1 itself, at the attack sites. Recently, we reported that the ARF-GEF GNOM also is required for penetration resistance, mediating transport of recycled material, including PEN1, to the site of attack. The close relative of PEN1, SYP122, does not accumulate at the sites of attack nor does it affect penetration resistance. In support of this, we show here that in contrast to PEN1, SYP122 does not continuously recycle. Furthermore, by using a PEN1 transgene that is only transcribed in dividing cells, we show that papillary PEN1 accumulation is not dependent on de-novo protein synthesis. This emphasizes the involvement of recycling in penetration resistance, which possibly relates to the differences in function of the two syntaxins.     

Self-consuming innate immunity in Arabidopsis

Programmed cell death (PCD) associated with the pathogen-induced hypersensitive response (HR) is a hallmark of plant innate immunity. HR PCD is triggered upon recognition of pathogen effector molecules by host immune receptors either directly or indirectly via effector modulation of host targets. However, it has been unclear by which molecular mechanisms plants execute PCD during innate immune responses. We recently examined HR PCD in autophagy-deficient Arabidopsis knockout mutants (atg) and find that PCD conditioned by one class of plant innate immune receptors is suppressed in atg mutants. Intriguingly, HR triggered by another class of immune receptors with different genetic requirements is not compromised, indicating that only a specific subset of immune receptors engage the autophagy pathway for HR execution. Thus, our work provides a primary example of autophagic cell death associated with innate immune responses in eukaryotes as well as of pro-death functions for the autophagy pathway in plants.     

New insights into innate immunity in Arabidopsis

The term innate immunity has been described as '. . . the surveillance system that detects the presence and nature of the infection and provides the first line of host defense . . .' (Medzhitov, 2001; Nat Rev Immunol 1: 135-145). The strategy of innate immunity is based on the recognition of constitutive and conserved molecules from pathogens by specific receptors, triggering defence responses (Medzhitov and Janeway, 2002; Science 296: 298-300). It has been only within the past few years that studies of plant innate immunity, especially in Arabidopsis, have provided important insights into molecular details that define innate immunity in plants. Here we review the innate immune response in Arabidopsis, where leucine-rich repeat (LRR) cell surface receptors play central roles in monitoring the presence of pathogen (microbe) associated molecules to initiate the rapid expression of defence genes. The PAMPS also activate the expression of genes encoding a family of endogenous peptides (AtPep1 paralogues) and their receptor (PEPR1) that amplify defence signalling through a feedback loop initiated by PAMPS. The concept of innate immunity has provided a valuable framework for researchers to re-evaluate the roles of exogenous and endogenous signals that regulate the expression of plant defensive genes.     

The ubiquitin pathway is required for innate immunity in Arabidopsis

Plant defences require a multitude of tightly regulated resistance responses. In Arabidopsis, the unique gain-of-function mutant suppressor of npr1-1 constitutive 1 ( snc1 ) carries a point mutation in a Resistance ( R )-gene, resulting in constitutive activation of defence responses without interaction with pathogens. This has allowed us to identify various downstream signalling components essential in multiple defence pathways. One mutant that suppresses snc1 -mediated constitutive resistance is modifier of snc1 5 ( mos5 ), which carries a 15-bp deletion in UBA1 , one of two ubiquitin-activating enzyme genes in Arabidopsis. A mutation in UBA2 does not suppress snc1 , suggesting that these two genes are not equally required in Arabidopsis disease resistance. On the other hand, a mos5 uba2 double mutant is lethal, implying partial redundancy of the two homologues. Apart from affecting snc1 -mediated resistance, mos5 also exhibits enhanced disease susceptibility to a virulent pathogen and is impaired in response to infection with avirulent bacteria carrying the protease elicitor AvrRpt2. The mos5 mutation in the C-terminus of UBA1 might affect binding affinity of the downstream ubiquitin-conjugating enzymes, thus perturbing ubiquitination of target proteins. Furthermore, SGT1b and RAR1, which are necessary for resistance conferred by the SNC1 -related R -genes RPP4 and RPP5 , are dispensable in snc1 -mediated resistance. Our data reveal the definite requirement for the ubiquitination pathway in the activation and downstream signalling of several R-proteins.     

Effector-triggered innate immunity contributes Arabidopsis resistance to Xanthomonas campestris

Xanthomonas campestris pv. campestris, the causal agent of black rot disease, depends on its type III secretion system (TTSS) to infect cruciferous plants, including Brassica oleracea, B. napus and Arabidopsis. Previous studies on the Arabidopsis-Pseudomonas syringae model pathosystem have indicated that a major function of TTSS from virulent bacteria is to suppress host defences triggered by pathogen-associated molecular patterns. Similar analyses have not been made for the Arabidopsis-X. campestris pv. campestris pathosystem. In this study, we report that X. campestris pv. campestris strain 8004, which is modestly pathogenic on Arabidopsis, induces strong defence responses in Arabidopsis in a TTSS-dependent manner. Furthermore, the induction of defence responses and disease resistance to X. campestris pv. campestris strain 8004 requires NDR1 (NON-RACE-SPECIFIC DISEASE RESISTANCE1), RAR1 (required for Mla12 resistance) and SGT1b (suppressor of G2 allele of skp1), suggesting that effector-triggered immunity plays a large role in resistance to this strain. Consistent with this notion, AvrXccC, an X. campestris pv. campestris TTSS effector protein, induces PR1 expression and confers resistance in Arabidopsis in a RAR1- and SGT1b-dependent manner. In rar1 and sgt1b mutants, AvrXccC acts as a virulence factor, presumably because of impaired resistance gene function.     

模式识别受体Dectin-1与天然免疫反应

在免疫反应早期,抗原递呈细胞表达的模式识别受体通过识别内、外源配体在清除衰老宿主细胞、分子以及防御感染中发挥重要作用。新近发现的树突状细胞相关性C型植物血凝素-1(dendritic cell—associated C-type lectin-1,Dectin-1)作为β-葡聚糖的主要受体,广泛分布于单核巨噬细胞系统、树突状细胞(dendritic cell,DC)及中性粒细胞等。本文对Dectin-1分子结构、细胞组织分布及其在诱导、调控局部和全身免疫、炎症反应中的作用进行概述。     

Salicylic acid-mediated innate immunity in Arabidopsis is regulated by SIZ1 SUMO E3 ligase

Reversible modifications of target proteins by small ubiquitin-like modifier (SUMO) proteins are involved in many cellular processes in yeast and animals. Yet little is known about the function of sumoylation in plants. Here, we show that the SIZ1 gene, which encodes an Arabidopsis SUMO E3 ligase, regulates innate immunity. Mutant siz1 plants exhibit constitutive systemic-acquired resistance (SAR) characterized by elevated accumulation of salicylic acid (SA), increased expression of pathogenesis-related (PR) genes, and increased resistance to the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) DC3000. Transfer of the NahG gene to siz1 plants results in reversal of these phenotypes back to wild-type. Analyses of the double mutants, npr1 siz1, pad4 siz1 and ndr1 siz1 revealed that SIZ1 controls SA signalling. SIZ1 interacts epistatically with PAD4 to regulate PR expression and disease resistance. Consistent with these observations, siz1 plants exhibited enhanced resistance to Pst DC3000 expressing avrRps4, a bacterial avirulence determinant that responds to the EDS1/PAD4-dependent TIR-NBS-type R gene. In contrast, siz1 plants were not resistant to Pst DC3000 expressing avrRpm1, a bacterial avirulence determinant that responds to the NDR1-dependent CC-NBS-type R gene. Jasmonic acid (JA)-induced PDF1.2 expression and susceptibility to Botrytis cinerea were unaltered in siz1 plants. Taken together, these results demonstrate that SIZ1 is required for SA and PAD4-mediated R gene signalling, which in turn confers innate immunity in Arabidopsis.     

Molecular determinants of syntaxin 1 modulation of N-type calcium channels

We have previously reported that syntaxin 1A, a component of the presynaptic SNARE complex, directly modulates N-type calcium channel gating in addition to promoting tonic G-protein inhibition of the channels, whereas syntaxin 1B affects channel gating but does not support G-protein modulation (Jarvis, S. E., and Zamponi, G. W. (2001) J. Neurosci. 21, 2939-2948). Here, we have investigated the molecular determinants that govern the action of syntaxin 1 isoforms on N-type calcium channel function. In vitro evidence shows that both syntaxin 1 isoforms physically interact with the G-protein beta subunit and the synaptic protein interaction (synprint) site contained within the N-type calcium channel domain II-III linker region. Moreover, in vitro evidence suggests that distinct domains of syntaxin participate in each interaction, with the COOH-terminal SNARE domain (residues 183-230) binding to Gbeta and the N-terminal (residues 1-69) binding to the synprint motif of the channel. Electrophysiological analysis of chimeric syntaxin 1A/1B constructs reveals that the variable NH(2)-terminal domains of syntaxin 1 are responsible for the differential effects of syntaxin 1A and 1B on N-type calcium channel function. Because syntaxin 1 exists in both "open" and "closed" conformations during exocytosis, we produced a constitutively open form of syntaxin 1A and found that it still promoted G-protein inhibition of the channels, but it did not affect N-type channel availability. This state dependence of the ability of syntaxin 1 to mediate N-type calcium channel availability suggests that syntaxin 1 dynamically regulates N-type channel function during various steps of exocytosis. Finally, syntaxin 1A appeared to compete with Ggamma for the Gbeta subunit both in vitro and under physiological conditions, suggesting that syntaxin 1A may contain a G-protein gamma subunit-like domain.     

The Pseudomonas syringae type III effector tyrosine phosphatase HopAO1 suppresses innate immunity in Arabidopsis thaliana

The bacterial pathogen Pseudomonas syringae pv. tomato (Pst) strain DC3000 infects tomato and Arabidopsis plants, and is a model for studying the molecular basis of bacterial disease. Pst DC3000 secretes a battery of largely uncharacterized effector proteins into host cells via a type-III secretion system (TTSS). Little is currently known about the molecular mechanisms by which individual TTSS effectors promote virulence. The effector HopAO1 has similarity to protein tyrosine phosphatases, including a conserved catalytic site, and suppresses the hypersensitive response (HR) in some non-host plants. Whether HopAO1 has a similar effect in the host Arabidopsis is not clear. Here, we show that transgenic expression of HopAO1 in Arabidopsis suppresses callose deposition elicited by the Pst DC3000 hrpA mutant, and allows the normally non-pathogenic hrpA mutant to multiply within the leaf tissue. HopAO1 also suppresses resistance to Pst DC3000 induced by flg22, a pathogen-associated molecular pattern (PAMP). However, HopAO1 does not suppress the HR triggered by several classical avirulence genes. These results suggest that HopAO1 targets primarily PAMP-induced innate immunity in Arabidopsis. The virulence function of HopAO1 is dependent on an intact phosphatase catalytic site, as transgenic plants expressing a catalytically inactive derivative do not show these effects. Intriguingly, expression of the catalytically inactive HopAO1 has a dominant-negative effect on the function of the wild-type HopAO1. Analysis of mitogen-activated protein kinase (MAPK) activity suggests that HopAO1 targets a step downstream or independent of MAPK activation. Genome-wide expression analysis revealed that expression of several well-known defense genes was suppressed in hrpA mutant-infected HopAO1 transgenic plants.     

A dual regulatory role of Arabidopsis calreticulin-2 in plant innate immunity

Calreticulin (CRT) is an endoplasmic reticulum-resident calcium-binding molecular chaperone that is highly conserved in multi-cellular eukaryotes. Higher plants contain two distinct groups of CRTs: CRT1/CRT2 and CRT3 isoforms. Previous studies have shown that bacterial elongation factor Tu receptor (EFR), a pattern-recognition receptor that is responsible for pathogen-associated molecular pattern-triggered immunity, is a substrate for Arabidopsis CRT3, suggesting a role for CRT3 in regulating plant defense against pathogens. Here we report that Arabidopsis CRT2 is another regulator of plant innate immunity. Despite significantly increased salicylic acid levels and constitutive expression of the systemic acquired resistance-associated marker genes PR1, PR2 and PR5, transgenic plants over-expressing CRT2 displayed reduced resistance to virulent Pseudomonas syringae pv. tomato DC3000 (PstDC3000). A (45)Ca(2+) overlay assay and a domain-swapping experiment further demonstrated that the negatively charged C-terminal tail of CRT2 is responsible for its high calcium-binding capacity and function in regulating the endogenous salicylic acid level. In addition, over-expression of the His173 mutant of CRT2 greatly enhanced plant defense against PstDC3000, supporting the existence of a self-inhibition mechanism that can counteract the effects of salicylic acid-dependent immune responses. These results suggest that CRT2 functions through its N-terminal domain(s) as a self-modulator that can possibly prevent the salicylic acid-mediated runaway defense responses triggered by its C-terminal calcium-buffering activity in response to pathogen invasion.     

A plasmodesmata-localized protein mediates crosstalk between cell-to-cell communication and innate immunity in Arabidopsis

Plasmodesmata (PD) are thought to play a fundamental role in almost every aspect of plant life, including normal growth, physiology, and developmental responses. However, how specific signaling pathways integrate PD-mediated cell-to-cell communication is not well understood. Here, we present experimental evidence showing that the Arabidopsis thaliana plasmodesmata-located protein 5 (PDLP5; also known as HOPW1-1-INDUCED GENE1) mediates crosstalk between PD regulation and salicylic acid-dependent defense responses. PDLP5 was found to localize at the central region of PD channels and associate with PD pit fields, acting as an inhibitor to PD trafficking, potentially through its capacity to modulate PD callose deposition. As a regulator of PD, PDLP5 was also essential for conferring enhanced innate immunity against bacterial pathogens in a salicylic acid-dependent manner. Based on these findings, a model is proposed illustrating that the regulation of PD closure mediated by PDLP5 constitutes a crucial part of coordinated control of cell-to-cell communication and defense signaling.     

ZmPep1, an ortholog of Arabidopsis elicitor peptide 1, regulates maize innate immunity and enhances disease resistance

ZmPep1 is a bioactive peptide encoded by a previously uncharacterized maize (Zea mays) gene, ZmPROPEP1. ZmPROPEP1 was identified by sequence similarity as an ortholog of the Arabidopsis (Arabidopsis thaliana) AtPROPEP1 gene, which encodes the precursor protein of elicitor peptide 1 (AtPep1). Together with its receptors, AtPEPR1 and AtPEPR2, AtPep1 functions to activate and amplify innate immune responses in Arabidopsis and enhances resistance to both Pythium irregulare and Pseudomonas syringae. Candidate orthologs to the AtPROPEP1 gene have been identified from a variety of crop species; however, prior to this study, activities of the respective peptides encoded by these orthologs were unknown. Expression of the ZmPROPEP1 gene is induced by fungal infection and treatment with jasmonic acid or ZmPep1. ZmPep1 activates de novo synthesis of the hormones jasmonic acid and ethylene and induces the expression of genes encoding the defense proteins endochitinase A, PR-4, PRms, and SerPIN. ZmPep1 also stimulates the expression of Benzoxazineless1, a gene required for the biosynthesis of benzoxazinoid defenses, and the accumulation of 2-hydroxy-4,7-dimethoxy-1,4-benzoxazin-3-one glucoside in leaves. To ascertain whether ZmPep1-induced defenses affect resistance, maize plants were pretreated with the peptide prior to infection with fungal pathogens. Based on cell death and lesion severity, ZmPep1 pretreatment was found to enhance resistance to both southern leaf blight and anthracnose stalk rot caused by Cochliobolis heterostrophus and Colletotrichum graminicola, respectively. We present evidence that peptides belonging to the Pep family have a conserved function across plant species as endogenous regulators of innate immunity and may have potential for enhancing disease resistance in crops.     

The major leaf ferredoxin Fd2 regulates plant innate immunity in Arabidopsis

Ferredoxins, the major distributors for electrons to various acceptor systems in plastids, contribute to redox regulation and antioxidant defence in plants. However, their function in plant immunity is not fully understood. In this study, we show that the expression of the major leaf ferredoxin gene Fd2 is suppressed by Pseudomonas syringae pv. tomato (Pst) DC3000 infection, and that knockout of Fd2 (Fd2‐KO) in Arabidopsis increases the plant's susceptibility to both Pst DC3000 and Golovinomyces cichoracearum. On Pst DC3000 infection, the Fd2‐KO mutant accumulates increased levels of jasmonic acid and displays compromised salicylic acid‐related immune responses. Fd2‐KO also shows defects in the accumulation of reactive oxygen species induced by pathogen‐associated molecular pattern‐triggered immunity. However, Fd2‐KO shows enhanced R‐protein‐mediated resistance to Pst DC3000/AvrRpt2 infection, suggesting that Fd2 plays a negative role in effector‐triggered immunity. Furthermore, Fd2 interacts with FIBRILLIN4 (FIB4), a harpin‐binding protein localized in chloroplasts. Interestingly, Fd2, but not FIB4, localizes to stromules that extend from chloroplasts. Taken together, our results demonstrate that Fd2 plays an important role in plant immunity.