Arabidopsis thaliana- the model plant to study host-pathogen interactions.

In the last twenty years, the use of Arabidopsis as a model plant sped up discoveries at the molecular levels in different plant-parasite interactions. Nowadays, we know of probably hundreds of genes that are involved in the one or the other defence reaction, offering hundreds of targets for drug development. Even more interesting, identifying crucial regulatory components might allow to influence the various defence pathways as needed. Moreover, since some pathogenic strategies are conserved between animal and plant pathogens, results obtained with one system might be applicable to the other.

Allicin disrupts the cell's electrochemical potential and induces apoptosis in yeast.

The volatile substance allicin gives crushed garlic (Allium sativum) its characteristic odor and is a pro-oxidant that undergoes thiol-disulfide exchange reactions with -SH groups in proteins and glutathione. The antimicrobial activity of allicin is suspected to be due to the oxidative inactivation of essential thiol-containing enzymes. We investigated the hypothesis that at threshold inhibitory levels allicin can shunt yeast cells into apoptosis by altering their overall redox status. Yeast cells were treated either with chemically synthesized, pure allicin or with allicin in garlic juice. Allicin-dependent cell oxidation was demonstrated with a redox-sensitive GFP construct and the shift in cellular electrochemical potential (E(hc)) from less than -215 to -181mV was calculated using the Nernst equation after the glutathione/glutathione disulfide couple (2GSH/GSSG) in the cell was quantified. Caspase activation occurred after allicin treatment, and yeast expressing a human antiapoptotic Bcl-XL construct was rendered more resistant to allicin. Also, a yeast apoptosis-inducing factor deletion mutant was more resistant to allicin than wild-type cells. We conclude that allicin in garlic juice can activate apoptosis in yeast cells through its oxidizing properties and that this presents an alternative cell-killing mechanism to the previously proposed specific oxidative inactivation of essential enzymes.

Imbalanced lignin biosynthesis promotes the sexual reproduction of homothallic oomycete pathogens.

Lignin is incorporated into plant cell walls to maintain plant architecture and to ensure long-distance water transport. Lignin composition affects the industrial value of plant material for forage, wood and paper production, and biofuel technologies. Industrial demands have resulted in an increase in the use of genetic engineering to modify lignified plant cell wall composition. However, the interaction of the resulting plants with the environment must be analyzed carefully to ensure that there are no undesirable side effects of lignin modification. We show here that Arabidopsis thaliana mutants with impaired 5-hydroxyguaiacyl O-methyltransferase (known as caffeate O-methyltransferase; COMT) function were more susceptible to various bacterial and fungal pathogens. Unexpectedly, asexual sporulation of the downy mildew pathogen, Hyaloperonospora arabidopsidis, was impaired on these mutants. Enhanced resistance to downy mildew was not correlated with increased plant defense responses in comt1 mutants but coincided with a higher frequency of oomycete sexual reproduction within mutant tissues. Comt1 mutants but not wild-type Arabidopsis accumulated soluble 2-O-5-hydroxyferuloyl-L-malate. The compound weakened mycelium vigor and promoted sexual oomycete reproduction when applied to a homothallic oomycete in vitro. These findings suggested that the accumulation of 2-O-5-hydroxyferuloyl-L-malate accounted for the observed comt1 mutant phenotypes during the interaction with H. arabidopsidis. Taken together, our study shows that an artificial downregulation of COMT can drastically alter the interaction of a plant with the biotic environment.

Flavin-containing monooxygenases in plants: looking beyond detox.

Flavin-containing monooxygenases (FMOs) are known in bacteria, yeast and mammals where they catalyze the transfer of one atom of molecular O(2) to low molecular weight substrates. The predominant physiological function of animal FMOs appears to be detoxification of a vast spectrum of xenobiotics but until recently very little was known about the function of FMOs in plants. In the last two to three years, genetic and biochemical characterization has shown that plant FMOs can catalyze specific steps in the biosynthesis of auxin or in the metabolism of glucosinolates, and, furthermore, have a role in pathogen defence. Thus, plant FMOs hint that further FMO functions might be identified also in non-plant organisms and could stimulate novel research in this area.

A role for a flavin-containing mono-oxygenase in resistance against microbial pathogens in Arabidopsis.

Using activation tagging in the Arabidopsis Col-0 rps2-101C background, we identified a mutant (FMO1-3D) that showed virtually no symptoms after inoculation with virulent Pseudomonas syringae pv. tomato DC3000 bacteria. The dominant, gain-of-function phenotype of the FMO1-3D mutant is due to over-expression of a class 3 flavin-containing mono-oxygenase (FMO). We recapitulated the FMO1-3D mutant phenotype in independent transgenic Col-0 lines over-expressing the FMO1 cDNA under the control of the 35S CaMV promoter. The increased basal resistance observed in the FMO1-3D mutant was also effective against the taxonomically unrelated downy mildew-causing pathogen Hyaloperonospora parasitica. By investigating the progeny from crosses of the FMO1-3D mutant with the NahG transgenic line, we showed that the enhanced basal resistance phenotype was dependent on the accumulation of salicylic acid. FMO1-3D plants showed wild-type resistant reactions after inoculation with avirulent bacteria, indicating that the R-gene-mediated defence physiology was not compromised by FMO1 over-expression. Transcripts of the class 3 FMO1 gene accumulated within 6 h after inoculation of wild-type Col-0 plants with avirulent Pst + avrRpt2 cells. Moreover, a T-DNA insertion into the FMO1 gene resulted in enhanced susceptibility to virulent Pseudomonas and Hyaloperonospora parasitica, suggesting that expression of the FMO1 gene is a hitherto undescribed component of the plant's resistance repertoire. We discuss the possibility that the FMO may participate in the detoxification of virulence factors produced by pathogens.

PCC1: a merging point for pathogen defence and circadian signalling in Arabidopsis.

Using a cDNA-array we identified expressed sequence tag 163B24T7 as rapidly up-regulated in Arabidopsis thaliana (L.) Heynh. after pathogen exposure. Detailed expression analysis revealed that the corresponding gene is up-regulated not only after exposure to avirulent Pseudomonas syringae pv. tomato but also to virulent strains. This up-regulation is dependent on functional salicylic acid defence-signalling pathways. Moreover, we found the gene was circadian-regulated, showing peaks of expression at the end of the day. Using plants overexpressing the clock component CCA1, we showed that the PCC1 gene is regulated by the inner clock of Arabidopsis. Accordingly, we named the gene PCC1, for pathogen and circadian controlled. PCC1 is a member of a novel family of six small polypeptides in Arabidopsis. A functional role for PCC1 in plant defence was demonstrated since plants overexpressing PCC1 are resistant against normally virulent oomycetes. Thus, PCC1 demonstrates a potential interrelationship between pathogen and circadian signalling pathways.

Organ-specificity in a plant disease is determined independently of R gene signaling.

The molecular basis of organ specificity in plant diseases is little characterized. Downy mildew of Arabidopsis caused by the oomycete Hyaloperonospora parasitica (formerly Peronospora parasitica) is characteristically a leaf disease. Resistant host genotypes recognize the pathogen in a gene-for-gene dependent manner and respond with the production of H2O2 and the execution of a genetically programmed hypersensitive cell death (HR). We inoculated the roots of Arabidopsis genotypes Col-0, Ws-0, and Wei-0 with the NOCO and WELA races of the pathogen and compared the responses with those observed in leaves. Combinations of incompatible genotypes of host and pathogen showed the expected responses of an oxidative burst and the HR in leaves, but surprisingly, roots showed no signs of active defense and appeared completely susceptible to all the H. parasitica isolates tested. Reverse transcriptase-polymerase chain reaction showed that the R gene RPP1, which mediates resistance in leaves of accession Ws-0 to the H. parasitica isolate NOCO, was expressed in leaves as well as in roots. Similarly, NDR1 and EDS1, two components of R gene-mediated signaling pathways, are also expressed in both tissues. To our knowledge, it has not been previously demonstrated that expression of R genes and downstream components of the signaling cascade are not sufficient for the induction of avirulence gene-mediated defense mechanisms in roots.

Downy mildew of Arabidopsis thaliana caused by Hyaloperonospora parasitica (formerly Peronospora parasitica).

UNLABELLED: SUMMARY Downy mildew of Arabidopsis is not a hugely destructive disease of an important crop plant, neither is it of any economic importance. The most obvious symptom, the aerial conidiophores, might, at a glance to the casual observer, be mistaken for the trichomes normally present on the leaves. However, a huge research effort is being devoted to this humble pathosystem which became established as a laboratory model in the 1990s. Since then, enormous progress has been made in cloning and characterizing major genes for resistance (RPP genes) and in defining many of their downstream signalling components, some of them RPP-gene specific. Resistance is generally associated with an oxidative burst and a salicylic acid dependent hypersensitive reaction type of programmed cell death. Biological and chemical induction of systemic acquired resistance (SAR) in Arabidopsis protecting against downy mildew were demonstrated early on, and investigations of mutants have contributed fundamentally to our understanding of host-pathogen interactions and the mechanisms of plant defence. This review will attempt to collate the wealth of information which has accrued with this pathosystem in the last decade and will attempt to predict future research directions by drawing attention to some still unanswered questions. TAXONOMY: Hyaloperonospora Constant. parasitica (Pers.:Fr) Fr. (formerly Peronospora parasitica), Kingdom Chromista, Phylum Oomycota, Order Peronosporales, Family Peronosporaceae, Genus Hyaloperonospora, of which it is the type species. The taxonomy of the group of organisms causing downy mildew of brassicas has undergone a number of revisions since Corda (1837) originally coined the genus Peronospora. All isolates pathogenic on brassicas were described initially as P. parasitica but Gäumann (1918) classified isolates from different brassicaceous hosts distinctly and thus defined 52 new species based on conidial dimensions and host range. After much debate it was decided to revert to the aggregate species of P. parasitica for all brassica-infecting downy mildews, whilst recognizing that these show some isolate-specific differences (Yerkes and Shaw, 1959). The latest re-examination of P. parasitica by Constantinescu and Fatehi (2002) has placed isolates of P. parasitica and five other downy mildew species in a clear new subgroup on the basis of their hyaline conidiospores, recurved conidiophore branch tips and ITS1, ITS2 and 5.8S rDNA sequence comparisons; meriting the coining of the new genus 'Hyaloperonospora Constant'. The class Oomycetes in the Kingdom Chromista (Straminipila) comprises fungus-like organisms with heterokont zoospores (i.e. possessing two types of flagellae, whiplash and tinsel). The Oomycetes have non-septate hyphae with cellulose-based walls containing very little or no chitin. The latter is regarded as a major distinction separating the Oomycetes from the true fungi, and reports of the presence of chitin had generally been regarded as due to small amounts of contamination (Gams et al., 1998). However, in view of recent studies by Werner et al. (2002) showing a chitin synthase gene in an Oomycete and demonstrating the presence of the polymer itself by an interaction with wheat germ agglutinin (WGA), it is perhaps safe to say that we have not seen the last taxonomic revision which will affect this group! The families within the Oomycetes show a clear evolutionary trend to a lesser absolute dependence on an aqueous environment and some members of the Peronosporales, e.g. H. parasitica, have no zoosporic stage in the life cycle. HOST RANGE: Isolates infecting Arabidopsis thaliana have so far proven to be non-pathogenic on other crucifers tested but exist in a clear gene-for-gene relationship with different host ecotypes. Disease symptoms: Infections are first apparent to the naked eye as a carpet or 'down' of conidiophores covering the upper and lower surfaces of leaves and petioles. This symptom is characteristic of this group of diseases and lends it its name. USEFUL WEBSITES: (links to references on Oomycetes), (TAIR, The Arabidopsis Information Resource).

Monitoring the switch from housekeeping to pathogen defense metabolism in Arabidopsis thaliana using cDNA arrays.

Plants respond to pathogen attack by deploying several defense reactions. Some rely on the activation of preformed components, whereas others depend on changes in transcriptional activity. Using cDNA arrays comprising 13,000 unique expressed sequence tags, changes in the transcriptome of Arabidopsis thaliana were monitored after attempted infection with the bacterial plant pathogen Pseudomonas syringae pv. tomato carrying the avirulence gene avrRpt2. Sampling at four time points during the first 24 h after infiltration revealed significant changes in the steady state transcript levels of approximately 650 genes within 10 min and a massive shift in gene expression patterns by 7 h involving approximately 2,000 genes representing many cellular processes. This shift from housekeeping to defense metabolism results from changes in regulatory and signaling circuits and from an increased demand for energy and biosynthetic capacity in plants fighting off a pathogenic attack. Concentrating our detailed analysis on the genes encoding enzymes in glycolysis, the Krebs cycle, the pentose phosphate pathway, the biosynthesis of aromatic amino acids, phenylpropanoids, and ethylene, we observed interesting differential regulation patterns. Furthermore, our data showed potentially important changes in areas of metabolism, such as the glyoxylate metabolism, hitherto not suspected to be components of plant defense.