The CURLY LEAF gene controls both division and elongation of cells during the expansion of the leaf blade in Arabidopsis thaliana

The CURLY LEAF (CLF ) gene in Arabidopsis thaliana (L.) Heynh. is required for stable repression of a floral homeotic gene, AGAMOUS in leaves and stems To clarify the function of CLF in organ development, we characterized clf mutants using an anatomical and genetic approach. The clf mutants had normal roots, hypocotyls, and cotyledons, but the foliage leaves and the stems had reduced dimensions. A decrease both in the extent of cell elongation and in the number of cells was evident in the clf mutant leaves, suggesting that the CLF gene might be involved in the division and elongation of cells during leaf morphogenesis. An analysis of the development of clf mutant leaves revealed that the period during which cell division or cell elongation occurred was of normal duration, while the rates of both cell production and cell elongation were lower than in the wild type. Two phases in the elongation of cells were also recognized from this analysis. From analysis of an angustifolia clf double mutant, we found that the two phases of elongation of leaf cells were regulated independently by each gene. Thus, the CLF gene appears to affect cell division at an earlier stage and cell elongation throughout the development of leaf primordia. Received: 19 February 1998 / Accepted: 24 March 1998     

Glutathione and tryptophan metabolites are key players in Arabidopsis nonhost resistance against Colletotrichum gloeosporioides

The nonhost resistance of Arabidopsis against hemibiotrophic fungi in the genus Colletotrichum consists of pre- and post-invasive immune responses. Previously, we reported EDR1 and PEN2 as important components of Arabidopsis pre-invasive resistance toward non-adapted Colletotrichum gloeosporioides (Cg). However, despite their defect in entry control pen2 and edr1 mutants terminated further growth of this pathogen by activating the post-invasive hypersensitive response (HR) accompanied by plant cell death. In the present study, we showed that γ-glutamylcysteine synthetase (GSH1), which is required for glutathione biosynthesis, and tryptophan (Trp) metabolism contribute to pre- and post-invasive non-host resistance against Cg. We found GSH1 to be involved in the PEN2-dependent entry control of Cg. Opposite to pen2 and edr1, gsh1 mutants failed to restrict the invasive growth of the pathogen, which demonstrated the requirement for GSH1 during post-invasive non-host resistance. Based on the infection and metabolic phenotypes of Arabidopsis mutants defective in Trp metabolism, we showed that the biosynthesis of Trp-derived phytochemicals is also essential for resistance to Cg during the post-invasive HR. By contrast, GSH1 and these metabolites are dispensable for the induction of HR cell death, which is triggered in the non-invaded mesophyll cells adjacent to the Cg-invaded epidermal cells.     

Bacterial non-host resistance: interactions of Arabidopsis with non-adapted Pseudomonas syringae strains

Although interactions of plants with virulent and avirulent host pathogens are under intensive study, relatively little is known about plant interactions with non-adapted pathogens and the molecular events underlying non-host resistance. Here we show that two Pseudomonas syringae strains for which Arabidopsis is a non-host plant, P. syringae pathovar (pv.) glycinea (Psg) and P. syringae pv. phaseolicola (Psp),induce salicylic acid (SA) accumulation and pathogenesis-related gene expression at inoculation sites, and that induction of these defences is largely dependent on bacterial type III secretion. The defence signalling components activated by non-adapted bacteria resemble those initiated by host pathogens, including SA, non-expressor of PR-1, non-race specific disease resistance 1, phytoalexin-deficient 4 and enhanced disease susceptibility 1. However, some differences in individual defence pathways induced by Psg and Psp exist, suggesting that for each strain, distinct sets of type III effectors are recognized by the plant. Although induction of SA-related defences occurs, it does not directly contribute to bacterial non-host resistance, because Arabidopsis mutants compromised in SA signalling and other classical defence pathways do not permit enhanced survival of Psg or Psp in leaves. The finding that numbers of non-adapted bacteria in leaf extracellular spaces rapidly decline after inoculation suggests that they fail to overcome toxic or structural defence barriers preceding SA-related responses. Consistent with this hypothesis, rapid, type III secretion system-independent upregulation of the lignin biosynthesis genes, PAL1 and BCB, which might contribute to an early induced, cell wall-based defence mechanism, occurs in response to non-adapted bacteria. Moreover, knockout of PAL1 permits increased leaf survival of non-host bacteria. In addition, different survival rates of non-adapted bacteria in leaves from Arabidopsis accessions and mutants with distinct glucosinolate composition or hydrolysis exist. Possible roles for early inducible, cell wall-based defences and the glucosinolate/myrosinase system in bacterial non-host resistance are discussed.     

RNAi of the sesquiterpene cyclase gene for phytoalexin production impairs pre- and post-invasive resistance to potato blight pathogens

Potato antimicrobial sesquiterpenoid phytoalexins lubimin and rishitin have been implicated in resistance to the late blight pathogen, Phytophthora infestans and early blight pathogen, Alternaria solani. We generated transgenic potato plants in which sesquiterpene cyclase, a key enzyme for production of lubimin and rishitin, is compromised by RNAi to investigate the role of phytoalexins in potato defence. The transgenic tubers were deficient in phytoalexins and exhibited reduced post-invasive resistance to an avirulent isolate of P. infestans, resulting in successful infection of the first attacked cells without induction of cell death. However, cell death was observed in the subsequently penetrated cells. Although we failed to detect phytoalexins and antifungal activity in the extract from wild-type leaves, post-invasive resistance to avirulent P. infestans was reduced in transgenic leaves. On the other hand, A. solani frequently penetrated epidermal cells of transgenic leaves and caused severe disease symptoms presumably from a deficiency in unidentified antifungal compounds. The contribution of antimicrobial components to resistance to penetration and later colonization may vary depending on the pathogen species, suggesting that sesquiterpene cyclase-mediated compounds participate in pre-invasive resistance to necrotrophic pathogen A. solani and post-invasive resistance to hemibiotrophic pathogen P. infestans.     

Arabidopsis non-host resistance PSS30 gene enhances broad-spectrum disease resistance in the soybean cultivar Williams 82

Non-host resistance (NHR), which protects all members of a plant species from non-adapted or non-host plant pathogens, is the most common form of plant immunity. NHR provides the most durable and robust form of broad-spectrum immunity against non-adaptive pathogens pathogenic to other crop species. In a mutant screen for loss of Arabidopsis (Arabidopsis thaliana) NHR against the soybean (Glycine max (L.) Merr.) pathogen Phytophthora sojae, the Phytophthora sojae-susceptible 30 (pss30) mutant was identified. The pss30 mutant is also susceptible to the soybean pathogen Fusarium virguliforme. PSS30 encodes a folate transporter, AtFOLT1, which was previously localized to chloroplasts and implicated in the transport of folate from the cytosol to plastids. We show that two Arabidopsis folate biosynthesis mutants with reduced folate levels exhibit a loss of non-host immunity against P. sojae. As compared to the wild-type Col-0 ecotype, the steady-state folate levels are reduced in the pss1, atfolt1 and two folate biosynthesis mutants, suggesting that folate is required for non-host immunity. Overexpression of AtFOLT1 enhances immunity of transgenic soybean lines against two serious soybean pathogens, the fungal pathogen F. virguliforme and the soybean cyst nematode (SCN) Heterodera glycines. Transgenic lines showing enhanced SCN resistance also showed increased levels of folate accumulation. This study thus suggests that folate contributes to non-host plant immunity and that overexpression of a non-host resistance gene could be a suitable strategy for generating broad-spectrum disease resistance in crop plants.     

Antagonistic roles of SEPALLATA3, FT and FLC genes as targets of the polycomb group gene CURLY LEAF

In Arabidopsis, mutations in the Pc-G gene CURLY LEAF (CLF) give early flowering plants with curled leaves. This phenotype is caused by mis-expression of the floral homeotic gene AGAMOUS (AG) in leaves, so that ag mutations largely suppress the clf phenotype. Here, we identify three mutations that suppress clf despite maintaining high AG expression. We show that the suppressors correspond to mutations in FPA and FT, two genes promoting flowering, and in SEPALLATA3 (SEP3) which encodes a co-factor for AG protein. The suppression of the clf phenotype is correlated with low SEP3 expression in all case and reveals that SEP3 has a role in promoting flowering in addition to its role in controlling floral organ identity. Genetic analysis of clf ft mutants indicates that CLF promotes flowering by reducing expression of FLC, a repressor of flowering. We conclude that SEP3 is the key target mediating the clf phenotype, and that the antagonistic effects of CLF target genes masks a role for CLF in promoting flowering.     

The genetics of stamenoid petal production in oilseed rape (Brassica napus) and equivalent variation in Arabidopsis thaliana

 The agronomic potential of a Brassica napus variant with petalless flowers was compromised by an associated detrimental change in leaf morphology. Genetic analysis demonstrated the cosegregation of genes controlling both morphologies. Two STAP loci controlling the production of flowers with stamenoid petals were mapped to homoeologous locations in the genome of B. napus. The STAP loci were probably duplicate genes because they exhibited an epistatic interaction such that only plants homozygous for recessive stap alleles at both loci expressed the variant phenotype. The CURLY LEAF (CLF) gene of Arabidopsis thaliana pleiotropically influences both flower and leaf morphologies. The cloned CLF gene of Arabidopsis was homologous to a polymorphic B. napus locus coincident with one of the B. napus STAP loci. The possibility that CLF is a candidate gene for STAP suggests that the variant stap alleles of B. napus exert pleiotropic effects over both flower and leaf morphologies. Received: 26 August 1996 / Accepted: 20 September 1996     

The Arabidopsis homologs of trithorax (ATX1) and enhancer of zeste (CLF) establish 'bivalent chromatin marks' at the silent AGAMOUS locus

Tightly balanced antagonism between the Polycomb group (PcG) and the Trithorax group (TrxG) complexes maintain Hox expression patterns in Drosophila and murine model systems. Factors belonging to the PcG/TrxG complexes control various processes in plants as well but whether they participate in mechanisms that antagonize, balance or maintain each other's effects at a particular gene locus is unknown. CURLY LEAF (CLF), an Arabidopsis homolog of enhancer of zeste (EZ) and the ARABIDOPSIS HOMOLOG OF TRITHORAX (ATX1) control the expression of the flower homeotic gene AGAMOUS (AG). Disrupted ATX1 or CLF function results in misexpression of AG, recognizable phenotypes and loss of H3K4me3 or H3K27me3 histone H3-tail marks, respectively. A novel idea suggested by our results here, is that PcG and TrxG complexes function as a specific pair generating bivalent chromatin marks at the silent AG locus. Simultaneous loss of ATX1 and CLF restored AG repression and normalized leaf phenotypes. At the molecular level, disrupted ATX1 and CLF functions did not lead to erasure of the CLF- and ATX1-generated epigenetic marks, as expected: instead, in the double mutants, H3K27me3 and H3K4me3 tags were partially restored. We demonstrate that ATX1 and CLF physically interact linking mechanistically the observed effects.     

Evaluation of foliar fungus‐mediated interactions with below and aboveground enemies of the invasive plant Ageratina adenophora

Plant–fungal associations are frequently key drivers of plant invasion success. Foliar fungi can benefit their invasive hosts by enhancing growth promotion, disease resistance and environmental stress tolerance. However, the roles of foliar fungi may vary when a given invasive plant faces different stresses. In this study, we designed three independent experiments to evaluate the effects of a foliar fungus, Colletotrichum sp., on the growth performance of the invasive plant Ageratina adenophora under different soil conditions, as well as the responses of A. adenophora to the foliar fungal pathogen Diaporthe helianthi and to herbivory. We found that the soil type was the most influential factor for the growth of A. adenophora. The role of the foliar fungus Colletotrichum sp. varied in the different soil types but generally adversely affected leaf development in A. adenophora. Colletotrichum sp. may be a weak latent foliar pathogen that can enhance the pathogenicity of D. helianthi on leaves of A. adenophora and marginally reduce signs of herbivory by natural insects in the wild on A. adenophora seedlings. In general, the benefits of the foliar fungus Colletotrichum to the fitness of A. adenophora are not significant in the context of this experimental design. However, our data highlight the need to consider both aboveground and belowground biota in different soil habitats when evaluating the effects of foliar fungi.     

IDL6‐HAE/HSL2 impacts pectin degradation and resistance to Pseudomonas syringae pv tomato DC3000 in Arabidopsis leaves

Plant cell walls undergo dynamic structural and chemical changes during plant development and growth. Floral organ abscission and lateral root emergence are both accompanied by cell‐wall remodeling, which involves the INFLORESCENCE DEFICIENT IN ABSCISSION (IDA)‐derived peptide and its receptors, HAESA (HAE) and HAESA‐LIKE2 (HSL2). Plant cell walls also act as barriers against pathogenic invaders. Thus, the cell‐wall remodeling during plant development could have an influence on plant resistance to phytopathogens. Here, we identified IDA‐like 6 (IDL6), a gene that is prominently expressed in Arabidopsis leaves. IDL6 expression in Arabidopsis leaves is significantly upregulated when the plant is suffering from attacks of the bacterial Pseudomonas syringae pv. tomato (Pst) DC3000. IDL6 overexpression and knockdown lines respectively decrease and increase the Arabidopsis resistance to Pst DC3000, indicating that the gene promotes the Arabidopsis susceptibility to Pst DC3000. Moreover, IDL6 promotes the expression of a polygalacturonase (PG) gene, ADPG2, and increases PG activity in Arabidopsis leaves, which in turn reduces leaf pectin content and leaf robustness. ADPG2 overexpression restrains Arabidopsis resistance to Pst DC3000, whereas ADPG2 loss‐of‐function mutants increase the resistance to the bacterium. Pst DC3000 infection elevates the ADPG2 expression partially through HAE and HSL2. Taken together, our results suggest that IDL6‐HAE/HSL2 facilitates the ingress of Pst DC3000 by promoting pectin degradation in Arabidopsis leaves, and Pst DC3000 might enhance its infection by manipulating the IDL6‐HAE/HSL2‐ADPG2 signaling pathway.     

Pathogen-associated molecular pattern recognition rather than development of tissue necrosis contributes to bacterial induction of systemic acquired resistance in Arabidopsis

Systemic acquired resistance (SAR) is usually described as a phenomenon whereby localized inoculation with a necrotizing pathogen renders a plant more resistant to subsequent pathogen infection. Here we show that Pseudomonas syringae strains for which Arabidopsis thaliana represents a non-host plant systemically elevate resistance although the underlying interactions neither trigger a hypersensitive response nor cause necrotic disease symptoms. A similar enhancement of systemic resistance was observed when elicitor-active preparations of two typical bacterial pathogen-associated molecular patterns (PAMPs), flagellin and lipopolysaccharides (LPS), were applied in a localized manner. Several lines of evidence indicate that the observed systemic resistance responses are identical to SAR. Localized applications of non-adapted bacteria, flagellin or LPS elevate levels of the SAR regulatory metabolite salicylic acid (SA) and pathogenesis-related (PR) gene expression not only in treated but also in distant leaves. All treatments also systemically increase expression of the SAR marker gene FLAVIN-DEPENDENT MONOOXYGENASE 1. Further, a whole set of SAR-deficient Arabidopsis lines, including mutants in SA biosynthesis and signalling, are impaired in establishing the systemic resistance response triggered by non-host bacteria or PAMPs. We also show that the magnitude of defence reactions such as SA accumulation, PR gene expression or camalexin accumulation induced at sites of virulent or avirulent P. syringae inoculation but not the extent of tissue necrosis during these interactions determines the extent of SAR in distant leaves. Our data indicate that PAMPs significantly contribute to SAR initiation in Arabidopsis and that tissue necroses at inoculation sites are dispensable for SAR activation.     

Leaf buds, a factor in host selection by Battus philenor butterflies

ABSTRACT.

• 1 Field and laboratory experiments identified a character intrinsic to Aristolochia reticulata Nutt. host plants, the terminal leaf bud, that is involved in host-selection behaviour by female pipevine swallowtail butterflies (Battus philenor L.) searching for oviposition sites.
• 2 In the field, the frequency with which females landed on non-host buds declined seasonally as the proportion of host foliage that consisted of buds decreased. Female butterflies did not land on non-host species in proportion to their abundance; rather, females landed on those non-host species whose buds resembled those of A.reticulata.
• 3 A.reticulata plants whose terminal leaf bud was concealed by plastic tape were less susceptible to oviposition in the field than were control plants.
• 4 Female butterflies released in a large, outdoor enclosure were conditioned to search for leaf buds only when exposed to a host species bearing a prominent terminal leaf bud.
• 5 The significance of conditioning of leaf-bud searching behaviour is discussed with respect to discrimination between hosts and non-hosts, between host species, and among plants within a host species.

     

Why species delimitation matters for fungal ecology: Colletotrichum diversity on wild and cultivated cashew in Brazil

Anthracnose is one of the most important plant diseases globally, occurring on a wide range of cultivated and wild host species. This study aimed to identify the Colletotrichum species associated with cashew anthracnose in Brazil, determine their phylogenetic relationships and geographical distribution, and provide some insight into the factors that may be influencing community composition. Colletotrichum isolates collected from symptomatic leaves, stems, inflorescences, and fruit of cultivated and wild cashew, across four Brazilian biomes, were identified as Colletotrichum chrysophilum, Colletotrichum fragariae, Colletotrichum fructicola, Colletotrichum gloeosporioides sensu stricto, Colletotrichum queenslandicum, Colletotrichum siamense and Colletotrichum tropicale. Colletotrichum siamense was the most dominant species. The greatest species richness was associated with cultivated cashew; leaves harbored more species than the other organs; the Atlantic Forest encompassed more species than the other biomes; and Pernambuco was the most species-rich location. However, accounting for the relative abundance of Colletotrichum species and differences in sample size across strata, the interpretation of which community is most diverse depends on how species are delimited. The present study provides valuable information about the Colletotrichum/cashew pathosystem, sheds light on the causal agents identification,and highlights the impact that species delimitation can have on ecological studies of fungi.