bluuebunny/crossref_metadata_2025 huggingface dataset:
DctBD-Dependent and -Independent Expression of the Sinorhizobium (Rhizobium) meliloti C4-Dicarboxylate Transport Gene (dctA) During Symbiosis
Authors: Boesten Bert , Batut Jacques , Boistard Pierre
Published: May 2007

The Sinorhizobium meliloti C4-dicarboxylate transport gene (dctA) is essential for symbiotic nitrogen fixation. Under free-living conditions, the expression of dctA is fully dependent on the cognate regulatory genes dctBD. However, during symbiosis with the Medicago sativa host plant, the dctA gene is efficiently expressed even in the absence of the dctBD genes. The spatial expression of the dctA gene has been monitored in situ in mature nitrogen-fixing nodules formed by wild-type and dctD mutant strains. In nodules induced by a wild-type strain, expression was observed in both the infection zone and the nitrogen-fixing zone of the nodule. DctD-independent expression of dctA was observed with a previously described dctA∷lacZ fusion (pCU700) and was found to be confined to the fixation zone (zone III) of mature nodules. Therefore, the operation of the alternative system of symbiotic dctA activation (ASA) is concomitant with the onset of nitrogen fixation, which could be consistent with an increased need for transport of (C)4(-dicarboxylic acids by the nitrogen-fixing) bacteroids. Sequences in the 5′ part of the dctA coding region were found to be essential for the activation of the dctA∷lacZ gene fusions by the ASA. Deletion of these sequences resulted in gene fusions that were found to be strictly dependent on dctBD for expression, under all conditions tested including symbiosis. Such gene fusions allowed us to establish that the DctBD-dependent dctA expression was occurring throughout the whole nodule.
Four Classes of Salicylate-Induced Tobacco Genes
Authors: Horvath Diana M. , Huang Dorothy J. , Chua Nam-Hai
Published: May 2007

We have identified and characterized fragments of 15 salicylic acid (SA) early response genes. The kinetics of induction and response to cycloheximide (CHX) treatment allowed classification of genes into four groups. Classes I–III are characterized by immediate-early responses, showing increased accumulation of mRNA within 30 min of SA treatment. Moreover, CHX did not block induction of these genes, indicating that latent cellular factors mediate the SA response. Class IV genes were induced more slowly, but still within 2 to 3 h of SA treatment, and required protein synthesis for expression. Although identified in this study as SA-responsive genes, several could also be induced by other compounds. Two genes were characterized in more detail, including isolation of cDNA sequences and additional analysis of gene expression. Sequence analysis revealed that the class I gene, C18-1, is the previously identified ethylene response element binding protein 1 (EREBP1), an ethylene-induced transcription factor for basic pathogenesis-related (PR) genes, whereas the class III gene, G8-1, is a novel sequence. G8-1 was found to be strongly induced only by SA and its active analogs and was exquisitely sensitive to low SA concentrations. These and other genes were found to be activated at early times following tobacco mosaic virus infection of resistant tobacco genotypes.
Wheat Genes Encoding Two Types of PR-1 Proteins Are Pathogen Inducible, but Do Not Respond to Activators of Systemic Acquired Resistance
Authors: Molina Antonio , Görlach Jörn , Volrath Sandra , Ryals John
Published: May 2007

Wheat cDNAs that encode proteins PR-1.1 and PR-1.2 were cloned. Deduced amino acid sequences were homologous to those of pathogen-induced, basic PR-1 proteins from plants. Although expression of PR1.1 and PR1.2 genes was induced upon infection with either compatible or incompatible isolates of the fungal pathogen Erysiphe graminis, these genes did not respond to activators of systemic acquired resistance (SAR), such as salicylic acid (SA), benzothiadiazole (BTH), or isonicotinic acid (INA)
The Plant-Growth-Promoting Rhizobacterium Paenibacillus polymyxa Induces Changes in Arabidopsis thaliana Gene Expression: A Possible Connection Between Biotic and Abiotic Stress Responses
Authors: Timmusk Salme , Wagner E. Gerhart H.
Published: May 2007

This paper addresses changes in plant gene expression induced by inoculation with plant-growth-promoting rhizobacteria (PGPR). A gnotobiotic system was established with Arabidopsis thaliana as model plant, and isolates of Paenibacillus polymyxa as PGPR. Subsequent challenge by either the pathogen Erwinia carotovora (biotic stress) or induction of drought (abiotic stress) indicated that inoculated plants were more resistant than control plants. With RNA differential display on parallel RNA preparations from P. polymyxa- treated or untreated plants, changes in gene expression were investigated. From a small number of candidate sequences obtained by this approach, one mRNA segment showed a strong inoculation-dependent increase in abundance. The corresponding gene was identified as ERD15, previously identified to be drought stress responsive. Quantification of mRNA levels of several stress-responsive genes indicated that P. polymyxa induced mild biotic stress. This suggests that genes and/or gene classes associated with plant defenses against abiotic and biotic stress may be co-regulated. Implications of the effects of PGPR on the induction of plant defense pathways are discussed.
BWMK1, a Novel MAP Kinase Induced by Fungal Infection and Mechanical Wounding in Rice
Authors: He Chaozu , Fong Steven Haw Tien , Yang Daichang , Wang Guo-Liang
Published: May 2007

The activation of the mitogen-activated protein (MAP) kinases by different environmental stresses has been previously observed in several dicot plant species. Here, we report the isolation of a novel MAP kinase in rice that is induced during infection by the blast fungus Magnaporthe grisea or upon mechanical wounding. The gene is designated as BWMK1 for blast- and wound-induced MAP kinase. The cDNA of BWMK1 was isolated from rice leaves challenged by the blast pathogen. Transcripts of the corresponding gene accumulated in rice leaves 4 h after blast inoculation and 30 min after mechanical wounding. This gene encodes a 506 amino acid protein that contains a new dual-phosphorylation activation motif TDY and about 150 unique amino acids on its C terminus. In-gel kinase activity and immunoprecipitation assays confirmed that BWMK1 is a functional MAP kinase. These results show that BWMK1 is a new member of the plant MAP kinase family and may mediate both defense and wound signaling in rice.
The 6K2 Protein and the VPg of Potato Virus A Are Determinants of Systemic Infection in Nicandra physaloides
Authors: Rajamäki Minna-Liisa , Valkonen Jari P. T.
Published: May 2007

Infection with the isolate PVA-M of potato virus A (PVA; genus Potyvirus) is restricted to the inoculated leaves of Nicandra physaloides (Solanaceae), whereas the isolate PVA-B11 infects plants systemically by 10 days post inoculation. Resistance to systemic infection was shown to develop during plant growth. A recombinant virus (B11-M) in which a 1,208-nucleotide sequence of the full-length cDNA clone of PVA-B11 was replaced with the corresponding sequence from PVA-M displayed a phenotype similar to that of PVA-M. The replaced sequence contained four amino acid differences between the two isolates: one in the 6K2 protein and three in the viral genome-linked protein (VPg). Site-directed mutagenesis of the cDNA clones and inoculation of the mutants to N. physaloides indicated that the amino acid substitutions of Met5Val in the 6K2 protein or Leu185Ser in the VPg permitted vascular movement and systemic infection. However, resistance was only partially overcome by these changes, since systemic infection proceeded at a slower rate than with PVA-B11. The amino acid substitution Val116Met in the VPg alone was sufficient to overcome resistance and recover the phenotype of the isolate PVA-B11. These data indicated that both the 6K2 protein and the VPg were avirulence determinants of PVA-M in N. physaloides and suggested a possibly coordinated function of them in the vascular movement of PVA.
Restoration of Wild-Type Virus by Double Recombination of Tombusvirus Mutants with a Host Transgene
Authors: Borja Marise , Rubio Teresa , Scholthof Herman B. , Jackson Andrew O.
Published: May 2007

Nicotiana benthamiana plants transformed with the coat protein gene of tomato bushy stunt virus (TBSV) failed to elicit effective virus resistance when inoculated with wild-type virus. Subsequently, R1 and R2 progeny from 13 transgenic lines were inoculated with a TBSV mutant containing a defective coat protein gene. Mild symptoms typical of those elicited in nontransformed plants infected with the TBSV mutant initially appeared. However, within 2 to 4 weeks, up to 20% of the transgenic plants sporadically began to develop the lethal syndrome characteristic of wild-type virus infections. RNA hybridization and immunoblot analyses of these plants and nontransformed N. benthamiana inoculated with virus from the transgenic lines indicated that wild-type virus had been regenerated by a double recombination event between the defective virus and the coat protein transgene. Similar results were obtained with a TBSV deletion mutant containing a nucleotide sequence marker, and with a chimeric cucumber necrosis virus (CNV) containing the defective TBSV coat protein gene. In both cases, purified virions contained wild-type TBSV RNA or CNV chimeric RNA derived by recombination with the transgenic coat protein mRNA. These results thus demonstrate that recombinant tombusviruses can arise frequently from viral genes expressed in transgenic plants.
Novel Genes Induced During an Arbuscular Mycorrhizal (AM) Symbiosis Formed Between Medicago truncatula and Glomus versiforme
Authors: van Buuren Marianne L. , Maldonado-Mendoza Ignacio E. , Trieu Anthony T. , Blaylock Laura A. , Harrison Maria J.
Published: May 2007

Many terrestrial plant species are able to form symbiotic associations with arbuscular mycorrhizal fungi. Here we have identified three cDNA clones representing genes whose expression is induced during the arbuscular mycorrhizal symbiosis formed between Medicago truncatula and an arbuscular mycorrhizal fungus, Glomus versiforme. The three clones represent M. truncatula genes and encode novel proteins: a xyloglucan endotransglycosylaserelated protein, a putative arabinogalactan protein (AGP), and a putative homologue of the mammalian p110 subunit of initiation factor 3 (eIF3). These genes show little or no expression in M. truncatula roots prior to formation of the symbiosis and are significantly induced following colonization by G. versiforme. The genes are not induced in roots in response to increases in phosphate. This suggests that induction of expression during the symbiosis is due to the interaction with the fungus and is not a secondary effect of improved phosphate nutrition. In situ hybridization revealed that the putative AGP is expressed specifically in cortical cells containing arbuscules. The identification of two mycorrhiza-induced genes encoding proteins predicted to be involved in cell wall structure is consistent with previous electron microscopy data that indicated major alterations in the extracellular matrix of the cortical cells following colonization by mycorrhizal fungi.
Mutation in GDP-Fucose Synthesis Genes of Sinorhizobium fredii Alters Nod Factors and Significantly Decreases Competitiveness to Nodulate Soybeans
Authors: Lamrabet Youssef , Bellogín Ramón A. , Cubo Teresa , Espuny Rosario , Gil Antonio , Krishnan Hari B. , Megias Manuel , Ollero Francisco Javier , Pueppke Steven G. , Ruiz-Sainz José E. , Spaink Herman P. , Tejero-Mateo Pilar , Thomas-Oates Jane , Vinardell José M.
Published: May 2007

We mutagenized Sinorhizobium fredii HH103-1 with Tn5- B20 and screened about 2,000 colonies for increased β-galactosidase activity in the presence of the flavonoid naringenin. One mutant, designated SVQ287, produces lipochitooligosaccharide Nod factors (LCOs) that differ from those of the parental strain. The nonreducing N-acetylglucosamine residues of all of the LCOs of mutant SVQ287 lack fucose and 2-O-methylfucose substituents. In addition, SVQ287 synthesizes an LCO with an unusually long, C20:1 fatty acyl side chain. The transposon insertion of mutant SVQ287 lies within a 1.1-kb HindIII fragment. This and an adjacent 2.4-kb HindIII fragment were sequenced. The sequence contains the 3′ end of noeK, nodZ, and noeL (the gene interrupted by Tn5-B20), and the 5′ end of nolK, all in the same orientation. Although each of these genes has a similarly oriented counterpart on the symbiosis plasmid of the broad-host-range Rhizobium sp. strain NGR234, there are significant differences in the noeK/nodZ intergenic region. Based on amino acid sequence homology, noeL encodes GDP-D-mannose dehydratase, an enzyme involved in the synthesis of GDP-Lfucose, and nolK encodes a NAD-dependent nucleotide sugar epimerase/dehydrogenase. We show that expression of the noeL gene is under the control of NodD1 in S. fredii and is most probably mediated by the nod box that precedes nodZ. Transposon insertion into noeL has two impacts on symbiosis with Williams soybean: nodulation rate is reduced slightly and competitiveness for nodulation is decreased significantly. Mutant SVQ287 retains its ability to form nitrogen-fixing nodules on other legumes, but final nodule number is attenuated on Cajanus cajan.
Comparison of Characteristics of the nodX Genes from Various Rhizobium leguminosarum Strains
Authors: Ovtsyna Alexandra O. , Rademaker Geert-Jan , Esser Edwin , Weinman Jeremy , Rolfe Barry G. , Tikhonovich Igor A. , Lugtenberg Ben J. J. , Thomas-Oates Jane E. , Spaink Herman P.
Published: May 2007

We have analyzed the nucleotide sequences of the nodX genes from two strains of Rhizobium leguminosarum bv. viciae able to nodulate Afghan peas (strains A1 and Himalaya) and from two strains of R. leguminosarum bv. trifolii (ANU843 and CSF). The nodX genes of strains A1 and ANU843 were shown to be functional for the induction of nodules on Afghan peas. To analyze the cause of phenotypic differences of strain A1 and strain TOM we have studied the composition of the lipochitin-oligosaccharides (LCOs) produced by strain A1 after induction by the flavonoid naringenin or various pea root exudates. The structural analysis of the LCOs by mass spectrometry revealed that strain A1 synthesizes a family of at least 23 different LCOs. The use of exudates instead of naringenin resulted only in quantitative differences in the ratios of various LCOs produced.
Aspartate Aminotransferase in Alfalfa Nodules: Localization of mRNA During Effective and Ineffective Nodule Development and Promoter Analysis
Authors: Yoshioka Hirofumi , Gregerson Robert G. , Samac Deborah A. , Hoevens Kim C. M. , Trepp Gian , Gantt J. Stephen , Vance Carroll P.
Published: May 2007

Aspartate aminotransferase (AAT) plays a critical role in the assimilation of symbiotically fixed nitrogen into aspartate and asparagine in legume root nodules. The enzyme occurs as a cytosolic form (AAT1) and a plastid form (AAT2) in alfalfa nodules. To elucidate the functional role of each isozyme in root nodule metabolism further, in situ hybridization was used to determine the pattern of transcript accumulation from the two genes. AAT2 transcripts were localized to infected cells throughout the symbiotic zone of effective alfalfa nodules; however, expression was reduced in ineffective nodules. The AAT1 gene was expressed in the uninfected cells of the invasion zone and symbiotic zone, the nodule parenchyma, and nodule vascular bundles of both effective and ineffective nodules. The AAT1 and AAT2 promoters were evaluated in transgenic alfalfa plants containing promoter β-glucuronidase (GUS) gene fusions. Histochemical staining patterns agreed with results from in situ localization. The distribution pattern of gene transcripts suggests that AAT1 has a role in maintenance of the O2diffusion barrier in nodules and that AAT2 plays a major role in assimilation of recently fixed nitrogen. Promoter deletion analysis of the AAT2 promoter revealed that nodule-specific expression was retained in a promoter fragment of 300 bp.
Colletotrichum trifoliiMutants Disrupted in the Catalytic Subunit of cAMP-Dependent Protein Kinase Are Nonpathogenic
Authors: Yang Zhonghui , Dickman Martin B.
Published: May 2007

Colletotrichum trifolii is the fungal pathogen of alfalfa that causes anthracnose disease. For successful plant infection, this fungus must undergo a series of morphological transitions following conidial attachment, including germination and subsequent differentiation, resulting in appressorium formation. Our previous studies with pharmacological effectors of signaling pathways have suggested the involvement of cyclic AMP (cAMP)-dependent protein kinase (PKA) during these processes. To more precisely evaluate the role of PKA in C. trifolii morphogenesis, the gene encoding the catalytic (C) subunit of PKA (Ct-PKAC) was isolated, sequenced, and inactivated by gene replacement. Southern blot analysis with C. trifolii genomic DNA suggested that Ct-PKAC is a single-copy gene. Northern (RNA) blot analysis with total RNA from different fungal growth stages indicated that the expression of this gene was developmentally regulated. When Ct-PKAC was insertionally inactivated by gene replacement, the transformants showed a small reduction in growth relative to the wild type and conidiation patterns were altered. Importantly, PKA-deficient strains were unable to infect intact alfalfa (host) plants, though only a slight delay was observed in the timing for conidial germination and appressorial formation in the Ct-PKAC disruption mutants. Moreover, these mutants were able to colonize host tissues following artificial wounding, resulting in typical anthracnose disease lesions. Coupled with microscopy, these data suggest that the defect in pathogenicity is likely due to a failure in penetration. Our results demonstrate that PKA has an important role in regulating the transition between vegetative growth and conidiation, and is essential for pathogenic development in C. trifolii.
The Ambivalence of the Barley Mlo Locus: Mutations Conferring Resistance Against Powdery Mildew (Blumeria graminis f. sp. hordei) Enhance Susceptibility to the Rice Blast Fungus Magnaporthe grisea
Authors: Jarosch Birgit , Kogel Karl-Heinz , Schaffrath Ulrich
Published: May 2007

Recessive alleles of the barley Mlo locus confer non-race-specific resistance against the powdery mildew fungus Blumeria graminis f. sp. hordei (Bgh). Recently the Mlo gene has been isolated and it was suggested that the Mlo product is a negative regulator of cell death. Thus, loss of function can precondition cells to a higher responsiveness for the onset of multiple defense functions. Here, we document an enhanced susceptibility of barley mlo mutants to the rice blast fungus Magnaporthe grisea. The disease phenotype is independent of the barley cultivar in which the mlo allele has been introgressed and occurs in equal amounts in barley backcross lines of cv. Ingrid carrying the mlo-1, mlo-3, or mlo-5 allele. Ror genes, which are required for the full expression of mlo resistance in barley against Bgh, do not affect the specific mlo-mediated phenotype observed after M. grisea infection. Formation of an effective papilla restricts blast development in epidermal cells of Mlo plants. In contrast, papillae are mostly penetrated in mlo mutants and, as a consequence, the fungus spreads into adjacent mesophyll cells. Both wild-type plants and mlo mutants did not differ in perception of a purified elicitor derived from M. grisea. Thus, we hypothesize that a functional Mlo protein is a prerequisite for penetration resistance of barley to fungal pathogens like M. grisea. The benefit of mlo alleles for durable resistance in barley and a proposed role of mlo-type-mutations in rice are discussed.
MtENOD20, a Nod Factor-Inducible Molecular Marker for Root Cortical Cell Activation
Authors: Vernoud Vanessa , Journet Etienne-Pascal , Barker David G.
Published: May 2007

The spatio-temporal expression pattern of the Medicago truncatula ENOD20 gene during early stages of nodulation has been analyzed with transgenic alfalfa (M. varia) expressing a pMtENOD20-GUS chimeric fusion. Our results show that transcriptional activation of this gene occurs initially in dividing inner cortical cells corresponding to sites of nodule primordium formation and subsequently in root hairs containing infection threads. Use of Sinorhizobium meliloti nod gene mutants that uncouple nodule organogenesis from infection has confirmed that early MtENOD20 transcription is tightly linked to cortical cell activation (CCA). Furthermore, these experiments have revealed that an S. meliloti nodH mutant, defective in Nod factor sulfation and epidermal cell activation, is nevertheless able to elicit low-level CCA. Purified S. meliloti Nod factors trigger MtENOD20 transcription very rapidly (within 12 to 24 h) in the root cortex, and studies with transgenic alfalfa show that Nod factors are able to elicit gene expression coupled to CCA at concentrations as low as 10-11 M. Finally, we have made use of a range of synthetic lipo-chitooligosaccharides to show that fatty acid chain length is an important structural parameter in the capacity of Nod factors to elicit CCA. Taken together, our results suggest that pMtENOD20-GUS transgenic lines should prove valuable tools in future studies of Rhizobium and Nod factor-elicited CCA.
The Rate of Cell-to-Cell Movement in Squash of Cucumber Mosaic Virus Is Affected by Sequences of the Capsid Protein
Authors: Wong Sek-Man , Thio Sharon Swee-Chin , Shintaku Michael H. , Palukaitis Peter
Published: May 2007

The M strain of cucumber mosaic virus (CMV) does not infect squash plants systemically and moves very slowly in inoculated cotyledons. Systemic infection and an increase in the rate of local movement were observed when amino acids 129 or 214 of the M-CMV capsid protein (CP) were altered to those present in the Fny strain of CMV. While the opposite alterations to the CP of Fny-CMV inhibited systemic infection of squash, they did not show the same effects on the rates of both cell-to-cell and long-distance movement. However, the ability of CMV to infect squash systemically was affected by the rate of cell-to-cell movement.
Insertional Mutagenesis and Cloning of the Genes Required for Biosynthesis of the Host-Specific AK-Toxin in the Japanese Pear Pathotype of Alternaria alternata
Authors: Tanaka Aiko , Shiotani Hiroshi , Yamamoto Mikihiro , Tsuge Takashi
Published: May 2007

The Japanese pear pathotype of Alternaria alternata causes black spot of Japanese pear by producing a host-specific toxin known as AK-toxin. Restriction enzyme-mediated integration (REMI) mutagenesis was used to tag genes required for toxin biosynthesis. Protoplasts of a wild-type strain were treated with a linearized plasmid along with the restriction enzyme used to linearize the plasmid. Of 984 REMI transformants recovered, three produced no detectable AK-toxin and lost pathogenicity on pear leaves. Genomic DNA flanking the integrated plasmid was recovered from one of the mutants. With the recovered DNA used as a probe, a cosmid clone of the wild-type strain was isolated. Structural and functional analyses of an 8.0-kb region corresponding to the tagged site indicated the presence of two genes. One, designated AKT1, encodes a member of the class of carboxyl-activating enzymes. The other, AKT2, encodes a protein of unknown function. The essential roles of these two genes in both AK-toxin production and pathogenicity were confirmed by transformation-mediated gene disruption experiments. DNA gel blot analysis detected AKT1 and AKT2 homologues not only in the Japanese pear pathotype strains but also in strains from the tangerine and strawberry pathotypes. The host-specific toxins of these two pathotypes are similar in structure to AK-toxin. Homologues were not detected in other pathotypes or in non-pathogenic strains of A. alternata, suggesting acquisition of AKT1 and AKT2 by horizontal transfer.
syrM Is Involved in the Determination of the Amount and Ratio of the Two Forms of the Acidic Exopolysaccharide EPSI in Rhizobium meliloti
Authors: Dusha Ilona , Oláh Boglárka , Szegletes Zsolt , Erdei László , Kondorosi Ádám
Published: May 2007

For the invasion of root nodule cells of alfalfa by its symbiotic partner Rhizobium meliloti, the bacterial succinoglycan (EPSI) is required. Nitrogen starvation of R. meliloti results in a considerable increase of EPSI production. We have shown that the presence of nitrogen also altered the ratio of high and low molecular weight (HMW and LMW, respectively) forms of EPSI, favoring the production of LMW molecules. The transcription of two exo genes (exoF, exoP) was found to be under the control of combined nitrogen. In contrast, exoH and exoK were expressed independently of the nitrogen level. ntrC and syrM, regulatory genes for the nitrogen control of nod gene expression, were shown to be involved in the regulation of EPSI production. In an syrM mutant both the amount and the ratio of the two main forms of EPSI were altered, compared with the wild-type strain: a reduced level of EPSI and the dominance of the LMW form in nitrogen-starved cultures, while an increased level of EPSI and the accumulation of the HMW fraction in nitrogen-containing samples was observed. An enhanced expression level of the exoK gene and the higher ratio of the LMW form to the total EPSI in nitrogen-free samples of the syrM mutant suggest a regulatory role for syrM in exoK expression and a contribution of exoK to the accumulation of the LMW form of EPSI.
Genetic and Physical Mapping of the RPP13 Locus, in Arabidopsis, Responsible for Specific Recognition of Several Peronospora parasitica (Downy Mildew) Isolates
Authors: Bittner-Eddy Peter , Can Canan , Gunn Nick , Pinel Matthieu , Tör Mahmut , Crute Ian , Holub Eric B. , Beynon Jim
Published: May 2007

Fifteen isolates of the biotrophic oomycete Peronospora parasitica (downy mildew) were obtained from a population of Arabidopsis thaliana plants that established naturally in a garden the previous year. They exhibited phenotypic variation in a set of 12 Arabidopsis accessions that suggested that the parasite population consisted of at least six pathotypes. One isolate, Maks9, elicited an interaction phenotype of flecking necrosis and no sporulation (FN) in the Arabidopsis accession Nd-1, and more extensive pitting necrosis with no sporulation (PN) in the accession Ws-2. RPP13 was designated as the locus for a single dominant resistance gene associated with the resistance in Nd-1 and mapped to an interval of approximately 60 kb on a bacterial artificial chromosome (BAC) contig on the lower arm of chromosome 3. This locus is approximately 6 cM telomeric to RPP1, which was previously described as the locus for the PN interaction with five Peronospora isolates, including resistance to Maks9 in Ws-2. New Peronospora isolates were obtained from four other geographically distinct populations of P. parasitica. Four isolates were characterized that elicited an FN phenotype in Nd-1 and mapped resistance to the RPP13 locus. This suggests that the RPP13 locus contains either a single gene capable of multiple isolate recognition or a group of tightly linked genes. Further analysis suggests that the RPP11 gene in the accession Rld-0 may be allelic to RPP13 but results in a different recognition capability.
Casuarina glauca Prenodule Cells Display the Same Differentiation as the Corresponding Nodule Cells
Authors: Laplaze Laurent , Duhoux Emile , Franche Claudine , Frutz Thierry , Svistoonoff Sergio , Bisseling Ton , Bogusz Didier , Pawlowski Katharina
Published: May 2007

Recent phylogenetic studies have implied that all plants able to enter root nodule symbioses with nitrogen-fixing bacteria go back to a common ancestor (D. E. Soltis, P. S. Soltis, D. R. Morgan, S. M. Swensen, B. C. Mullin, J. M. Dowd, and P. G. Martin, Proc. Natl. Acad. Sci. USA, 92:2647–2651, 1995). However, nodules formed by plants from different groups are distinct in nodule organogenesis and structure. In most groups, nodule organogenesis involves the induction of cortical cell divisions. In legumes these divisions lead to the formation of a nodule primordium, while in non-legumes they lead to the formation of a so-called prenodule consisting of infected and un-infected cells. Nodule primordium formation does not involve prenodule cells, and the function of prenodules is not known. Here, we examine the differentiation of actinorhizal prenodule cells in comparison to nodule cells with regard to both symbionts. Our findings indicate that prenodules represent primitive symbiotic organs whose cell types display the same characteristics as their nodule counterparts. The results are discussed in the context of the evolution of root nodule symbioses.
Characterization of a Casuarina glauca Nodule-Specific Subtilisin-like Protease Gene, a Homolog of Alnus glutinosa ag12
Authors: Laplaze Laurent , Ribeiro Ana , Franche Claudine , Duhoux Emile , Auguy Florence , Bogusz Didier , Pawlowski Katharina
Published: May 2007

In search of plant genes expressed during early interactions between Casuarina glauca and Frankia, we have isolated and characterized a C. glauca gene that has strong homology to subtilisin-like protease gene families of several plants including the actinorhizal nodulin gene ag12 of another actinorhizal plant, Alnus glutinosa. Based on the expression pattern of cg12 in the course of nodule development, it represents an early actinorhizal nodulin gene. Our results suggest that subtilisin-like proteases may be a common element in the process of infection of plant cells by Frankia in both Betulaceae (Alnus glutinosa) and Casuarinaceae (Casuarina glauca) symbioses.
Molecular Cloning and Characterization of a Tobacco MAP Kinase Kinase That Interacts with SIPK
Authors: Liu Yanhong , Zhang Shuqun , Klessig Daniel F.
Published: May 2007

A tobacco MAP kinase termed SIPK (salicylic acid-induced protein kinase) is activated in response to a variety of stress signals, including pathogen attack and wounding (S. Zhang and D. F. Klessig, Proc. Natl. Acad. Sci. USA 95:7225–7230, 1998; S. Zhang and D. F. Klessig, Proc. Natl. Acad. Sci. USA 95:7433–7438, 1998). Using the yeast two-hybrid system, we have identified a gene encoding a protein that interacts with SIPK but not the wounding induced protein kinase (WIPK), which is another tobacco MAP kinase. Sequence analysis indicated that this SIPK-interacting protein is a member of the MAP kinase kinase family; thus, it was named SIPK kinase (SIPKK). Co-immunoprecipitation experiments demonstrated that SIPKK and SIPK interact in vitro. Consistent with its putative function as a kinase, SIPKK phosphorylated myelin basic protein in vitro. Interestingly, SIPKK was induced at the mRNA level after Tobacco mosaic virus (TMV) infection or wounding, albeit with kinetics that are too slow to account for the activation of SIPK following these stimuli.
MucR Is Necessary for Galactoglucan Production in Sinorhizobium meliloti EFB1
Authors: Martín Marta , Lloret Javier , Sánchez-Contreras María , Bonilla Ildefonso , Rivilla Rafael
Published: May 2007

Sinorhizobium meliloti can produce two types of acidic exopolysaccharides, succinoglycan and galactoglucan, that are interchangeable for infection of alfalfa nodules. Strain SU47 and derivatives produce only succinoglycan, unless it grows under phosphate limitation or carries a mutation in either of two regulatory loci, mucR or expR. It has been proposed that MucR acts as a transcriptional repressor that blocks the expression of the exp genes responsible for galactoglucan production. Strain EFB1 simultaneously produces both exopolysaccharides. Heterologous expression of lacZ transcriptional fusions of the expE promoters has shown that genetic background is more important that promoter sequence for exp gene expression, since expE promoters from both strains are expressed at high level in EFB1 and not in SU47. We have found that mucR is present in mucoid and nonmucoid strains, and in EFB1 differs from SU47 in only one conservative amino acid change. MucR proteins from both strains are interchangeable. An mucR mutant of EFB1 cannot produce galactoglucan and does not express mucS.
Specific Binding Sites for an Antifungal Plant Defensin from Dahlia (Dahlia merckii) on Fungal Cells Are Required for Antifungal Activity
Authors: Thevissen Karin , Osborn Rupert W. , Acland David P. , Broekaert Willem F.
Published: May 2007

Dm-AMP1, an antifungal plant defensin from seeds of dahlia (Dahlia merckii), was radioactively labeled with t-butoxycarbonyl-[35S]-L-methionine N-hydroxy-succinimi-dylester. This procedure yielded a 35S-labeled peptide with unaltered antifungal activity. [35S]Dm-AMP1 was used to assess binding on living cells of the filamentous fungus Neurospora crassa and the unicellular fungus Saccharomyces cerevisiae. Binding of [35S]Dm-AMP1 to fungal cells was saturable and could be competed for by preincubation with excess, unlabeled Dm-AMP1 as well as with Ah-AMP1 and Ct-AMP1, two plant defensins that are highly homologous to Dm-AMP1. In contrast, binding could not be competed for by more distantly related plant defensins or structurally unrelated antimicrobial peptides. Binding of [35S]Dm-AMP1 to either N. crassa or S. cerevisiae cells was apparently irreversible. In addition, whole cells and microsomal membrane fractions from two independently obtained S. cerevisiae mutants selected for resistance to Dm-AMP1 exhibited severely reduced binding affinity for [35S]Dm-AMP1, compared with wild-type yeast. This finding suggests that binding of Dm-AMP1 to S. cerevisiae plasma membranes is required for antifungal activity of this protein.
Resistance of Tomato Line Hawaii7996 to Ralstonia solanacearum Pss4 in Taiwan Is Controlled Mainly by a Major Strain-Specific Locus
Authors: Wang Jaw-Fen , Olivier Jocelyne , Thoquet Philippe , Mangin Brigitte , Sauviac Laurent , Grimsley Nigel H.
Published: May 2007

Bacterial wilt caused by the soilborne bacterium Ralstonia solanacearum attacks hundreds of plant species, including many agriculturally important crops. Natural resistance to this disease has been found in some species and is usually inherited as a polygenic trait. In tomato, a model crop plant, genetic analysis previously revealed the involvement of several QTL (quantitative trait loci) controlling resistance and, in all of these studies with different strains of the pathogen, loci on chromosome 6 played the predominant role in controlling this trait. Using quantitative data collected from a greenhouse test F3 population, we identified a new locus on chromosome 12 that appears to be active specifically against a race 1 biovar 3 Pss4 bacterial strain endemic to Taiwan. Chromosome 6 still contributes significantly to the control of the resistance, and weaker associations of the trait to other regions of the genome are observed. These results are discussed in the context of current molecular knowledge about the strain specificity of disease resistance genes.
Contribution of Fungal Loline Alkaloids to Protection from Aphids in a Grass-Endophyte Mutualism
Authors: Wilkinson Heather H. , Siegel Malcolm R. , Blankenship Jimmy D. , Mallory Allison C. , Bush Lowell P. , Schardl Christopher L.
Published: May 2007

Fungal endophytes provide grasses with enhanced protection from herbivory, drought, and pathogens. The loline alkaloids (saturated 1-aminopyrrolizidines with an oxygen bridge) are fungal metabolites often present in grasses with fungal endophytes of the genera Epichloë or Neotyphodium. We conducted a Mendelian genetic analysis to test for activity of lolines produced in plants against aphids feeding on those plants. Though most loline-producing endophytes are asexual, we found that a recently described sexual endophyte, Epichloë festucae, had heritable variation for loline alkaloid expression (Lol+) or non-expression (Lol¯). By analyzing segregation of these phenotypes and of linked DNA polymorphisms in crosses, we identified a single genetic locus controlling loline alkaloid expression in those E. festucae parents. We then tested segregating Lol+ and Lol¯ full-sibling fungal progeny for their ability to protect host plants from two aphid species, and observed that alkaloid expression cosegregated with activity against these insects. The in planta loline alkaloid levels correlated with levels of anti-aphid activity. These results suggested a key role of the loline alkaloids in protection of host plants from certain aphids, and represent, to our knowledge, the first Mendelian analysis demonstrating how a fungal factor contributes protection to plant-fungus mutualism.
Both Induction and Morphogenesis of Cyst Nematode Feeding Cells Are Mediated by Auxin
Authors: Goverse Aska , Overmars Hein , Engelbertink Jan , Schots Arjen , Bakker Jaap , Helder Johannes
Published: May 2007

Various lines of evidence show that local changes in the auxin concentration are involved in the initiation and directional expansion of syncytia induced by cyst nematodes. Analysis of nematode infections on auxin-insensitive tomato and Arabidopsis mutants revealed various phenotypes ranging from complete inhibition of syncytium development to a decrease in hypertrophy and lateral root formation at the infection site. Specific activation of an auxin-responsive promoter confirmed the role of auxin and pointed at a local accumulation of auxin in developing syncytia. Disturbance of auxin gradients by inhibiting polar auxin transport with N-(1-naphthyl)phtalamic acid (NPA) resulted in abnormal feeding cells, which were characterized by extreme galling, massive disordered cell divisions in the cortex, and absence of radial expansion of the syncytium initial toward the vascular bundle. The role of auxin gradients in guiding feeding cell morphogenesis and the cross-talk between auxin and ethylene resulting in a local activation of cell wall degrading enzymes are discussed.
Site-Directed Mutagenesis of the magB Gene Affects Growth and Development in Magnaporthe grisea
Authors: Fang Eric G. C. , Dean Ralph A.
Published: May 2007

G protein signaling is commonly involved in regulating growth and differentiation of eukaryotic cells. We previously identified MAGB, encoding a Gα subunit, from Magnaporthe grisea, and disruption of MAGB led to defects in a number of cellular responses, including appres-sorium formation, conidiation, sexual development, mycelial growth, and surface sensing. In this study, site-directed mutagenesis was used to further dissect the pleiotropic effects controlled by MAGB. Conversion of glycine 42 to arginine was predicted to abolish GTPase activity, which in turn would constitutively activate G protein signaling in magBG42R. This dominant mutation caused autolysis of aged colonies, misscheduled melanization, reduction in both sexual and asexual reproduction, and reduced virulence. Furthermore, magBG42R mutants were able to produce appressoria on both hydrophobic and hydrophilic surfaces, although development on the hydrophilic surface was delayed. A second dominant mutation, magBG203R (glycine 203 converted to arginine), was expected to block dissociation of the Gβγ from the Gα subunit, thus producing a constitutively inactive G protein complex. This mutation did not cause drastic phenotypic changes in the wild-type genetic background, other than increased sensitivity to repression of conidiation by osmotic stress. However, magBG203R is able to complement phenotypic defects in magB mutants. Comparative analyses of the phenotypical effects of different magB mutations are consistent with the involvement of the Gβγ subunit in the signaling pathways regulating cellular development in M. grisea.
TheIGene of Bean: A Dosage-Dependent Allele Conferring Extreme Resistance, Hypersensitive Resistance, or Spreading Vascular Necrosis in Response to the PotyvirusBean common mosaic virus
Authors: Collmer Candace Whitmer , Marston Marcia Fisher , Taylor Jessica C. , Jahn Molly
Published: May 2007

The resistance to the potyvirus Bean common mosaic virus (BCMV) conferred by the I allele in cultivars of Phaseolus vulgaris has been characterized as dominant, and it has been associated with both immunity and a systemic vascular necrosis in infected bean plants under field, as well as controlled, conditions. In our attempts to understand more fully the nature of the interaction between bean with the I resistance allele and the pathogen BCMV, we carefully varied both I allele dosage and temperature and observed the resulting, varying resistance responses. We report here that the I allele in the bean cultivars we studied is not dominant, but rather incompletely dominant, and that the system can be manipulated to show in plants a continuum of response to BCMV that ranges from immunity or extreme resistance, to hypersensitive resistance, to systemic phloem necrosis (and subsequent plant death). We propose that the particular phenotypic outcome in bean results from a quantitative interaction between viral pathogen and plant host that can be altered to favor one or the other by manipulating I allele dosage, temperature, viral pathogen, or plant cultivar.
Effects of Targeted Replacement of the Tomatinase Gene on the Interaction ofSeptoria lycopersiciwith Tomato Plants
Authors: Martin-Hernandez A. M. , Dufresne M. , Hugouvieux V. , Melton R. , Osbourn A.
Published: May 2007

Many plants produce constitutive antifungal molecules belonging to the saponin family of secondary metabolites, which have been implicated in plant defense. Successful pathogens of these plants must presumably have some means of combating the chemical defenses of their hosts. In the oat root pathogen Gaeumannomyces graminis, the saponin-detoxifying enzyme avenacinase has been shown to be essential for pathogenicity. A number of other phytopathogenic fungi also produce saponin-degrading enzymes, although the significance of these for saponin resistance and pathogenicity has not yet been established. The tomato leaf spot pathogen Septoria lycopersici secretes the enzyme tomatinase, which degrades the tomato steroidal glycoalkaloid α-tomatine. Here we report the isolation and characterization of tomatinase-deficient mutants of S. lycopersici following targeted gene disruption. Tomatinase-minus mutants were more sensitive to α-tomatine than the wild-type strain. They could, however, still grow in the presence of 1 mM α-tomatine, suggesting that nondegra-dative mechanisms of tolerance are also important. There were no obvious effects of loss of tomatinase on macroscopic lesion formation on tomato leaves, but trypan blue staining of infected tissue during the early stages of infection revealed more dying mesophyll cells in leaves that had been inoculated with tomatinase-minus mutants. Expression of a defense-related basic β-1,3 glucanase gene was also enhanced in these leaves. These differences in plant response may be associated with subtle differences in the growth of the wild-type and mutant strains during infection. Alternatively, tomatinase may be involved in suppression of plant defense mechanisms.
Xv4-vrxv4: A New Gene-for-Gene Interaction Identified BetweenXanthomonas campestrispv.VesicatoriaRace T3 and the Wild Tomato RelativeLycopersicon pennellii
Authors: Astua-Monge Gustavo , Minsavage Gerald V. , Stall Robert E. , Vallejos C. Eduardo , Davis Michael J. , Jones Jeffrey B.
Published: May 2007

Strains of tomato race 3 (T3) of Xanthomonas campestris pv. vesicatoria elicit a hypersensitive response (HR) in leaves of Lycopersicon pennellii LA716. Geneticsegregation of the resistance exhibited ratios near 3:1 in F2populations, which confirmed that a single dominant gene controlled the inheritance of this trait. With the aid of a collection of introgression lines, restriction fragment length polymorphism, and cleaved amplified polymorphic sequence markers, the resistance locus was located on chromosome 3 between TG599 and TG134. An avirulence gene named avrXv4 was also isolated by mobilizing a total of 600 clones from a genomic DNA library of the T3 strain 91–118 into the X. campestris pv. vesicatoria strain ME90, virulent on L. pennellii. One cosmid clone, pXcvT3-60 (29-kb insert), induced HR in resistant plants. The avirulent phenotype of pXcvT3-60 was confirmed by comparing growth rates in planta and electrolyte leakages among transconjugants carrying a mutated or intact clone with the wild-type T3 strain 91–118. A 1.9-kb DNA fragment contained within a 6.8-kb active subclone was sequenced and was determined to carry an open reading frame of 1,077 bp. The predicted AvrXv4 protein exhibits high similarity to members of an emerging new family of bacterial proteins from plant and mammalian pathogens comprising AvrRxv, AvrBsT, YopJ, YopP, AvrA, and YL4O.
Nitric Oxide Inhibition of Tobacco Catalase and Ascorbate Peroxidase
Authors: Clark Daniel , Durner Jörg , Navarre Duroy A. , Klessig Daniel F.
Published: May 2007

We used a variety of nitric oxide (NO) donors to demonstrate that NO inhibits the activities of tobacco catalase and ascorbate peroxidase (APX). This inhibition appears to be reversible because removal of the NO donor led to a significant recovery of enzymatic activity. In contrast, APX and catalase were irreversibly inhibited by peroxynitrite. The ability of NO and peroxynitrite to inhibit the two major H2O2-scavenging enzymes in plant cells suggests that NO may participate in redox signaling during the activation of defense responses following pathogen attack.
Lipochito-Oligosaccharide Nodulation Factors Stimulate Cytoplasmic Polarity with Longitudinal Endoplasmic Reticulum and Vesicles at the Tip in Vetch Root Hairs
Authors: Miller Deborah D. , Klooster Hetty B. Leferink-ten , Emons Anne Mie C.
Published: May 2007

Vetch root hair development has four stages: bulge, growing, growth terminating, and full-grown hair. In the assay we used, the nodulation factor induced swellings and outgrowths in growth-terminating hairs. Bulges, swellings, and full-grown hairs have transverse endoplasmic reticulum (ER) and no tip-accumulated vesicles. Growing hairs and outgrowths show vesicle accumulation in the tip and longitudinal subapical ER. Bulge walls and walls of swellings appear mottled.
Hypaphorine from the Ectomycorrhizal Fungus Pisolithus tinctorius Counteracts Activities of Indole-3-Acetic Acid and Ethylene but Not Synthetic Auxins in Eucalypt Seedlings
Authors: Ditengou Franck Anicet , Lapeyrie Frédéric
Published: May 2007

Very little is known about the molecules regulating the interaction between plants and ectomycorrhizal fungi during root colonization. The role of fungal auxin in ectomycorrhiza has repeatedly been suggested and questioned, suggesting that, if fungal auxin controls some steps of colonized root development, its activity might be tightly controlled in time and in space by plant and/or fungal regulatory mechanisms. We demonstrate that fungal hypaphorine, the betaine of tryptophan, counteracts the activity of indole-3-acetic acid (IAA) on eucalypt tap root elongation but does not affect the activity of the IAA analogs 2,4-D ((2,4-dichlorophenoxy)acetic acid) or NAA (1-naphthaleneacetic acid). These data suggest that IAA and hypaphorine interact during the very early steps of the IAA perception or signal transduction pathway. Furthermore, while seedling treatment with 1-amincocyclopro-pane-1-carboxylic acid (ACC), the precursor of ethylene, results in formation of a hypocotyl apical hook, hypaphorine application as well as root colonization by Pisolithus tinctorius, a hypaphorine-accumulating ectomycorrhizal fungus, stimulated hook opening. Hypaphorine counteraction with ACC is likely a consequence of hypaphorine interaction with IAA. In most plant-microbe interactions studied, the interactions result in increased auxin synthesis or auxin accumulation in plant tissues. The P. tinctorius / eucalypt interaction is intriguing because in this interaction the microbe down-regulates the auxin activity in the host plant. Hypaphorine might be the first specific IAA antagonist identified.