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Query: EC:3.2.1.15 (pectinase)
 2,440 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The exudates or liquid droplets on various structures of a number of fungi were examined. The droplets were enveloped in membranous material and were associated with actively growing mycelia, including fruiting structures. Osmium tetroxide vapour-fixed droplets of Claviceps purpurea, Myrothecium roridum, Sclerotinia sclerotiorum, Sclerotium rolfsii, and Thanathephorus cucumeris did not dry to a powder but remained intact as spheres when freeze-dried. Fractured spheres, examined with the scanning electron microscope, showed the presence of a membranous structure similar to that of rapidly frozen colloidal solutions with the ice crystals removed by sublimation. Locules or cavities within the freeze-dried droplets are thought to be due to the entrapment of air when droplets coalesce. Biochemical analyses of the exudates showed that acid phosphatase, beta-glucosidase, acid and alkaline protease. RNase polygalacturonase and cellulase enzymes as well as oxalic acid and ammonia were present.
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PMID:Fungal exudates. 72 49

Sclerotinia sclerotiorum acidifies its ambient environment by producing oxalic acid. This production of oxalic acid during plant infection has been implicated as a primary determinant of pathogenicity in this and other phytopathogenic fungi. We found that ambient pH conditions affect multiple processes in S. sclerotiorum. Exposure to increasing alkaline ambient pH increased the oxalic acid accumulation independent of carbon source, sclerotial development was favored by acidic ambient pH conditions but inhibited by neutral ambient pH, and transcripts encoding the endopolygalacturonase gene pg1 accumulated maximally under acidic culture conditions. We cloned a putative transcription factor-encoding gene, pac1, that may participate in a molecular signaling pathway for regulating gene expression in response to ambient pH. The three zinc finger domains of the predicted Pac1 protein are similar in sequence and organization to the zinc finger domains of the A. nidulans pH-responsive transcription factor PacC. The promoter of pac1 contains eight PacC consensus binding sites, suggesting that this gene, like its homologs, is autoregulated. Consistent with this suggestion, the accumulation of pac1 transcripts paralleled increases in ambient pH. Pac1 was determined to be a functional homolog of PacC by complementation of an A. nidulans pacC-null strain with pac1. Our results suggest that ambient pH is a regulatory cue for processes linked to pathogenicity, development, and virulence and that these processes may be under the molecular regulation of a conserved pH-dependent signaling pathway analogous to that in the nonpathogenic fungus A. nidulans.
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PMID:pH signaling in Sclerotinia sclerotiorum: identification of a pacC/RIM1 homolog. 1113 30

The synergistic activities of oxalic acid and endopolygalacturonases are thought to be essential for full virulence of Sclerotinia sclerotiorum and other oxalate-producing plant pathogens. Both oxalic acid production and endopolygalacturonase activity are regulated by ambient pH. Since many gene products with pH-sensitive activities are regulated by the PacC transcription factor in Aspergillus nidulans, we functionally characterized a pacC gene homolog, pac1, from S. sclerotiorum. Mutants with loss-of-function alleles of the pac1 locus were created by targeted gene replacement. In vitro mycelial growth of these pac1 mutants was normal at acidic pH, but growth was inhibited as culture medium pH was increased. Development and maturation of sclerotia in culture was also aberrant in these pac1 replacement mutants. Although oxalic acid production remained alkaline pH-responsive, the kinetics and magnitude of oxalate accumulation were dramatically altered. Additionally, maximal accumulation of endopolygalacturonase gene transcripts (pg1) was shifted to higher ambient pH. Virulence in loss-of-function pac1 mutants was dramatically reduced in infection assays with tomato and Arabidopsis. Based on these results, pac1 appears to be necessary for the appropriate regulation of physiological processes important for pathogenesis and development of S. sclerotiorum.
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PMID:The Sclerotinia sclerotiorum pac1 gene is required for sclerotial development and virulence. 1297 2

The necrotrophic fungal pathogen Sclerotinia sclerotiorum secretes oxalic acid and endo-polygalacturonase (endo-PG) in host plants. Oxalic acid acidifies the plant tissue to values more suitable to endo-PG activity. However, we observed that the infected soybean seedlings possessed a pH of 3.8, which is below that optimal for endo-PG activity (4.5 to 5.0). We investigated, therefore, the effects of pH (from 5.0 to 3.6) and oxalate (5 to 20 mM) on the activity of the major basic endo-PG (PGb) and towards an acidic endo-PG (PGa) secreted by S. sclerotiorum during soybean infection. We verified that only PGb activity is stimulated by oxalate, while at the lowest pH levels, PGa escapes the inhibition of a soybean polygalacturonase-inhibiting protein (PGIP). These results, performed on polygalacturonic acid, were apparently consistent with data obtained from studies with soybean hypocotyl segments, in which PGb activity was increased by oxalate and PGa maintained its activity also at pH 3.6, possibly because at this pH the PGIP contained in the plant tissue is inactive. Reverse transcription-polymerase chain reaction analysis showed that, during soybean infection, the expression of the putative pga gene is delayed in comparison to the basic one. The different temporal expressions of the two endo-PGs and their differing responses to pH, oxalate, and PGIP seem to be consistent with a possible maximization of the fungal PG activity in the host tissue.
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PMID:Relationships among endo-polygalacturonase, oxalate, pH, and plant polygalacturonase-inhibiting protein (PGIP) in the interaction between Sclerotinia sclerotiorum and soybean. 1559 46

Sclerotinia sclerotiorum is unusual among necrotrophic pathogens in its requirement for senescent tissues to establish an infection and to complete the life cycle. A model for the infection process has emerged whereby the pathogenic phase is bounded by saprophytic phases; the distinction being that the dead tissues in the latter are generated by the actions of the pathogen. Initial colonization of dead tissue provides nutrients for pathogen establishment and resources to infect healthy plant tissue. The early pathogenicity stage involves production of oxalic acid and the expression of cell wall degrading enzymes, such as specific isoforms of polygalacturonase (SSPG1) and protease (ASPS), at the expanding edge of the lesion. Such activities release small molecules (oligo-galacturonides and peptides) that serve to induce the expression of a second wave of degradative enzymes that collectively bring about the total dissolution of the plant tissue. Oxalic acid and other metabolites and enzymes suppress host defences during the pathogenic phase, while other components initiate host cell death responses leading to the formation of necrotic tissue. The pathogenic phase is followed by a second saprophytic phase, the transition to which is effected by declining cAMP levels as glucose becomes available and further hydrolytic enzyme synthesis is repressed. Low cAMP levels and an acidic environment generated by the secretion of oxalic acid promote sclerotial development and completion of the life cycle. This review brings together histological, biochemical and molecular information gathered over the past several decades to develop this tri-phasic model for infection. In several instances, studies with Botrytis species are drawn upon for supplemental and supportive evidence for this model. In this process, we attempt to outline how the interplay between glucose levels, cAMP and ambient pH serves to coordinate the transition between these phases and dictate the biochemical and developmental events that define them.
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PMID:Sclerotinia sclerotiorum: when "to be or not to be" a pathogen? 1611 22

Botrytis cinerea is a necrotrophic pathogen that attacks more than 200 plant species. Here, the nonpathogenic mutant A336, obtained via insertional mutagenesis, was characterized. Mutant A336 was nonpathogenic on leaves and fruits, on intact and wounded tissue, while still able to penetrate the host plant. It grew normally in vitro on rich media but its conidiation pattern was altered. The mutant did not produce oxalic acid and exhibited a modified regulation of the production of some secreted proteins (acid protease 1 and endopolygalacturonase 1). Culture filtrates of the mutant triggered an important oxidative burst in grapevine (Vitis vinifera) suspension cells, and the mutant-plant interaction resulted in the formation of hypersensitive response-like necrosis. Genetic segregation analyses revealed that the pathogenicity phenotype was linked to a single locus, but showed that the mutated gene was not tagged by the plasmid pAN7-1. Mutant A336 is the first oxalate-deficient mutant to be described in B. cinerea and it differs from all the nonpathogenic B. cinerea mutants described to date.
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PMID:Characterization of a new, nonpathogenic mutant of Botrytis cinerea with impaired plant colonization capacity. 1662 75

The object of this study was to determine if calcium cross-linking of pectin in the pit membrane of xylem parenchyma restricts water movement which results in deep supercooling. Current year shoots of ;Loring' peach (Prunus persica) were infiltrated with oxalic acid or EGTA solutions for 24 or 48 hours and then either prepared for ultrastructural analysis or subjected to differential thermal analysis. The effect of 0.25 to 1.0% pectinase (weight/volume) on deep supercooling was also investigated. The use of 5 to 50 millimolar oxalic acid and pectinase resulted in a significant reduction (flattening) of the low temperature exotherm and a distinct swelling and partial degradation of the pit membrane. EGTA (10 millimolar) for 24 or 48 hours shifted the low temperature exotherm to warmer temperatures and effected the outermost layer of the pit membrane. A hypothesis is presented on pectin-mediated regulation of deep supercooling of xylem parenchyma.
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PMID:Effect of Macerase, Oxalic Acid, and EGTA on Deep Supercooling and Pit Membrane Structure of Xylem Parenchyma of Peach. 1666 41

Abstract The fungal pathogen Botrytis cinerea is capable of developing on a wide variety of host plants that differ greatly in their pH values and biochemical defences. To evaluate whether the pH of the host tissue can regulate the production of pathogenicity factors by this fungus, we examined the ability of two isolates of B. cinerea that originated from different plant species to secrete putative virulence elements on synthetic media buffered at pH 2.0 to pH 7.0. Even though differing in the intensity of their responses, both isolates reacted similarly to their ambient pH. The production of extracellular polysaccharides and oxalic acid was detectable above pH 4.0 and pH 5.0 respectively. Conversely, the production of aspartic acid proteases could only be seen between pH 3.0 and 4.0. Finally, the secretion of polygalacturonase and laccase activity was found to exhibit two maxima, one around pH 3.1 and one around pH 6.0. Thus, pathogenicity factor production was found to be minimal between pH 4.5 and 5.5 and a different set of factors was produced at pH 3.1 and 6.0, two values that were found to correspond respectively to the average host fruit and leaf pH. These results demonstrate that ambient pH differentially regulates the synthesis of pathogenicity factors by Botrytis and may act as a novel regulatory element to assist this fungus in tuning its virulence machinery to the composition of its host tissue.
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PMID:Differential regulation by ambient pH of putative virulence factor secretion by the phytopathogenic fungus Botrytis cinerea. 1971 67

SUMMARY The production of oxalic acid by Sclerotinia sclerotiorum is regulated by the ambient pH environment. This regulation and that of a few investigated pH-responsive genes is mediated in part by the zinc finger transcription factor encoded by pac1, an orthologue of the Aspergillus nidulans pacC gene. We manipulated the pac1 sequence by site-directed mutagenesis to create a dominant activating pac1(c) mutation and introduced this allele into a pac1 loss-of-function (Deltapac1) strain. Consistent with a constitutive activation of Pac1 function, oxalic acid accumulation in recovered Deltapac1-pac1(c) strains was largely independent of ambient pH. Likewise, all three Deltapac1-pac1(c) strains accumulated detectable pac1 transcripts in a pH 3 environment; however, accumulation of pac1 transcripts remained alkaline-inducible, but much reduced relative to wild-type in two of the three Deltapac1-pac1(c) strains. Surprisingly, the accumulation of pg1 and acp1 transcripts, normally favoured by low pH conditions, were up-regulated across the range of ambient pH conditions examined (pH 3.4-7.2). Accumulation of neutral pH-expressed endopolygalacturonase-6 (pg6) transcripts, however, did not differ from wild-type. In pathogenicity assays using Arabidopsis and detached tomato leaflets, Deltapac1-pac1(c) strains were reduced in virulence despite the ability to accumulate oxalic acid independent of the prevailing ambient pH environment. These results support the hypothesis that appropriate gene regulation in response to ambient pH is important for full S. sclerotiorum virulence independent of oxalic acid accumulation.
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PMID:An activating mutation of the Sclerotinia sclerotiorum pac1 gene increases oxalic acid production at low pH but decreases virulence. 2050 25

Extracellular polygalacturonase (PG) production was estimated in vitro, using liquid cultures of three species of brown-rot decay fungi (Postia placenta, Gloeophyllum trabeum and Serpula incrassata), by cup-plate assay, assay of reducing sugars, and decrease in viscosity. Although all three experimental assays demonstrated that PG was induced by pectin in all three fungi, decrease in viscosity gave the best correlation with decay capacity in soil block tests. PG activity, determined as an increase in reducing sugar activity, was greatest in G. trabeum and weakest in S. incrassata. The optimum pH for PG activity was between pH 2.5 and 4.5. Oxalic acid production was also enhanced by pectin and functioned synergistically with PG activity. We conclude that these fungi produce PG that is best induced by pectin and that PG activity exceeds production of xylanase and endoglucanase activity in vitro. Polygalacturonase is likely to act synergistically with oxalic acid to solubilize and hydrolyse the pectin in pit membranes and middle lamellae. Thus, production of PG and oxalic acid should facilitate early spread of hyphae and enhance the lateral flow of wood-decay enzymes and agents into adjacent tracheids and the wood cell wall, thus initiating the diffuse decay caused by brown-rot fungi.
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PMID:Induction of polygalacturonase and the formation of oxalic acid by pectin in brown-rot fungi. 2441 6


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