Dataset Information

Morphological Characterization and Quantification of the Mycelial Growth of the Brown-Rot Fungus Postia placenta for Modeling Purposes.

ABSTRACT: Continuous observation was performed using confocal laser scanning microscopy to visualize the three-dimensional microscopic growth of the brown-rot fungus, Postia placenta, for seventeen days. The morphological characterization of Postia placenta was quantitatively determined, including the tip extension rate, branch angle and branching length, (hyphal length between two adjacent branch sites). A voxel method has been developed to measure the growth of the biomass. Additionally, the tip extension rate distribution, the branch angle distribution and the branching length distribution, which quantified the hyphal growth characteristics, were evaluated. Statistical analysis revealed that the extension rate of tips was randomly distributed in space. The branch angle distribution did not change with the development of the colony, however, the branching length distribution did vary with the development of the colony. The experimental data will be incorporated into a lattice-based model simulating the growth of Postia placenta.

SUBMITTER: Du H

PROVIDER: S-EPMC5014427 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Publications

Morphological Characterization and Quantification of the Mycelial Growth of the Brown-Rot Fungus Postia placenta for Modeling Purposes.

Du Huan H Lv Pin P Ayouz Mehdi M Besserer Arnaud A Perré Patrick P

PloS one 20160907 9

Continuous observation was performed using confocal laser scanning microscopy to visualize the three-dimensional microscopic growth of the brown-rot fungus, Postia placenta, for seventeen days. The morphological characterization of Postia placenta was quantitatively determined, including the tip extension rate, branch angle and branching length, (hyphal length between two adjacent branch sites). A voxel method has been developed to measure the growth of the biomass. Additionally, the tip extensi ...[more]

PMID: 27602575

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Project description:Wood-degrading brown rot fungi are essential recyclers of plant biomass in forest ecosystems. Their efficient cellulolytic systems, which have potential biotechnological applications, apparently depend on a combination of two mechanisms: lignocellulose oxidation by reactive oxygen species (ROS) and polysaccharide hydrolysis by a limited set of glycoside hydrolases (GHs). Given that ROS are strongly oxidizing and non-selective, these two steps are likely segregated. A common hypothesis has been that brown rot fungi use a concentration gradient of chelated metal ions to confine ROS generation inside wood cell walls before enzymes can infiltrate. We examined an alternative: that lignocellulose-oxidation (LOX) components involved in ROS production are differentially expressed by brown rot fungi ahead of GH components. We used spatial mapping to resolve a temporal sequence in Postia placenta, sectioning thin wood wafers colonized directionally. Among sections, we measured gene expression by whole transcriptome sequencing (RNAseq) and assayed relevant enzyme activities. We found a marked pattern of LOX upregulation in a narrow (5-mm; 48-hr) zone at the hyphal front, which included many genes likely involved in ROS generation. Upregulation of GH5 endoglucanases and many other GHs clearly occurred later, behind the hyphal front, with notable exceptions of two likely expansins and a GH28 pectinase. Our results support a staggered mechanism for brown rot that is controlled by differential expression rather than microenvironmental gradients. This mechanism likely results in an oxidative pretreatment of lignocellulose, possibly facilitated by expansin- and pectinase-assisted cell wall swelling, before cellulases and hemicellulases are deployed for polysaccharide depolymerization. We sequenced mRNA from 9 Postia placenta samples taken from 3 wood sections of wafer design, with 3 bioreplicates for each wood section, to compare the gene expression during brown rot processes. Three wood sections of the wafer are representing early to late decay stages.

Project description:Brown rot fungi dominate wood decomposition in coniferous forests, and their carbohydrate-selective mechanisms are of commercial interest. Brown rot was recently described as a two-step, sequential mechanism orchestrated by fungi using differentially expressed genes (DEGs) and consisting of oxidation via reactive oxygen species (ROS) followed by enzymatic saccharification. There have been indications, however, that the initial oxidation step itself might require induction. To capture this early gene regulation event, here, we integrated fine-scale cryosectioning with whole-transcriptome sequencing to dissect gene expression at the single-hyphal-cell scale (tens of micrometers). This improved the spatial resolution 50-fold, relative to previous work, and we were able to capture the activity of the first 100 μm of hyphal front growth by Rhodonia placenta in aspen wood. This early decay period was dominated by delayed gene expression patterns as the fungus ramped up its mechanism. These delayed DEGs included many genes implicated in ROS pathways (lignocellulose oxidation [LOX]) that were previously and incorrectly assumed to be constitutively expressed. These delayed DEGs, which include those with and without predicted functions, also create a focused subset of target genes for functional genomics. However, this delayed pattern was not universal, with a few genes being upregulated immediately at the hyphal front. Most notably, this included a gene commonly implicated in hydroquinone and iron redox cycling: benzoquinone reductase. IMPORTANCE Earth's aboveground terrestrial biomass is primarily wood, and fungi dominate wood decomposition. Here, we studied these fungal pathways in a common "brown rot"-type fungus, Rhodonia placenta, that selectively extracts sugars from carbohydrates embedded within wood lignin. Using a space-for-time design to map fungal gene expression at the extreme hyphal front in wood, we made two discoveries. First, we found that many genes long assumed to be "on" (constitutively expressed) from the very beginning of decay were instead "off" before being upregulated, when mapped (via transcriptome sequencing [RNA-seq]) at a high resolution. Second, we found that the gene encoding benzoquinone reductase was "on" in incipient decay and quickly downregulated, implying a key role in "kick-starting" brown rot.

Dataset Information

Morphological Characterization and Quantification of the Mycelial Growth of the Brown-Rot Fungus Postia placenta for Modeling Purposes.

Publications

Morphological Characterization and Quantification of the Mycelial Growth of the Brown-Rot Fungus Postia placenta for Modeling Purposes.

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