Biology of Pleurotus eryngii and role in biotechnological processes: a review

Pleurotus eryngii is considered a complex species owing to a perplexed structure within species and a wide geographical distribution. Due to its remarkable flavor, high nutritional value, and numerous medicinal features, P. eryngii is commercially cultivated on various raw plant materials. Its efficacy in using nutrients from lignocellulose residues is based on possession of a potent ligninolytic enzyme system, constituted of laccase, Mn-oxidizing peroxidases, and aryl-alcohol oxidase, which successfully degrade different aromatic compounds. Similarly, due to the ability of these enzymes, P. eryngii plays a very important role in many biotechnological processes, such as food production (edible basidiomata), biotransformation of raw plant materials to feed, biopulping and biobleaching of paper pulp, as well as bioremediation of soil and industrial waters.     

EAAT2 density at the astrocyte plasma membrane and Ca2 + -regulated exocytosis

We studied whether regulated exocytosis affects the glutamate transporter density in cultured astrocytes, in which the expression of a fluorescently labeled excitatory amino acid transporter 2 (EAAT2-EGFP) predominantly labeled the plasma membrane. The addition of ionomycin that elevates cytosolic Ca²⁺ strongly increased the fluorescence of FM 4-64 membrane area dye, confirming the presence of regulated exocytosis in transfected astrocytes. However, concomitant with Ca²⁺-dependent FM 4-64 fluorescence increase, ionomycin induced a significant steady-state decrease in EAAT2-EGFP fluorescence. This is likely due to a secondary inner filter effect since,(i) in the absence of FM 4-64, ionomycin stimulation was ineffective in changing the EAAT2-EGFP fluorescence, and (ii) fluorescence changes in FM 4-64 and EAAT2-EGFP were inversely correlated. To test whether subcellular EAAT2-EGFP structures are translocated from the cytoplasm to the plasma membrane during ionomycin stimulation, EAAT2-EGFP fluorescence was monitored locally at the plasma membrane and a few microns away in the adjacent cytoplasm. Measurements revealed sites with an increase in EAAT2-EGFP plasma membrane fluorescence correlated with a fluorescence decrease beneath the plasma membrane, and sites with plasma membrane fluorescence decrease correlated with fluorescence increase within the adjacent cytoplasm. The sites of rapid translocation/retrieval of EAAT2-EGFP structures to/from the plasma membrane appeared to be distributed in a punctuate pattern around the cell perimeter. The density of EAAT2-EGFP was regulated in a Ca²⁺-dependent manner, since in the absence of extracellular Ca²⁺ local translocation/retrieval events were absent, revealing rapid surface density regulation of EAAT2 in astrocytes by regulated exo/endocytosis.     

Isomaltose formation by free and immobilized dextransucrase

The acceptor reaction of dextransucrase from Leuconostoc mesenteroides NRRL-B512F with glucose as acceptor is of technical interest for isomaltooligosaccharide (IMOs) synthesis. Different experimental conditions were investigated for free and immobilized enzyme. The data for oligosaccharide formation up to a degree of polymerization 4 were correlated with a model developed earlier, and optimal reaction conditions for immobilized dextransucrase design and application were identified for later continuous application. Furthermore, stability was investigated for free and immobilized enzyme including stabilization by sugars.     

Surface modification of polyacrylonitrile with nitrile hydratase and amidase from Agrobacterium tumefaciens

A new strain of Agrobacterium tumefaciens (BST05) was found to grow on polyacrylonitrile (PAN; ¹³C labelled) converting the polymer to polyacrylic acid as shown by solid state NMR. When cultivated in a medium containing acetonitrile the bacterium produced nitrile hydratase and amidase activity. Activity recovery after lyophilisation and enzyme stability was significantly enhanced in the presence of 5% sorbitol leading to half life times of 12, 72 and 154 days at 25°C, 4°C and –20°C. The enzymes were able to convert 1.1% of the nitrile groups of PAN-powder to the corresponding acids. PAN fabrics were mainly converted to the amides as shown by an 80% increase of the O/C ratio in ESCA analysis. These data were confirmed by cationic dyeing and FTIR-ATR analysis.     

Genetic architecture of resistance to Septoria tritici blotch (Mycosphaerella graminicola) in European winter wheat

Genome-wide marker–trait associations (MTA) were established in a population of 358 European winter wheat cultivars and 14 spring wheat cultivars (Triticum aestivum L.) for resistance to Septoria tritici blotch caused by the fungal pathogen Mycosphaerella graminicola. The MTA were based on field data in two consecutive years and genotypic data on 732 microsatellite markers. Best linear unbiased estimations (BLUEs) for resistance were calculated across the trials and ranged from 0.67 (most resistant) to 19.63 (most susceptible) with an average value of 4.93. A total of 115 MTA relating to 68 molecular markers was discovered for the two trials and BLUEs by using a mixed linear model corrected by a kinship matrix. In addition, two candidate genes, Ppd-D1 for photoperiodism and the dwarfing gene Rht-D1, were significantly associated with resistance to Septoria tritici blotch. Several MTA co-located with known resistance genes, e.g. Stb1, 3, 4, 6 and 8, while multiple additional MTA were discovered on several chromosomes, such as 2A, 2D, 3A, 5B, 7A and 7D. The results provide proof of concept for the method of genome-wide association analysis and indicate the presence of further Stb resistance genes in the European winter wheat pool.     

Phylogenetic and functional relationships in pastures and meadows from the North Adriatic Karst

Probably due to its phytogeographic position, the grasslands in the North Adriatic Karst are among the richest grasslands in the world and harbour the highest small-scale density of plant species found in terrestrial habitats. Different moisture and soil conditions determine distinct vegetation types, such as meadows, composed by mesophyllous plants and on mesic conditions, and semi-natural pastures, composed by sclerophyllous plants and on oligotrophic conditions. Even though plants in these two vegetation types differ in some of their attributes, their functional and phylogenetic relationships remain to be tested. We used a dataset comprising 180 species, in which 48 plots in the meadows and 52 plots in the pastures had been sampled, and tested the phylogenetic and functional relationships between the two vegetation types. The pastures contained more original species, but both the pastures and the meadows are expected to be important to increase biodiversity at landscape level. Different community assembly processes occurred in the two vegetation types, with limiting similarity leading to functional overdispersion in the meadows and environmental filtering leading to functional clustering in the pastures. Overall, traits were convergent, leading to a clustered phylogenetic structure in the meadows, probably due to pair-wise competition, and an overdispersed phylogenetic structure in the pastures, where species from different clades were filtered by the oligotrophic conditions. Transhumance may have contributed to the random pattern of trait diversity we found across the nodes of the phylogenetic tree.     

High plant species richness indicates management-related disturbances rather than the conservation status of forests

There is a wealth of smaller-scale studies on the effects of forest management on plant diversity. However, studies comparing plant species diversity in forests with different management types and intensity, extending over different regions and forest stages, and including detailed information on site conditions are missing. We studied vascular plants on 1500 20 m × 20 m forest plots in three regions of Germany (Schwäbische Alb, Hainich-Dün, Schorfheide-Chorin). In all regions, our study plots comprised different management types (unmanaged, selection cutting, deciduous and coniferous age-class forests, which resulted from clear cutting or shelterwood logging), various stand ages, site conditions, and levels of management-related disturbances. We analyzed how overall richness and richness of different plant functional groups (trees, shrubs, herbs, herbaceous species typically growing in forests and herbaceous light-demanding species) responded to the different management types. On average, plant species richness was 13% higher in age-class than in unmanaged forests, and did not differ between deciduous age-class and selection forests. In age-class forests of the Schwäbische Alb and Hainich-Dün, coniferous stands had higher species richness than deciduous stands. Among age-class forests, older stands with large quantities of standing biomass were slightly poorer in shrub and light-demanding herb species than younger stands. Among deciduous forests, the richness of herbaceous forest species was generally lower in unmanaged than in managed forests, and it was even 20% lower in unmanaged than in selection forests in Hainich-Dün. Overall, these findings show that disturbances by management generally increase plant species richness. This suggests that total plant species richness is not suited as an indicator for the conservation status of forests, but rather indicates disturbances.     

Structure, dynamics, lipid binding, and physiological relevance of the putative GTPase-binding domain of Dictyostelium formin C

Dictyostelium Formin C (ForC) is involved in the regulation of local actin cytoskeleton reorganization (e.g. during cellular adhesion or migration). ForC contains formin homology 2 and 3 (FH2 and -3) domains and an N-terminal putative GTPase-binding domain (GBD) but lacks a canonical FH1 region. To better understand the role of the GBD, its structure, dynamics, lipid-binding properties, and cellular functions were analyzed by NMR and CD spectroscopy and by in vivo fluorescence microscopy. Moreover, the program CS-Rosetta was tested for the structure prediction based on chemical shift data only. The ForC GBD adopts an ubiquitin-like α/β-roll fold with an unusually long loop between β-strands 1 and 2. Based on the lipid-binding data, the presence of DPC micelles induces the formation of α-helical secondary structure and a rearrangement of the tertiary structure. Lipid-binding studies with a mutant protein and a peptide suggest that the β1-β2 loop is not relevant for these conformational changes. Whereas small amounts of negatively charged phosphoinositides (1,2-dioctanoyl-sn-glycero-3-(phosphoinositol 4,5-bisphosphate) and 1,2-dihexanoyl-sn-glycero-3-(phosphoinositol 3,4,5-trisphosphate)) lower the micelle concentration necessary to induce the observed spectral changes, other negatively charged phospholipids (1,2-dihexanoyl-sn-glycero-3-(phospho-L-serine) and 1,2-dihexanoyl-sn-glycero-3-phospho-(1'-rac-glycerol)) had no such effect. Interestingly, bicelles and micelles composed of diacylphosphocholines had no effect on the GBD structure. Our data suggest a model in which part of the large positively charged surface area of the GBD mediates localization to specific membrane patches, thereby regulating interactions with signaling proteins. Our cellular localization studies show that both the GBD and the FH3 domain are required for ForC targeting to cell-cell contacts and early phagocytic cups and macropinosomes.     

Impairment of protein trafficking by direct interaction of gliadin peptides with actin

Intestinal celiac disease (CD) is triggered by peptic-tryptic digest of gluten, known as Frazer's Fraction (FF), in genetically predisposed individuals. Here, we investigate the immediate effects of FF on the actin cytoskeleton and the subsequent trafficking of actin-dependent and actin-independent proteins in COS-1 cells. Morphological alterations in the actin filaments were revealed concomitant with a drastic reduction in immunoprecipitated actin from cells incubated with FF. These alterations elicit impaired protein trafficking of intestinal sucrase–isomaltase, a glycoprotein that follows an actin-dependent vesicular transport to the cell surface. However, the actin-independent transport of intestinal lactase phlorizin hydrolase remains unaffected. Moreover, the morphological alteration in actin is induced by direct interaction of this protein with gliadin peptides carrying the QQQPFP epitope revealed by co-immunoprecipitation utilizing a monoclonal anti-gliadin antibody. Finally, stimulation of cells with FF directly influences the binding of actin to Arp2. Altogether, our data demonstrate that FF directly interacts with actin and alters the integrity of the actin cytoskeleton thus leading to an impaired trafficking of intestinal proteins that depend on an intact actin network. This direct interaction could be related to the endocytic segregation of gliadin peptides as well as the delayed endocytic vesicle trafficking and maturation in gliadin-positive intestinal epithelial cells and opens new insights into the pathogenesis of CD.