Fungal behavior and recent developments in biopulping technology.

Biological pretreatment of wood chips by fungi is a well-known approach prior to mechanical- or chemical pulp production. For this biological approach, a limited number of white-rot fungi with an ability to colonize and selectively degrade lignin are used to pretreat wood chips allowing the remaining cellulose to be processed for further applications. Biopulping is an environmentally friendly technology that can reduce the energy consumption of traditional pulping processes. Fungal pretreatment also reduces the pitch content in the wood chips and improves the pulp quality in terms of brightness, strength, and bleachability. The bleached biopulps are easier to refine compared to pulps produced by conventional methodology. In the last decades, biopulping has been scaled up with pilot trials towards industrial level, with optimization of several intermediate steps and improvement of economic feasibility. Nevertheless, fundamental knowledge on the biochemical mechanisms involved in biopulping is still lacking. Overall, biopulping technology has advanced rapidly during recent decades and pilot mill trials have been implemented. The use of fungi as pretreatment for pulp production is in line with modern circular economy strategies and can be implemented in existing production plants. In this review, we discuss some recent advances in biopulping technology, which can improve mechanical-, chemical-, and organosolv pulping processes along with their mechanisms.

Functional Nanostructures from Sol-Gel Synthesis Using Keggin Polyoxometallate Phosphotungstic Acid as a Precursor.

Subjecting phosphotungstic acid solutions to low pH in combination with introduction of polyvalent cations led to the formation of nanostructured microspheres of approximately 2 µm in size, as shown by scanning electron microscopy, which were almost insoluble and resistant to degradation at neutral and high pH. These microspheres were composed of secondary nanospheres with diameters around 20 nm as revealed by transmission electron microscopy and atomic force microscopy. Investigations of the crystal structure of a potential intermediate of this process, namely, acidic lanthanum phosphotungstate, [La(H2O)9](H3O)3[PW12O40]2(H2O)19, showed a tight network of hydrogen bonding, permitting closer packing of phosphotungstic acid anions, thereby confirming the mechanism of the observed self-assembly process. The new material demonstrated promising electrochemical properties in oxygen evolution reactions with the high stability of the obtained electrode material.

Insights into asynchronous changes of cell wall polymers accumulated in different cell types during conifer xylem differentiation.

An improved understanding of the events involved in cell wall polymers deposition during xylem development could provide new scientific ways for molecular regulation and biomass utilization. Axial and radial cells are spatially heterogeneous and have highly cross-correlated developmental behavior, whereas the deposition of corresponding cell wall polymers during xylem differentiation is less studied. To clarify our hypothesis that cell wall polymers of two cell types accumulated asynchronously, we performed hierarchical visualization, including label-free in situ spectral imaging of different polymer compositions during the development of Pinus bungeana. In axial tracheids, the deposition of cellulose and glucomannan was observed on earlier stages of secondary wall thickening than that of xylan and lignin, while xylan distribution was strongly related to spatial distribution of lignin during differentiation. The content of lignin and polysaccharides increased by over 130 % and 60 % respectively when the S3 layer was formed, compared to the S2 stage. In ray cells, the deposition of crystalline cellulose, xylan, and lignin was generally lagged compared to that in corresponding axial tracheids, although the process followed a similar order. The concentration of lignin and polysaccharides in ray cells was only approximately 50 % of that in the axial tracheids during secondary wall thickening.

Electron tomography unravels new insights into fiber cell wall nanostructure; exploring 3D macromolecular biopolymeric nano-architecture of spruce fiber secondary walls.

Lignocellulose biomass has a tremendous potential as renewable biomaterials for fostering the "bio-based society" and circular bioeconomy paradigm. It requires efficient use and breakdown of fiber cell walls containing mainly cellulose, hemicellulose and lignin biopolymers. Despite their great importance, there is an extensive debate on the true structure of fiber walls and knowledge on the macromolecular nano-organization is limited and remains elusive in 3D. We employed dual-axis electron tomography that allows visualization of previously unseen 3D macromolecular organization/biopolymeric nano-architecture of the secondary S2 layer of Norway spruce fiber wall. Unprecedented 3D nano-structural details with novel insights into cellulose microfibrils (~ 2 nm diameter), macrofibrils, nano-pore network and cell wall chemistry (volume %) across the S2 were explored and quantified including simulation of structure related permeability. Matrix polymer association with cellulose varied between microfibrils and macrofibrils with lignin directly associated with MFs. Simulated bio-nano-mechanical properties revealed stress distribution within the S2 and showed similar properties between the idealized 3D model and the native S2 (actual tomogram). Present work has great potential for significant advancements in lignocellulose research on nano-scale understanding of cell wall assembly/disassembly processes leading to more efficient industrial processes of functionalization, valorization and target modification technologies.

Immunothrombosis and vascular heterogeneity in cerebral cavernous malformation.

Cerebral cavernous malformation (CCM) is a neurovascular disease that results in various neurological symptoms. Thrombi have been reported in surgically resected CCM patient biopsies, but the molecular signatures of these thrombi remain elusive. Here, we investigated the kinetics of thrombi formation in CCM and how thrombi affect the vasculature and contribute to cerebral hypoxia. We used RNA sequencing to investigate the transcriptome of mouse brain endothelial cells with an inducible endothelial-specific Ccm3 knock-out (Ccm3-iECKO). We found that Ccm3-deficient brain endothelial cells had a higher expression of genes related to the coagulation cascade and hypoxia when compared with wild-type brain endothelial cells. Immunofluorescent assays identified key molecular signatures of thrombi such as fibrin, von Willebrand factor, and activated platelets in Ccm3-iECKO mice and human CCM biopsies. Notably, we identified polyhedrocytes in Ccm3-iECKO mice and human CCM biopsies and report it for the first time. We also found that the parenchyma surrounding CCM lesions is hypoxic and that more thrombi correlate with higher levels of hypoxia. We created an in vitro model to study CCM pathology and found that human brain endothelial cells deficient for CCM3 expressed elevated levels of plasminogen activator inhibitor-1 and had a redistribution of von Willebrand factor. With transcriptomics, comprehensive imaging, and an in vitro CCM preclinical model, this study provides experimental evidence that genes and proteins related to the coagulation cascade affect the brain vasculature and promote neurological side effects such as hypoxia in CCMs. This study supports the concept that antithrombotic therapy may be beneficial for patients with CCM.

The contribution of G-layer glucose in Salix clones for biofuels: comparative enzymatic and HPLC analysis of stem cross sections.

BACKGROUND: Interest on the use of short rotation willow as a lignocellulose resource for liquid transport fuels has increased greatly over the last 10 years. Investigations have shown the advantages and potential of using Salix spp. for such fuels but have also emphasized the wide variations existing in the compositional structure between different species and genotypes in addition to their effects on overall yield. The present work studied the importance of tension wood (TW) as a readily available source of glucose in 2-year-old stems of four Salix clones (Tora, Björn, Jorr, Loden). Studies involved application of a novel approach whereby TW-glucose and residual sugars and lignin were quantified using stem cross sections with results correlated with HPLC analyses of milled wood. Compositional analyses were made for four points along stems and glucose derived from enzyme saccharification of TW gelatinous (G) layers (G-glucose), structural cell wall glucose (CW-glucose) remaining after saccharification and total glucose (T-glucose) determined both theoretically and from HPLC analyses. Comparisons were also made between presence of other characteristic sugars as well as acid-soluble and -insoluble lignin. RESULTS: Preliminary studies showed good agreement between using stem serial sections and milled powder from Salix stems for determining total sugar and lignin. Therefore, sections were used throughout the work. HPLC determination of T-glucose in Salix clones varied between 47.1 and 52.8%, showing a trend for higher T-glucose with increasing height (Björn, Tora and Jorr). Using histochemical/microscopy and image analysis, Tora (24.2%) and Björn (28.2%) showed greater volumes of % TW than Jorr (15.5%) and Loden (14.0%). Total G-glucose with enzyme saccharification of TW G-layers varied between 3.7 and 14.7% increasing as the total TW volume increased. CW-glucose measured after enzyme saccharification showed mean values of 41.9-49.1%. Total lignin between and within clones showed small differences with mean variations of 22.4-22.8% before and 22.4-24.3% after enzyme saccharification. Calculated theoretical and quantified values for CW-glucose at different heights for clones were similar with strong correlation: T-glucose = G-glucose + CW-glucose. Pearson's correlation displayed a strong and positive correlation between T-glucose and G-glucose, % TW and stem height, and between G-glucose with % TW and stem height. CONCLUSIONS: The use of stem cross sections to estimate TW together with enzyme saccharification represents a viable approach for determining freely available G-glucose from TW allowing comparisons between Salix clones. Using stem sections provides for discrete morphological/compositional tissue comparisons between clones with results consistent with traditional wet chemical analysis approaches where entire stems are milled and analyzed. The four clones showed variable TW and presence of total % G-glucose in the order Björn > Tora > Jorr > Loden. Calculated in terms of 1 m3, Salix stems Tora and Björn would contain ca. 0.24 and 0.28 m3 of tension wood representing a significant amount of freely available glucose.

Palmdelphin Regulates Nuclear Resilience to Mechanical Stress in the Endothelium.

BACKGROUND: PALMD (palmdelphin) belongs to the family of paralemmin proteins implicated in cytoskeletal regulation. Single nucleotide polymorphisms in the PALMD locus that result in reduced expression are strong risk factors for development of calcific aortic valve stenosis and predict severity of the disease. METHODS: Immunodetection and public database screening showed dominant expression of PALMD in endothelial cells (ECs) in brain and cardiovascular tissues including aortic valves. Mass spectrometry, coimmunoprecipitation, and immunofluorescent staining allowed identification of PALMD partners. The consequence of loss of PALMD expression was assessed in small interferring RNA-treated EC cultures, knockout mice, and human valve samples. RNA sequencing of ECs and transcript arrays on valve samples from an aortic valve study cohort including patients with the single nucleotide polymorphism rs7543130 informed about gene regulatory changes. RESULTS: ECs express the cytosolic PALMD-KKVI splice variant, which associated with RANGAP1 (RAN GTP hydrolyase activating protein 1). RANGAP1 regulates the activity of the GTPase RAN and thereby nucleocytoplasmic shuttling via XPO1 (Exportin1). Reduced PALMD expression resulted in subcellular relocalization of RANGAP1 and XPO1, and nuclear arrest of the XPO1 cargoes p53 and p21. This indicates an important role for PALMD in nucleocytoplasmic transport and consequently in gene regulation because of the effect on localization of transcriptional regulators. Changes in EC responsiveness on loss of PALMD expression included failure to form a perinuclear actin cap when exposed to flow, indicating lack of protection against mechanical stress. Loss of the actin cap correlated with misalignment of the nuclear long axis relative to the cell body, observed in PALMD-deficient ECs, Palmd-/- mouse aorta, and human aortic valve samples derived from patients with calcific aortic valve stenosis. In agreement with these changes in EC behavior, gene ontology analysis showed enrichment of nuclear- and cytoskeleton-related terms in PALMD-silenced ECs. CONCLUSIONS: We identify RANGAP1 as a PALMD partner in ECs. Disrupting the PALMD/RANGAP1 complex alters the subcellular localization of RANGAP1 and XPO1, and leads to nuclear arrest of the XPO1 cargoes p53 and p21, accompanied by gene regulatory changes and loss of actin-dependent nuclear resilience. Combined, these consequences of reduced PALMD expression provide a mechanistic underpinning for PALMD's contribution to calcific aortic valve stenosis pathology.

A modified CUT&RUN protocol and analysis pipeline to identify transcription factor binding sites in human cell lines.

CUT&RUN is a recently developed in situ chromatin profiling technique that enables high-resolution chromatin mapping and probing. Herein, we describe our adapted CUT&RUN protocol for transcription factors (TFs). Our protocol outlines all necessary steps for TF profiling including the procedure to obtain proteinA-Mnase, while also outlining the bioinformatic pipeline steps required to process, analyze, and identify novel binding sites and sequences. Due to the small number of cells required, this method will allow the elucidation of cell context-dependent functions of many TFs. For details on the use and execution of this protocol, please refer to Kong et al. (2021).

Enzymatic hydrolysis of the gelatinous layer in tension wood of Salix varieties as a measure of accessible cellulose for biofuels.

BACKGROUND: Salix (willow) species represent an important source of bioenergy and offer great potential for producing biofuels. Salix spp. like many hardwoods, produce tension wood (TW) characterized by special fibres (G-fibres) that produce a cellulose-rich lignin-free gelatinous (G) layer on the inner fibre cell wall. Presence of increased amounts of TW and G-fibres represents an increased source of cellulose. In the present study, the presence of TW in whole stems of different Salix varieties was characterized (i.e., physical measurements, histochemistry, image analysis, and microscopy) as a possible marker for the availability of freely available cellulose and potential for releasing D-glucose. Stem cross sections from different Salix varieties (Tora, Björn) were characterized for TW, and subjected to cellulase hydrolysis with the free D-glucose produced determined using a glucose oxidase/peroxidase (GOPOD) assay. Effect of cellulase on the cross sections and progressive hydrolysis of the G-layer was followed using light microscopy after staining and scanning electron microscopy (SEM). RESULTS: Tension wood fibres with G-layers were developed multilaterally in all stems studied. Salix TW from varieties Tora and Björn showed fibre G-layers were non-lignified with variable thickness. Results showed: (i) Differences in total % TW at different stem heights; (ii) that using a 3-day incubation period at 50 °C, the G-layers could be hydrolyzed with no apparent ultrastructural effects on lignified secondary cell wall layers and middle lamellae of other cell elements; and (iii) that by correlating the amount of D-glucose produced from cross sections at different stem heights together with total % TW and density, an estimate of the total free D-glucose in stems can be derived and compared between varieties. These values were used together with a literature value (45%) for estimating the contribution played by G-layer cellulose to the total cellulose content. CONCLUSIONS: The stem section-enzyme method developed provides a viable approach to compare different Salix varieties ability to produce TW and thus freely available D-glucose for fermentation and biofuel production. The use of Salix stem cross sections rather than comminuted biomass allows direct correlation between tissue- and cell types with D-glucose release. Results allowed correlation between % TW in cross sections and entire Salix stems with D-glucose production from digested G-layers. Results further emphasize the importance of TW and G-fibre cellulose as an important marker for enhanced D-glucose release in Salix varieties.

γ-Tubulin Complexes and Fibrillar Arrays: Two Conserved High Molecular Forms with Many Cellular Functions.

Higher plants represent a large group of eukaryotes where centrosomes are absent. The functions of γ-tubulin small complexes (γ-TuSCs) and γ-tubulin ring complexes (γ-TuRCs) in metazoans and fungi in microtubule nucleation are well established and the majority of components found in the complexes are present in plants. However, plant microtubules are also nucleated in a γ-tubulin-dependent but γ-TuRC-independent manner. There is growing evidence that γ-tubulin is a microtubule nucleator without being complexed in γ-TuRC. Fibrillar arrays of γ-tubulin were demonstrated in plant and animal cells and the ability of γ-tubulin to assemble into linear oligomers/polymers was confirmed in vitro for both native and recombinant γ-tubulin. The functions of γ-tubulin as a template for microtubule nucleation or in promoting spontaneous nucleation is outlined. Higher plants represent an excellent model for studies on the role of γ-tubulin in nucleation due to their acentrosomal nature and high abundancy and conservation of γ-tubulin including its intrinsic ability to assemble filaments. The defining scaffolding or sequestration functions of plant γ-tubulin in microtubule organization or in nuclear processes will help our understanding of its cellular roles in eukaryotes.

Propranolol Reduces the Development of Lesions and Rescues Barrier Function in Cerebral Cavernous Malformations: A Preclinical Study.

[Figure: see text].

Comparison of the Decay Behavior of Two White-Rot Fungi in Relation to Wood Type and Exposure Conditions.

Fungal wood decay strategies are influenced by several factors, such as wood species, moisture content, and temperature. This study aims to evaluate wood degradation characteristics of spruce, beech, and oak after exposure to the white-rot fungi Pleurotusostreatus and Trametesversicolor. Both fungi caused high mass losses in beech wood, while spruce and oak wood were more resistant to decay. The moisture content values of the decayed wood correlated with the mass losses for all three wood species and incubation periods. Combined microscopic and chemical studies indicated that the two fungi differed in their decay behavior. While T. versicolor produced a decay pattern (cell wall erosion) typical of white-rot fungi in all wood species, P. ostreatus caused cell wall erosion in spruce and beech and soft-rot type I (cavity formation) decay in oak wood. These observations suggest that P. ostreatus may have the capacity to produce a wider range of enzymes/radicals triggered by the chemical composition of wood cell walls and/or local compositional variability within the cell wall.

Removal of Diclofenac, Paracetamol, and Carbamazepine from Model Aqueous Solutions by Magnetic Sol-Gel Encapsulated Horseradish Peroxidase and Lignin Peroxidase Composites.

Sustainable and green synthesis of nanocomposites for degradation of pharmaceuticals was developed via immobilization and stabilization of the biological strong oxidizing agents, peroxidase enzymes, on a solid support. Sol-gel encapsulated enzyme composites were characterized using electron microscopy (TEM, SEM), atomic force microscopy, FTIR spectroscopy, and thermogravimetric analysis. Horseradish peroxidase (HRP) and lignin peroxidase (LiP) were adsorbed onto magnetite nanoparticles and sol-gel encapsulated in a surface silica layer. Encapsulation enhanced the stability of the biocatalysts over time and thermal stability. The biocatalysts showed appreciable selectivity in oxidation of the organic drinking water pollutants diclofenac, carbamazepine, and paracetamol with improved activity being pharmaceutical specific for each enzyme. In particular, sol-gel encapsulated LiP- and HRP-based nanocomposites were active over 20 consecutive cycles for 20 days at 55 °C (24 h/cycle). The stability of the sol-gel encapsulated catalysts in acidic medium was also improved compared to native enzymes. Carbamazepine and diclofenac were degraded to 68% and 64% by sol-gel LiP composites respectively at pH 5 under elevated temperature. Total destruction of carbamazepine and diclofenac was achieved at pH 3 (55 °C) within 3 days, in the case of both immobilized HRP and LiP. Using NMR spectroscopy, characterization of the drug decomposition products, and decomposition pathways by the peroxidase enzymes suggested.

Titanium phosphonate oxo-alkoxide "clusters": solution stability and facile hydrolytic transformation into nano titania.

Titanium (oxo-) alkoxide phosphonate complexes were synthesized using different titanium precursors and tert-butylphosphonic acid (tBPA) as molecular models for interaction between phosphonates and titania surfaces and to investigate the solution stability of these species. Reflux of titanium(iv) ethoxide or titanium(iv)(diisopropoxide)bis(2,4-pentadionate) with tert-butylphosphonic acid in toluene-ethanol mixture or acetone yielded seven titanium alkoxide phosphonate complexes; [Ti5(µ3-O)(µ2-O)(µ-HOEt)2(µ-OEt)3(µ2-OEt)(µ3-tBPA)3(µ3-HtBPA)(µ2-tBPA)2(µ2-HtBPA)]·3EtOH, 1, [Ti4O(µ-OEt)5(µ2-OEt)7(µ3-tBPA)], 2, [Ti4(µ2-O)2(µ-OEt)2(µ-HOEt)2(µ2-tPBA)2(µ2-HtPBA)6]·4EtOH, 3, [Ti4(µ2-O)2(µ-OEt)2(µ-HOEt)2(µ2-tPBA)2(µ2-HtPBA)6]·2EtOH, 4, [Ti6(µ2-O)(µ3-O)2(µ2-OEt)5(µ-OEt)6(µ3-tBPA)3(µ3-HtBPA)], 5, [Ti4(µ-iOPr)4(acac)4(µ2-tBPA)4], 6 and [Ti5(µ4-O)(µ2-O)3(µ2-OEt)4(µ-OEt)6(µ-HOEt)(µ3-tBPA)]2, 7. The binding mode of tBPA to the titanium oxo-core were either double or triple bridging or a combination of the two. No monodentate or chelating coordination was observed. 31P NMR spectrometry of dissolved single crystals indicates that 1 and 5 retain their solid-state structures in solution, the latter even on moderate heating, while 6 and 7 dissolved into several other forms. The complexes were found to be sensitive towards hydrolysis, proceeding in a topotactic fashion with densification of the material into plates and lamellae resulting finally in "core-shell" nanoparticles with a crystalline core (anatase) and an amorphous outer shell upon contact with water at room temperature as observed by HRTEM and AFM analyses. 31P NMR data supported degradation after addition of water to solutions of the complexes. Hydrolysis under different conditions affords complex oxide structures of different morphologies.

Basic Medium Heterogeneous Solution Synthesis of α-MnO2 Nanoflakes as an Anode or Cathode in Half Cell Configuration (vs. Lithium) of Li-Ion Batteries.

Nano α-MnO2 is usually synthesized under hydrothermal conditions in acidic medium, which results in materials easily undergoing thermal reduction and offers single crystals often over 100 nm in size. In this study, α-MnO2 built up of inter-grown ultra-small nanoflakes with 10 nm thickness was produced in a rapid two-step procedure starting via partial reduction in solution in basic medium subsequently followed by co-proportionation in thermal treatment. This approach offers phase-pure α-MnO2 doped with potassium (cryptomelane type K0.25Mn8O16 structure) demonstrating considerable chemical and thermal stability. The reaction pathways leading to this new morphology and structure have been discussed. The MnO2 electrodes produced from obtained nanostructures were tested as electrodes of lithium ion batteries delivering initial discharge capacities of 968 mAh g-1 for anode (0 to 2.0 V) and 317 mAh g-1 for cathode (1.5 to 3.5 V) at 20 mA g-1 current density. At constant current of 100 mA g-1, stable cycling of anode achieving 660 mAh g-1 and 145 mAh g-1 for cathode after 200 cycles is recorded. Post diagnostic analysis of cycled electrodes confirmed the electrode materials stability and structural properties.

γ-Tubulin has a conserved intrinsic property of self-polymerization into double stranded filaments and fibrillar networks.

γ-Tubulin is essential for microtubule nucleation and also plays less understood roles in nuclear and cell-cycle-related functions. High abundancy of γ-tubulin in acentrosomal Arabidopsis cells facilitated purification and biochemical characterization of large molecular species of γ-tubulin. TEM, fluorescence, and atomic force microscopy of purified high molecular γ-tubulin forms revealed the presence of linear filaments with a double protofilament substructure, filament bundles and aggregates. Filament formation from highly purified γ-tubulin free of γ-tubulin complex proteins (GCPs) was demonstrated for both plant and human γ-tubulin. Moreover, γ-tubulin associated with porcine brain microtubules formed oligomers. Experimental evidence on the intrinsic ability of γ-tubulin to oligomerize/polymerize was supported by conservation of α- and ß-tubulin interfaces for longitudinal and lateral interactions for γ-tubulins. STED (stimulated emission depletion) microscopy of Arabidopsis cells revealed fine, short γ-tubulin fibrillar structures enriched on mitotic microtubular arrays that accumulated at polar regions of acentrosomal spindles and the outer nuclear envelope before mitosis, and were also present in nuclei. Fine fibrillar structures of γ-tubulin representing assemblies of higher order were localized in cell-cycle-dependent manner at sites of dispersed γ-tubulin location in acentrosomal plant cells as well as at sites of local γ-tubulin enrichment after drug treatment. Our findings that γ-tubulin preserves the capability of prokaryotic tubulins to self-organize into filaments assembling by lateral interaction into bundles/clusters help understanding of the relationship between structure and multiple cellular functions of this protein species and suggest that besides microtubule nucleation and organization, γ-tubulin may also have scaffolding or sequestration functions.

Modification of the nanostructure of lignocellulose cell walls via a non-enzymatic lignocellulose deconstruction system in brown rot wood-decay fungi.

Wood decayed by brown rot fungi and wood treated with the chelator-mediated Fenton (CMF) reaction, either alone or together with a cellulose enzyme cocktail, was analyzed by small angle neutron scattering (SANS), sum frequency generation (SFG) spectroscopy, Fourier transform infrared (FTIR) analysis, X-ray diffraction (XRD), atomic force microscopy (AFM), and transmission electron microscopy (TEM). Results showed that the CMF mechanism mimicked brown rot fungal attack for both holocellulose and lignin components of the wood. Crystalline cellulose and lignin were both depolymerized by the CMF reaction. Porosity of the softwood cell wall did not increase during CMF treatment, enzymes secreted by the fungi did not penetrate the decayed wood. The enzymes in the cellulose cocktail also did not appear to alter the effects of the CMF-treated wood relative to enhancing cell wall deconstruction. This suggests a rethinking of current brown rot decay models and supports a model where monomeric sugars and oligosaccharides diffuse from the softwood cell walls during non-enzymatic action. In this regard, the CMF mechanism should not be thought of as a "pretreatment" used to permit enzymatic penetration into softwood cell walls, but instead it enhances polysaccharide components diffusing to fungal enzymes located in wood cell lumen environments during decay. SANS and other data are consistent with a model for repolymerization and aggregation of at least some portion of the lignin within the cell wall, and this is supported by AFM and TEM data. The data suggest that new approaches for conversion of wood substrates to platform chemicals in biorefineries could be achieved using the CMF mechanism with >75% solubilization of lignocellulose, but that a more selective suite of enzymes and other downstream treatments may be required to work when using CMF deconstruction technology. Strategies to enhance polysaccharide release from lignocellulose substrates for enhanced enzymatic action and fermentation of the released fraction would also aid in the efficient recovery of the more uniform modified lignin fraction that the CMF reaction generates to enhance biorefinery profitability.

Comparison of traditional field retting and Phlebia radiata Cel 26 retting of hemp fibres for fibre-reinforced composites.

Classical field retting and controlled fungal retting of hemp using Phlebia radiata Cel 26 (a mutant with low cellulose degrading ability) were compared with pure pectinase treatment with regard to mechanical properties of the produced fibre/epoxy composites. For field retting a classification of the microbial evolution (by gene sequencing) and enzyme profiles were conducted. By phylogenetic frequency mapping, different types of fungi, many belonging to the Ascomycota phylum were found on the fibres during the first 2 weeks of field retting, and thereafter, different types of bacteria, notably Proteobacteria, also proliferated on the field retted fibres. Extracts from field retted fibres exhibited high glucanase activities, while extracts from P. radiata Cel 26 retted fibres showed high polygalacturonase and laccase activities. As a result, fungal retting gave a significantly higher glucan content in the fibres than field retting (77 vs. 67%) and caused a higher removal of pectin as indicated by lower galacturonan content of fibres (1.6%) after fibres were retted for 20 days with P. radiata Cel 26 compared to a galacturonan content of 3.6% for field retted fibres. Effective fibre stiffness increased slightly after retting with P. radiata Cel 26 from 65 to 67 GPa, while it decreased after field retting to 52 GPa. Effective fibre strength could not be determined similarly due to variations in fibre fracture strain and fibre-matrix adhesion. A maximum composite strength with 50 vol% fibres of 307 MPa was obtained using P. radiata Cel 26 compared to 248 MPa with field retting.

Continual removal of H3K9 promoter methylation by Jmjd2 demethylases is vital for ESC self-renewal and early development.

Chromatin-associated proteins are essential for the specification and maintenance of cell identity. They exert these functions through modulating and maintaining transcriptional patterns. To elucidate the functions of the Jmjd2 family of H3K9/H3K36 histone demethylases, we generated conditional Jmjd2a/Kdm4a, Jmjd2b/Kdm4b and Jmjd2c/Kdm4c/Gasc1 single, double and triple knockout mouse embryonic stem cells (ESCs). We report that while individual Jmjd2 family members are dispensable for ESC maintenance and embryogenesis, combined deficiency for specifically Jmjd2a and Jmjd2c leads to early embryonic lethality and impaired ESC self-renewal, with spontaneous differentiation towards primitive endoderm under permissive culture conditions. We further show that Jmjd2a and Jmjd2c both localize to H3K4me3-positive promoters, where they have widespread and redundant roles in preventing accumulation of H3K9me3 and H3K36me3. Jmjd2 catalytic activity is required for ESC maintenance, and increased H3K9me3 levels in knockout ESCs compromise the expression of several Jmjd2a/c targets, including genes that are important for ESC self-renewal. Thus, continual removal of H3K9 promoter methylation by Jmjd2 demethylases represents a novel mechanism ensuring transcriptional competence and stability of the pluripotent cell identity.

Improved material properties of solution-cast starch films: Effect of varying amylopectin structure and amylose content of starch from genetically modified potatoes.

High-amylose potato starches were produced through genetic modification resulting in changed granule morphology and composition, with higher amylose content and increased chain length of amylopectin. The increased amylose content and structural changes in amylopectin enhanced film-forming behavior and improved barrier and tensile properties in starch films. The molecular structure in these starches was related to film-forming properties. Solution-cast films of high-amylose starch revealed a homogeneous structure with increasing surface roughness at higher amylose content, possibly due to amylose aggregation. Films exhibited significantly higher stress and strain at break compared with films of wild-type starch, which could be attributable to the longer chains of amylopectin being involved in the interconnected network and more interaction between chains, as shown using transmission electron microscopy. The oxygen permeability of high-amylose starch films was significantly decreased compared with wild-type starch. The nature of the modified starches makes them an interesting candidate for replacement of non-renewable oxygen and grease barrier polymers used today.