Complete mitochondrial genomes of edible mushrooms: features, evolution, and phylogeny.

Edible mushrooms are an important food source with high nutritional and medicinal value. They are a useful source for studying phylogenetic evolution and species divergence. The exploration of the evolutionary relationships among these species conventionally involves analyzing sequence variations within their complete mitochondrial genomes, which range from 31,854 bp (Cordyceps militaris) to 197,486 bp (Grifolia frondosa). The study of the complete mitochondrial genomes of edible mushrooms has emerged as a critical field of research, providing important insights into fungal genetic makeup, evolution, and phylogenetic relationships. This review explores the mitochondrial genome structures of various edible mushroom species, highlighting their unique features and evolutionary adaptations. By analyzing these genomes, robust phylogenetic frameworks are constructed to elucidate mushrooms lineage relationships. Furthermore, the exploration of different variations of mitochondrial DNA presents novel opportunities for enhancing mushroom cultivation biotechnology and medicinal applications. The mitochondrial genomic features are essential for improving agricultural practices and ensuring food security through improved crop productivity, disease resistance, and nutritional qualities. The current knowledge about the mitochondrial genomes of edible mushrooms is summarized in this review, emphasising their significance in both scientific research and practical applications in bioinformatics and medicine.

A Novel Bacillus amyloliquefaciens Specifically Improving the Solubility and Antioxidant Activities of Edible Bird's Nest.

Edible bird's nest (EBN), a most highly priced and valuable foodstuff, contains high percentage of proteins and carbohydrates. However, proteins adhering to these carbohydrates make the EBN hard and tough, which need to be boiled as the bird's nest soup to make the Chinese cuisine. To overcome the hard and tough texture of EBN and improve the digestion degrees, the present study screened and identified a probiotic strain Bacillus amyloliquefaciens YZW02 from 5-year stored EBN sample completely solubilizing EBN for the first time. The 24-h B. amyloliquefaciens fermented EBN contained 20.30-21.48 mg/mL of the soluble protein contents with a recovery rate of 98-100%, DPPH radical scavenging rate of 84.76% and ABTS radical scavenging capacity of 41.05%. The mixed fermentation of B. amyloliquefaciens YZW02 and Bacillus natto BN1 were further applied to improve the low-MW peptide percentages and antioxidant activities. The mixed-fermentation of B. natto BN1 with 4-h cultured B. amyloliquefaciens YZW02 had the lowest percentage (82.23%) of >12-kDa proteins/peptides and highest percentages of 3-12 kDa, 1-3 kDa and 0.1-1 kDa peptides of 8.6% ± 0.08, 7.57% ± 0.09, 1.77% ± 0.05 and 0.73% ± 0.05, with the highest DPPH, ABTS and •OH scavenging capacity of 90.23%, 46.45% and 49.12%, respectively. These findings would provide an efficient strategy for improving the solubility and antioxidant activities of EBNs.

Neutron Activation Analysis Based on AB-BNCT Treatment Room.

ABSTRACT: Boron neutron capture therapy (BNCT) is an ideal binary targeted radiotherapy for treating refractory tumors. An accelerator-based BNCT (AB-BNCT) neutron source has attracted more and more attention due to its advantages such as higher neutron yield in the keV energy region, less gamma radiation, and higher safety. In addition to 10B, neutrons also react with other elements in the treatment room during BNCT to produce many activation products. Due to the long half-life of some activation products, there will be residual radiation after the end of treatment and the shutdown of the accelerator, which has adverse effects on radiation workers. Therefore, the ambient dose equivalent rate in the treatment room needs to be evaluated. The AB-BNCT neutron source model proposed by Li is studied in this paper. Based on the Monte Carlo method, the Geant4 platform was used to simulate the dose induced by radionuclides near the Beam Shaping Assembly (BSA) of the source. It is concluded that the concrete wall contributed the most to the radiation dose. The dose rate of 2.45 µSv h-1 after 13 min of shutdown meets the dose rate limit of 2.5 µSv h-1, at which point it is safe for workers to enter the treatment room area.

Exploration of the Correlation Between GRHL1 Expression and Tumor Microenvironment in Endometrial Cancer and Immunotherapy.

Introduction: GRHL1 belongs to the family of Grainyhead-like (GRHL). Previous studies have shown that dysregulation of growth and survival pathways is associated with the GRHL family of gene cancers. Immunotherapy with checkpoint inhibitors has changed the treatment paradigm for many tumors, including endometrial cancer (EC). However, the effect of GRHL1 on immunotherapy in EC and its relationship with immune cell infiltration are poorly understood. Methods: Differential expression of GRHL1 between EC and normal EC tissues was analyzed by searching the TCGA database, and the results were verified utilizing immunohistochemistry analyses. Next, the relationship between GRHL1, CD8+ T cells and tumor microenvironment (TME) was also investigated, and the effect of GRHL1 expression on immunotherapy in EC was evaluated. Results: According to the findings, EC tissues had elevated expression levels of GRHL1 relative to normal tissues. Patients with EC who expressed GRHL1 at high levels experienced worse overall survival (OS) and Progression-free survival (PFS) than those whose expression was lower. In addition, GRHL1 expression was negatively correlated with CD8+ T cells, and patients with high GRHL1 expression were less effective in receiving immunotherapy. Conclusion: The expression of GRHL1 was high in EC patients, and high expression of GRHL1 inhibits the proliferation of CD8+ T cells in the tumor microenvironment of EC and affect the efficacy of immunotherapy.

Recent Advances of Oxalate Decarboxylase: Biochemical Characteristics, Catalysis Mechanisms, and Gene Expression and Regulation.

Oxalate decarboxylase (OXDC) is a typical Mn2+/Mn3+ dependent metal enzyme and splits oxalate to formate and CO2 without any organic cofactors. Fungi and bacteria are the main organisms expressing the OXDC gene, but with a significantly different mechanism of gene expression and regulation. Many articles reported its potential applications in the clinical treatment of hyperoxaluria, low-oxalate food processing, degradation of oxalate salt deposits, oxalate acid diagnostics, biocontrol, biodemulsifier, and electrochemical oxidation. However, some questions still remain to be clarified about the role of substrate binding and/or protein environment in modulating the redox properties of enzyme-bound Mn(II)/Mn(III), the nature of dioxygen involved in the catalytic mechanism, and how OXDC acquires Mn(II) /Mn(III). This review mainly summarizes its biochemical and structure characteristics, gene expression and regulation, and catalysis mechanism. We also deep-mined oxalate decarboxylase gene data from National Center for Biotechnology Information to give some insights to explore new OXDC with diverse biochemical properties.

Overall quality changes and deterioration mechanism of fragrant rapeseed oils during 6-Month storage.

The strong-fragrant rapeseed oil (SFRO) is a popular rapeseed oil in China with a low refining degree only degumming with hot water, which remarkably affects its storage stability. The present study compared the overall changes of physical/chemical/nutrient quality of FROs at various temperatures, light wavelengths and headspace volumes. Results showed that red light (680 nm) had a most significant adverse effect on the overall quality of SFRO with the higher correlation coefficients to PV and TOTOX of 0.71 and 0.70, and lower correlation coefficients to chlorophyll and tocopherol of -0.95 and -0.53, respectively. Further studies revealed that red light accelerated the oxidation of fragrant rapeseed oils by degrading chlorophyll to initiate the photo-oxidation process and synthesize high amount of secondary oxidation products including aliphatic and aromatic oxidized compounds from linolenic acid. These findings provided a reference to control the deterioration of FROs by preventing the transmittance of red light.

Grifola frondosa polysaccharides: A review on structure/activity, biosynthesis and engineering strategies.

Polysaccharides are the main polymers in edible fungi Grifola frondosa, playing a crucial role in the physiology and representing the healthy benefits for humans. Recent efforts have well elucidated the fine structures and biological functions of G. frondosa polysaccharides. The recently-rapid developments and increasing availability in fungal genomes also accelerated the better understanding of key genes and pathways involved in biosynthesis of G. frondosa polysaccharides. Herein, we provide a brief overview of G. frondosa polysaccharides and their activities, and comprehensively outline the complex process, genes and proteins corresponding to G. frondosa polysaccharide biosynthesis. The regulation strategies including strain improvement, process optimization and genetic engineering were also summarized for maximum production of G. frondosa polysaccharides. Some remaining unanswered questions in describing the fine synthesis machinery were also pointed out to open up new avenues for answering the structure-activity relationship and improving polysaccharide biosynthesis in G. frondosa. The review hopefully presents a reasonable full picture of activities, biosynthesis, and production regulation of polysaccharide in G. frondosa.

Recent insights into glucans biosynthesis and engineering strategies in edible fungi.

Fungal α/ß-glucans have significant importance in cellular functions including cell wall structure, host-pathogen interactions and energy storage, and wide application in high-profile fields, including food, nutrition, and pharmaceuticals. Fungal species and their growth/developmental stages result in a diversity of glucan contents, structures and bioactivities. Substantial progresses have been made to elucidate the fine structures and functions, and reveal the potential molecular synthesis pathway of fungal α/ß-glucans. Herein, we review the current knowledge about the biosynthetic machineries, including: precursor UDP-glucose synthesis, initiation, elongation/termination and remodeling of α/ß-glucan chains, and molecular regulation to maximally produce glucans in edible fungi. This review would provide future perspectives to biosynthesize the targeted glucans and reveal the catalytic mechanism of enzymes associated with glucan synthesis, including: UDP-glucose pyrophosphate phosphorylases (UGP), glucan synthases, and glucanosyltransferases in edible fungi.

Biochemical and structural characterization of a glucan synthase GFGLS2 from edible fungus Grifola frondosa to synthesize ß-1, 3-glucan.

BACKGROUND: Grifola frondosa is a Basidiomycete fungus belonging to the family of Grifolaceae and the order of Polyporales. ß-Glucans are the main polymers in G. frondosa, playing a crucial role in the physiology and representing the healthy benefits for humans. The membrane-integrated ß-1, 3-glucan synthase (GLS) is responsible for glucan synthesis, cell wall assembly, differentiation and growth of the edible fungi. However, the structural/catalytic characteristics and mechanisms of ß-1, 3-glucan synthases in G. frondosa are still unknown due to their extremely complex structures with multi-transmembranes and large molecular masses. RESULTS: Herein, a ß-1, 3-glucan synthase (GFGLS2) was purified and identified from the cultured mycelia with a specific activity of 60.01 pmol min-1 µg-1 for the first time. The GFGLS2 showed a strict specificity to UDP-glucose with a Vmax value of 1.29 ± 0.04 µM min-1 at pH 7.0 and synthesized ß-1, 3-glucan with a maximum degree of polymerization (DP) of 62. Sequence Similarity Network (SSN) analysis revealed that GFGLS2 has a close relationship with others in Ganoderma sinense, Trametes coccinea, Polyporus brumalis, and Trametes pubescens. With the assistance of 3D structure modelling by AlphaFold 2, molecular docking and molecular dynamics simulations, the central hydrophilic domain (Class III) in GFGLS2 was the main active sites through binding the substrate UDP-glucose to 11 amino acid residues via hydrogen bonds, π-stacking and salt bridges. CONCLUSIONS: The biochemical, 3D structural characterization and potential catalytic mechanism of a membrane-bound ß-1, 3-glucan synthase GFGLS2 from cultured mycelia of G. frondosa were well investigated and would provide a reasonable full picture of ß-1, 3-glucan synthesis in fungi.

Receptor-interacting protein kinase 1 (RIPK1) regulates cervical cancer cells via NF-κB-TNF-α pathway: An in vitro study.

INTRODUCTION: Cervical cancer (CC) is associated with high morbidity and mortality rates in women. Members of the receptor-interacting protein kinase (RIPK) family are important regulators of inflammation and cell death. However, the characteristics, molecular functions, and expression mechanisms of RIPK1 in CC remain unclear. MATERIAL AND METHODS: To determine whether RIPK1 can be used for targeted therapy of CC, we assessed the clinical importance, biological function, and potential impact of RIPK1 in CC in 50 patients with CC. We utilized immunohistochemical staining, transfection, western blotting, cell counting kit-8 assay, colony formation assay, and wound healing assays among others, to elucidate the role of RIPK1 in CC. RESULTS: RIPK1 expression was higher in tumor tissues than in paracancerous tissues. Poor prognosis of CC was linked to RIPK1 upregulation. Furthermore, silencing RIPK1 significantly inhibited the proliferation, migration, and invasion of CC cells in vitro. We also established that RIPK1 increased cell migration, invasion, and multiplication by regulating nuclear factor kappa-B (NF-κB) and tumor necrosis factor (TNF). DISCUSSION: RIPK1 activates NF-κB and regulates TNF release to enhance the proliferation and spread of CC cells while suppressing their apoptosis. Therefore, RIPK1 plays a key role in the formation and progression of CC and is a potential target for CC treatment.

Facile one-step synthesis of gold nanoparticles using Viscum album and evaluation of their antibacterial potential.

Nanostructure gold nanoparticles (Au NPs) are well-known biological active materials, synthesised under different environment-friendly approaches that has gained significant interest in the field of biomedicine. This study investigated a novel, fast, easy, cost-effective and the eco-friendly method to synthesise Au NPs from mediated Viscum album Linn plant extract, where the plant metabolites act as stabilising and reducing agents. The synthesised Au NPs were analysed by UV/Vis spectroscopy that gave strong signals and a sharp absorption peak at 545nm due to the presence of surface plasmon resonance (SPR) bands. In addition, energy dispersive X-ray spectroscopy (EDX) showed that strong signals of Au NPs appeared at 9.7 and 2.3keV, as the rays of light passed. X-ray diffraction recognised the crystalline material and provided information on the cell unit that the synthesised Au NPs are face-centreed cubic in structure. The diffraction of X-ray spectra showed intense peaks at 38.44°, 44.7°, 44.9° and 77.8°. The mediated V. album plant extracts and synthesised Au NPs were screened against gram-positive and gram-negative (Enterobacter , Salmonella typhi , Escheria coli and Bacillus subtilis ) bacterial strains, confirming their antibacterial potential. Au NPs showed strong antibacterial activity due to its unique steric configuration. Au NPs damaged bacterial cell membrane leading to the leakage of the cytoplasm and death of the cell.

[Study on the sensitivity of a volumetric modulated arc therapy plan verification equipment on multi-leaf collimator opening and closing errors and its gamma pass rate limit].

To investigate the γ pass rate limit of plan verification equipment for volumetric modulated arc therapy (VMAT) plan verification and its sensitivity on the opening and closing errors of multi-leaf collimator (MLC), 50 cases of nasopharyngeal carcinoma VMAT plan with clockwise and counterclockwise full arcs were randomly selected. Eight kinds of MLC opening and closing errors were introduced in 10 cases of them, and 80 plans with errors were generated. Firstly, the plan verification was conducted in the form of field-by-field measurement and true composite measurement. The γ analysis with the criteria of 3% dose difference, distance to agreement of 2 mm, 10% dose threshold, and absolute dose global normalized conditions were performed for these fields. Then gradient analysis was used to investigate the sensitivity of field-by-field measurement and true composite measurement on MLC opening and closing errors, and the receiver operating characteristic curve (ROC) was used to investigate the optimal threshold of γ pass rate for identifying errors. Tolerance limits and action limits for γ pass rates were calculated using statistical process control (SPC) method for another 40 cases. The error identification ability using the tolerance limit calculated by SPC method and the universal tolerance limit (95%) were compared with using the optimal threshold of ROC. The results show that for the true composite measurement, the clockwise arc and the counterclockwise arc, the descent gradients of the γ passing rate with per millimeter MLC opening error are 10.61%, 7.62% and 6.66%, respectively, and the descent gradients with per millimeter MLC closing error are 9.75%, 7.36% and 6.37%, respectively. The optimal thresholds obtained by the ROC method are 99.35%, 97.95% and 98.25%, respectively, and the tolerance limits obtained by the SPC method are 98.98%, 97.74% and 98.62%, respectively. The tolerance limit calculated by SPC method is close to the optimal threshold of ROC, both of which could identify all errors of ±2 mm, while the universal tolerance limit can only partially identify them, indicating that the universal tolerance limit is not sensitive on some large errors. Therefore, considering the factors such as ease of use and accuracy, it is suggested to use the true composite measurement in clinical practice, and to formulate tolerance limits and action limits suitable for the actual process of the institution based on the SPC method. In conclusion, it is expected that the results of this study can provide some references for institutions to optimize the radiotherapy plan verification process, set appropriate pass rate limit, and promote the standardization of plan verification.

Sensitivity of Three Patient-Specific Quality Assurance Systems to MLC Aperture Errors With Volumetric Modulated Arc Therapy.

Purpose: To compare the sensitivity of ArcCHECK (AC), portal dosimetry (PD), and an in-house logfile-based system (LF) to multileaf collimators (MLC) aperture errors and the ability to identify these errors. Methods and Materials: For 12 retrospective original head and neck volumetric modulated arc therapy (VMAT) plans, MLC aperture errors of ± 0.4mm, ± 1.2mm, ± 2mm, and ± 3mm were introduced for each plan, resulting in 96 plans with errors. AC, PD, and LF were used for the gamma evaluation at 3%/3mm, 3%/2mm, and 2%/2mm criteria. Gradient analysis was used to evaluate the sensitivity to MLC aperture errors. The area under the curve (AUC) obtained from the receiver operating characteristic (ROC) curve was used to evaluate the ability to identify MLC aperture errors and dose errors, and the optimal cut-off value to identify the error was obtained. Results: The gamma pass rate (%GP) of LF had the smallest descent gradient as the MLC error increases in any case. The descent gradient of PD was larger than AC, except for the case at the 2%/2mm criteria. For the 3%/3mm criteria, the MLC aperture errors that can be perfectly identified by AC, PD, and LF were ± 3mm, ± 2mm, and ± 1.2mm, respectively, and the average percent dose error (%DEs) of dose metrics in targets that can be perfectly identified were 4% to 5%, 3% to 4%, and 2% to 3%, respectively. For the 3%/2mm criteria, the errors that AC, PD, and LF can perfectly identify were the same as the 3%/3mm criteria. For the 2%/2mm criteria, AC can perfectly identify the MLC error of ± 2mm and the %DE of 3% to 4%. PD and LF can identify the MLC error of ± 1.2mm and the %DE of 2% to 3%. Conclusion: Different patient-specific quality assurance (PSQA) systems have different sensitivity and recognition abilities to MLC aperture errors. Institutions should formulate their own customized %GP limits based on their PSQA process through ROC or other methods.

Spatio-Temporal Modification of Lignin Biosynthesis in Plants: A Promising Strategy for Lignocellulose Improvement and Lignin Valorization.

Lignin is essential for plant growth, structural integrity, biotic/abiotic stress resistance, and water transport. Besides, lignin constitutes 10-30% of lignocellulosic biomass and is difficult to utilize for biofuel production. Over the past few decades, extensive research has uncovered numerous metabolic pathways and genes involved in lignin biosynthesis, several of which have been highlighted as the primary targets for genetic manipulation. However, direct manipulation of lignin biosynthesis is often associated with unexpected abnormalities in plant growth and development for unknown causes, thus limiting the usefulness of genetic engineering for biomass production and utilization. Recent advances in understanding the complex regulatory mechanisms of lignin biosynthesis have revealed new avenues for spatial and temporal modification of lignin in lignocellulosic plants that avoid growth abnormalities. This review explores recent work on utilizing specific transcriptional regulators to modify lignin biosynthesis at both tissue and cellular levels, focusing on using specific promoters paired with functional or regulatory genes to precisely control lignin synthesis and achieve biomass production with desired properties. Further advances in designing more appropriate promoters and other regulators will increase our capacity to modulate lignin content and structure in plants, thus setting the stage for high-value utilization of lignin in the future.

Functional Characterization and Structural Basis of the ß-1,3-Glucan Synthase CMGLS from Mushroom Cordyceps militaris.

ß-1,3-Glucan synthases play key roles in glucan synthesis, cell wall assembly, and growth of fungi. However, their multi-transmembrane domains (over 14 TMHs) and large molecular masses (over 100 kDa) significantly hamper understanding of their catalytic characteristics and mechanisms. In the present study, the 5841-bp gene CMGLS encoding the 221.7 kDa membrane-bound ß-1,3-glucan synthase CMGLS in Cordyceps militaris was cloned, identified, and structurally analyzed. CMGLS was partially purified with a specific activity of 87.72 pmol/min/µg, a purification fold of 121, and a yield of 10.16% using a product-entrapment purification method. CMGLS showed a strict specificity to UDP-glucose with a Km value of 84.28 µM at pH 7.0 and synthesized ß-1,3-glucan with a maximum degree of polymerization (DP) of 70. With the assistance of AlphaFold and molecular docking, the 3D structure of CMGLS and its binding features with substrate UDP-glucose were proposed for the first time to our knowledge. UDP-glucose potentially bound to at least 11 residues via hydrogen bonds, π-stacking ,and salt bridges, and Arg 1436 was predicted as a key residue directly interacting with the moieties of glucose, phosphate, and the ribose ring on UDP-glucose. These findings would open an avenue to recognize and understand the glucan synthesis process and catalytic mechanism of ß-1,3-glucan synthases in mushrooms.

The ß-1,3-glucan synthase gene GFGLS2 plays major roles in mycelial growth and polysaccharide synthesis in Grifola frondosa.

ß-1,3-Glucans are well-known biological and health-promoting compounds in edible fungi. Our previous results characterized a glucan synthase gene (GFGLS) of Grifola frondosa for the first time to understand its role in mycelial growth and glucan biosynthesis. In the present study, we identified and functionally reannotated another glucan synthase gene, GFGLS2, based on our previous results. GFGLS2 had a full sequence of 5944 bp including 11 introns and 12 exons and a coding information for 1713 amino acids of a lower molecular weight (195.2 kDa) protein with different conserved domain sites than GFGLS (5927 bp with also 11 introns and a coding information for 1781 aa). Three dual-promoter RNA-silencing vectors, pAN7-iGFGLS-dual, pAN7-iGFGLS2-dual, and pAN7-CiGFGLS-dual, were constructed to downregulate GFGLS, GFGLS2, and GFGLS/GFGLS2 expression by targeting their unique exon sequence or conserved functional sequences. Silencing GFGLS2 resulted in higher downregulation efficiency than silencing GFGLS. Cosilencing GFGLS and GFGLS2 had a synergistic downregulation effect, with slower mycelial growth and glucan production by G. frondosa. These findings indicated that GFGLS2 plays major roles in mycelial growth and polysaccharide synthesis and provides a reference to understand the biosynthesis pathway of mushroom polysaccharides. KEY POINTS: • The 5944-bp glucan synthase gene GFGLS2 of G. frondosa was cloned and reannotated • GFGLS2 showed identity and significant differences with the previously identified GFGLS • GFGLS2 played a major role in fermentation and glucan biosynthesis.

Changes of structures and biosynthesis/hydrolysis-associated genes expression of glucans at different Volvariella volvacea maturity stages.

In the present study, effects of maturity stage on structural characteristics and biosynthesis/hydrolysis-associated genes expression of glucans from Volvariella volvacea fruit body were well investigated. Elongation and pileus expansion stages decreased total soluble carbohydrate and protein contents to 17.09 mg/g and 8.33 mg/g, and significantly accumulated the total amino acids contents to 32.37 mg/g. Yields of crude polysaccharides significantly increased to 8.12% at egg stage and decreased to 3.72% at pileus expansion stage. Purified VVP I-a and VVP I-b were proved to be α-glucans. The maturity process affected the monosaccharide compositions, decreased the molecular weights of VVP I-a and VVP I-b with decreased transcription levels of glucan biosynthesis-associated enzyme genes vvugp and vvgls and increased glucan hydrolysis-associated glucanase gene vvexg2 expression with no significant effects on backbone structures including glycosidic linkages and configurations. The findings would benefit for understanding change patterns of V. volvacea glucan structures and their biosynthesis/hydrolysis-associated genes expression at maturity stages.

Improved glucose and xylose co-utilization by overexpression of xylose isomerase and/or xylulokinase genes in oleaginous fungus Mucor circinelloides.

Most of the oleaginous microorganisms cannot assimilate xylose in the presence of glucose, which is the major bottleneck in the bioconversion of lignocellulose to biodiesel. Our present study revealed that overexpression of xylose isomerase (XI) gene xylA or xylulokinase (XK) gene xks1 increased the xylose consumption by 25 to 37% and enhanced the lipid content by 8 to 28% during co-fermentation of glucose and xylose. In xylA overexpressing strain Mc-XI, the activity of XI was 1.8-fold higher and the mRNA level of xylA at 24 h and 48 h was 11- and 13-fold higher than that of the control, respectively. In xks1 overexpressing strain Mc-XK, the mRNA level of xks1 was 4- to 11-fold of that of the control strain and the highest XK activity of 950 nmol min-1 mg-1 at 72 h which was 2-fold higher than that of the control. Additionally, expression of a translational fusion of xylA and xks1 further enhanced the xylose utilization rate by 45%. Our results indicated that overexpression of xylA and/or xks1 is a promising strategy to improve the xylose and glucose co-utilization, alleviate the glucose repression, and produce lipid from lignocellulosic biomass in the oleaginous fungus M. circinelloides. KEY POINTS: • Overexpressing xylA or xks1 increased the xylose consumption and the lipid content. • The xylose isomerase activity and the xylA mRNA level were enhanced in strain Mc-XI. • Co-expression of xylA and xks1 further enhanced the xylose utilization rate by 45%.

Characterization of a transcriptional regulator PtxS from Pseudomonas plecoglossicida for regulating 2-ketogluconic acid metabolism.

Homologs of PtxS are ubiquitous transcriptional regulators controlling the expression of the glucose dehydrogenase and kgu operon to globally regulate the 2-ketogluconic acid (2KGA) metabolism in Pseudomonas. In the present study, a PtxS from a 2KGA industrial producer Pseudomonas plecoglossicida JUIM01 (PpPtxS) was heterologously expressed in E. coli BL21(DE3), then structurally and functionally characterized. The obtained results showed that PpPtxS was a 36.65-kDa LacI-family transcriptional regulator. 2KGA was the sole effector of PpPtxS. Glucose negatively affected the molecular binding of PpPtxS and 2KGA, and gluconic acid inhibited the PpPtxS-2KGA binding reaction. PpPtxS in water solution mainly existed as a dimer and bound to two molecules of 2KGA. The effector 2KGA mainly bound to the region close to the C-terminal of PpPtxS by interacting with the 299th to the 301st amino acids (Ala, Gln, Pro, Thr, Glu and Arg). PpPtxS specifically recognized and bound to a 14-bp palindrome sequence (5'-TGAAACCGGTTTCA-3') due to its conserved HTH motif at the N-terminal. The characterization of PpPtxS in this study would provide a theoretical guidance for the industrial production of 2KGA.

The role of Rho1 gene in the cell wall integrity and polysaccharides biosynthesis of the edible mushroom Grifola frondosa.

Grifola frondosa polysaccharides, especially ß-glucans, showed the significant antitumor, hypoglycemic, and immune-stimulating activities. In the present study, a predominant regulatory subunit gfRho1p of ß-1,3-glucan synthase in G. frondosa was identified with a molecular weight of 20.79 kDa and coded by a putative 648-bp small GTPase gene gfRho1. By constructing mutants of RNA interference and over-expression gfRho1, the roles of gfRho1 in the growth, cell wall integrity and polysaccharide biosynthesis were well investigated. The results revealed that defects of gfRho1 slowed mycelial growth rate by 22% to 33%, reduced mycelial polysaccharide and exo-polysaccharide yields by 4% to 7%, increased sensitivity to cell wall stress, and down-regulated gene transcriptions related to PKC-MAPK signaling pathway in cell wall integrity. Over-expression of gfRho1 improved mycelial growth rate and polysaccharide production of G. frondosa. Our study supports that gfRho1 is an essential regulator for polysaccharide biosynthesis, cell growth, cell wall integrity and stress response in G. frondosa.