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Fungi, as eukaryotic organisms, contain two genomes, the mitochondrial genome and the nuclear genome, in their cells. How the two genomes evolve and correlate to each other is debated. Herein, taking the gourmet pine mushroom Tricholoma matsutake as an example, we performed comparative mitogenomic analysis using samples collected from diverse locations and compared the evolution of the two genomes. The T. matsutake mitogenome encodes 49 genes and is rich of repetitive and non-coding DNAs. Six genes were invaded by up to 11 group I introns, with one cox1 intron cox1P372 showing presence/absence dynamics among different samples. Bioinformatic analyses suggested limited or no evidence of mitochondrial heteroplasmy. Interestingly, hundreds of mitochondrial DNA fragments were found in the nuclear genome, with several larger than 500 nt confirmed by PCR assays and read count comparisons, indicating clear evidence of transfer of mitochondrial DNA into the nuclear genome. Nuclear DNA of T. matsutake showed a higher mutation rate than mitochondrial DNA. Furthermore, we found evidence of incongruence between phylogenetic trees derived from mitogenome and nuclear DNA sequences. Together, our results reveal the dynamic genome evolution of the gourmet pine mushroom.

Afzelia africana, Entada africana, and Pterocarpus erinaceus are economically important tropical tree species in Africa. In this study, we performed morphological and molecular analyses to examine the mycorrhizal status of these species in their natural habitat in Benin, West Africa. Fine roots were examined using trypan blue staining. Putative arbuscular mycorrhizal fungi (AMF) were identified by extracting root DNA and sequencing the large subunit region of ribosomal DNA. Operational taxonomic units (OTUs) were positioned on a consensus phylogenetic tree produced using maximum likelihood and neighbor-joining models. Morphological analyses revealed vesicles, arbuscules, and inter- and intracellular hyphae in roots of all three tree species. Molecular analyses revealed 39 AMF OTUs, among which 18 were from roots of E. africana, 15 from P. erinaceus, and 9 from A. africana. These OTUs belonged to Glomus, Nanoglomus, Rhizoglomus, Sclerocystis, Septoglomus, and Scutellospora; two unidentified species belonged to the family Glomeraceae. This is the first study to report AMF in roots of A. africana, E. africana, and P. erinaceus in Benin, and opens new avenues for future studies.

RNF8 (ring finger protein 8), a RING finger E3 ligase best characterized for its role in DNA repair and sperm formation via ubiquitination, has been found to promote tumor metastasis in breast cancer recently. However, whether RNF8 also plays a role in other types of cancer, especially in lung cancer, remains unknown. We show here that RNF8 expression levels are markedly increased in human lung cancer tissues and negatively correlated with the survival time of patients. Overexpression of RNF8 promotes the EMT process and migration ability of lung cancer cells, while knockdown of RNF8 demonstrates the opposite effects. In addition, overexpression of RNF8 activates the PI3K/Akt signaling pathway, knockdown of RNF8 by siRNA inhibits this activation, and pharmacologic inhibition of PI3K/Akt in RNF8-overexpressing cells also reduces the expression of EMT markers and the ability of migration. Furthermore, RNF8 is found to directly interact with Slug and promoted the K63-Ub of Slug, and knockdown of Slug disrupts RNF8-dependent EMT in A549 cells, whereas overexpression of Slug rescues RNF8-dependent MET in H1299 cells, and depletion of RNF8 expression by shRNA inhibits metastasis of lung cancer cells in vivo. Taken together, these results indicate that RNF8 is a key regulator of EMT process in lung cancer and suggest that inhibition of RNF8 could be a useful strategy for lung cancer treatment. Implications: This study provides a new mechanistic insight into the novel role of RNF8 and identifies RNF8 as a potential new therapeutic target for the treatment of lung cancer.

Salmonella is a significant food-borne pathogen that infects a large number of people worldwide. In this study, a lytic bacteriophage vB_SenS_SE1 capable of infecting Salmonella is isolated from municipal wastewater in Beijing, and its biological and genomic features are analyzed. Transmission electron micrograph shows that vB_SenS_SE1 is likely a Siphoviridae virus, with an icosahedral head and a long non-contracted tail. The stability test in vitro reveals that it is stable at 4-50 °C and pH 4-12. Based on the one-step growth curve, vB_SenS_SE1 has a 60-min exponential phase and a low burst size (19 PFU per cell). Bioinformatics analysis reveals that vB_SenS_SE1 consists of a circular, double-stranded DNA molecule of 40,987 bp with a GC content of 51.2%. Its genome carries 63 predicted open reading frames (orfs), with 22 orfs encoding known proteins. Phylogenetic analysis of the large terminase subunit shows that vB_SenS_SE1 exhibits strong homology to Salmonella phage St161, St162, VSiP, and FSL SP-031. The CoreGenes analysis shows that it is a member of the virus genus Cornellvirus. The features of phage vB_SenS_SE1 suggest that it has the potential to be an agent to control Salmonella.

Long non-coding RNAs (lncRNAs) regulate gene expression in a variety of ways at epigenetic, chromatin remodeling, transcriptional, and translational levels. Accumulating evidence suggests that lncRNA X-inactive specific transcript (lncRNA Xist) serves as an important regulator of cell growth and development. Despites its original roles in X-chromosome dosage compensation, lncRNA Xist also participates in the development of tumor and other human diseases by functioning as a competing endogenous RNA (ceRNA). In this review, we comprehensively summarized recent progress in understanding the cellular functions of lncRNA Xist in mammalian cells and discussed current knowledge regarding the ceRNA network of lncRNA Xist in various diseases. Long non-coding RNAs (lncRNAs) are transcripts that are more than 200 nt in length and without an apparent protein-coding capacity (Furlan and Rougeulle, 2016; Maduro et al., 2016). These RNAs are believed to be transcribed by the approximately 98–99% non-coding regions of the human genome (Derrien et al., 2012; Fu, 2014; Montalbano et al., 2017; Slack and Chinnaiyan, 2019), as well as a large variety of genomic regions, such as exonic, tronic, and intergenic regions. Hence, lncRNAs are also divided into eight categories: Intergenic lncRNAs, Intronic lncRNAs, Enhancer lncRNAs, Promoter lncRNAs, Natural antisense/sense lncRNAs, Small nucleolar RNA-ended lncRNAs (sno-lncRNAs), Bidirectional lncRNAs, and non-poly(A) lncRNAs (Ma et al., 2013; Devaux et al., 2015; St Laurent et al., 2015; Chen, 2016; Quinn and Chang, 2016; Richard and Eichhorn, 2018; Connerty et al., 2020). A range of evidence has suggested that lncRNAs function as key regulators in crucial cellular functions, including proliferation, differentiation, apoptosis, migration, and invasion, by regulating the expression level of target genes via epigenomic, transcriptional, or post-transcriptional approaches (Cao et al., 2018). Moreover, lncRNAs detected in body fluids were also believed to serve as potential biomarkers for the diagnosis, prognosis, and monitoring of disease progression, and act as novel and potential drug targets for therapeutic exploitation in human disease (Jiang W. et al., 2018; Zhou et al., 2019a). Long non-coding RNA X-inactive specific transcript (lncRNA Xist) are a set of 15,000–20,000 nt sequences localized in the X chromosome inactivation center (XIC) of chromosome Xq13.2 (Brown et al., 1992; Debrand et al., 1998; Kay, 1998; Lee et al., 2013; da Rocha and Heard, 2017; Yang Z. et al., 2018; Brockdorff, 2019). Previous studies have indicated that lncRNA Xist regulate X chromosome inactivation (XCI), resulting in the inheritable silencing of one of the X-chromosomes during female cell development. Also, it serves a vital regulatory function in the whole spectrum of human disease (notably cancer) and can be used as a novel diagnostic and prognostic biomarker and as a potential therapeutic target for human disease in the clinic (Liu et al., 2018b; Deng et al., 2019; Dinescu et al., 2019; Mutzel and Schulz, 2020; Patrat et al., 2020; Wang et al., 2020a). In particular, lncRNA Xist have been demonstrated to be involved in the development of multiple types of tumors including brain tumor, Leukemia, lung cancer, breast cancer, and liver cancer, with the prominent examples outlined in Table 1. It was also believed that lncRNA Xist (Chaligne and Heard, 2014; Yang Z. et al., 2018) contributed to other diseases, such as pulmonary fibrosis, inflammation, neuropathic pain, cardiomyocyte hypertrophy, and osteoarthritis chondrocytes, and more specific details can be found in Table 2. This review summarizes the current knowledge on the regulatory mechanisms of lncRNA Xist on both chromosome dosage compensation and pathogenesis (especially cancer) processes, with a focus on the regulatory network of lncRNA Xist in human disease.

Forest trees are colonised by different species of ectomycorrhizal (ECM) fungi that interact competitively or mutualistically with one another. Most ECM fungi can produce sporocarps. To date, the effects of co-colonising fungal species on sporocarp formation in ECM fungi remain unknown. In this study, we examined host plant growth, mycorrhizal colonisation, and sporocarp formation when roots of Pinus densiflora are colonised by Laccaria japonica and three other ECM fungal species (Cenococcum geophilum, Pisolithus sp., and Suillus luteus). Sporocarp numbers were recorded throughout the experimental period. The biomass, photosynthetic rate, and mycorrhizal colonisation rate of the seedlings were also measured at 45days, 62days, and 1year after seedlings were transplanted. Results indicated that C. geophilum and S. luteus may negatively impact mycorrhizal colonisation and sporocarp formation in L. japonica. Sporocarp formation in L. japonica was positively correlated with conspecific mycorrhizal colonisation but negatively correlated with the biomass of seedlings of P. densiflora. The co-occurring ECM fungi largely competed with L. japonica, resulting in various effects on mycorrhizal colonisation and sporocarp formation in L. japonica. A variety of mechanisms may be involved in the competitive interactions among the different ECM fungal species, including abilities to more rapidly colonise root tips, acquire soil nutrients, or produce antibiotics. These mechanisms need to be confirmed in further studies.

Background Pinene is a monoterpene, that is used in the manufacture of fragrances, insecticide, fine chemicals, and renewable fuels. Production of pinene by metabolic-engineered microorganisms is a sustainable method. Purple non-sulfur photosynthetic bacteria belong to photosynthetic chassis that are widely used to synthesize natural chemicals. To date, researches on the synthesis of pinene by purple non-sulfur photosynthetic bacteria has not been reported, leaving the potential of purple non-sulfur photosynthetic bacteria synthesizing pinene unexplored. Results Rhodobacter sphaeroides strain was applied as a model and engineered to express the fusion protein of heterologous geranyl diphosphate synthase (GPPS) and pinene synthase (PS), hence achieving pinene production. The reaction condition of pinene production was optimized and 97.51 μg/L of pinene was yielded. Then, genes of 1-deoxy-d-xylulose 5-phosphate synthase, 1-deoxy-d-xylulose 5-phosphate reductoisomerase and isopentenyl diphosphate isomerase were overexpressed, and the ribosome binding site of GPPS-PS mRNA was optimized, improving pinene titer to 539.84 μg/L. Conclusions In this paper, through heterologous expression of GPPS-PS, pinene was successfully produced in R. sphaeroides, and pinene production was greatly improved by optimizing the expression of key enzymes. This is the first report on pinene produce by purple non-sulfur photosynthetic bacteria, which expands the availability of photosynthetic chassis for pinene production.

Most edible ectomycorrhizal (ECM) mushrooms are currently harvested from nature and many of them are high-priced. Demand for the wild mushrooms as a culinary delicacy has stimulated research that aims to understand (1) the puzzled role that the ECM fungi play in the forest ecosystem, and (2) nutritional and other requirements for fruiting, which is highly variable. In this review, we focus on understanding of the ECM fungi mediated carbon and nitrogen movement between the symbiotic partners and on the interactions with other fungi in forest ecosystems. Thereby, we better understand the diverse nitrogen requirements for edible ECM fungal growth and mushroom fruiting. We attempt to provide a theoretical basis for the future research of edible ECM mushrooms in wild and controlled conditions.

Background Lung cancer is one of the most malignant types of cancer worldwide. Recently, the pivotal role of miRNAs in carcinogenesis and tumor metastasis have been reported for their direct regulation of specific target genes. However, the precise roles of miR-654-5p in lung adenocarcinoma have been poorly investigated. In this study, we designed a series of closed-loop integrated bioinformatic analyses and in vitro experimental validation to explore the main function and regulation pattern of miR-654-5p, and elucidate its roles in lung adenocarcinoma metastasis and prognosis both in silico and in vitro. Method Data from The Cancer Genome Atlas (TCGA) were used to analyze the pan-cancer prognostic value of miR-654-5p. miRanda, TargetScan, RNA22 and miRWalks were utilized to screen the targets of miR-654-5p. Metascape and miRWalks were used to perform gene ontology analyses and multiple enrichment analyses. STRING database and Cytoscape were used to construct protein-protein network and identify hub genes. GEPIA 2.0 were used to perform pan-cancer expression and survival analyses of hub genes. The regulation between miRNAs and predicted genes were verified by qPCR, western blotting and dual-luciferase system. In addition, EMT hallmarks detection, cell proliferation assay and wound healing assay were used to demonstrate the predicted functions of miR-654-5p experimentally. Results We identified 275 potential targets of miR-654-5p and validated the direct regulatory effects of miR-654-5p on RNF8 in vitro as a proof of concept. Furthermore, we revealed the pivotal functions and disease association of miR-654-5p and experimentally validated the tumor suppressor roles of miR-645-5p that inhibited lung cancer cell epithelial-mesenchymal transition process, cell proliferation, and migration capacity. Among these 275 targets, 11 highly interconnected hub genes in lung adenocarcinoma were selected and studied through pan-cancer expression and pan-cancer survival analysis, in order to assess these targets’ clinical value. Conclusions our research not only identified 275 hub target genes of miR-654-5p in silico and performed experimental validation but also revealed the tumor-suppressive roles of miR-654-5p in lung adenocarcinoma metastasis in vitro for the first time.

Protein expression and degradation are fundamental to cell function and physiological status of organisms. Interfering with protein expression not only provides powerful strategies to analyze the function of proteins but also inspires effective treatment methods for diseases caused by protein dysfunction. Recently, harnessing the power of the ubiquitin-proteasome system for targeted protein degradation (TPD) has become the focus of researches. Over the past two decades, TPD technologies, such as E3 ligase modification, PROTACs, and the Trim-Away method, have successfully re-oriented the ubiquitin-proteasome pathway and thus degraded many pathogenic proteins and even "undruggable" targets. However, A low-cost, convenient, and modularized TPD method is currently not available. Herein, we proposed a synthetic biology TPD method, termed Predator, by integrating the classic function of E3 ligase Trim21 and the expression of a bifunctional fusion protein that links Trim21 and the target protein, which leads to the formation of a ternary complex inside mammalian cells and therefore induce the ubiquitination and subsequent proteasome-dependent degradation of the target protein. We first proved this concept by using nanobody and scFv as the targeting module for the Predator system to degrade free GFP and membrane protein ErbB3, respectively. Then, we give an example of how the engineered Predator system can be developed towards biomedical solutions in the context of diabetes mellitus. Ligands-receptor interaction and adenovirus-mediated gene delivery were introduced to the Predator system, and we found this bifunctional fusion protein, in which glucagon was selected to function as the targeting module, downregulated the endogenous glucagon receptor (GCGR) and attenuated glucagon-stimulated glucose production in primary hepatocytes. Although preliminarily, our results showed that this Predator system is a highly modularized and convenient TPD method with good potential for both fundamental researches and clinical usage.Graphic abstract

Tricholoma matsutake is known to be the dominant fungal species in matsutake fruitbody neighboring (shiro) soil. To understand the mechanisms behind matsutake dominance, we studied the bacterial communities in matsutake dominant shiro soil and non-shiro soil, isolated the strains of Streptomyces from matsutake mycorrhizal root tips both from shiro soil and from the Pinus densiflora seedlings cultivated in shiro soil. Further, we investigated three Streptomyces spp. for their ability to inhibit fungal growth and Pinus densiflora seedling root elongation as well as two strains for their antifungal and antioxidative properties.Our results showed that Actinobacteria was the most abundant phylum in shiro soil. However, the differences in the Actinobacterial community composition (phylum or order level) between shiro and non-shiro soils were not significant, as indicated by PERMANOVA analyses. A genus belonging to Actinobacteria, Streptomyces, was present on the matsutake mycorrhizas, although in minority. The two antifungal assays revealed that the broths of three Streptomyces spp. had either inhibitory, neutral or promoting effects on the growth of different forest soil fungi as well as on the root elongation of the seedlings. The extracts of two strains, including one isolated from the P. densiflora seedlings, inhibited the growth of either pathogenic or ectomycorrhizal fungi. The effect depended on the medium used to cultivate the strains, but not the solvent used for the extraction. Two Streptomyces spp. showed antioxidant activity in one out of three assays used, in a ferric reducing antioxidant power assay. The observed properties seem to have several functions in matsutake shiro soil and they may contribute to the protection of the shiro area for T. matsutake dominance.

The human gastrointestinal tract is estimated to be colonised by over 1014 bacteria, approximately 10-fold of the total number of cells in the human body. Recently, an important role of the gut microbiota composition in many areas, such as normal development, immune system maturation, CNS functions, has been discovered. Also, accumulating evidence suggests that gut bacteria play critical roles in maintaining human health in many aspects. Gut microbiota dysbiosis may lead to a number of diseases, including gastrointestinal disorders, obesity, cardiovascular diseases, allergy and central nervous system-related diseases. In this review, we propose that novel effective and safe therapy methods for human diseases based on modulation of gut microbiota would attract more attention from both scientists and clinicians.

Background: Single nucleotide polymorphisms (SNPs) of small non-coding RNAs (sncRNAs) can influence sncRNA function and target gene expression to mediate the risk of certain diseases. The aim of the present study was to evaluate the prognostic relevance of sncRNA SNPs for colorectal cancer, which has not been well characterized to date. Methods: We comprehensively examined 31 common SNPs of sncRNAs, and assessed the impact of these variants on survival in a cohort of 188 patients with colorectal cancer. Results: Three SNPs were significantly associated with survival of patients with colorectal cancer after correction for multiple testing, and two of the SNPs (hsa-mir-196a-2 rs11614913 and U85 rs714775) remained significant in multivariate analyses. Additional in silico analysis provided further evidence of this association, since the expression levels of the target genes of the hsa-miR-196a (HOXA7, HOXB8, and AKT1) were significantly correlated with colorectal cancer progression. Furthermore, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses indicated that hsa-miR-196a is associated with well-known oncogenic pathways, including cellular protein modification process, mitotic cell cycle, adherens junction, and extracellular matrix receptor interaction pathways. Conclusion: Our results suggest that SNPs of sncRNAs could play a critical role in cancer progression, and that hsa-miR-196a might be a valuable biomarker or therapeutic target for colorectal cancer patients.

The androgen receptor (AR) poly‐glutamine polymorphism (AR‐Q) was reported to play role in endometrial cancer (EMCA) development, yet controversial. Environmental factors interact with genetic variation have been reported in EMCA. Aerosol toxins, polycyclic aromatic hydrocarbon benzo[a]pyrene (BaP), are EMCA facilitators. This report examined the interplay between AR‐Qs and BaP in EMCA. During analysing patient AR‐Q polymorphism and Aryl hydrocarbon Receptor (AhR) expressions, we found overall survival (OS) benefit is ascending with AR‐Q lengths (5‐year OS of 61.3% in Q length 23). And AhR is higher expressed in short AR‐Q tumour compared to that in long AR‐Q patient. In vitro study found androgen‐response element (ARE) activity descends with AR‐Qs length (Q13 > Q25 > Q35), whereas BaP suppresses ARE activities in EMCA cells. Furthermore, AR‐Q13 (but not AR‐Q25, or ‐35) enhances BaP‐induced dioxin‐responsive element (DRE) activity. Lastly, AR‐Q13 exerts higher colony‐forming capacity than other AR‐Qs, and knock‐down AhR abolished AR‐Q13‐mediated colony numbers. This study demonstrated a possible interaction of gene (AR‐Q polymorphism) and environmental toxins (e.g. BaP) to affect cancer progression. A large‐scale epidemiology and public health survey on the interaction of environmental toxin and AR poly‐Q in EMCA is suggested.

Jingzhaotoxin-III (JZTX-III), a 36-residue peptide cardiotoxin containing three pairs of disulphide bonds, has been characterized from the venom of the Chinese tarantula Chilobrachys jingzhao. JZTX-III is a promising target for drug development and clinical application, due to its specific inhibitory effects on the human voltage-gated potassium channel subtype hKv2.1 and sodium channel subtype hNav1.5, which are mainly expressed in the cardiac myocytes. The most direct way to obtain JZTX-III is by extraction from the native venom of the tarantula jingzhao, but the amount is often insufficient to meet research and clinical demands. Therefore, there is need for an efficient expression system that can provide a larger quantity of JZTX-III. In this paper, we utilized a galactose auto-induction system to assist the Escherichia coli strain SHuffle T7 Express to express recombinant JZTX-III, followed by in situ purification on a Ni-nitrilotriacetic acid (Ni-NTA) column. Subsequent experiments were performed to demonstrate the advantages of the galactose auto-induction method and to optimize the incubation conditions. Under the optimal expression conditions, the product of the purified bioactive recombinant JZTX-III reached 12.1 mg/L. Furthermore, it is expected that this expression method can be widely applied to the heterologous expression of other disulphide-bond-rich peptides.

The genome sequence, morphology, and genetic features of a novel phage, named SSE1, is reported here. Phage SSE1 that infects Shigella dysenteriae (China General Microbiological Culture Collection Center number: 1.1869) was isolated from the aeration tank water of a sewage treatment plant. SSE1 showed morphological features associated with those of phages in Myoviridae. The whole genome sequence of phage SSE1 is composed of 169,744 bp with the GC content of 37.51%. The double-stranded DNA of SSE1 contains 270 open reading frameworks (ORFs). Phylogenetically, phage SSE1 showed a stronger homology (whole genome and terminase large subunit protein sequence) to Escherichia phages than other Shigella phages in the NCBI database, but SSE1 did not infect Escherichia stains. This indicates that phage SSE1 should be a novel phage infecting Shigella dysenteriae. Besides, the result of this study provided a new idea for phage therapy. SSE1 may become a candidate for potential therapy against Shigella dysenteriae infection in clinical applications.

Breast cancer is the leading cause of cancer-associated deaths among females. In recent decades, microRNAs (miRNAs), a type of short non-coding RNA that regulates gene expression at the post-transcription level, have been reported to participate in the regulation of many hub genes associated with tumorigenesis, tumor progression, and metastasis. However, the precise mechanism by which miRNAs regulate breast cancer metastasis remains poorly discussed, which limits the opportunity for the development of novel, effective therapeutic targets. Here, we aimed to determine the miR-622-related principal regulatory mechanism in cancer. First, we found that miR-622 was significantly related to a poor prognosis in various cancers. By utilizing an integrated miRNA prediction process, we identified 77 promising targets and constructed a protein-protein interaction network. Furthermore, enrichment analyses, including GO and KEGG pathway analyses, were performed to determine the potential function of miR-622, which revealed regulation networks and potential functions of miR-622. Then, we identified a key cluster comprised of six hub genes in the protein-protein interaction network. These genes were further chosen for pan-cancer expression, prognostic and predictive marker analyses based on the TCGA and GEO datasets to mine the potential clinical values of these hub genes. To further validate our bioinformatic results, the regulatory axis of miR-622 and RNF8, one of the hub genes recently reported to promote breast cancer cell EMT process and breast cancer metastasis, was selected as in vitro proof of concept. In vitro, we demonstrated the direct regulation of RNF8 by miR-622 and found that the predicted miR-622-RNF8 axis could regulate RNF8-induced epithelial-mesenchymal transition, cell migration, and cell viability. These results were further demonstrated with rescue experiments. We established a closed-loop miRNA-target-phenotype research model that integrated the bioinformatic analysis of the miRNA target genes and experimental validation of the identified key miRNA-target-phenotype axis. We not only identified the hub target genes of miR-622 in silico but also revealed the regulatory mechanism of miR-622 in breast cancer cell EMT process, viability, and migration in vitro for the first time.

A virulent phage, named ST20, infecting Escherichia coli O165:H8 was isolated from wastewater and subjected to genomic sequencing using the Illumina HiSeq system. Genomic analysis revealed that it contains double-stranded DNA, and its complete genome consists of 44,517 nucleotides with an average GC content of 50.81%. Morphological observations showed that phage ST20 belongs to the order Caudovirales and the family Siphoviridae due to its characteristic icosahedral capsid and a long noncontractile tail. This phage was further characterized by one-step growth curve analysis and measurement of its stability at 4 °C. The study has implications for the development of potential biocontrol agents.

MicroRNAs (miRNAs) are a class of endogenous noncoding genes that regulate gene expression at the posttranscriptional level. In recent decades, miRNAs have been reported to play important roles in tumor growth and metastasis, while some reported functions of a specific miRNA in tumorigenesis are contradictory. In this study, we reevaluated the role of miR-214, which has been reported to serve as an oncogene or anti-oncogene in breast cancer metastasis. We found that miR-214 inhibited breast cancer via targeting RNF8, a newly identified regulator that could promote epithelial-mesenchymal transition (EMT). Specifically, the survival rate of breast cancer patients was positively correlated with miR-214 levels and negatively correlated with RNF8 expression. The overexpression of miR-214 inhibited cell proliferation and invasion of breast cancer, while suppression of miR-214 by chemically modified antagomir enhanced the proliferation and invasion of breast cancer cells. Furthermore, miR-214 could modulate the EMT process via downregulating RNF8. To our knowledge, this is the first report that reveals the role of the miR-214-RNF8 axis in EMT, and our results demonstrate a novel mechanism for miR-214 acting as a tumor suppressor through the regulation of EMT.

Premise of the Study Novel and cost-effective microsatellite markers were developed to explore the population genetics, biogeographic structure, and evolutionary history of the prized Euro-Asian wild edible ectomycorrhizal fungus Tricholoma matsutake (Tricholomataceae). Methods and results Eighteen new polymorphic simple sequence repeat loci, detected from a microsatellite-enriched genomic library, were used to characterize 131 individuals from eight T. matsutake populations. The number of alleles ranged from two to 10, with averages of 1.42 to 3.22. Levels of observed and expected heterozygosity ranged from 0.00-1.00 and from 0.00-0.83, with mean values of 0.21 and 0.26, respectively. In total, 50% of the loci showed interspecific transferability and polymorphism in the related species T. equestre. Conclusions These newly developed markers will aid research into the genetic diversity and population structure of T. matsutake. They can also be used in other species of Tricholoma.

For years, microbes have been widely applied as chassis in the construction of synthetic metabolic pathways. However, the lack of in vivo enzyme clustering of heterologous metabolic pathways in these organisms often results in low local concentrations of enzymes and substrates, leading to a low productive efficacy. In recent years, multiple methods have been applied to the construction of small metabolic clusters by spatial organization of heterologous metabolic enzymes. These methods mainly focused on using engineered molecules to bring the enzymes into close proximity via different interaction mechanisms among proteins and nucleotides and have been applied in various heterologous pathways with different degrees of success while facing numerous challenges. In this paper, we mainly reviewed some of those notable advances in designing and creating approaches to achieve spatial organization using different intermolecular interactions. Current challenges and future aspects in the further application of such approaches are also discussed in this paper.

Tricholoma matsutake is an ectomycorrhizal (ECM) fungus capable of in vitro saprotrophic growth, but the sources of C and N used to generate sporocarps in vivo are not well understood. We examined natural abundance isotope data to investigate this phenomenon. For this purpose, C, N and their stable isotopes (13C, 15N) content of fungal sporocarps and their potential nutrient sources (i.e., foliage, litter, fine roots, wood, and soil) were investigated from two well-studied sites in Finland and Japan. Our results show that δ13C values of T. matsutake and other fungal groups are consistent with those of most studies, but a very high δ15N value (16.8‰ ± 2.3) is observed in T. matsutake. Such isotopic pattern of fungal δ15N suggests that matsutake has a greater proteolytic potential to digest chemically complex 15N-enriched organic matter and hydrophobic hyphae. This assumption is further supported by a significant and positive correlation between δ13Ccap–stipe and δ15Ncap–stipe exclusively in T. matsutake, which suggests common C and N sources (protein) possible for isotopically enriched cap. The 13C increase of caps relative to stipe presumably reflects greater contents of 13C-enriched protein than 13C-depleted chitin. We conclude that T. matsutake is a typical ECM fungus which obtains for its sporocarp development for both C and N from a common protein source (vs. photosynthetic carbon) present in soil organic matter. Electronic supplementary material The online version of this article (10.1007/s00572-018-0870-8) contains supplementary material, which is available to authorized users.

Commercial harvesting of matsutake in Finland has driven the need to understand how fruiting is affected by climate and habitat. A productive site in Finland was studied in this respect. Productivity, duration, and early onset of fruiting were examined in relation to mean air temperature, soil temperature, accumulated degree-days, and precipitation. We monitored the fruiting of matsutake and other macrofungi during 2008–2013. As a pilot study, our results showed the timing and yield of the annual fruiting of Tricholoma matsutake varies in Southern Finland and can occur from the middle of July to middle of September; although we did not find a clear and simple correlation between fruiting and rainfall, an average amount of precipitation (i.e. 90–110% of the long-term average precipitation) prior to the first fruiting was associated with high yield; we did not identify any threshold soil temperature associated with the onset of fruiting, however, we found that soil temperature might provide a means to monit...

Tricholoma matsutake (S. Ito & S. Imai) Singer is an ectomycorrhizal basidiomycete that produces highly prized mushrooms known as ‘true matsutake’. Recent research has shown that T. matsutake has a wide but patchy distribution in temperate and boreal forests of Eurasia and subtropical China in association with Pinus, Picea, Tsuga, Abies and even fagaceous broadleaves. Molecular analyses of the microbial communities living in shiro soil have been made in certain locations, but their generality has yet to be determined systematically and across the entire range. Variation in fruiting in relation to climate and geography has improved our understanding of matsutake phenology, and important in-roads have been made into its ecology over the past 15 years. T. matsutake is a commercially-important fungal species that plays a significant role in the functional diversity of forests in the Northern Hemisphere, but much remains to be learned about this enigmatic taxon.

Endophytic fungi and ectomycorrhizal fungi co-exist in the mycorrhizal root tips of boreal forest trees.However, very little is known about the functional role they play in their host’s biology.The activity of enzymesresponsible for important biochemical processes is used to determine the functional role of root-associatedmycorrhizal fungi. However, enzyme activity is never studied in the presence of endophytic fungi in-planta. Theaims of this study were to investigate the effect of Scots pine (Pinus sylvestris L.) root-isolated fungal endophytes on the host plant root morphology, to determine their functional effects using host root-excreted enzyme activitymeasurements, and to compare them with roots colonized by decomposer and ectomycorrhizal fungal strains andnoncolonised Scots pine root tips.Our results show that endophytic fungi did not damage the pine roots incontrast to the decomposer fungi. The endophytic fungi penetrated the cortical cells of the host plant. The rootscolonised by endophytic fungi produce different exo-enzymes compared with those produced by roots colonizedby other fungal groups or noncolonized control root tips.Our results indicate that endophytic fungi are clearlya distinctive ecological group of fungi that have functional traits different from those of ectomycorrhizal anddecomposer fungi.; Les champignons endophytes et ectomycorhiziens coexistent dans les extrémités radiculaires mycorhi-ziennes des arbres de la forêt boréale.Cependant, on sait vraiment peu de choses du rôle fonctionnel qu’ils jouent dans la biologie de leur hôte. L’activité des enzymes responsables de processus biochimiques importants est utilisée pour expliquer le rôle fonctionnel des champignons mycorhiziens associés aux racines. Toutefois, l’activité enzymatique n’est jamais étudiée en présence de champignons endophytes « in planta ». Les buts de l’étude présente étaient d’examiner l’effet d’endophytes fongiques isolés de racines du pin sylvestre (Pinus sylvestris L.) sur la morphologie de la racine de la plante–hôte, de déterminer leur effet fonctionnel en utilisant des mesures de l’activité enzymatique excrétée par les racines de l’hôte et de les comparer aux racines colonisées par des souches de champignons décomposeurs et ectomycorhiziens et aux extrémités radiculaires de pin sylvestre non colonisées.Les résultats des auteurs montrent que les champignons endophytes n’endommagent pas les racines de pin comparativement aux champignons décomposeurs. Les champignons endophytes pénétraient les cellules corticales de la plante-hôte. Les racines colonisées par les champignons endophytes produisent différentes exo-enzymes comparativement a celles produites par les racines colonisées par d’autres groupes de champignons ou des extrémités radiculaires contrôles non colonisées.Leurs résultats indiquent que les champignons endophytes constituent clairement un groupe écologique distinctif de champignons qui possèdent des traits fonctionnels différents de ceux des champignons ectomycorhiziens et décomposeurs.

Tricholoma matsutake is an ectomycorrhizal fungus that forms commercially important mushrooms in coniferous forests. In this study, we explored the ability of T. matsutake to form mycorrhizae with Pinus sylvestris by inoculating emblings produced through somatic embryogenesis (SE) in an aseptic culture system. Two months after inoculation, clones with less phenolic compounds in the tissue culture phase formed mycorrhizae with T. matsutake, while clones containing more phenols did not. Effects of inoculation on embling growth varied among clones; two of the four tested showed a significant increase in biomass and two had a significant increase in root density. In addition, results suggest that clones forming well-developed mycorrhizae absorbed more Al, Fe, Na, P, and Zn after 8 weeks of inoculation. This study illustrates the value of SE materials in experimental work concerning T. matsutake as well as the role played by phenolic compounds in host plant response to infection by mycorrhizal fungi.

Regulatory T (Treg) cells play a critical role in Helicobacter pylori immune evasion and persistent infection. In addition to Treg cells, it is still unknown whether a newly defined B-cell subset, interleukin (IL)-10-producing B cells, is involved. Using a mouse model of H. pylori infection, we investigated the dynamic changes of IL-10-producing B cells and Foxp3(+) Treg cells in gastrointestinal mucosa, spleen and mesenteric lymph nodes following H. pylori infection. We observed that in addition to Foxp3(+) Treg cells, IL-10-producing B cells could also be induced after H. pylori infection and they expanded earlier than Foxp3(+) Treg cells did. Moreover, the regulatory immune responses induced by H. pylori were not limited to gastric mucosa. Our findings may provide new clues for further research on H. pylori immune evasion and diseases associated with H. pylori infection.

Malignant immature ovarian teratomas (IOTs) most often occur in women of reproductive age. It is unclear, however, what roles estrogenic signaling plays in the development of IOT. In this study, we examined whether estrogen receptors (ERα and β) promote the cellular malignancy of IOT. Estradiol (E2), PPT (propylpyrazole), and DPN (diarylpropionitrile) (ERα- and β-specific agonists, respectively), as well as ERα- or ERβ-specific short hairpin (sh)RNA were applied to PA-1 cells, a well-characterized IOT cell line. Cellular tumorigenic characteristics, for example, cell migration/invasion, expression of the cancer stem/progenitor cell marker CD133, and evidence for epithelial-mesenchymal transition (EMT) were examined. In PA-1 cells that expressed ERα and ERβ, we found that ERα promoted cell migration and invasion. We also found that E2/ERα signaling altered cell behavior through non-classical transactivation function. Our data show non-genomic E2/ERα activations of focal adhesion kinase-Ras homolog gene family member A (FAK-RhoA) and ERK governed cell mobility capacity. Moreover, E2/ERα signaling induces EMT and overexpression of CD133 through upregulation micro-RNA 21 (miR21; IOT stem/progenitor promoter), and ERK phosphorylations. Furthermore, E2/ERα signaling triggers a positive feedback regulatory loop within miR21 and ERK. At last, expression levels of ERα, CD133, and EMT markers in IOT tissue samples were examined by immunohistochemistry. We found that cytosolic ERα was co-expressed with CD133 and mesenchymal cell markers but not epithelial cell markers. In conclusion, estrogenic signals exert malignant transformation capacity of cancer cells, exclusively through non-genomic regulation in female germ cell tumors.

Thrombomodulin (TM), a natural anticoagulation factor, maintains circulation homeostasis in endothelial cells. TM has additional roles in modulating inflammation, thrombosis, and carcinogenesis. However, there is little information on the role of TM in the progression and metastasis of ovarian cancer. RNA silencing and cDNA expression vectors were used to manipulate target gene expression in ovarian cancer cells. Cell growth and migration were evaluated by an MTT assay, a wound-healing migration assay, a transwell migration assay, and a biosensor system. In this study, we found that TM silencing may enhance the growth rate of cells. The migratory ability of ovarian cancer cells was enhanced dramatically after TM silencing. TM overexpression in ovarian cells suppressed the proliferation and migration capability. Furthermore, we found that skov-3 cells treated with TM shRNA expressed high levels of fibronectin and vimentin and that the expression of these markers correlated positively with their migratory ability. Our results demonstrate that TM expression may regulate cell growth and migration in ovarian cancer cells. This finding suggests that TM may be a novel prognostic and therapeutic target for ovarian cancer.

This study aimed to understand the interaction between temperature and genotype in terms of the effect on early seedling development of European aspen (Populus tremula) and various F2-aspen hybrids. We evaluated the response of 16 different European aspen and F2-hybrid families on seed germination, survival rate and seedling height in one- and two-family trials under three different temperature regimes. In one-family trials, higher germination and higher survival rates were observed in higher temperature (C1700), leading to taller seedlings. European aspen × hybrid aspen individuals (Asp × Hyb) had a higher survival rate and taller seedlings than Asp, Hyb × Asp or Hyb × Hyb individuals. The difference between Asp × Hyb and Hyb × Asp was pronounced. Both growth conditions and genotype (i.e., hybrid cross) had strong effects on germination, survival rate and height of European aspen and F2-hybrid seedlings. However, the interaction of genotype and growth conditions also had a significant influence on survival rate and seedling height, but not on germination. Two-family trials involving European aspen and F2-hybrids led to significant negative effects on germination and survival rate and also facilitation effects on seedling height. Similarly, genotype had strong effects on germination and survival rate. Different genotypes and traits affected survival rate and seedling height differently in different growth conditions. These differences were more pronounced in the warmest environment, i.e., 1,700 degree days. We conclude that introgression between F2-hybrids and local European aspen is likely in the current climate, and any warming will likely favor certain F2-hybrids, especially the most probable types (P. tremula × (P. tremula × P. tremuloides)).

The Surendra background update and novel fast model matching were mixed which can reduce the matching region. A Yuntai tracking system was present for people tracking, the fuzzy control tracking based on polar coordinates was also present, which makes the tracking of people object always in the video range and the Yuntai neednt move frequency. Results indicate that the algorithm is superior to the previously published variants of the model matching and the Yuntai system track people in real time.

Removing the litter layer from an area known to contain matsutake (Tricholoma matsutake) is believed to enhance mushroom productivity in Asian countries where it occurs. This study aimed to understand the effect of understorey vegetation and litter cover on matsutake fruitbody formation and to evaluate the utility of litter derived dissolved organic matter (DOM) as a growth medium. An inventory of understorey vegetation and litter cover was conducted in a productive forest site mainly including conifers that contained 15 plots in which fruitbodies had formed in successive years and 15 nearby control plots without fruitbodies. Litter composition of fruiting plots was investigated. Two T. matsutake isolates were incubated for one month in DOM derived from decomposing spruce or birch litter and mycelial growth was estimated by monitoring the concentration of ergosterol. Although plant diversity was rather uniform in the study site, total vegetation cover was substantially lower and litter cover was somewhat higher in fruiting plots. We concluded that litter derived carbon has a positive impact on T. matsutake fruitbody formation. This was confirmed by in vitro culture experiments where T. matsutake was able to grow in spruce litter derived DOM.

Clinical investigations highlight the increased incidence of metabolic syndrome in prostate cancer (PCa) patients receiving androgen deprivation therapy (ADT). Studies using global androgen receptor (AR) knockout mice demonstrate that AR deficiency results in the development of insulin resistance in males. However, mechanisms by which AR in individual organs coordinately regulates insulin sensitivity remain unexplored. Here we tested the hypothesis that functional AR in the brain contributes to whole-body insulin sensitivity regulation and to the metabolic abnormalities developed in AR-deficient male mice. The mouse model selectively lacking AR in the central nervous system and AR-expressing GT1-7 neuronal cells were established and used to delineate molecular mechanisms in insulin signaling modulated by AR. Neuronal AR deficiency leads to reduced insulin sensitivity in middle-aged mice. Neuronal AR regulates hypothalamic insulin signaling by repressing nuclear factor-κB (NF-κB)–mediated induction of protein-tyrosine phosphatase 1B (PTP1B). Hypothalamic insulin resistance leads to hepatic insulin resistance, lipid accumulation, and visceral obesity. The functional deficiency of AR in the hypothalamus leads to male mice being more susceptible to the effects of high-fat diet consumption on PTP1B expression and NF-κB activation. These findings suggest that in men with PCa undergoing ADT, reduction of AR function in the brain may contribute to insulin resistance and visceral obesity. Pharmacotherapies targeting neuronal AR and NF-κB may be developed to combat the metabolic syndrome in men receiving ADT and in elderly men with age-associated hypogonadism.

Objectives Both overproduction of nitric oxide and oxidative injury to the cardiovascular and pulmonary systems contribute to fatal pathophysiology during endotoxemia. We investigated the effect of propofol on oxidative stress-related enzymes in lung (L2), heart (H9C2) and macrophage (NR8383) cells during endotoxemia. Methods Experimental endotoxemia was induced by co-culture of Escherichia coli lipopolysaccharide (15 μg/mL) in the abovementioned three types of cells that were under the effect of propofol (15 or 30 μM for 1 or 4 hours). Cellular expression of induced nitric oxide synthase (iNOS), superoxide dismutase (SOD) 1 and 2, and p47phox (representing NADPH oxidase) were determined by immunoblotting. The cellular oxidative burst activity was determined using a dihydroethidium method via flow cytometry to represent the level of reactive oxygen species. The in vivo endotoxemia model was also employed for comparison using a systemic injection of lipopolysaccharide (15 mg/kg) under propofol maintenance (15 or 30 mg/kg/h). The Student t test (two groups) was used for statistical evaluation among the means, and the Newman–Keuls test was used for analysis of variance in the statistical analysis. Results In lung L2 cells, propofol significantly reduced the expression of iNOS, SOD1, SOD2, and p47phox under LPS-induced endotoxemia. However, in H9C2 cardiac cells and NR8383 macrophages, only the expression of iNOS was significantly suppressed, but not that of SOD1, SOD2, or p47phox. The level of reactive oxygen species was suppressed in all three kinds of cell. In in vivo animal tissue, except for the suppression of iNOS expression in lung and heart cells, propofol in lung cells produced only SOD1 suppression, but in rat heart the expression of both SOD1 and SOD2 was suppressed. Conclusion These results suggest that propofol may have a protective role for lung cells. This effect is associated with its suppression of oxidative-related enzymes, including iNOS, SOD1, SOD2, and p47phox. In cardiac myocytes and macrophages, propofol also provides an antioxidative effect, probably via its inhibition of iNOS. The overall effect of propofol in the organs may be a combination of its effects on various cells. In addition, a reduction in reactive oxygen species plays a major role in the beneficial effect of propofol on experimental endotoxemia.

Background: MicroRNAs (miRNAs) mediate negative regulation of target genes through base pairing, and aberrant miRNA expression has been described in cancers. We hypothesized that single nucleotide polymorphisms (SNPs) within miRNA target sites might influence clinical outcomes in patients with colorectal cancer. Methods: Sixteen common SNPs within miRNA target sites were identified, and the association between these SNPs and overall survival was assessed in colorectal cancer patients using Kaplan-Meier analysis, Cox regression model, and survival tree analysis. Results: Survival tree analysis identified a higher-order genetic interaction profile consisting of the RPS6KB1 rs1051424 and ZNF839 rs11704 that was significantly associated with overall survival. The 5-year survival rates were 74.6%, 62.7%, and 57.1% for the low-, medium-, and high-risk genetic profiles, respectively (P = 0.006). The genetic interaction profile remained significant even after adjusting for potential risk factors. Additional in silico analysis provided evidence that rs1051424 and rs11704 affect RPS6KB1 and ZNF839 expressions, which in turn is significantly correlated with prognosis in colorectal cancer. Conclusion: Our results suggest that the genetic interaction profiles among SNPs within miRNA target sites might be prognostic markers for colorectal cancer survival.

In this study, to investigate the effects of naringin on vascular endothelial cell (VEC) function, proliferation, apoptosis, and angiogenesis, rat VECs were cultured in vitro and randomly divided into four groups: control, serum‑starved, low‑concentration naringin treatment, and high‑concentration naringin treatment. MTT assay was used to detect cell proliferation while Hoechst 33258 staining and flow cytometry were used to detect apoptosis. Changes in the expression of apoptosis‑associated proteins [GRP78, CHOP, caspase‑12, and cytochrome c (Cyt.c)] were detected using western blotting. JC‑1 staining was employed to detect changes in mitochondrial membrane potential. Intracellular caspase‑3, ‑8, and ‑9 activity was determined by spectrophotometry. ELISA was used to detect endothelin (ET), and a Griess assay was used to detect changes in the expression of nitric oxide (NO) in culture medium. The study further divided an ovariectomized (OVX) rat model of osteoporosis randomly into four groups: OVX, sham‑operated, low‑concentration naringin treatment (100 mg/kg), and high‑concentration naringin treatment (200 mg/kg). After 3 months of treatment, changes in serum ET and NO expression, bone mineral density (BMD), and microvessel density of the distal femur (using CD34 labeling of VECs) were determined. At each concentration, naringin promoted VEC proliferation in a time‑ and dose‑dependent manner. Naringin also significantly reduced serum starvation‑induced apoptosis in endothelial cells, inhibited the expression of GRP78, CHOP, caspase‑12, and Cyt.c proteins, and reduced mitochondrial membrane potential as well as reduced the activities of caspase‑3 and ‑9. Furthermore, naringin suppressed ET in vitro and in vivo while enhancing NO synthesis. Distal femoral microvascular density assessment showed that the naringin treatment groups had a significantly higher number of microvessels than the OVX group, and that microvascular density was positively correlated with BMD. In summary, naringin inhibits apoptosis in VECs by blocking the endoplasmic reticulum (ER) stress‑ and mitochondrial‑mediated pathways. Naringin also regulates endothelial cell function and promotes angiogenesis to exert its anti‑osteoporotic effect.

Structure-activity relationship (SAR) investigations of a novel class of triazolopyridazinone p38α mitogen activated protein kinase (MAPK) inhibitors are disclosed. From these studies, increased in vitro potency was observed for 2,6-disubstituted phenyl moieties and N-ethyl triazolopyridazinone cores due to key contacts with Leu108, Ala157 and Val38. Further investigation led to the identification of three compounds, 3g, 3j and 3m that are highly potent inhibitors of LPS-induced MAPKAP kinase 2 (MK2) phosphorylation in 50% human whole blood (hWB), and possess desirable in vivo pharmacokinetic and kinase selectivity profiles.

Cell proliferation and tumor metastasis are considered as the main reasons for death in colorectal carcinoma (CRC). IRE1α-XBP1 pathway is the most conserved UPR pathways, which are activated during ER stress caused by the accumulation of unfolded or misfolded protein in the lumen of ER. Here, we demonstrated the critical role of IRE1α-XBP1 pathway and underlying molecular mechanism in cell proliferation and tumor metastasis in CRC. By the use of tissue microarray analysis of samples from 119 patients with CRC, IRE1α was determined to be an independent predictor of overall survival as higher expression of IRE1α in CRC patients showed lower survival rates (p = 0.0041). RNA interference and ectopic expression of IRE1α were applied to determine the molecular effects of IRE1α in CRC cells. The silencing of IRE1α inhibited the proliferation and blocked the invasion of CRC cells in vitro, while ectopic expression of IRE1α in turn promoted cell proliferation and invasion. IRE1α-XBP1 pathway regulated the mitosis of CRC cells through the directly binding of XBP1s to Cyclin D1 promoter to activate Cyclin D1 expression. Our results reveal that IRE1α-XBP1 pathway plays an important role in tumor progression and epithelial-to-mesenchymal transition (EMT), and IRE1α could be employed as a novel prognostic marker and a promising therapeutic target for CRC.

UV irradiation is one of the major external insults to cells and can cause skin aging and cancer. In response to UV light-induced DNA damage, the nucleotide excision repair (NER) pathways are activated to remove DNA lesions. We report here that testicular nuclear receptor 4 (TR4), a member of the nuclear receptor family, modulates DNA repair specifically through the transcription-coupled (TC) NER pathway but not the global genomic NER pathway. The level of Cockayne syndrome B protein (CSB), a member of the TC-NER pathway, is 10-fold reduced in TR4-deficient mouse tissues, and TR4 directly regulates CSB at the transcriptional level. Moreover, restored CSB expression rescues UV hypersensitivity of TR4-deficient cells. Together, these results indicate that TR4 modulates UV sensitivity by promoting the TC-NER DNA repair pathway through transcriptional regulation of CSB. These results may lead to the development of new treatments for UV light-sensitive syndromes, skin cancer, and aging.

Tricholoma matsutake is an economically important ectomycorrhizal fungus of coniferous woodlands. Mycologists suspect that this fungus is also capable of saprotrophic feeding. In order to evaluate this hypothesis, enzyme and chemical assays were performed in the field and laboratory. From a natural population of T. matsutake in southern Finland, samples of soil-mycelium aggregate (shiro) were taken from sites of sporocarp formation and nearby control (PCR-negative) spots. Soil organic carbon and activity rates of hemicellulolytic enzymes were measured. The productivity of T. matsutake was related to the amount of utilizable organic carbon in the shiro, where the activity of xylosidase was significantly higher than in the control sample. In the laboratory, sterile pieces of bark from the roots of Scots pine were inoculated with T. matsutake and the activity rates of two hemicellulolytic enzymes (xylosidase and glucuronidase) were assayed. Furthermore, a liquid culture system showed how T. matsutake can utilize hemicellulose as its sole carbon source. Results linked and quantified the general relationship between enzymes secreted by T. matsutake and the degradation of hemicellulose. Our findings suggest that T. matsutake lives mainly as an ectomycorrhizal symbiont but can also feed as a saprotroph. A flexible trophic ecology confers T. matsutake with a clear advantage in a heterogeneous environment and during sporocarp formation.

Early studies suggest that TR4 nuclear receptor is a key transcriptional factor regulating various biological activities, including reproduction, cerebella development, and metabolism. Here we report that mice lacking TR4 ( TR4−/−) exhibited increasing genome instability and defective oxidative stress defense, which are associated with premature aging phenotypes. At the cellular level, we observed rapid cellular growth arrest and less resistance to oxidative stress and DNA damage in TR4−/− mouse embryonic fibroblasts (MEFs) in vitro. Restoring TR4 or supplying the antioxidant N-acetyl-l-cysteine (NAC) to TR4−/− MEFs reduced the DNA damage and slowed down cellular growth arrest. Focused qPCR array revealed alteration of gene profiles in the DNA damage response (DDR) and anti-reactive oxygen species (ROS) pathways in TR4−/− MEFs, which further supports the hypothesis that the premature aging in TR4−/− mice might stem from oxidative DNA damage caused by increased oxidative stress or compromised genome integrity. Together, our finding identifies a novel role of TR4 in mediating the interplay between oxidative stress defense and aging.

OBJECTIVE TR4 is a nuclear receptor without clear pathophysiological roles. We investigated the roles of hepatic TR4 in the regulation of lipogenesis and insulin sensitivity in vivo and in vitro. RESEARCH DESIGN AND METHODS TR4 activity and phosphorylation assays were carried out using hepatocytes and various TR4 wild-type and mutant constructs. Liver tissues from TR4 knockout, C57BL/6, and db/db mice were examined to investigate TR4 target gene stearoyl-CoA desaturase (SCD) 1 regulation. RESULTS TR4 transactivation is inhibited via phosphorylation by metformin-induced AMP-activated protein kinase (AMPK) at the amino acid serine 351, which results in the suppression of SCD1 gene expression. Additional mechanistic dissection finds TR4-transactivated SCD1 promoter activity via direct binding to the TR4-responsive element located at −243 to −255 on the promoter region. The pathophysiological consequences of the metformin→AMPK→TR4→SCD1 pathway are examined via TR4 knockout mice and primary hepatocytes with either knockdown or overexpression of TR4. The results show that the suppression of SCD1 via loss of TR4 resulted in reduced fat mass and increased insulin sensitivity with increased β-oxidation and decreased lipogenic gene expression. CONCLUSIONS The pathway from metformin→AMPK→TR4→SCD1→insulin sensitivity suggests that TR4 may function as an important modulator to control lipid metabolism, which sheds light on the use of small molecules to modulate TR4 activity as a new alternative approach to battle the metabolic syndrome.

We investigated the developmental expression pattern of AmphiCaM in cephalochordate amphioxus (Branchiostoma belcheri tsingtauense). We cultured and sampled the animals at different developmental stages (eggs and larvae), and used in-situ hybridization and northern blotting to document the spatial and temporal changes in AmphiCaM expression. The alimentary tract dominates the development from the late neurula stage to the adult stage. AmphiCaM expression increased significantly in the alimentary tract during the late neurula stage and remained elevated in the adults. Our results indicate that AmphiCaM is involved in the differentiation of the alimentary tract in amphioxus; and furthermore, provide an insight into the change in function of CaM genes during evolution.

The p38α mitogen-activated protein (MAP) kinase is a central signaling molecule in many proinflammatory pathways, regulating the cellular response to a multitude of external stimuli including heat, ultraviolet radiation, osmotic shock, and a variety of cytokines especially interleukin-1β and tumor necrosis factor α. Thus, inhibitors of this enzyme are postulated to have significant therapeutic potential for the treatment of rheumatoid arthritis, inflammatory bowel disease, and Crohn’s disease, as well as other diseases where aberrant cytokine signaling is the driver of disease. In this communication, we describe a novel class of 7-alkyl-1,5-bis-aryl-pyrazolopyridinone-based p38α inhibitors. In particular, compound 3f is highly potent in the enzyme and cell-based assays, selective in an Ambit kinase screen, and efficacious (ED50 ≤ 0.01 mg/kg) in the rat collagen induced arthritis (CIA) model.

Over a 5 year period, we examined the influence of substrate and fertilization on nursery growth and outplanting performance of Norway spruce (Picea abies (L.) Karst.). We focused on the relative growth and development of roots and shoots and the colonization intensity and diversity of ectomycorrhizal (ECM) fungi. In the nursery, a conventional substrate (low-humified Sphagnum peat) supplemented with woody material (wood fibre and pine bark) and either mineral or organic fertilizers yielded shorter seedlings than those grown on the unmodified substrate. However, after outplanting, the growth rate of seedlings cultivated on modified substrates was higher than that of seedlings grown on the unmodified substrate. Seedlings cultivated in modified substrates had significantly higher root/shoot ratios and ECM diversity; the latter remained significant after ≥3 years of outplanting. Seedlings grown on a substrate containing 50% woody material and supplemented with organic fertilizer had the highest growth rate among all seedlings during the 3 year period of outplanting. Colonization intensity of ECM fungi was high in all seedlings except for those grown in heavily fertilized substrate. This study suggests that nursery techniques that produce seedlings with higher root/shoot ratios and ECM diversities could improve plantation success and growth rate for at least the first 3 years of outplanting.

The p38 mitogen-activated protein kinase (MAPK) is a central signaling molecule in many proinflammatory pathways, regulating the cellular response to a multitude of external stimuli including heat, ultraviolet radiation, osmotic shock, and a variety of cytokines especially interleukin-1beta and tumor necrosis factor alpha. Thus, inhibitors of this enzyme are postulated to have significant therapeutic potential for the treatment of rheumatoid arthritis, inflammatory bowel disease, osteoporosis, and many other diseases where aberrant cytokine signaling is the driver of disease. Herein, we describe a novel class of 3-amino-7-phthalazinylbenzoisoxazole-based inhibitors. With relatively low molecular weight, these compounds are highly potent in enzyme and cell-based assays, with minimal protein shift in 50% human whole blood. Compound 3c was efficacious (ED 50 = 0.05 mg/kg) in the rat collagen induced arthritis (CIA) model.

Androgen and the androgen receptor (AR) have been shown to play critical roles in male fertility. Our previous data demonstrated that mice lacking AR (AR(-/y)) revealed incomplete germ cell development and lowered serum testosterone levels, which resulted in azoospermia and infertility. However, the consequences of AR loss in Leydig cells remain largely unknown. Using a Cre-LoxP conditional knockout strategy, we generated a tissue-specific knockout mouse (L-AR(-/y)) with the AR gene deleted by the anti-Müllerian hormone receptor-2 (Amhr2) promoter driven Cre expressed in Leydig cells. Phenotype analyses show that the outside appearance of L-AR(-/y) mice was indistinguishable from wild type mice (AR(+/y)), but with atrophied testes and epididymis. L-AR(-/y) mice were infertile, with spermatogenic arrest predominately at the round spermatid stage and no sperm could be detected in the epididymis. L-AR(-/y) mice also have lower serum testosterone concentrations and higher serum leuteinizing hormone and follicle-stimulating hormone concentrations than AR(+/y) mice. Further mechanistic studies demonstrated that hypotestosteronemia in L-AR(-/y) mice is not caused by reducing numbers of Leydig cells, but instead by the alterations of several key steroidogenic enzymes, including 17beta-HSD3, 3beta-HSD6, and P450c17. Together, L-AR(-/y) mice provide in vivo evidence that functional AR in Leydig cells is essential to maintain normal spermatogenesis, testosterone production, and required for normal male fertility.

An in vitro system was used for ectomycorrhizal synthesis of Cenococcum geophilum Fr. with Cathaya argyrophylla Chun et Kuang, an endangered species. Calli initiated from stem segments and adventitious roots differentiated from young seedlings were removed and cocultured with Cenococcum geophilum on a modified Murashige-Skoog medium. Fungal hyphae were visible within intercellular spaces of the callus 4 weeks after inoculation, but definite and well-developed Hartig net structures did not form in the calli 8 weeks after inoculation. The typical ectomycorrhizal structures (i.e. hyphal mantle and intracortical Hartig net) were observed in root segments 8 weeks after inoculation. This is the first report of aseptic ectomycorrhizal-like formation/infection between root organ/callus of Cathaya argyrophylla and the ectomycorrhizal fungus Cenococcum geophilum. This culture system is useful for further investigation of mycorrhizal synthesis in Cathaya trees. (Managing editor: Ya-Qin Han)

One type of soil collected from Maoer-shan in Heilongjiang Province, China was selected to induce hyphal growth of Tricholoma matsutake by a soil screening experiment. It was confirmed that hyphal growth of all the tested T. matsutake isolates was significantly stimulated in soil by supplemented with 0.5%∼2.0% olive oil. The aggregation of hyphae and soil resembled natural Shiro. The biomass of hyphae in the soil increases with increasing olive oil concentrations. Moreover, seedlings of Pinus densiflora grew well in the soil containing 0.5%∼1.0% olive oil and were also successfully infected by T. matsutake isolate A in the soil containing 1.0% olive oil. This study established a culture system of artificial Shiro formation and also provided a premise for formulation of culture substratum for fruit body formation of T. matsutake.