Chitinase Is Involved in the Fruiting Body Development of Medicinal Fungus Cordyceps militaris.

Cordyceps militaris is a famous traditional edible and medicinal fungus in Asia, and its fruiting body has rich medicinal value. The molecular mechanism of fruiting body development is still not well understood in C. militaris. In this study, phylogenetically analysis and protein domains prediction of the 14 putative chitinases were performed. The transcription level and enzyme activity of chitinase were significant increased during fruiting body development of C. militaris. Then, two chitinase genes (Chi1 and Chi4) were selected to construct gene silencing strain by RNA interference. When Chi1 and Chi4 genes were knockdown, the differentiation of the primordium was blocked, and the number of fruiting body was significantly decreased approximately by 50% compared to wild-type (WT) strain. The length of the single mature fruiting body was shortened by 27% and 38% in Chi1- and Chi4-silenced strains, respectively. In addition, the chitin content and cell wall thickness were significantly increased in Chi1- and Chi4-silenced strains. These results provide new insights into the biological functions of chitinase in fruiting body development of C. militaris.

Multi-omics analysis unravels positive effect of rotenone on the cordycepin biosynthesis in submerged fermentation of Cordyceps militaris.

Cordycepin is the key pharmacologically active compound of Cordyceps militaris, and various fermentation strategies have been developed to increase cordycepin production. This study aimed to investigate the effect of rotenone on cordycepin biosynthesis in submerged fermentation of C. militaris, and also to explore its possible induction mechanisms via multi-omics analysis. Adding 5 mg/L rotenone significantly increased the cordycepin production by 316.09 %, along with mycelial growth inhibition and cell wall destruction. Moreover, transcriptomic analysis and metabolomic analysis revealed the accumulation of cordycepin was promoted by alterations in energy metabolism and amino acid metabolism pathways. Finally, the integration analysis of the two omics confirmed rotenone altered the nucleotide metabolism pathway toward adenosine and up-regulated the cordycepin synthesis genes (cns1-3) to convert adenosine to cordycepin. This work reports, for the first time, rotenone could act as an effective inducer of cordycepin synthesis.

Dual antioxidant activity and the related mechanisms of a novel pentapeptide GLP4 from the fermented mycelia of Ganoderma lingzhi.

Oxidative stress causes chronic inflammation, and mediates various diseases. The discovery of antioxidants from natural sources is important to research. Here we identified a novel antioxidant peptide (GLP4) from Ganoderma lingzhi mycelium and investigated its antioxidant type and potential protective mechanisms. Through free radical scavenging assay, active site shielding validation, superoxide dismutase (SOD) activity assay, and lipid peroxidation assay, we demonstrated that GLP4 was a novel protective agent with both direct and indirect antioxidant activities. GLP4 could directly enter human umbilical vein endothelial cells (HUVECs) as an exogenous substance. Meanwhile, GLP4 promoted the nuclear translocation of nuclear factor erythroid-2-related factor 2 (Nrf2) and activated the Nrf2/antioxidant response element (ARE) signaling pathway, exhibiting antioxidant and anti-apoptotic cytoprotective effects on hydrogen peroxide (H2O2)-induced HUVECs. Pull-down experiments of GLP4 target proteins, bioinformatics analysis and molecular docking further revealed that GLP4 mediated Nrf2 activation through binding to phosphoglycerate mutase 5 (PGAM5). The results suggested that GLP4 is a novel peptide with dual antioxidant activity and has promising potential as a protective agent in preventing oxidative stress-related diseases.

Roles of FERONIA-like receptor genes in regulating grain size and quality in rice.

Grain yield and quality are critical factors that determine the value of grain crops. In this study, we analyzed the functions of 12 FERONIA-like receptor (FLR) family members in rice and investigated their effects on grain size and quality. We found that FLR1, FLR2 and FLR8 negatively regulated grain size, and FLR15 positively regulated grain size. flr1 mutants had a higher cell number and an accelerated rate of grain filling compared to wild-type plants, which led to grains with greater widths. A mechanism underlying the regulation of grain size by FLR1 is that FLR1 is associated with OsRac1 Rho-like GTPase, a positive regulator of grain size. Regarding grain quality, the flr1 mutant had a higher percentage of chalkiness compared with wild-type plants, and seeds carrying mutations in flr3 and flr14 had endosperms with white floury cores. To elucidate the possible mechanism underlying this phenomenon, we found that FLR1 was constitutively expressed during endosperm development. RNA-seq analysis identified 2,367 genes that were differentially expressed in the flr1 mutant, including genes involved in starch and sucrose metabolism and carbon fixation. In this study, we identified the roles played by several FLR genes in regulating grain size and quality in rice and provided insights into the molecular mechanism governing the FLR1-mediated regulation of grain size.

Analysis of reference genes stability and histidine kinase expression under cold stress in Cordyceps militaris.

The development of fungal fruiting bodies from a hyphal thallus is inducible under low temperature (cold stress). The molecular mechanism has been subject to surprisingly few studies. Analysis of gene expression level has become an important means to study gene function and its regulation mechanism. But identification of reference genes (RGs) stability under cold stress have not been reported in famous medicinal mushroom-forming fungi Cordyceps militaris. Herein, 12 candidate RGs had been systematically validated under cold stress in C. militaris. Three different algorithms, geNorm, NormFinder and BestKeeper were applied to evaluate the expression stability of the RGs. Our results showed that UBC and UBQ were the most stable RGs for cold treatments in short and long periods, respectively. 2 RGs (UBC and PP2A) and 3 RGs (UBQ, TUB and CYP) were the suitable RGs for cold treatments in short and long periods, respectively. Moreover, target genes, two-component-system histidine kinase genes, were selected to validate the most and least stable RGs under cold treatment, which indicated that use of unstable expressed genes as RGs leads to biased results. Our results provide a good starting point for accurate reverse transcriptase quantitative polymerase chain reaction normalization by using UBC and UBQ in C. militaris under cold stress and better support for understanding the mechanism of response to cold stress and fruiting body formation in C. militaris and other mushroom-forming fungi in future research.

RALF1-FERONIA complex affects splicing dynamics to modulate stress responses and growth in plants.

The environmentally responsive signaling pathways that link global transcriptomic changes through alternative splicing (AS) to plant fitness remain unclear. Here, we found that the interaction of the extracellular rapid alkalinization FACTOR 1 (RALF1) peptide with its receptor FERONIA (FER) triggered a rapid and massive RNA AS response by interacting with and phosphorylating glycine-rich RNA binding protein7 (GRP7) to elevate GRP7 nuclear accumulation in Arabidopsis thaliana. FER-dependent GRP7 phosphorylation enhanced its mRNA binding ability and its association with the spliceosome component U1-70K to enable splice site selection, modulating dynamic AS. Genetic reversal of a RALF1-FER-dependent splicing target partly rescued mutants deficient in GRP7. AS of GRP7 itself induced nonsense-mediated decay feedback to the RALF1-FER-GRP7 module, fine-tuning stress responses, and cell growth. The RALF1-FER-GRP7 module provides a paradigm for regulatory mechanisms of RNA splicing in response to external stimuli.

Interdependent nitric oxide and hydrogen peroxide independently regulate the coix seed oil-induced triterpene acid accumulation in Ganoderma lingzhi.

Recent progress has been made in adding exogenous vegetable oils in culture media to promote bioactive metabolite production in several medicinal mushrooms, but the mechanism is still unclear. In this study, we found that the vegetable oil coix seed oil (CSO) could induce the biosynthesis of triterpene acids (TAs) and also significantly increase cytoplasmic nitric oxide (NO) and hydrogen peroxide (H2O2) concentrations in the mycelium of Ganoderma lingzhi. The change in TA biosynthesis caused by CSO could be reversed by adding NO scavenger or H2O2 scavenger, and adding NO scavenger or H2O2 scavenger resulted in the reduction of the cytoplasmic H2O2 or NO concentration under CSO treatment, respectively. Moreover, adding NO scavenger or H2O2 scavenger reversed TA biosynthesis, which could be rescued by H2O2 or NO donor, respectively. Taken together, our study indicated that both NO and H2O2 were involved in the regulation of TA biosynthesis, and CSO-activated NO and H2O2 were interdependent but independently regulated the TA biosynthesis under CSO treatment in G. lingzhi.

Research Progress on Signaling Pathway-Associated Oxidative Stress in Endothelial Cells.

Studying the mechanisms of oxidative stress in endothelial cells is vital to the discovery of novel drugs for the treatment of cardiovascular disease. This article reviews the progress within the field of the role of oxidative responses in the physiology and growth of endothelial cells and emphasizes the effects of several main signal pathways involved in the oxidative stress of endothelial cells. Herein, we aim to provide scientific direction that can serve as a basis for researchers specializing in the signaling pathway of oxidative stress.