Lignocellulose Degradation Efficiency of Agaricus bisporus Strains Grown on Wheat Straw-Based Compost.

The cultivation of Agaricus bisporus was investigated with two commercial strains, A15 and W192. Nitrogen and lignocellulose were analyzed in absolute amounts based on mass balance to accurately compare the degradation efficiency of the compost, and the correlation between the degradation efficiency and extracellular enzyme activity of the mycelium was analyzed. Lignocellulose utilization efficiency positively correlated with mushroom yield. For the same strain, the compost with high lignocellulose content resulted in high utilization efficiency, which increased the yield of A. bisporus. For the same compost, the lignocellulose utilization efficiency of A15 was higher than that of W192. The activities of manganese-dependent peroxidase and ß-glucosidase indicated that W192 may have a higher demand for lignin and cellulose. Therefore, a higher yield of W192 was obtained with high-lignocellulose compost. The metabolism of cellulose and hemicellulose in the mycelial growth stage seemed to be conducive to high mushroom yield.

Transcriptome and Differentially Expressed Gene Profiles in Mycelium, Primordium and Fruiting Body Development in Stropharia rugosoannulata.

Stropharia rugosoannulata uses straw as a growth substrate during artificial cultivation and has been widely promoted in China. However, its fruiting body formation and development processes have not been elucidated. In this study, the developmental transcriptomes were analyzed at three stages: the mycelium (G-S), primordium (P-S) and fruiting body (M-F) stages. A total of 9690 differentially expressed genes (DEGs) were identified in the different developmental stages. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses showed that these DEGs were involved mainly in hydrolase activity, structural molecule activity and oxidoreductase activity as well as xenobiotic biodegradation and metabolism and energy metabolism pathways. We further found that the higher expression of most carbohydrate enzyme (i.e., GH, CE, CBM, AA and PL) genes in the hyphal (i.e., G-S) stage was related mainly to substrate degradation, while the upregulation of glycosyltransferase (GT) gene expression in the P-S and M-F stages may be related to cell wall synthesis. In addition, we found that CO2-sensing-related genes (i.e., CA-2, CA-3, PKA-1 and PKA-2) were upregulated in the P-S and M-F stages, heat shock protein genes (HSP60 and HSP90) were significantly downregulated in the P-S stage and upregulated in the M-F stage and the transcription factors (i.e., steA, MYB, nosA, HAP1, and GATA-4/5/6) involved in growth and development were significantly upregulated in the P-S stage. These results suggest that environmental factors (i.e., CO2 and temperature) and transcription factors may play a key role in primordium formation. In short, this study provides new insights into the study of stimulating primordia formation affecting the development of fruiting bodies of S. rugosoannulata.

The physical structure of compost and C and N utilization during composting and mushroom growth in Agaricus bisporus cultivation with rice, wheat, and reed straw-based composts.

The cultivation of Agaricus bisporus with compost made from wheat (Triticum aestivum L.), rice (Oryza sativa L.), and reed (Phragmites australis Trin.) straw was investigated. Straw degradation was analyzed at the microscopic level, and the corresponding changes in the breakdown of different lignocellulose components during different phases of composting and mushroom production helped in understanding the yield-limiting factors of using different straws to grow mushrooms. The wheat straw compost resulted in the highest mushroom production and had the highest bioconversion efficiency. The rice straw was limited by the softer texture, which resulted in low-porosity and overdecomposed compost in the composting process and decreased the amount of available lignocellulose during mycelial growth. Although reed straw had the largest carbon resources, its utilization rate was the lowest. The hard structure, low water holding capacity, and high porosity increased the recalcitrance of reed straw to degradation and prolonged the composting time, which resulted in large N and C losses and an increased C/N ratio. Moreover, reed straw failed to transform into "ready-to-consume C" in composting. Therefore, a high C/N ratio and deficiency of available nutrition decreased the utilization efficiency of the lignocellulosic components by A. bisporus during mycelial colonization and mushroom production. The investigation revealed that degradability by and availability to microbiota and A. bisporus seemed to be the overriding factors for optimizing the composting process with different straw types. KEY POINTS: • The physical structure of compost has a significant influence on the composting process. • Degradability and availability are key factors in compost quality evaluation. • Lignocellulose utilization efficiency positively correlated with mushroom yield.

Exogenous l-ascorbic acid regulates the antioxidant system to increase the regeneration of damaged mycelia and induce the development of fruiting bodies in Hypsizygus marmoreus.

Hypsizygus marmoreus is an important commercial edible fungus, but the lack of basic studies on this fungus has hindered further development of its commercial value. In this study, we found that the treatment of damaged vegetative mycelia with 1 mM l-ascorbic acid (ASA) significantly increased the antioxidant enzyme activities (GPX, GR, CAT and SOD) and antioxidant contents (GSH and ASA) and reduced the ROS levels (H2O2 and O2-) in mechanically damaged mycelia. Additionally, this treatment increased mycelial biomass. At the reproductive stage, our results demonstrated that the treatment of damaged H. marmoreus mycelia with 2.24 mM ASA significantly increased the antioxidant enzyme activities (GPX, GR, GST, TRXR and CAT), endogenous ASA contents and GSH/GSSG ratios in different developmental stages and significantly decreased the MDA and H2O2 contents. Furthermore, this study showed that the expression levels of the antioxidant enzyme genes were consistent with the enzyme activities. Damaged mycelia treated with ASA regenerated 2-3 d earlier than the control group and showed significantly enhanced fruiting body production. These results suggested that exogenous ASA regulated mycelia intracellular ASA content to increase mycelial antioxidant abilities, induce the regeneration of damaged mycelia and regulate the development of fruiting bodies in H. marmoreus.

Comparative transcriptome analysis reveals potential fruiting body formation mechanisms in Morchella importuna.

Morchella importuna has been artificially cultivated, but stable production remains difficult because its mechanisms of fruiting body formation are unclear. To investigate the fruiting body formation mechanisms, we sequenced the transcriptomes of Morchella importuna at the mycelial and young fruiting body stages. Among the 12,561 differentially expressed genes (DEGs), 9215 were upregulated, and 3346 were downregulated. DEG enrichment analysis showed that these genes were enriched in the "generation of precursor metabolites and energy", "carbohydrate catabolic process", and "oxidoreductase activity" Gene Ontology (GO) functional categories. Enzyme activity assay results indicated that the activity levels of CAZymes (carbohydrate-active enzymes), oxidoreductases (SOD (superoxide dismutase), CAT (catalase)) and mitochondrial complex (complex I, II, III) proteins were significantly increased from the mycelial stage to the young fruiting body stage. In addition, the genes encoding CAZymes, mitochondrial proteins, oxidoreductases and heat shock proteins had higher expression levels in the young fruiting body stage than in the mycelial stage, and the qRT-PCR results showed similar trends to the RNA-Seq results. In summary, these results suggest that carbohydrate catabolism and energy metabolism are significantly enhanced in the young fruiting body stage and that growth environment temperature changes affect the formation of fruiting bodies.

Exogenous glycine inhibits root elongation and reduces nitrate-N uptake in pak choi (Brassica campestris ssp. Chinensis L.).

Nitrogen (N) supply, including NO3--N and organic N in the form of amino acids can influence the morphological attributes of plants. For example, amino acids contribute to plant nutrition; however, the effects of exogenous amino acids on NO3--N uptake and root morphology have received little attention. In this study, we evaluated the effects of exogenous glycine (Gly) on root growth and NO3--N uptake in pak choi (Brassica campestris ssp. Chinensis L.). Addition of Gly to NO3--N agar medium or hydroponic solution significantly decreased pak choi seedling root length; these effects of Gly on root morphology were not attributed to the proportion of N supply derived from Gly. When pak choi seedlings were exposed to mixtures of Gly and NO3--N in hydroponic culture, Gly significantly reduced 15NO3--N uptake but significantly increased the number of root tips per unit root length, root activity and 15NO3--N uptake rate per unit root length. In addition, 15N-Gly was taken up into the plants. In contrast to absorbed NO3--N, which was mostly transported to the shoots, a larger proportion of absorbed Gly was retained in the roots. Exogenous Gly enhanced root 1-aminocyclopropane-1-carboxylic acid synthase (ACS) and oxidase (ACO) activities and ethylene production. The ethylene antagonists aminoethoxyvinylglycine (0.5 µM AVG) and silver nitrate (10 µM AgNO3) partly reversed Gly-induced inhibition of primary root elongation on agar plates and increased the NO3--N uptake rate under hydroponic conditions, indicating exogenous Gly exerts these effects at least partly by enhancing ethylene production in roots. These findings suggest Gly substantially affects root morphology and N uptake and provide new information on the specific responses elicited by organic N sources.

The transmembrane protein TMEPAI induces myeloma cell apoptosis by promoting degradation of the c-Maf transcription factor.

TMEPAI (transmembrane prostate androgen-induced protein, also called prostate transmembrane protein, androgen-induced 1 (PMEPA1)) is a type I transmembrane (TM) protein, but its cellular function is largely unknown. Here, studying factors influencing the stability of c-Maf, a critical transcription factor in multiple myeloma (MM), we found that TMEPAI induced c-Maf degradation. We observed that TMEPAI recruited NEDD4 (neural precursor cell expressed, developmentally down-regulated 4), a WW domain-containing ubiquitin ligase, to c-Maf, leading to its degradation through the proteasomal pathway. Further investigation revealed that TMEPAI interacts with NEDD4 via its conserved PY motifs. Alanine substitution or deletion of these motifs abrogated the TMEPAI complex formation with NEDD4, resulting in failed c-Maf degradation. Functionally, TMEPAI suppressed the transcriptional activity of c-Maf. Of note, increased TMEPAI expression was positively associated with the overall survival of MM patients. Moreover, TMEPAI was down-regulated in MM cells, and re-expression of TMEPAI induced MM cell apoptosis. In conclusion, this study highlights that TMEPAI decreases c-Maf stability by recruiting the ubiquitin ligase NEDD4 to c-Maf for proteasomal degradation. Our findings suggest that the restoration of functional TMEPA1 expression may represent a promising complementary therapeutic strategy for treating patients with MM.

Inhibition of the deubiquitinase USP5 leads to c-Maf protein degradation and myeloma cell apoptosis.

The deubiquitinase USP5 stabilizes c-Maf, a key transcription factor in multiple myeloma (MM), but the mechanisms and significance are unclear. In the present study, USP5 was found to interact with c-Maf and prevented it from degradation by decreasing its polyubiquitination level. Specifically, the 308th and 347th lysine residues in c-Maf were critical for USP5-mediated deubiquitination and stability. There are five key domains in the USP5 protein and subsequent studies revealed that the cryptic ZnF domain and the C-box domain interacted with c-Maf but the UBA1/UBA2 domain partly increased its stability. Notably, MafA and MafB are also members of the c-Maf family, however, USP5 failed to deubiquitinate MafA, suggesting its substrate specificity. In the functional studies, USP5 was found to promoted the transcriptional activity of c-Maf. Consistent with the high level of c-Maf protein in MM cells, USP5 was also highly expressed. When USP5 was knocked down, c-Maf underwent degradation. Interestingly, USP5 silence led to apoptosis of MM cells expressing c-Maf but not MM cells lacking c-Maf, indicating c-Maf is a key factor in USP5-mediated MM cell proliferation and survival. Consistent with this finding, WP1130, an inhibitor of several Dubs including USP5, suppressed the transcriptional activity of c-Maf and induced MM cell apoptosis. When c-Maf was overexpressed, WP1130-induced MM cell apoptosis was abolished. Taken together, these findings suggest that USP5 regulates c-Maf stability and MM cell survival. Targeting the USP5/c-Maf axis could be a potential strategy for MM treatment.

Discrimination of plant root zone water status in greenhouse production based on phenotyping and machine learning techniques.

Plant-based sensing on water stress can provide sensitive and direct reference for precision irrigation system in greenhouse. However, plant information acquisition, interpretation, and systematical application remain insufficient. This study developed a discrimination method for plant root zone water status in greenhouse by integrating phenotyping and machine learning techniques. Pakchoi plants were used and treated by three root zone moisture levels, 40%, 60%, and 80% relative water content. Three classification models, Random Forest (RF), Neural Network (NN), and Support Vector Machine (SVM) were developed and validated in different scenarios with overall accuracy over 90% for all. SVM model had the highest value, but it required the longest training time. All models had accuracy over 85% in all scenarios, and more stable performance was observed in RF model. Simplified SVM model developed by the top five most contributing traits had the largest accuracy reduction as 29.5%, while simplified RF and NN model still maintained approximately 80%. For real case application, factors such as operation cost, precision requirement, and system reaction time should be synthetically considered in model selection. Our work shows it is promising to discriminate plant root zone water status by implementing phenotyping and machine learning techniques for precision irrigation management.

The ubiquitin ligase HERC4 mediates c-Maf ubiquitination and delays the growth of multiple myeloma xenografts in nude mice.

The transcription factor c-Maf is extensively involved in the pathophysiology of multiple myeloma (MM), a fatal malignancy of plasma cells. In the present study, affinity chromatography and mass spectrometry were used to identify c-Maf ubiquitination-associated proteins, from which the E3 ligase HERC4 was found to interact with c-Maf and catalyzed its polyubiquitination and subsequent proteasome-mediated degradation. HERC4 mediated polyubiquitination at K85 and K297 in c-Maf, and this polyubiquitination could be prevented by the isopeptidase USP5. Further analysis on the NCI-60 cell line collection revealed that RPMI 8226, a MM-derived cell line, expressed the lowest level of HERC4. Primary bone marrow analysis revealed HERC4 expression was high in normal bone marrow, but was steadily decreased during myelomagenesis. These findings suggested HERC4 played an important role in MM progression. Moreover, ectopic HERC4 expression decreased MM proliferation in vitro, and delayed xenograft tumor growth in vivo. Therefore, modulation of c-Maf ubiquitination by targeting HERC4 may represent a new therapeutic modality for MM.

Poly-(allylamine hydrochloride)-coated but not poly(acrylic acid)-coated upconversion nanoparticles induce autophagy and apoptosis in human blood cancer cells.

Upconversion nanoparticles (UCNPs) have gained increased attention due to their various medical applications such as drug delivery, detection, imaging, and photodynamic therapy. But little is known about their direct biological activity. In the present study, we synthesized NaYF4:Yb,Er UCNPs coated with poly-(allylamine hydrochloride) (PAH) or poly(acrylic acid) (PAA) and investigated their effects in human myeloma and leukemia cells. When these cells were incubated with both types of UCNPs, we found that PAH-UCNPs but not PAA-UCNPs increased the expression of LC3-II, a hallmark of autophagy. This effect was confirmed by the accumulation of LC3 puncta as analyzed by immunofluorescence microscopy. Induction of LC3-II could be blocked by 3-methyl adenosine (3-MA), an autophagy inhibitor. Consistent with this observation, PAH-UCNPs also inhibited both AKT and mTOR activation, the key step in autophagy activation. Further studies demonstrated that PAH-UCNPs also decreased Bcl-2 but increased Beclin1 and Atg14 expression, suggesting that PAH-UCNPs-induced autophagy was associated with increased PI3KC3-Beclin1 activity. Moreover, PAH-UCNPs induced apoptosis in myeloma cells and enhanced apoptosis induced by bortezomib and doxorubicin, two major anti-myeloma drugs. Therefore, our study suggested that PAH-UCNPs alone can induce both apoptosis and autophagy in human blood cancer cells by modulating the AKT-mTOR and PI3KC3-Beclin1-Atg14 signaling pathways. This study implies the potential application of PAH-UCNPs in blood cancer-cell killing.

Clioquinol induces pro-death autophagy in leukemia and myeloma cells by disrupting the mTOR signaling pathway.

Clioquinol is an anti-microbial drug, and it was recently found to induce cancer cell death. In the present study, clioquinol was found to trigger autophagy by inducing LC3 lipidation and autophagosome formation which was abolished by an autophagy inhibitor 3-methyladenine. Further study showed clioquinol displayed no effects on PI3KC3 or Beclin 1 expression but downregulated the expression and the enzymatic activity of mammalian target of Rapamycin (mTOR), a critical modulator of autophagy. Moreover, clioquinol inhibited the catalytic activity of the mTOR complex 1, thus suppressing phosphorylation of P70S6K and 4E-BP1, two major proteins associated with autophagy in the mTORC1 signaling pathway. Clioquinol induced leukemia and myeloma cell apoptosis, however, addition of autophagy inhibitor 3-methyladenine attenuated this kind of cell death. Therefore, this study demonstrated that clioquinol induces autophagy in associated with apoptosis in leukemia and myeloma cells by disrupting mTOR signaling pathway.