This study investigated the secondary metabolites in the product of rice fermentation with Alternaria alternata and their activity of down-regulating thymosin beta 10(TMSB10) protein expression. The secondary metabolites of A. alternata were separated and purified by various chromatographic separation techniques, including silica gel column chromatography, Sephadex LH-20 column chromatography, and semi-preparative high-performance liquid chromatography. Their structures were identified by spectral techniques such as nuclear magnetic resonance spectroscopy(NMR), infrared spectroscopy(IR), and high-resolution electrospray ionization mass spectrometry(HR-ESI-MS) and comparison with literature data. A total of 10 compounds were isolated and identified, including two new compounds(1S,3S)-2,3-dihydro-3,6-dihydroxy-8-methoxy-1-methylcyclopenta[c][2]benzopyran-5(1H)-one(1), 3,3a, 6-trihydroxy-8-methoxy-1-methyl-2,3,3a, 9b-tetrahydrocyclopenta[c]isochromen-5(1H)-one(2), and the eight known compounds are alternariol 9-methyl ether(3), 1-deoxyrubralactone(4),(3aR)-3,3a-dihydro-1,6-dihydroxy-8-methoxy-3a-methylcyclopenta[c][2]benzopyran-2,5-dione(5), altechromone A(6), alternariol(7), altenuene(8), altenusin(9), and 3'-hydroxyalternariol 5-O-methyl ether(10). The effects of these compounds on TMSB10 expression were tested. Compound 7 showed a significant down-regulation effect on TMSB10 expression with an inhibition rate of 40.5%. The results showed that benzopyrone compounds of A. alternata have the activity of down-regulating the expression of TMSB10 protein, providing theoretical basis and research value for the study of non-small cell lung cancer.
Alternaria/metabolism* ; Down-Regulation/drug effects* ; Humans ; Molecular Structure

Guided by molecular networking, nine novel curvularin derivatives (1-9) and 16 known analogs (10-25) were isolated from the hydrothermal vent sediment fungus Penicillium sp. HL-50. Notably, compounds 5-7 represented a hybrid of curvularin and purine. The structures and absolute configurations of compounds 1-9 were elucidated via nuclear magnetic resonance (NMR) spectroscopy, X-ray diffraction, electronic circular dichroism (ECD) calculations, ¹³C NMR calculation, modified Mosher's method, and chemical derivatization. Investigation of anti-inflammatory activities revealed that compounds 7-9, 11, 12, 14, 15, and 18 exhibited significant suppressive effects against lipopolysaccharide (LPS)-induced nitric oxide (NO) production in murine macrophage RAW264.7 cells, with IC₅₀ values ranging from 0.44 to 4.40 μmol·L^-1. Furthermore, these bioactive compounds were found to suppress the expression of inflammation-related proteins, including inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), NLR family pyrin domain-containing protein 3 (NLRP3), and nuclear factor kappa-B (NF-κB). Additional studies demonstrated that the novel compound 7 possessed potent anti-inflammatory activity by inhibiting the transcription of inflammation-related genes, downregulating the expression of inflammation-related proteins, and inhibiting the release of inflammatory cytokines, indicating its potential application in the treatment of inflammatory diseases.
Penicillium/chemistry* ; Mice ; Animals ; Anti-Inflammatory Agents/isolation & purification* ; RAW 264.7 Cells ; Nitric Oxide/metabolism* ; Hydrothermal Vents/microbiology* ; Macrophages/immunology* ; Molecular Structure ; Nitric Oxide Synthase Type II/immunology* ; Cyclooxygenase 2/immunology* ; Geologic Sediments/microbiology* ; NF-kappa B/immunology* ; NLR Family, Pyrin Domain-Containing 3 Protein/immunology*

Five novel nor-eremophilane-type sesquiterpenoids, peniroqueforins E-H and J (1-4 and 7), two new eremophilane-type sesquiterpenoids, peniroqueforins I and K (5 and 8), and a new eudesmane-type sesquiterpenoid, peniroqueforin L (9), along with four known compounds (6 and 10-12), were isolated and characterized from fungus Penicillium roqueforti (P. roqueforti). The structures and absolute configurations of these compounds were determined through comprehensive spectroscopic analyses, electronic circular dichroism (ECD) data analyses, and single-crystal X-ray diffraction methods. The anti-multi-drug resistance (MDR) cancer activity of these compounds was evaluated using SW620/Ad300 cells. Notably, the half maximal inhibitory concentration (IC₅₀) value of paclitaxel (PTX) combined with 1 in SW620/Ad300 cells was 50.36 nmol·L^-1, which was 65-fold more potent than PTX alone (IC₅₀ 3.26 μmol·L^-1). Subsequent molecular docking studies revealed an affinity between compound 1 and P-glycoprotein (P-gp), suggesting that this nor-eremophilane-type sesquiterpenoid (1) could serve as a potential lead for MDR reversal in cancer cells through P-gp inhibition.
Penicillium/chemistry* ; Humans ; Sesquiterpenes/isolation & purification* ; Cell Line, Tumor ; Molecular Structure ; Drug Resistance, Neoplasm/drug effects* ; Antineoplastic Agents/pharmacology* ; Drug Resistance, Multiple/drug effects* ; Molecular Docking Simulation

(±)-Penicithrones A-D (1a/1b-4a/4b), four novel pairs of anthrone-cyclopentenone heterodimers characterized by a distinctive bridged 6/6/6-5 tetracyclic core skeleton, together with three previously identified compounds (5-7), were isolated from the crude extract of the mangrove-derived fungus Penicillium sp., guided by heteronuclear single quantum correlation (HSQC)-based small molecule accurate recognition technology (SMART 2.0) and liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based molecular networking. The structural elucidation of new compounds was accomplished through comprehensive spectroscopic analysis, and their absolute configurations were determined using DP4+ ¹³C nuclear magnetic resonance (NMR) calculations and electronic circular dichroism (ECD) calculations. Compounds 1a/1b-4a/4b demonstrated moderate cytotoxicity against three human cancer cell lines HeLa, HCT116 and MCF-7 with half maximal inhibitory concentration (IC₅₀) values ranging from 15.95 ± 1.64 to 28.56 ± 2.59 μmol·L^-1.
Humans ; Penicillium/chemistry* ; Molecular Structure ; Cyclopentanes/isolation & purification* ; Cell Line, Tumor ; Antineoplastic Agents/pharmacology* ; Tandem Mass Spectrometry ; Dimerization ; HeLa Cells ; Magnetic Resonance Spectroscopy

Hormesis effect has been observed in the secondary metabolite synthesis of microorganisms induced by rare earth elements. However, the underlying molecular mechanism remains unclear. To analyze the molecular mechanism of the regulatory effect of Rhodotorula mucilaginosa in the presence of lanthanum chloride, different concentrations of lanthanum chloride were added to the fermentation medium of Rhodotorula mucilaginosa, and the carotenoid content was subsequently measured. It was found that the concentrations of La³⁺ exerting the promotional and inhibitory effects were 0-100 mg/L and 100-400 mg/L, respectively. Furthermore, the expression of 33 genes and the synthesis of 55 metabolites were observed to be up-regulated, while the expression of 85 genes and the synthesis of 123 metabolites were found to be down-regulated at the concentration range of the promotional effect. Notably, the expression of carotenoid synthesis-related genes except AL1 was up-regulated. Additionally, the content of β-carotene, lycopene, and astaxanthin demonstrated increases of 10.74%, 5.02%, and 3.22%, respectively. The expression of 5 genes and the synthesis of 91 metabolites were up-regulated, while the expression of 35 genes and the synthesis of 138 metabolites were down-regulated at the concentration range of the inhibitory effect. Meanwhile, the content of β-carotene, lycopene, and astaxanthin decreased by 21.73%, 34.81%, and 35.51%, respectively. In summary, appropriate concentrations of rare earth ions can regulate the synthesis of secondary metabolites by modulating the activities of various enzymes involved in metabolic pathways, thereby exerting the hormesis effect. The findings of this study not only contribute to our comprehension for the mechanism of rare earth elements in organisms but also offer a promising avenue for the utilization of rare earth elements in diverse fields, including agriculture, pharmaceuticals, and healthcare.
Lanthanum/pharmacology* ; Rhodotorula/genetics* ; Carotenoids/metabolism* ; Hormesis/drug effects* ; Fermentation ; Multiomics

Protein tyrosine phosphatases (PTPs, EC 3.1.3.48) are key regulators of cellular processes, with the catalytic activity attributed to the conserved motif (H/V)CX₅R(S/T), where cysteine and arginine residues are critical. Previous studies revealed that alternative splicing of extracellular phosphatase mRNA precursors in Metarhizium anisopliae generated two distinct transcripts, with the longer sequence containing a novel HCPTPMLS motif resembling PTP signatures but lacking the arginine residue. To identify the novel signature motif and characterize its enzymatic properties, we heterologously expressed and purified both proteins in Pichia pastoris and comprehensively characterized their enzymatic properties. The protein containing the HCPTPMLS motif (designated as L-protein) exhibited the highest activity at pH 5.5 and a strong preference for pTyr substrates. Its phosphatase activity was inhibited by Ag⁺, Zn²⁺, Cu²⁺, molybdate, and tungstate, but enhanced by Ca²⁺ and EDTA. AcP101 (lacking HCPTPMLS) showed the maximal activity at pH 6.5 and a strong preference toward pNPP (P < 0.05), with the activity inhibited by NaF and tartrate, but enhanced by Mg²⁺ and Mn²⁺. Functional analysis confirmed that the L-protein retained the PTP activity despite the absence of arginine in its signature motif, while AcP101 functioned as an acid phosphatase. This study provides the first functional validation of an arginine-deficient PTP motif, expanding the definition of PTP signature motifs and offering new insights for phosphatase classification.
Metarhizium/genetics* ; Protein Tyrosine Phosphatases/chemistry* ; Amino Acid Motifs ; Recombinant Proteins/biosynthesis* ; Amino Acid Sequence ; Pichia/metabolism* ; Fungal Proteins/chemistry* ; Substrate Specificity ; Saccharomycetales

Xanthocillin is a unique natural product with an isonitrile group and shows remarkable antibacterial activity. In this study, the genome of an endophytic fungus Penicillium chrysogenum MT-40 isolated from Huperzia serrata was sequenced, and the gene clusters with the potential to synthesize xanthocillin analogues were mined by local BLAST and various bioinformatics analysis tools. As a result, a biosynthetic gene cluster (named for) responsible for the biosynthesis of xanthocillin analogues was identified by further heterologous expression of the key genes in Aspergillus oryzae NSAR1. Specifically, the ForB catalyzes the synthesis of 2-formamido-3-(4-hydroxyphenyl) acrylic acid, and the ForG catalyzes the dimerization of 2-formamido-3-(4-hydroxyphenyl) acrylic acid to produce the xanthocillin analogue N, N'-(1, 4-bis (4-hydroxyphenyl) buta-1, 3-diene-2, 3-diyl) diformamide. The results reported here provide a reference for further discovery of xanthocillin analogues from fungi.
Penicillium chrysogenum/genetics* ; Huperzia/microbiology* ; Acrylates ; Multigene Family

It is unclear how VvLaeA functions in regulating the growth and development of Volvariella volvacea (Bull. ex. Fr.) Sing.. Firstly, bioinformatics analysis of VvLaeA was carried out in this study. Subsequently, the Vvgpd promoter and the open reading frame (ORF) fragment of VvlaeA were amplified and fused by polymerase chain reaction (PCR). The fusion fragment was cloned into the pK2 (bar) plasmid. The recombinant construct pK2(bar)-OEVvlaeA was transfected into Beauveria bassiana by Agrobacterium tumefaciens-mediated transformation. Finally, the growth and development of the transformants were examined. The results showed that VvLaeA shared a low homology with similar proteins in other fungi. Compared with the wild type, the colony diameter of the transformant was significantly increased. However, the pigment deposition, conidial yields and germination rates were significantly decreased. The overexpression strains were more sensitive to stresses than that of the wild type. Further studies showed the conidial cell wall properties of the transformants were altered, and the expressions of genes related to the conidial development were significantly down-regulated. Collectively, VvLaeA increased the growth rate of B. bassiana strains and negatively regulated the pigmentation and conidial development, which shed a light for the functional identification of straw mushroom genes.
Beauveria/metabolism* ; Spores, Fungal ; Growth and Development

Rhodotorula toruloides is a non-conventional red yeast that can synthesize various carotenoids and lipids. It can utilize a variety of cost-effective raw materials, tolerate and assimilate toxic inhibitors in lignocellulosic hydrolysate. At present, it is widely investigated for the production of microbial lipids, terpenes, high-value enzymes, sugar alcohols and polyketides. Given its broad industrial application prospects, researchers have carried out multi-dimensional theoretical and technological exploration, including research on genomics, transcriptomics, proteomics and genetic operation platform. Here we review the recent progress in metabolic engineering and natural product synthesis of R. toruloides, and prospect the challenges and possible solutions in the construction of R. toruloides cell factory.
Gene Editing ; Metabolic Engineering ; Rhodotorula/metabolism* ; Lipids

In Zherong county, Fujian province, the black spot of Pseudostellaria heterophylla often breaks out in the rainy season from April to June every year. As one of the main leaf diseases of P. heterophylla, black spot seriously affects the yield and quality of the medicinal material. To identify and characterize the pathogens causing black spot, we isolated the pathogens, identified them as a species of Alternaria according to Koch's postulates, and then tested their pathogenicity and biological characteristics. The results showed that the pathogens causing P. heterophylla black spot were A. gaisen, as evidenced by the similar colony morphology, spore characteristics, sporulation phenotype, and the same clade with A. gaisen on the phylogenetic tree(the maximum likelihood support rate of 100% and the Bayesian posterior probability of 1.00) built based on the tandem sequences of ITS, tef1, gapdh, endoPG, Alta1, OPA10-2, and KOG1077. The optimum conditions for mycelial growth of the pathogen were 25 ℃, pH 5-8, and 24 h dark culture. The lethal conditions for mycelia and spores were both treatment at 50 ℃ for 10 min. We reported for the first time the A. gaisen-caused black spot of P. heterophylla. The results could provide a theoretical basis for the diagnosis and control of P. heterophylla leaf spot diseases.
Bayes Theorem ; Phylogeny ; Caryophyllaceae ; Alternaria ; Mycelium