Heterogenetic mechanism in multidimensional pit mud with different fermentation years: From microbial structure dynamic succession to metabolism phenotypes.

Pit mud (PM) is fermenting agents in the strong-flavor baijiu (SFB) production. In this paper, the discrepancies in fermentation parameters, microbial community succession patterns and metabolic phenotypes were compared in multidimensional PMs. The results showed that pyruvic acid, succinic acid, S-Acetyldihydrolipoamide-E, glycerol and glyceric acid were the key metabolites responsible for the metabolic differences between the 2-, 30-,100- and 300-year multidimensional PMs, while the butanoic acid, heptyl, heptanoic acid, heptanoic acid ethyl ester, hexanoic acid and octanoic acid were the key differential flavor compounds in the 2-, 30-,100- and 300-year multidimensional PMs. Concurrently, the diversity and abundance of microbial community also exhibited significant differences between the new and old multidimensional PMs, the assembly pattern of bacterial communities changed from deterministic to stochasticity from lower (bottom of the pit and under the huangshui fluid) to upper PM (up the huangshui fluid and top of the pit). Key microorganisms related to the succession process of the lower PM were Clostridium, Methanobacterium, Petrimonas, Lactobacillus, Methanobrevibacter, Bellilinea, Longilinea, Bacillus. In contrast, the upper PM were Caproicibacter, Longilinea, Lactobacillus, Proteinphilum, Methanobrevibacter, Methanobacterium, Methanobacteriaceae, Petrimonas, Bellilinea and Atopobium. Redundancy analysis (RDA) indicated that the key environmental factors regulating the succession of microbial in upper PM were lactic acid, moisture, pH and available phosphorus. In contrast, the lower was lactic acid, acetic acid and ammonia N. Based on these results, heterogeneous mechanisms between new and old multidimensional PMs were explored, providing a theoretical support for improving the quality of new PM.

Multicopper Oxidase from Lactobacillus hilgardii: Mechanism of Degradation of Tyramine and Phenylethylamine in Fermented Food.

Elevated levels of biogenic amines (BAs) in fermented food can have negative effects on both the flavor and health. Mining enzymes that degrade BAs is an effective strategy for controlling their content. The study screened a strain of Lactobacillus hilgardii 1614 from fermented food system that can degrade BAs. The multiple copper oxidase genes LHMCO1614 were successfully mined after the whole genome protein sequences of homologous strains were clustered and followed by homology modeling. The enzyme molecules can interact with BAs to stabilize composite structures for catalytic degradation, as shown by molecular docking results. Ingeniously, the kinetic data showed that purified LHMCO1614 was less sensitive to the substrate inhibition of tyramine and phenylethylamine. The degradation rates of tyramine and phenylethylamine in huangjiu (18% vol) after adding LHMCO1614 were 41.35 and 40.21%, respectively. Furthermore, LHMCO1614 demonstrated universality in degrading tyramine and phenylethylamine present in other fermented foods as well. HS-SPME-GC-MS analysis revealed that, except for aldehydes, the addition of enzyme treatment did not significantly alter the levels of major flavor compounds in enzymatically treated fermented foods (p > 0.05). This study presents an enzymatic approach for regulating tyramine and phenylethylamine levels in fermented foods with potential applications both targeted and universal.

Polyphyllin VII Promotes Apoptosis in Breast Cancer by Inhibiting MAPK/ERK Signaling Pathway through Downregulation of SOS1.

Polyphyllin VII is a biologically active herbal monomer extracted from the traditional Chinese herbal medicine Chonglou. Many studies have demonstrated the anticancer activity of polyphyllin VII against various types of cancers, such as colon, liver, and lung cancer, but its effect on breast cancer has not been elucidated. In this study, we demonstrate that polyphyllin VII inhibited proliferation, increased production of intracellular reactive oxygen species, and decreased mitochondrial membrane potential in breast cancer cells. Notably, polyphyllin VII also induced apoptosis via the mitochondrial pathway. Transcriptome sequencing was used to analyze the targets of PPVII in regulating breast cancer cells. Mechanistic studies showed that polyphyllin VII downregulated Son of Sevenless1 (SOS1) and inhibited the MAPK/ERK pathway. Furthermore, PPVII exerted strong antitumor effects in vivo in nude mice injected with breast cancer cells. Our results suggest that PPVII may promote apoptosis through regulating the SOS1/MAPK/ERK pathway, making it a possible candidate target for the treatment of breast cancer.

DSNetax: a deep learning species annotation method based on a deep-shallow parallel framework.

Microbial community analysis is an important field to study the composition and function of microbial communities. Microbial species annotation is crucial to revealing microorganisms' complex ecological functions in environmental, ecological and host interactions. Currently, widely used methods can suffer from issues such as inaccurate species-level annotations and time and memory constraints, and as sequencing technology advances and sequencing costs decline, microbial species annotation methods with higher quality classification effectiveness become critical. Therefore, we processed 16S rRNA gene sequences into k-mers sets and then used a trained DNABERT model to generate word vectors. We also design a parallel network structure consisting of deep and shallow modules to extract the semantic and detailed features of 16S rRNA gene sequences. Our method can accurately and rapidly classify bacterial sequences at the SILVA database's genus and species level. The database is characterized by long sequence length (1500 base pairs), multiple sequences (428,748 reads) and high similarity. The results show that our method has better performance. The technique is nearly 20% more accurate at the species level than the currently popular naive Bayes-dominated QIIME 2 annotation method, and the top-5 results at the species level differ from BLAST methods by <2%. In summary, our approach combines a multi-module deep learning approach that overcomes the limitations of existing methods, providing an efficient and accurate solution for microbial species labeling and more reliable data support for microbiology research and application.

Research progress on oncoprotein hepatitis B X­interacting protein (Review).

Hepatitis B X­interacting protein (HBXIP) is a membrane protein located on the lysosomal surface and encoded by the Lamtor gene. It is expressed by a wide range of tumor types, including breast cancer, esophageal squamous cell carcinoma and hepatocellular carcinoma, and its expression is associated with certain clinicopathological characteristics. In the past decade, research on the oncogenic mechanisms of HBXIP has increased and the function of HBXIP in normal cells has been gradually elucidated. In the present review, the following was discussed: The normal physiological role of the HBXIP carcinogenic mechanism; the clinical significance of high levels of HBXIP expression in different tumors; HBXIP regulation of transcription, post­transcription and post­translation processes in tumors; the role of HBXIP in improving the antioxidant capacity of tumor cells; the inhibition of ferroptosis of tumor cells and regulating the metabolic reprogramming of tumor cells; and the role of HBXIP in promoting the malignant progression of tumors. In conclusion, the present review summarized the existing knowledge of HBXIP, established its carcinogenic mechanism and discussed future related research on HBXIP.

Malate, a natural inhibitor of 6PGD, improves the efficacy of chemotherapy in lung cancer.

OBJECTIVE: Metabolic reprogramming is an important coordinator of tumor development and resistance to therapy, such as the tendency of tumor cells to utilize glycolytic energy rather than oxidative phosphorylation, even under conditions of sufficient oxygen. Therefore, targeting metabolic enzymes is an effective strategy to overcome therapeutic resistance. MATERIALS AND METHODS: We explored the differential expression and growth-promoting function of MDH2 by immunohistochemistry and immunoblotting experiments in lung cancer patients and lung cancer cells. Pentose phosphate pathway-related phenotypes (including ROS levels, NADPH levels, and DNA synthesis) were detected intracellularly, and the interaction of malate and proteinase 6PGD was detected in vitro. In vivo experiments using implanted xenograft mouse models to explore the growth inhibitory effect and pro-chemotherapeutic function of dimethyl malate (DMM) on lung cancer. RESULTS: We found that the expression of malate dehydrogenase (MDH2) in the tricarboxylic acid cycle (TCA cycle) was increased in lung cancer. Biological function enrichment analysis revealed that MDH2 not only promoted oxidative phosphorylation, but also promoted the pentose phosphate pathway (PPP pathway). Mechanistically, it was found that malate, the substrate of MDH2, can bind to the PPP pathway metabolic enzyme 6PGD, inhibit its activity, reduce the generation of NADPH, and block DNA synthesis. More importantly, DMM can improve the sensitivity of lung cancer to the clinical drug cisplatin. CONCLUSION: We have identified malate as a natural inhibitor of 6PGD, which will provide new leads for the development of 6PGD inhibitors. In addition, the metabolic enzyme MDH2 and the metabolite malate may provide a backup option for cells to inhibit their own carcinogenesis, as the accumulated malate targets 6PGD to block the PPP pathway and inhibit cell cycle progression.

Excessive dietary iron exposure increases the susceptibility of largemouth bass (Micropterus salmoides) to Aeromonas hydrophila by interfering with immune response, oxidative stress, and intestinal homeostasis.

Iron is an essential cofactor in the fundamental metabolic pathways of organisms. Moderate iron intake can enhance animal growth performance, while iron overload increases the risk of pathogen infection. Although the impact of iron on the pathogen-host relationship has been confirmed in higher vertebrates, research in fish is extremely limited. The effects and mechanisms of different levels of iron exposure on the infection of Aeromonas hydrophila in largemouth bass (Micropterus salmoides) remain unclear. In this study, experimental diets were prepared by adding 0, 800, 1600, and 3200 mg/kg of FeSO4∙7H2O to the basal feed, and the impact of a 56-day feeding period on the mortality rate of largemouth bass infected with A. hydrophila was analyzed. Additionally, the relationships between mortality rate and tissue iron content, immune regulation, oxidative stress, iron homeostasis, gut microbiota, and tissue morphology were investigated. The results showed that the survival rate of largemouth bass infected with A. hydrophila decreased with increasing iron exposure levels. Excessive dietary iron intake significantly increased iron deposition in the tissues of largemouth bass, reduced the expression and activity of antioxidant enzymes superoxide dismutase, catalase, and glutathione peroxidase, increased the content of lipid peroxidation product malondialdehyde, and thereby induced oxidative stress. Excessive iron supplementation could influence the immune response of largemouth bass by upregulating the expression of pro-inflammatory cytokines in the intestine and liver, while downregulating the expression of anti-inflammatory cytokines. Additionally, excessive iron intake could also affect iron metabolism by inducing the expression of hepcidin, disrupt intestinal homeostasis by interfering with the composition and function of the gut microbiota, and induce damage in the intestinal and hepatic tissues. These research findings provide a partial theoretical basis for deciphering the molecular mechanisms underlying the influence of excessive iron exposure on the susceptibility of largemouth bass to pathogenic bacteria.

The antipsychotic drug pimozide promotes apoptosis through the RAF/ERK pathway and enhances autophagy in breast cancer cells.

The antipsychotic drug pimozide has been demonstrated to inhibit cancer. However, the precise anti-cancer mechanism of pimozide remains unclear. The purpose of this study was to investigate the effects of pimozide on human MCF-7 and MDA-MB-231 breast cancer cell lines, and the potential involvement in the RAF/ERK signaling. The effects of pimozide on cells were examined by 4,5-dimethylthiazol-2-yl-3,5-diphenylformazan, wound healing, colony formation, transwell assays, and caspase activity assay. Flow cytometry and acridine orange and ethidium bromide staining were performed to assess changes in cells. Transmission electron microscopy and monodansylcadaverine staining were used to observe autophagosomes. The cyclic adenosine monophosphate was evaluated using the FRET system. Immunohistochemistry, immunofluorescence, RNA interference, and western blot investigated the expression of proteins. Mechanistically, we focus on the RAF1/ERK signaling. We detected pimozide was docked to RAF1 by Schrodinger software. Pimozide down-regulated the phosphorylation of RAF1, ERK 1/2, Bcl-2, and Bcl-xl, up-regulated Bax, and cleaved caspase-9 to induce apoptosis. Pimozide might promote autophagy by up-regulating cAMP. The enhancement of autophagy increased the conversion of LC3-I to LC3-II and down-regulated p62 expression. But mTOR signaling was not involved in promoting autophagy. The knockdown of RAF1 expression induced autophagy and apoptosis in breast cancer cells, consistent with the results of pimozide or sorafenib alone. Blocked autophagy by chloroquine resulted in the impairment of pimozide-induced apoptosis. These data showed that pimozide inhibits breast cancer by regulating the RAF/ERK signaling pathway and might activate cAMP-induced autophagy to promote apoptosis and it may be a potential drug for breast cancer treatment.

Antioxidant curcumin induces oxidative stress to kill tumor cells (Review).

Curcumin is a plant polyphenol in turmeric root and a potent antioxidant. It binds to antioxidant response elements for gene regulation by nuclear factor erythroid 2-related factor 2, thereby suppressing reactive oxygen species (ROS) and exerting anti-inflammatory, anti-infective and other pharmacological effects. Of note, curcumin induces oxidative stress in tumors. It binds to several enzymes in tumors, such as carbonyl reductases, glutathione S-transferase P1 and nicotinamide adenine dinucleotide phosphate to induce mitochondrial damage, increase ROS production and ultimately induce tumor cell death. However, the instability and poor pharmacokinetic profile of curcumin in vivo limit its clinical application. Therefore, the effects of curcumin in vivo may be enhanced through its combination with drugs, derivative development and nanocarriers. In the present review, the mechanisms of curcumin that induce tumor cell death through oxidative stress are discussed. In addition, the methods used to enhance the antitumor activity of curcumin are described. Finally, the existing knowledge on the functions of curcumin in tumors, particularly in terms of oxidative stress, are summarized to facilitate future curcumin research.

Environmental factors drive microbial community succession in biofortified wheat Qu and its improvement on the quality of Chinese huangjiu.

Wheat Qu plays the role of saccharification fermentation, providing microorganisms and flavor in the fermentation of huangjiu, and the use of functional microorganisms to fortify wheat Qu is becoming increasingly popular. Yet, the mechanisms promoting microbial successions of wheat Qu remain unclear. In this study, we first correlated microbial community succession with physicochemical factors (moisture, temperature, acidity, glucoamylase and amylase) in inoculated raw wheat Qu (IRWQ) with Saccharopolyspora rosea. The Mantel test was performed to investigate the significance and found that temperature (r = 0.759, P = 0.001), moisture (r = 0.732, P = 0.006), and acidity (r = 0.712, P = 0.017) correlated significantly with the bacterial community in phase 1 (0-40 h). Meanwhile, temperature correlated significantly with the fungal community in phases 1 and 2 (40-120 h). To confirm the effect of temperature on microbial communities, the artificial reduction of bio-heat (37°C) in IRWQ also reduced the relative abundance of heat-resistant microorganisms including Bacillus and Saccharopolyspora. A higher abundance of Saccharopolyspora (87%) in IRWQ was observed following biofortified inoculation of S. rosea, in which glucoamylase activity increased by 40% compared to non-inoculated raw wheat Qu (NIRWQ) (1086 U/g vs 776 U/g). Finally, the IRWQ was employed to mechanized huangjiu fermentation and it was found to reduce the bitter amino acid and higher alcohol content by 27% and 8%, respectively, improving the drinking comfort and quality of huangjiu.

Microbial-derived salt-tolerant proteases and their applications in high-salt traditional soybean fermented foods: a review.

Different microorganisms can produce different proteases, which can adapt to different industrial requirements such as pH, temperature, and pressure. Salt-tolerant proteases (STPs) from microorganisms exhibit higher salt tolerance, wider adaptability, and more efficient catalytic ability under extreme conditions compared to conventional proteases. These unique enzymes hold great promise for applications in various industries including food, medicine, environmental protection, agriculture, detergents, dyes, and others. Scientific studies on microbial-derived STPs have been widely reported, but there has been little systematic review of microbial-derived STPs and their application in high-salt conventional soybean fermentable foods. This review presents the STP-producing microbial species and their selection methods, and summarizes and analyzes the salt tolerance mechanisms of the microorganisms. It also outlines various techniques for the isolation and purification of STPs from microorganisms and discusses the salt tolerance mechanisms of STPs. Furthermore, this review demonstrates the contribution of modern biotechnology in the screening of novel microbial-derived STPs and their improvement in salt tolerance. It highlights the potential applications and commercial value of salt-tolerant microorganisms and STPs in high-salt traditional soy fermented foods. The review ends with concluding remarks on the challenges and future directions for microbial-derived STPs. This review provides valuable insights into the separation, purification, performance enhancement, and application of microbial-derived STPs in traditional fermented foods.

Enzymatic properties and inhibition tolerance analysis of key enzymes in ß-phenylethanol anabolic pathway of Saccharomyces cerevisiae HJ.

Huangjiu is known for its unique aroma, primarily attributed to its high concentration of ß-phenylethanol (ranging from 40 to 130 mg/L). Phenylalanine aminotransferase Aro9p and phenylpyruvate decarboxylase Aro10p are key enzymes in the ß-phenylethanol synthetic pathway of Saccharomyces cerevisiae HJ. This study examined the enzymatic properties of these two enzymes derived from S. cerevisiae HJ and S288C. After substrate docking, Aro9pHJ (-24.05 kJ/mol) and Aro10pHJ (-14.33 kJ/mol) exhibited lower binding free energies compared to Aro9pS288C (-21.93 kJ/mol) and Aro10pS288C (-12.84 kJ/mol). ARO9 and ARO10 genes were heterologously expressed in E. coli BL21. Aro9p, which was purified via affinity chromatography, showed inhibition by l-phenylalanine (L-PHE), but the reaction rate Vmax(Aro9pHJ: 23.89 µmol·(min∙g)-1) > Aro9pS288C: 21.3 µmol·(min∙g)-1) and inhibition constant Ki values (Aro9pHJ: 0.28 mol L-1>Aro9pS288C 0.26 mol L-1) indicated that Aro9p from S. cerevisiae HJ was more tolerant to substrate stress during Huangjiu fermentation. In the presence of the same substrate phenylpyruvate (PPY), Aro10pHJ exhibited a stronger affinity than Aro10pS288C. Furthermore, Aro9pHJ and Aro10pHJ were slightly more tolerant to the final metabolites ß-phenylethanol and ethanol, respectively, compared to those from S288C. The study suggests that the mutations in Aro9pHJ and Aro10pHJ may contribute to the increased ß-phenylethanol concentration in Huangjiu. This is the first study investigating enzyme tolerance mechanisms in terms of substrate and product, providing a theoretical basis for the regulation of the ß-phenylethanol metabolic pathway.