Transcript Levels Of Enzymes Research Articles

Stress-induced modification of Escherichia coli tRNA generates 5-methylcytidine in the variable loop

There has been recent interest in trying to understand the connection between transfer RNA (tRNA) posttranscriptional modifications and changes in-cellular environmental conditions. Here, we report on the identification of the modified nucleoside 5-methylcytidine (m5C) in Escherichia coli tRNAs. This modification was determined to be present at position 49 of tRNA Tyr-QUA-II. Moreover, m5C levels in this tRNA are significantly elevated under high reactive oxygen specieis (ROS) conditions in E. coli cells. We identified the known ribosomal RNA methyltransferase rsmF as the enzyme responsible for m5C synthesis in tRNA and enzyme transcript levels are responsive to elevated levels of ROS in the cell. We further find that changes in m5C levels in this tRNA are not specific to Fenton-like reaction conditions elevating ROS, but heat shock can also induce increased modification of tRNA Tyr-QUA-II. Altogether, this work illustrates how cells adapt to changing environmental conditions through variations in tRNA modification profiles.

Transcriptional Downregulation of Methanol Metabolism Key Genes During Yeast Death in Engineered Pichia pastoris.

Pichia pastoris possesses the unique ability to utilize methanol as its sole carbon source, which makes it a proper host for producing various high-value-added products via metabolic engineering. Nevertheless, cell death has been observed during the fermentation of modified P. pastoris, with limited literature elucidating the underlying causes and mechanisms. Understanding the death mechanisms during methanol-based fermentation is crucial for optimizing fermentation strategies, enhancing the accumulation of target products, and reducing production costs. Here, we first sought to eliminate the potential causes of cell death during fermentation, such as inadequate inorganic salts and toxic by-product accumulation. The elimination of these potential causes was achieved efficiently utilizing the high-throughput fermentation equipment. Subsequently, we established a correlation between yeast cell death and the duration of the methanol metabolism period by monitoring the growth of the yeast at different fermentation stages. A critical revelation from this work came from analyzing the yeast's transcriptomic data at various stages of methanol metabolism. It was observed that a significant characteristic of yeast cell death during fermentation was the marked down-regulation of transcript levels of key enzymes involved in the methanol assimilation pathway and genes related to their biosynthesis process. The findings of this work are crucial for better understanding the causes and mechanisms of cell death for engineered P. pastoris during methanol-utilized fermentation.

The impact of freeze-dried Baiyedancong-Oolong tea aqueous extract containing bioactive compounds on the activities of CYP450 enzymes, the transport capabilities of P-gp and OATs, and transcription levels in mice.

In this study, (-)-epigallocatechin gallate (EGCG) and caffeine extracted from freeze-dried autumn Baiyedancong Oolong tea (FBOT) were orally administered to mice for 7 consecutive days to explore the effects of BOT and its bioactive compounds on the activities and transcription levels of CYP450 enzymes, intestinal effluence transporter P-gp, and renal ingestion Organic Anion Transporters (OATs). Concurrently, EGCG and caffeine enhanced the activities of CYP3A, CYP2E1, and CYP2C37 in the liver of mice, while impairing the transport capabilities of P-gp and OATs. Reduced levels of MDR1 encoding P-gp transcription in the small intestine and renal OAT1 and OAT3 revealed that transcription was involved in the regulation of CYP450, P-gp, and OATs. The reduced transcription level of liver CYP2E1 suggested that CYP2E1 activity may have been elevated due to alternative mechanisms, but not through transcription. The absorption, metabolism, and excretion of drugs may be influenced by the daily consumption or high-dose administration of BOT and its related products, in which EGCG and caffeine may make great contributions.

UV-B Radiation Exhibited Tissue-Specific Regulation of Isoflavone Biosynthesis in Soybean Cell Suspension Cultures.

Isoflavones, a class of substances with high biological activity, are abundant in soybeans. This study investigated isoflavone biosynthesis in soybean cell suspension cultures under UV-B radiation. UV-B radiation enhanced the transcription level and activity of key enzymes involved in isoflavone synthesis in cell suspension cultures. As a result, the isoflavone contents significantly increased by 19.80% and 91.21% in hypocotyl and cotyledon suspension cultures compared with the control, respectively. Meanwhile, a significant difference was observed in the composition of isoflavones between soybean hypocotyl and cotyledon suspension cultures. Genistin was only detected in hypocotyl suspension cultures, whereas glycitin, daidzein, and genistein accumulated in cotyledon suspension cultures. Therefore, UV-B radiation exhibited tissue-specific regulation of isoflavone biosynthesis in soybean cell suspension cultures. The combination of suspension cultures and abiotic stress provides a novel technological approach to isoflavone accumulation.

Dehydroabietylamine exerts antitumor effects by affecting nucleotide metabolism in gastric cancer.

Nucleotide metabolism is the ultimate and most critical link in the self-replication process of tumors, including gastric cancer (GC). However, in clinical treatment, classic antitumor drugs such as 5-fluorouracil (5-FU) are mostly metabolic analogs of purines or pyrimidines, which lack specificity for tumor cells and therefore have significant side effects. It is unclear whether there are other drugs that can target nucleotide metabolism, except for nucleic acid analogs. Here, we found that a natural compound, dehydroabietylamine (DHAA), significantly reduced the viability and proliferation of GC cells and organoids. DHAA disrupts the purine and pyrimidine metabolism of GC cells, causing DNA damage and further inducing apoptosis. DHAA treatment decreased transcription and protein levels of key enzymes involved in the nucleotide metabolism pathway, with significant reductions in the expression of pyrimidine metabolism key enzymes CAD, DHODH, and purine metabolism key enzymes PAICS. We also found that DHAA directly binds to and reduces the expression of Forkhead box K2 (FOXK2), a common transcription factor for these metabolic enzymes. Ultimately, DHAA was shown to delay tumorigenesis in K19-Wnt1/C2mE transgenic mice model and reduce levels of CAD, DHODH, and PAICS in vivo. We demonstrate that DHAA exerts an anticancer effect on GC by targeting transcription factor FOXK2, reducing transcription of key genes for nucleotide metabolism and impairing nucleotide biosynthesis, thus DHAA is a promising candidate for GC therapy.

Exploring ocular fibulin-3 (EFEMP1): Anatomical, age-related, and species perspectives

Fibulin-3 (FBLN3, aka EFEMP1) is a secreted extracellular matrix (ECM) glycoprotein implicated in ocular diseases including glaucoma and age-related macular degeneration. Yet surprisingly, little is known about its native biology, expression patterns, and localization in the eye. To overcome these shortcomings, we conducted gene expression analysis and immunohistochemistry for FBLN3 in ocular tissues from mice, pigs, non-human primates, and humans. Moreover, we evaluated age-related changes in FBLN3 and FBLN3-related ECM remodeling enzymes/inhibitors in aging mice. We found that FBLN3 displayed distinct staining patterns consistent across the mouse retina, particularly in the ganglion cell layer and inner nuclear layer (INL). In contrast, human retinas exhibited a unique staining pattern, with enrichment of FBLN3 in the retinal pigment epithelium (RPE), INL, and outer nuclear layer (ONL) in the peripheral retina. This staining transitioned to the outer plexiform layer (OPL) in the central retina/macula, and was accompanied by reduced RPE immunoreactivity approaching the fovea. Surprisingly, we found significant age-related increases in FBLN3 expression and protein abundance in the mouse retina which was paralleled by reduced transcript levels of FBLN3-degrading enzymes (i.e., Mmp2 and Htra1). Our findings highlight important species-dependent, retinal region-specific, and age-related expression and localization patterns of FBLN3 which favor its accumulation during aging. These findings contribute to a better understanding of FBLN3's role in ocular pathology and provide valuable insights for future FBLN3 research.

Bacillus velezensis strain NA16 shows high poultry feather-degrading efficiency, protease and amino acid production

Isolated Bacillus velezensis strain NA16, which produces proteases, amino acids and the transcription levels of different keratinolytic enzymes and disulfide reductase genes in whole gene sequencing, was evaluated during feather degradation. The result shows under optimum fermentation conditions, chicken feather fermentation showed total amino acid concentration of 7599 mg/L, degradation efficiency of 99.3% at 72 h, and protease activity of 1058 U/mL and keratinase activity of 288 U/mL at 48 h. Goose feather fermentation showed total amino acid concentration of 4918 mg/L (96 h), and degradation efficiency was 98.9% at 120 h. Chicken feather fermentation broth at 72 h showed high levels of 17 amino acids, particularly phenylalanine (1050 ± 1.90 mg/L), valine (960 ± 1.04 mg/L), and glutamic (950 ± 3.00 mg/L). Scanning electron microscopy and Fourier transform infrared analysis revealed the essential role of peptide bond cleavage in structural changes and degradation of feathers. Protein purification and zymographic analyses revealed a key role in feather degradation of the 39-kDa protein encoded by gene1031, identified as an S8 family serine peptidase. Whole genome sequencing of NA16 revealed 26 metalloproteinase genes and 22 serine protease genes. Among the proteins, S8 family serine peptidase (gene1031, gene1428) and S9 family peptidase (gene3132) were shown by transcription analysis to play major roles in chicken feather degradation. These findings revealed the transcription levels of different families of keratinolytic enzymes in the degradation of feather keratin by microorganisms, and suggested potential applications of NA16 in feather waste management and amino acid production.

Evaluation of the dietary taurine requirement for early juvenile mud crab Scylla paramamosain

As a conditionally essential amino acid (CEAA), taurine plays an important role in the growth, development, and physiological metabolism of aquatic animals; however, its demand level in the mud crab Scylla paramamosain is still poorly understood. A 60-day growth trial was conducted to investigate the effects of different taurine levels (0% (T0.0), 0.2% (T0.2), 0.4% (T0.4), 0.8% (T0.8), and 1.6% (T1.6)) added to diet on the survival, growth, body composition, immunity, antioxidant and osmoregulatory capacity of early juvenile S. paramamosain (initial body weight 8.69 ± 0.19 mg). The results showed that dietary taurine levels >0.4% (inclusive) significantly increased the survival of early juvenile mud crabs. The growth performance (final body weight (FBW), weight gain (WG), specific growth rate (SGR)) of early juvenile mud crabs increased significantly with increasing dietary taurine levels, reaching a maximum in the T0.8 group, and the target of rapamycin (tor) gene transcription levels also peaked in the T0.8 group. The body composition of mud crabs was not affected by dietary treatments. Further, dietary taurine significantly increased digestive enzyme (trypsin, lipase, and amylase) activities and transcript levels of appetite-related genes (neuropeptide F1 (npf1) and neuropeptide F2 (npf2)) in early juvenile mud crabs. In addition, dietary taurine increased the expression levels of immune-related genes (prophenoloxidase (proPO), prophenoloxidase activating factor 1 (ppaf1), prophenoloxidase activating factor 2 (ppaf2), crustin antimicrobial peptide (crustin), lysozyme (lzm)) and antioxidant capacity (total antioxidant capacity (T-AOC), superoxide dismutase (SOD) and catalase (CAT) activities) in early juvenile mud crabs. Finally, the results of the acute salinity challenge showed that the early juvenile mud crabs' survival in the T0.8 group was significantly higher than that of the other groups, while significantly higher Na+/K+-ATPase (NKA) activity and Na+/K+-ATPase α-subunit (nka-α) and Na+/K+/2Cl−-cotransporter (nkcc) gene expression levels were also detected in the T0.8 group. Together with growth performance, immunity, antioxidant capacity and osmoregulatory capacity, the present study concluded that the appropriate dietary taurine level for early juvenile mud crabs was 1.08%–1.33%.

Integrative effects of phytohormones in the phenolic acids production in Salvia verticillata L. under multi-walled carbon nanotubes and methyl jasmonate elicitation

Salvia verticillata L. is a well-known herb rich in rosmarinic acid (RA) and with therapeutic values. To better understand the possible roles of phytohormones in the production of phenolic acids in S. verticillata, in this work, we investigated some physiological and biochemical responses of the species to methyl jasmonate (MJ) and multi-walled carbon nanotubes (MWCNTs) as two effective elicitors. The leaves were sprayed with aqueous solutions containing 100 mg L−1 MWCNTs and 100 µM MJ and then harvested during interval times of exposure up to 96 h. The level of abscisic acid, as the first effective phytohormone, was altered in the leaves in response to MJ and MWCNTs elicitation (2.26- and 3.06-fold more than the control, respectively), followed by significant increases (P ˂ 0.05) detected in jasmonic acid and salicylic acid contents up to 8 h after exposure. Obtained data revealed that simultaneously with changes in phytohormone profiles, significant (P ˂ 0.05) rises were observed in the content of H2O2 (8.85- and 9.74-folds of control), and the amount of lipid peroxidation (10.18- and 17.01-folds of control) during the initial times after exposure to MJ and MWCNTs, respectively. Later, the content of phenolic acids increased in the elicited leaves due to changes in the transcription levels of key enzymes involved in their biosynthesis pathways, so 2.71- and 11.52-fold enhances observed in the RA content of the leaves after exposure to MJ and MWCNTs, respectively. It is reasonable to conclude that putative linkages between changes in some phytohormone pools lead to the accumulation of phenolic acids in the leaves of S. verticillata under elicitation. Overall, the current findings help us improve our understanding of the signal transduction pathways of the applied stimuli that led to enhanced secondary metabolite production in medicinal plants.

Glutathione-dependent redox homeostasis is critical for chlorothalonil detoxification in tomato leaves

Glutathione plays a critical role in plant growth, development and response to stress. It is a major cellular antioxidant and is involved in the detoxification of xenobiotics in many organisms, including plants. However, the role of glutathione-dependent redox homeostasis and associated molecular mechanisms regulating the antioxidant system and pesticide metabolism remains unclear. In this study, endogenous glutathione levels were manipulated by pharmacological treatments with glutathione synthesis inhibitors and oxidized glutathione. The application of oxidized glutathione enriched the cellular oxidation state, reduced the activity and transcript levels of antioxidant enzymes, upregulated the expression level of nitric oxide and Ca2+ related genes and the content, and increased the residue of chlorothalonil in tomato leaves. Further experiments confirmed that glutathione-induced redox homeostasis is critical for the reduction of pesticide residues. RNA sequencing analysis revealed that miRNA156 and miRNA169 that target transcription factor SQUAMOSA-Promoter Binding Proteins (SBP) and NUCLEAR FACTOR Y (NFY) potentially participate in glutathione-mediated pesticide degradation in tomato plants. Our study provides important clues for further dissection of pesticide degradation mechanisms via miRNAs in plants.

Umami loss mechanism upon shiitake mushrooms under cold storage: Revisiting the role of energy metabolism via integrated physiological and transcriptomic analysis

Umami loss of shiitake mushrooms (Lentinula edodes) is still an inevitable developmental process and a significant challenge hampering its storage and marketing. The present study explores the physiological and molecular responses to umami quality loss involved in energy metabolism during postharvest storage under room temperature (RT) and low temperature (LT) in shiitake mushrooms. Results showed that LT storage prolonged the mushroom shelf life while inhibiting electrolyte leakage, malondialdehyde (MDA), mitochondrial permeability transition pore (MPTP), Ca2+, and H2O2 contents. Moreover, transcriptome analysis revealed that LT storage maintained high umami quality and sufficient energy supply by suppressing the degradation genes of umami nucleotides and the genes related to the mitochondrial energy metabolism pathway, including metabolic pathways and biosynthesis of secondary metabolites. Additionally, the depression activities and gene transcription levels of key enzymes related to umami nucleotide degradation contributed to maintaining umami taste in cold storage. A conceptual model was proposed to elucidate the molecular mechanism of cold storage alleviated umami loss in shiitake mushrooms. The results provide a theoretical foundation for the umami loss mechanism of mushrooms under cold storage.

Metabolome and Transcriptome Association Analysis Reveals the Link Between Pigmentation and Nutrition Utilization in the Juveniles of Sea Cucumber Holothuria leucospilota.

The sea cucumber Holothuria leucospilota is an economically and ecologically important tropical species. Following development into juveniles, H. leucospilota undergoes a color change from white to black, involving a pigmentation process for over a period of several months. In this study, a combination of liquid chromatography-tandem mass spectrometry (LC-MS/MS) and Next-Generation sequencing (NGS) were employed to investigate the changes in metabolomic and transcriptomic profiles during pigmentation in H. leucospilota juveniles. The metabolomic analysis identified a total of 341 metabolites, of which 52 were found to be differentially regulated (P < 0.05 and VIP > 1), with 27 being upregulated in white individuals and 25 in black individuals. Additionally, 632 differentially expressed genes (DEGs) were identified, with 380 genes upregulated in white samples and 252 genes upregulated in black samples. Interestingly, the melanin content and tyrosinase transcript levels did not display significant differences between the two groups. Metabolomic data suggested the involvement of the linoleic acid metabolic pathway in pigmentation. Transcriptomic analysis, coupled with realtime PCR validation, revealed a decrease in the transcript levels of digestive enzymes like α-amylase, maltase-glucoamylase, and trehalase after the juveniles changed to black. Furthermore, the mRNA expressions of major yolk proteins showed a decline, indicating a shift in the accumulation of protein nutrient sources. Overall, our findings suggest that during the pigmentation process in H. leucospilota, no significant changes were observed in the classical melanin pathway, while notable alterations were observed in their nutritional status. This study provides valuable insights into the regulatory mechanisms of pigmentation in marine organisms.

SAT263 Osteocalcin Promotes Cortisol Production In Human Adrenal Cells

Abstract Disclosure: R. Scheske: None. D. van Rooyen: None. W.E. Rainey: None. Background: Osteocalcin (OCN) is a peptide exclusively produced by osteoblasts in bone. In an uncarboxylated form, OCN has been shown to function as a hormone affecting physiological processes such as reproduction and energy metabolism in mice and humans. Murine studies have shown that uncarboxylated OCN upregulates fetal mouse glucocorticoid production through a unique regulatory axis between osteoblasts and adrenal cortex cells. The increase in adrenal steroid production has been attributed to OCN heightening adrenocorticotropic hormone (ACTH). The influence of adrenal glucocorticoids on glucose metabolism and bone composition is well studied but the effect of OCN on human adrenal steroidogenesis remains unknown. Herein, we hypothesized OCN would stimulate cortisol production in human adrenal cells. Methods: OCN’s effect on adrenal glucocorticoid production was investigated using primary cultures of human adrenal cells (PAC). Cells were treated for 24h with 1000 nM OCN with/without cAMP/protein kinase A (PKA) signaling pathway agonists (ACTH, forskolin, 8-bromo cAMP (8Br-cAMP), and dibutyryl cyclic AMP (dbcAMP)). Cortisol levels were quantified using an enzyme-linked immunosorbent assay. RNA was isolated and assessed by quantitative real-time PCR (RT-qPCR) to identify changes in transcript levels of enzymes catalyzing glucocorticoid production: cholesterol side-chain cleavage enzyme (CYP11A1), 17α-hydroxylase/17,20-lyase (CYP17A1), and 11β-hydroxylase (CYP11B1). Results: Cortisol production in PAC were significantly induced by all tested agonists; ACTH (22-fold), forskolin (27-fold), 8Br-cAMP (26-fold), and dbcAMP (30-fold) relative to basal. Although OCN, alone, did not significantly increase cortisol in this cell model, co-treatment with ACTH increased cortisol levels 1.4-fold (40.1 ng/mL; p&amp;lt;0.0001) compared to ACTH (28.2 ng/mL). Similarly, co-treatment with forskolin or 8Br-cAMP increased cortisol, relative to the respective agonists, by 1.3-fold (p&amp;lt;0.001) and dbcAMP by 1.2-fold (p&amp;lt;0.01). RT-qPCR analysis showed no changes in CYP11A1 or CYP17A1 transcript levels in co-treatments with OCN and ACTH relative to ACTH. However, CYP11B1 mRNA levels, relative to the respective controls, were increased in samples co-treated with OCN and ACTH/-agonist. OCN co-treatments with ACTH, forskolin, 8Br-cAMP and dbcAMP increased CYP11B1 levels by 1.8-, 1.4-, 1.3-, and 1.2-fold, respectively. Conclusion: Data presented in this study show stimulatory effect of OCN on human adrenal cells. OCN, in the presence of ACTH and other PKA agonists, increased cortisol production via upregulating CYP11B1 transcript levels. Defining the mechanism and receptor interaction through which OCN functions in human adrenal cells remain to be determined. Presentation: Saturday, June 17, 2023

Environmental factors drive the succession of microbial community structure during wheat Qu fermentation

Environmental factors significantly affected the succession of microbial communities during the fermentation of Huangjiu Raw wheat Qu (RWQ). Influence of environmental factors on microbial community during fermentation of Chinese Huangjiu RWQ. The interaction of environmental factors and microorganisms affects the changes of microbial communities and key enzyme transcript levels in different stages during the production of Huangjiu RWQ. This study combined environmental factor analysis and high-throughput sequencing technology to comprehensively analyze the specific changes of microbial communities and environmental factors at each stage of Huangjiu production. In addition, the influence of the interaction of environmental factors and microorganisms on microbial succession was investigated by correlation analysis, and the enzymes related to saccharification and ethanol fermentation during the production of RWQ were explored. The results showed that Bacillus, Weisseria, Thermophilic actinomycetes and Lactobacillus were the dominant bacteria in RWQ, while Aspergillus, Thermomyces, Fusarium and Alternaria were the dominant fungi. Fungi were active in the early stage of RWQ fermentation, while bacteria were active in the middle and late stage of fermentation. Microbial community succession was closely related to environmental humidity and CO2. These environmental factors were positively correlated with most fungi, but negatively correlated with Thermophilic actinomycetes, Glycopolyspora and Acinetobacter. In fungi, the enzymes related to saccharification and fermentation mainly came from Thermophilic ascomycetes and thermophilic fungi, while in Bacteria, these two enzymes mainly came from weisseria. This study revealed the relationship between RWQ microbe and environmental factors, which was helpful to clarify the succession law of RWQ microbe.

Increased serine synthesis in cumulus cells of young infertile women with diminished ovarian reserve.

What are the differences in gene expression of cumulus cells (CCs) between young women with diminished ovarian reserve (DOR) and those of similar age with normal ovarian reserve (NOR)? Gene expression and metabolome profiling analysis demonstrate that the de novo serine synthesis pathway (SSP) is increased in the CCs of young women with DOR. The incidence of DOR has risen, tending to present at younger ages. Its mechanisms and aetiologies are still poorly understood. Abnormal metabolism is present in luteinized CCs of patients with DOR. Previous studies have revealed that mitochondrial dysfunction and impaired oxidative phosphorylation in CCs are related to DOR in women of advanced age. The pathogenic mechanisms likely differ between young women with DOR and cases associated with advanced maternal age. Several studies have examined amino acid metabolism in the follicle, with a focus on embryo development, but less information is available about CCs. The physiological significance of de novo serine synthesis in follicles and oocytes remains largely unknown. CC samples were obtained from 107 young infertile women (age <38 years) undergoing ICSI, from July 2017 to June 2019, including 54 patients with DOR and 53 patients with NOR. Oocyte development data were analysed retrospectively. Comprehensive genome-wide transcriptomics of CCs was performed. Differentially expressed genes (DEGs) were identified. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to categorize the functions of the DEGs and identify significantly enriched pathways. The transcript and protein levels of key enzymes involved in serine synthesis were verified in additional samples using quantitative real-time PCR (qRT-PCR) (n = 10) and capillary western blotting (n = 36). Targeted metabolomics of amino acids in CC extracts was performed by ultrahigh-performance liquid MS (UHPLC-MS/MS). The number of oocytes (2.4 ± 2.2 versus 12.1 ± 5.3) and metaphase II oocytes (2.1 ± 2.0 versus 9.9 ± 4.9) retrieved was significantly decreased in the DOR versus the NOR group, respectively (P < 0.0001). The rates of fertilization (80.7% versus 78.8%), viable embryos (73.7% versus 72.5%), and high-quality embryos (42.8% versus 49.0%) did not differ between the DOR and NOR groups, respectively (P > 0.05). A total of 95 DEGs were found by transcriptome sequencing. GO and KEGG analyses demonstrated that the DEGs were linked to amino acid metabolism and suggested significantly higher activity of the de novo SSP in the CCs of young women with DOR. Further qRT-PCR and capillary western blotting revealed that key enzymes (PHGDH, PSAT1, PSPH, and SHMT2) involved in de novo serine synthesis were upregulated, and UHPLC-MS/MS analysis showed increases in serine and glycine (a downstream product of serine) levels in the CCs of young patients with DOR. Our data clearly demonstrate that the de novo SSP, which diverts 3-phosphoglycerate from glycolysis to serine synthesis, was upregulated in young DOR CCs. N/A. Regarding the reproductive capacity of young patients DOR, the pregnancy outcomes were not analysed. The sample size was limited, and only women undergoing ICSI were examined since this was a prerequisite for the acquisition of CCs, which may cause selection bias. The exact mechanisms by which the SSP in CCs regulates ovarian reserve still require further study. Our research presents new evidence that alterations of the SSP in CCs of young infertile women are associated with DOR. We believe this is a significant contribution to the field, which should be key for understanding the cause and mechanisms of ovarian hypofunction in young women. This work was supported by grants from the Ministry of Science and Technology of China (2018YFC1005001) and National Natural Science Foundation of China (31601197). There were no competing interests. N/A.

Strigolactone-Mediated Trehalose Enhances Salt Resistance in Tomato Seedlings

Strigolactones (SLs) are newly discovered plant hormones that modulate a variety of physiological and biochemical processes and plant stress responses. In this study, SLs’ synthetic analog, GR24, significantly improved the growth of tomato seedlings under salt stress, while SLs’ synthesis inhibitor, TIS108, inversed the positive role of SLs, indicating that SLs could effectively enhance salt-stress resistance in tomato. To further explore the mechanism of SL-modulated trehalose (Tre) in response to salt stress, Tre metabolism was analyzed during this process. GR24 increased the endogenous Tre and starch contents and decreased the glucose (Glu) level under salt-stress conditions. Additionally, the TPS and TPP activities were enhanced by GR24 and the activity of THL was inhibited by GR24 under salt stress; thereafter, Tre biosynthesis-related genes, including TPS1, TPS2, TPP1, and TPP2, were also upregulated by GR24 under salt stress. However, the function of GR24 in Tre metabolism was inhibited by TIS108. Thus, the results indicated that GR24 improved the expression levels or activities of Tre biosynthesis-related genes or enzymes and inhibited the transcript level or activity of genes or enzymes related to Tre degradation, respectively, resulting in an increase in the endogenous Tre level and, therefore, weakening the salt toxicity of tomato seedlings.

Endophytic fungus Colletotrichum sp. AP12 promotes growth physiology and andrographolide biosynthesis in Andrographis paniculata (Burm. f.) Nees.

Endophytic fungi can promote host plant growth, enhance antioxidant defense enzyme activity, and induce the biosynthesis and accumulation of secondarymetabolites. Therefore, using endophytic fungi to improve the quality and yield of medicinal plants or important crops is an effective means of regulation. Colletotrichum sp. AP12 has been reported to produce andrographolide compounds (ADCs). This study aimed to investigate the effects of AP12 and its elicitors on the growth, defense enzyme activity, accumulation, and transcription levels of key genes in Andrographis paniculata (Burm. f.) Nees (A. paniculata). Using fermentation method to prepare AP12 into the inactivated fermentation solution (IFS), fermentation solution (FS), inactivated mycelium solution (IMS), and mycelium solution (MS), and the results showed that all four fungal elicitor components (ECs) could promote A. paniculata growth, enhance antioxidant defense enzymes, and increase ADC content and yield, especially the IMS group that had the highest leaf area, whole plant dry weight, superoxide dismutase (SOD), catalase (CAT) enzyme activities, total lactone contents, and yields, which were 2.37-, 1.60-, 2.20-, 3.27-, 1.59-, and 2.65-fold of the control, respectively. The 14-deoxyandrographolide (NAD) in the host irrigated with MS was 3.35-fold that of the control. In addition, AP12-infected A. paniculata sterile seedlings could significantly increase ADC content and expression levels of key enzyme genes, especially on day 12, when the total lactone content of the host reached 88.881± 5.793 mg/g DW, while on day 6, CPS gene expression level reached 10.79-fold that of the control, in turn promoting the biosynthesis and accumulation of andrographolide. In conclusion, the endophytic fungus AP12 is beneficial to the growth and secondary metabolism of A. paniculata, which is helpful for the cultivation and application of the biological bacterial fertilizer in A. paniculata, providing a theoretical and research basis for the use of endophytic fungi as a microbial resource to improve the quality and yield of medicinal plants.

Redox basis of photosynthesis inhibition at supra-optimal bicarbonate in mesophyll protoplasts of Arabidopsis thaliana

We examined the patterns of photosynthetic O2 evolution at 1 mM (optimal) and 10 mM (supra-optimal) bicarbonate in mesophyll protoplasts of Arabidopsis thaliana. The photosynthetic rate of protoplasts reached the maximum at an optimal concentration of 1 mM bicarbonate and got suppressed at supra-optimal levels of bicarbonate. We examined the basis of such photosynthesis inhibition by mesophyll protoplasts at supra-optimal bicarbonate. The wild-type protoplasts exposed to supra-optimal bicarbonate showed up signs of oxidative stress. Besides the wild-type, two mutants were used: nadp-mdh (deficient in chloroplastic NADP-MDH) and vtc1 (deficient in mitochondrial ascorbate biosynthesis). The protoplasts of the nadp-mdh mutant exhibited a higher photosynthetic rate and greater sensitivity to supra-optimal bicarbonate than the wild-type. The ascorbate-deficient vtc1 mutant had a low photosynthetic rate and no significant inhibition at high bicarbonate. The nadp-mdh mutants had elevated activities, protein, and transcript levels of key antioxidant enzymes. On the other hand, the antioxidant enzyme systems in vtc1 mutants were not much affected at supra-optimal bicarbonate. We propose that the inhibition of photosynthesis at supra-optimal bicarbonate depends on the redox state of mesophyll protoplasts. The robust antioxidant enzyme systems in protoplasts of nadp-mdh mutant might be priming the plants to sustain high photosynthesis at supra-optimal bicarbonate.

Regulation of Carotenoid Biosynthesis and Degradation in Lettuce (Lactuca sativa L.) from Seedlings to Harvest.

Lettuce (Lactuca sativa L.) is one of the commercially important leafy vegetables worldwide. However, lettuce cultivars vary widely in their carotenoid concentrations at the time of harvest. While the carotenoid content of lettuce can depend on transcript levels of key biosynthetic enzymes, genes that can act as biomarkers for carotenoid accumulation at early stages of plant growth have not been identified. Transcriptomic and metabolomic analysis was performed on the inner and outer leaves of the six cultivars at different developmental stages to identify gene-to-metabolite networks affecting the accumulation of two key carotenoids, β-carotene and lutein. Statistical analysis, including principal component analysis, was used to better understand variations in carotenoid concentration between leaf age and cultivars. Our results demonstrate that key enzymes of carotenoid biosynthesis pathway can alter lutein and β-carotene biosynthesis across commercial cultivars. To ensure high carotenoids content in leaves, the metabolites sink from β-carotene and lutein to zeaxanthin, and subsequently, abscisic acid needs to be regulated. Based on 2-3-fold carotenoids increase at 40 days after sowing (DAS) as compared to the seedling stage, and 1.5-2-fold decline at commercial stage (60 DAS) compared to the 40 DAS stage, we conclude that the value of lettuce for human nutrition would be improved by use of less mature plants, as the widely-used commercial stage is already at plant senescence stage where carotenoids and other essential metabolites are undergoing degradation.

Analysis on driving factors of microbial community succession in Jiuyao of Shaoxing Huangjiu (Chinese yellow rice wine)

The microbial ecosystem of fermented food is greatly disturbed by human activities.Jiuyao is important saccharification starter for brewing huangjiu. The interaction between environmental factors and microorganisms significantly affected the microbial community structure at different stages of Jiuyao manufacturing. This study combined environmental factor analysis and high-throughput sequencing technology to comprehensively analyze the specific changes of microbial community and environmental factors in each fermentation stage of Jiuyao production and their correlation. The results showed that the activities of liquefaction enzyme, glycosylation enzyme and acid protease reached the highest value on the 8 th day (192 h) after the beginning of fermentation, and the cellulase activity reached the highest value at the end of fermentation. Pediococcus（37.5 %-58.2 %）, Weissella（9.2 %-27.0 %） and Pelomonas（0.1 %-12.1 %） were the main microbial genera in the genus bacteria, and Saccharomycopsis（37.1 %-52.0 %）, Rhizopus（12.5 %-31.0 %） and Saccharomyces（4.0 %-20.5 %） were the main microbial genera in the genus fungi. The results of correlation analysis showed that the microbial communities in Jiuyao were closely related to environmental factors. Most microbial communities were positively correlated with temperature, but negatively correlated with ambient humidity, CO2 concentration, acidity and water content of Jiuyao. In addition, the transcription levels of enzymes related to microbial glucose metabolism in Jiuyao were higher in the late stage of Jiuyao fermentation. Interestingly, these enzymes had high transcription levels in fungi such as Saccharomycopsis, Rhizopus and Saccharomyces, as well as in bacteria such as Pediococcus and Lactobacillus. This study provides a reference for revealing the succession rule of microbial community structure caused by environmental factors during the preparation of Jiuyao in Shaoxing Huangjiu.