Bidirectional fermentation of Monascus and Mulberry leaves enhances GABA and pigment contents: establishment of strategy, studies of bioactivity and mechanistic.

Bidirectional fermentation is a technology that utilizes fungi to ferment medicinal edible substrates, with synergistic and complementary advantages. In this work, a fermentation strategy was established to produce a high yield of γ-aminobutyric acid (GABA) and Monascus pigments (MPs) using Monascus and mulberry leaves (MLs). Firstly, the basic fermentation parameters were determined using single-factor experiments, followed by Plackett-Burman (PB) experimental design to identify MLs, glucose, peptone, and temperature as significant influencing factors. The fermentation parameters were optimized using an artificial neural network (ANN). Finally, the effects of bidirectional fermentation of MLs and Monascus were investigated by bioactivity analysis, microstructure observation, and RT-qPCR. The outcomes showed that the bidirectional fermentation significantly increased the bioactive content and promoted the secondary metabolism of Monascus. The established fermentation conditions were 44.2 g/L of MLs, 57 g/L of glucose, 15 g/L of peptone, 1 g/L of MgSO4, 2 g/L of KH2PO4, 8% (v/v) of inoculum, 180 rpm, initial pH 6, 32 °C and 8 days. The content of GABA reached 13.95 g/L and the color value of MPs reached 408.07 U/mL. This study demonstrated the feasibility of bidirectional fermentation of MLs and Monascus, providing a new idea for the application of MLs and Monascus.

Combined volatile compounds and non-targeted metabolomics analysis reveals variation in flavour characteristics, metabolic profiles and bioactivity of mulberry leaves after Monascus purpureus fermentation.

BACKGROUND: Mulberry leaves (MLs) are widely used in food because of their nutritional and functional characteristics. However, plant cell walls and natural bitterness influence nutrient release and the flavor properties of MLs. Liquid-state fermentation using Monascus purpureus (LFMP) is a common processing method used to improve food properties. The present study used headspace solid-phase micro extraction gas chromatography-mass spectrometry (HS-SPME-GC-MS) and non-targeted metabolomics to examine changes in volatile and non-volatile metabolites in MLs. The transformation mechanism of LFMP was investigated by microscopic observation and dynamic analysis of enzyme activity, and changes in the biological activity of MLs were analyzed. RESULTS: LFMP significantly increased total phenolics, total flavonoids, free amino acids and soluble sugars in MLs, at the same time as decreasing phytic acid levels. In total, 92 volatile organic compounds (VOCs) were identified and quantified. VOCs such as (2R,3R)-(-)-2,3-butanediol, terpineol and eugenol showed some improvement in the flavour characteristics of MLs. By using non-targeted metabolomics, 124 unique metabolites in total were examined. LFMP altered the metabolic profile of MLs, mainly in plant secondary metabolism, lipid metabolism and amino acid metabolism. Microscopic observation and dynamic analysis of enzyme activity indicated that LFMP promoted cell wall degradation and biotransformation of MLs. In addition, LFMP significantly increased the angiotensin I-converting enzyme and α-glucosidase inhibitory activity of MLs. CONCLUSION: LFMP altered the flavour characteristics, metabolite profile and biological activity of MLs. These findings will provide ideas for the processing of MLs into functional foods. In addition, they also provide useful information for biochemical studies of fermented MLs. © 2023 Society of Chemical Industry.

Multi-frequency ultrasound-assisted cellulase extraction of protein from mulberry leaf: Kinetic, thermodynamic, and structural properties.

The effects of different extraction methods (traditional extraction, ultrasound extraction, cellulase extraction, and ultrasound-assisted cellulase extraction) on the yield of mulberry leaf protein (MLP) were investigated, and the results revealed that multi-frequency ultrasound-assisted cellulase extraction was the most efficient extraction method. The mechanism of the synergistic extraction method used to efficiently extract protein from mulberry leave was investigated, focusing on the kinetics and thermodynamics of the enzymatic process. The results revealed that kinetic parameters KM decreased by 14.07% and kA increased by 5.02%, and the thermodynamic parameters Ea, ΔH, and ΔS decreased by 44.81%, 48.41%, and 21.12 %, respectively, following the process of multi-frequency ultrasound (MFU) pretreatment. The spectral analysis with fluorescence spectra manifested that ultrasound exposed hydrophobic groups and induced molecular unfolding of MLP. Atomic force microscope showed that ultrasound decreased particle size while increasing flexibility of MLP. The effect of ultrasound increases the binding frequency of cellulase and substrates, resulting in greater affinity between the two and promoting the solubilization of MLP. This study provides a theoretical basis to improve the application prospects of MLP.

Characteristics of enzymolysis of silkworm pupa protein after tri-frequency ultrasonic pretreatment: kinetics, thermodynamics, structure and antioxidant changes.

As a by-product of the sericulture industry, the utilization rate of silkworm pupa resources is currently not high. Proteins are converted into bioactive peptides through enzymatic hydrolysis. Not only can it solve the utilization problem, but it also creates more valuable nutritional additives. Silkworm pupa protein (SPP) was pretreated with tri-frequency ultrasonic (22/28/40 kHz). Effects of ultrasonic pretreatment on enzymolysis kinetics, enzymolysis thermodynamics, hydrolysate structure as well as hydrolysate antioxidant of SPP were investigated. Ultrasonic pretreatment significantly increased the hydrolysis efficiency, showing a 6.369% decrease in k m and a 16.746% increase in k A after ultrasonic action (p < 0.05). The SPP enzymolysis reaction followed a second-order rate kinetics model. Evaluation of enzymolysis thermodynamics revealed that Ultrasonic pretreatment markedly enhanced the SPP enzymolysis, leading to a 21.943% decrease in E a. Besides, Ultrasonic pretreatment significantly increased SPP hydrolysate's surface hydrophobicity, thermal stability, crystallinity, and antioxidant activities (DPPH radical scavenging activity, Fe2+ chelation ability, and reducing power). This study indicated that tri-frequency ultrasonic pretreatment could be an efficient approach to enhancing the enzymolysis and improving the functional properties of SPP. Therefore, tri-frequency ultrasound technology can be applied industrially to enhance enzyme reaction process.

Silk Fibroin-Induced Gadolinium-Functionalized Gold Nanoparticles for MR/CT Dual-Modal Imaging-Guided Photothermal Therapy.

The development of multifunction nanoplatforms integrating accurate diagnosis and efficient therapy is of great significance for the precise treatment of tumors. Gold nanoparticles (AuNPs) possessing hallmark features of computed tomography (CT) imaging and photothermal conversion capability hold great potential in tumor theranostics. In this study, taking the advantages of outstanding biocompatibility, interesting anti-inflammatory and immunomodulatory properties, and abundant amino acid residues of silk fibroin (SF), a multifunctional Gd-hybridized AuNP nanoplatform was constructed using SF as a stabilizer and reductant via a facile one-pot biomimetic method, denoted as Gd:AuNPs@SF. The obtained Gd:AuNPs@SF possessed fascinating biocompatibility and excellent photothermal conversion efficiency. Functionalized with Gd, Gd:AuNPs@SF exhibited super tumor-contrasted imaging performance in magnetic resonance (MR) and CT imaging modalities. Moreover, Gd:AuNPs@SF, with strong NIR absorbance, demonstrated that it could effectively kill tumor cells in vitro, and was also proved to successfully ablate tumor tissues through MR/CT imaging-guided photothermal therapy (PTT) without systemic toxicity in Pan02 xenograft C57BL/6 mouse models. We successfully synthesized Gd:AuNPs@SF for MR/CT dual-mode imaging-guided PTT via a facile one-pot biomimetic method, and this biomimetic strategy can also be used for the construction of other multifunction nanoplatforms, which is promising for precise tumor theranostics.

Functional and postoperative outcomes after high-intensity interval training in lung cancer patients: A systematic review and meta-analysis.

Objective: The study evaluated the effects of high-intensity interval training (HIIT) on postoperative complications and lung function in patients with lung cancer compared to usual care. Methods: We searched electronic databases in April 2022, including PubMed, Embase, the Cochrane Library, Web of Science, and the China National Knowledge Infrastructure (CNKI). Two authors independently applied the Cochrane Risk of Bias tool to assess the quality of RCTs. The postoperative complications, length of hospitalization, and cardiopulmonary functions from the studies were pooled for statistical analysis. Results: A total of 12 randomized controlled trials were eligible for inclusion and were conducted in the meta-analysis. HIIT significantly increased VO2peak (MD = 2.65; 95% CI = 1.70 to 3.60; I2 = 40%; P <0.001) and FEV1 (MD = 0.12; 95% CI = 0.04 to 0.20; I2 = 51%; P = 0.003) compared with usual care. A subgroup analysis of studies that applied HIIT perioperatively showed significant improvement of HIIT on FEV1 (MD = 0.14; 95% CI = 0.08 to 0.20; I2 = 36%; P <0.0001). HIIT significantly reduced the incidence of postoperative atelectasis in lung cancer patients compared with usual care (RD = -0.16; 95% CI = -0.24 to -0.08; I2 = 24%; P <0.0001). There was no statistically significant effect of HIIT on postoperative arrhythmias (RD = -0.05; 95% CI = -0.13 to 0.03; I2 = 40%; P = 0.22), length of hospitalization (MD = -1.64; 95% CI = -3.29 to 0.01; P = 0.05), and the six-minute walk test (MD = 19.77; 95% CI = -15.25 to 54.80; P = 0.27) compared to usual care. Conclusion: HIIT may enhance VO2peak and FEV1 in lung cancer patients and reduce the incidence of postoperative atelectasis. However, HIIT may not reduce the incidence of postoperative arrhythmia, shorten the length of hospitalization, or improve the exercise performance of patients with lung cancer. Systematic review registration: PROSPERO, CRD42022335441.

Solid state fermentation by Fomitopsis pinicola improves physicochemical and functional properties of wheat bran and the bran-containing products.

Wheat bran was solid state fermented by Fomitopsis pinicola. The results showed that the processing properties were increased by fermentation and the content of total phenol and alkylresorcinols was 5.91 and 1.55 times of the unfermented bran respectively by the 6th day. The total antioxidant capacity was 5.73 times of the unfermented sample by the 4th day. Electronic nose analysis showed that the fermented wheat bran had a special flavor. GC-MS analysis found that 4-ethyl-2-methoxy-phenol was the main flavor substance, which was sharply increased during the fermentation. Furthermore, the textural properties of the dough and bread containing fermented bran were significantly improved. The content of phytic acid in the bread was significantly decreased, while the protein, total phenol and alkylresorcinols contents were significantly increased. Results suggest that solid state fermentation by Fomitopsis pinicola is a promising way to improve wheat bran to a nutritious and flavorful cereal food ingredient.

Antioxidant and Hemolysis Protective Effects of Polyphenol-Rich Extract from Mulberry Fruits.

BACKGROUND: Mulberry fruits are a superior source of polyphenol, especially anthocyanins that contribute potentially to the beneficial effects which include reducing the risk of cardiovascular diseases and cancers with antioxidant, anti-inflammatory, and chemoprotective properties. OBJECTIVES: In this study, purification of the polyphenol-rich extract from mulberry fruit (MPE) was purified and assessed the activities of antioxidant and hemolysis protective in vivo and in vitro. MATERIALS AND METHODS: Antioxidant activities in vitro was measured by quantifying its 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity, reducing power and Fe2+-chelating ability. MPE was purified by high-pressure liquid chromatography (HPLC) and analyzed individual polyphenols using liquid chromatography-mass spectrometry (LC-MS)/MS. RESULTS: The total polyphenol content was 147.69 ± 0.02 mg gallic acid equivalents (GAE)/g dried weight (DW) in the extract and 403.55 ± 0.02 mg GAE/g DW in the purified extract. Further identification by HPLC-ultraviolet-visible and LC-MS/MS analysis indicated in MPE, an anthocyanin compound, cyanidin-3-O-glucoside. With regard to in vitro assays, MPE possessed antioxidant effect, especially in Fe2+ chelating ability with an IC50 value of 1.016 mg/mL. The protective effects on mouse red blood cell hemolysis and lipid peroxidation ex vivo were dose and time dependent. CONCLUSION: It indicates that MPE could be a good candidate for future biomedical applications to promote human health with limited side effects. SUMMARY: Mulberry fruit is an excellent source of polyphenols, in particular, anthocyanins, which has infinite health benefits. This study determined the predominant anthocyanin, cyanidin-3-glucoside, could possibly be the rationale behind the antioxidant and antihemolytic effect of MPE. Results indicate that MPE could be a good candidate for future biomedical applications to promote human health with limited side effects. Abbreviations used: MPE: Purification of the polyphenol-rich extract from mulberry fruit, LC-MS: Liquid chromatography-mass spectrometry, HPLC: High-pressure liquid chromatography, DPPH: 2,2-diphenyl-1-picrylhydrazyl scavenging activity, RBC: Red blood cell, GAE: Gallic acid equivalent, FeCl2: Ferrous chloride, H2O2: Hydrogen peroxide, EDTA-2Na: Ethylenediaminetetraacetic acid disodium salt, PBS: Phosphate-buffered saline, TCA: Trichloroacetic acid, TBA: 2-thiobarbituric acid, FeSO4: Ferrous sulphate, MDA: Malondialdehyde, VC: Vitamin C, DW: Dried weight.

Production of ACE inhibitory peptides from sweet sorghum grain protein using alcalase: Hydrolysis kinetic, purification and molecular docking study.

In this study, sweet sorghum grain protein (SSGP) was hydrolyzed using alcalase yielding ACE inhibitory peptides. A kinetic model was proposed to describe the enzymolysis process of SSGP. The kinetic parameters, a and b, were determined according to experimental data. It was found that the model was reliable to describe the kinetic behaviour for SSGP hydrolysis by alcalase. After hydrolysis, the SSGP hydrolysate with DH of 19% exhibited the strongest ACE inhibitory activity and the hydrolysate was then used to isolate ACE inhibitory peptides. A novel ACE inhibitory peptide was successfully purified from this hydrolysate by ultrafiltration, ion exchange chromatography, gel filtration chromatography, and reversed-phased high performance liquid chromatography (RP-HPLC). The amino acid sequence of the purified peptide was identified as Thr-Leu-Ser (IC50=102.1 µM). The molecular docking studies revealed that the ACE inhibition of the tripeptide was mainly attributed to its C-terminal Ser, which can effectively interact with the S1 and S2 pockets of ACE. Our studies suggest that the tripeptide from the SSGP hydrolysate can be utilized to develop functional food ingredients or pharmaceuticals for prevention of hypertension.

Characterization, antioxidant and antitumor activities of polysaccharides from purple sweet potato.

Three polysaccharides, PSPP1-1, PSPP2-1 and PSPP3-1, were isolated from purple sweet potato. The three polysaccharides belonged to ß-type polysaccharides and contained low proportions of proteins and uronic acids. PSPP1-1 and PSPP3-1 with molecular weights of 33.3 and 75.3 kDa, respectively, were composed of rhamnose, xylose, glucose and galactose, whereas PSPP2-1 with molecular weight of 17.8 kDa was composed of rhamnose and galactose. The three polysaccharides possessed in vitro antioxidant (scavenging DPPH radicals, chelating ferrous ions and reducing power) and antitumor activities (against SGC7901 and SW620 cells) in a dose-dependent manner. Among the three polysaccharides, PSPP2-1 exhibited the strongest reducing power, scavenging activity on DPPH radicals and chelating capability on ferrous ions. PSPP1-1 showed the strongest inhibitory activities on the growth of SGC7901 and SW620 cells. In addition, flow cytometry results showed that PSPP1-1 could induce apoptosis in SGC7901 and SW620 cells. These results suggest that polysaccharides from purple sweet potato are potential natural antioxidant and antitumor agents that can be used as drugs or functional food ingredients.

[Responses of ecosystem carbon budget to increasing nitrogen deposition in differently degraded Leymus chinensis steppes in Inner Mongolia, China].

Based on a field manipulative nitrogen (N) addition experiment, the effects of atmospheric N deposition level change on the plant biomass and net primary productivity (NPP), soil respiration (Rs) and net ecosystem exchange (NEE) were investigated respectively in 2009 and 2010 in two differently degraded Leymus chinensis steppes in Inner Mongolia of China, and the difference in the response of NEE to equal amount of N addition [10 g x (M2 x a)(-1), MN] between the two steppes was also discussed. The results indicated that for the light degraded Leymus chinensis steppe (site A) , the average plant aboveground biomass (AGB) in MN treatment were 21.5% and 46.8% higher than those of CK in these two years. But for the moderate degraded Leymus chinensis steppe (site B), the N addition decreased the plant AGB and ANPP in 2009, while showed positive effects in 2010. N addition increased the belowground biomass (BGB) of the both sites and belowground NPP (BNPP) of site B in both years, but decreased the BNPP of site A in 2010. The increase of N input in the two steppes did not change the seasonal variation of Rs. The cumulative annual soil C emissions in MN treatment in site A showed an increase of about 14.6% and 25.7% of those in the CK respectively for these two years, while were decreased by about 10.4% and 11.3%, respectively in site B. The NEE of MN treatments, expressed by C, for the two steppes were 59.22 g x (m2 x a)(1) and 166.68 g x (m2 x a)(-1), as well as 83.27 g x (m2 x a)(-1) and 117.47 g x (m2 x a)(-1), respectively in these two years. The increments in NEE originated from N addition for these two years were 15.79 g x (M2 x a)(-1) and 82.94 g x (M2 x a)(-1) in site A and 74.54 g x (M2 x a)(-1) and 101.23 g x (M2 x a)(-1) in site B. The N input per unit could obtain greater C sink effect in the steppe with lower initial N level.

A novel angiotensin-І converting enzyme (ACE) inhibitory peptide from gastrointestinal protease hydrolysate of silkworm pupa (Bombyx mori) protein: Biochemical characterization and molecular docking study.

Silkworm pupa (Bombyx mori) protein was hydrolyzed using gastrointestinal endopeptidases (pepsin, trypsin and α-chymotrypsin). Then, the hydrolysate was purified sequentially by ultrafiltration, gel filtration chromatography and RP-HPLC. A novel ACE inhibitory peptide, Ala-Ser-Leu, with the IC50 value of 102.15µM, was identified by IT-MS/MS. This is the first report of Ala-Ser-Leu from natural protein. Lineweaver-Burk plots suggest that the peptide is a competitive inhibitor against ACE. The molecular docking studies revealed that the ACE inhibition of Ala-Ser-Leu is mainly attributed to forming very strong hydrogen bonds with the S1 pocket (Ala354) and the S2 pocket (Gln281 and His353). The results indicate that silkworm pupa (B. mori) protein or its gastrointestinal protease hydrolysate could be used as a functional ingredient in auxiliary therapeutic foods against hypertension.

[Reponses of soil total organic carbon and dissolved organic carbon to simulated nitrogen deposition in temperate typical steppe in Inner Mongolia, China].

Based on a field manipulative nitrogen (N) addition experiment, the effects of atmospheric N deposition level change on the contents, inter-annual variation and profile distribution of soil total organic carbon (TOC) and dissolved organic carbon (DOC) were investigated from May, 2008 to October, 2011 in a temperate typical steppe in Inner Mongolia of China, and the relationship between TOC and DOC was also discussed. The treatments in the manipulative experiment included N additions at rates of 0, 5, 10, and 20 g x (m2 x a)(-1), representing the control (CK), low N (LN), medium N (MN), and high N (HN) treatment, respectively. The results indicated that the concentrations of soil TOC and DOC decreased progressively with soil depth in all cases except for the DOC at 10-20 cm depth in individual years. The increase of N input in typical steppe did not change the vertical distribution of soil TOC and DOC, but reduced the vertical variation of TOC and increased the vertical variation of DOC in the surface soil horizon. In addition, the contents of soil TOC and DOC at 0- 10 cm and 10- 20 cm soil layers changed insignificantly after the continuous increase in anthropogenic N input for four years. The soil organic C density of 0-20 cm soil layer for different N treatment levels varied between 3.9 kg x m(-2) and 5.6 kg x m(-2), and the soil organic C densities of fertilized treatments in the first two years were similar to or slightly lower than those of CK, while in the following two years, the increase in N deposition gradually played a positive role in increasing soil organic C density, but the differences in soil TOC and DOC contents between CK and fertilized plots were not significant (P > 0.05). The ratio of soil DOC to TOC (DOC/TOC) varied from 0.32% to 1.09%. The increase in N deposition generally lowered the proportion of DOC in soil TOC, which was conducive to the accumulation of soil organic C. The change of soil DOC was positively correlated with that of TOC (P < 0.01). The temporal variations of soil DOC in different N treatments were all far greater than those of TOC, and the soil DOC was the important sensitive indicator for predicting and evaluating the response of soil C pool to the change in atmospheric N deposition in the temperate grassland ecosystem.

Improved 1-Deoxynojirimycin (DNJ) production in mulberry leaves fermented by microorganism.

DNJ, an inhibitor of α-glucosidase, is used to suppress the elevation of postprandial hyperglycemia. In this study, we focus on screening an appropriate microorganism for performing fermentation to improve DNJ content in mulberry leaf. Results showed that Ganoderma lucidum was selected from 8 species and shown to be the most effective in improvement of DNJ production from mulberry leaves through fermentation. Based on single factor and three factor influence level tests by following the Plackett-Burman design, the optimum extraction yield was analyzed by response surface methodology (RSM). The extracted DNJ was determined by reverse-phase high performance liquid chromatograph equipped with fluorescence detector (HPLC-FD). The results of RSM showed that the optimal condition for mulberry fermentation was defined as pH 6.97, potassium nitrate content 0.81% and inoculums volume 2 mL. The extraction efficiency reached to 0.548% in maximum which is 2.74 fold of those in mulberry leaf.

Differential responses of short-term soil respiration dynamics to the experimental addition of nitrogen and water in the temperate semi-arid steppe of Inner Mongolia, China.

We examined the effects of simulated rainfall and increasing N supply of different levels on CO2 pulse emission from typical Inner Mongolian steppe soil using the static opaque chamber technique, respectively in a dry June and a rainy August. The treatments included NH4NO3 additions at rates of 0, 5, 10, and 20 g N/(m(2)·year) with or without water. Immediately after the experimental simulated rainfall events, the CO2 effluxes in the watering plots without N addition (WCK) increased greatly and reached the maximum value at 2 hr. However, the efflux level reverted to the background level within 48 hr. The cumulative CO2 effluxes in the soil rang ed from 5.60 to 6.49 g C/m(2) over 48 hr after a single water application, thus showing an increase of approximately 148.64% and 48.36% in the effluxes during both observation periods. By contrast, the addition of different N levels without water addition did not result in a significant change in soil respiration in the short term. Two-way ANOVA showed that the effects of the interaction between water and N addition were insignificant in short-term soil CO2 effluxes in the soil. The cumulative soil CO2 fluxes of different treatments over 48 hr accounted for approximately 5.34% to 6.91% and 2.36% to 2.93% of annual C emission in both experimental periods. These results stress the need for improving the sampling frequency after rainfall in future studies to ensure more accurate evaluation of the grassland C emission contribution.

Improved 1-Deoxynojirimycin (DNJ) production in mulberry leaves fermented by microorganism

DNJ, an inhibitor of α-glucosidase, is used to suppress the elevation of postprandial hyperglycemia. In this study, we focus on screening an appropriate microorganism for performing fermentation to improve DNJ content in mulberry leaf. Results showed that Ganoderma lucidum was selected from 8 species and shown to be the most effective in improvement of DNJ production from mulberry leaves through fermentation. Based on single factor and three factor influence level tests by following the Plackett-Burman design, the optimum extraction yield was analyzed by response surface methodology (RSM). The extracted DNJ was determined by reverse-phase high performance liquid chromatograph equipped with fluorescence detector (HPLC-FD). The results of RSM showed that the optimal condition for mulberry fermentation was defined as pH 6.97, potassium nitrate content 0.81% and inoculums volume 2 mL. The extraction efficiency reached to 0.548% in maximum which is 2.74 fold of those in mulberry leaf.

Core promoter regulates the expression of cathepsin B gene in the fat body of Bombyx mori.

Bombyx mori cathepsin B (BmCatB) is involved in the programmed cell death of the fat body during B. mori metamorphosis. For a better understanding of the functional regulatory mechanism, the promoter region of BmCatB in the transcriptional regulation has been identified and analyzed in the present study. BmCatB promoter region performed by the 5' truncation or mutagenesis of EcREs was inserted in the pFA3Luc-A3RL double fluorescence expression vector to activate the fireflies luciferase (FLuc) gene. The results indicated that the dual-luciferase activity of BmCatB gene in the silkworm larval fat body is regulated by the length of promoter. Site-directed mutagenesis of EcRE experiment has shown that the EcREs are up-regulated significantly in the regulation of the BmCatB promoter. A 142bp region (-1165 to -1023) and EcREs are the mainly fat-body tissue-specificity related region and could function as a core promoter element.

Identification and functional analysis of the cathepsin D gene promoter of Bombyx mori.

The gene encoding cathepsin D of silkworm, Bombyx mori (BmCatD) is specifically expressed in the larval fat body and pupal gut, and plays an important role in the programmed cell death during metamorphosis. To identify element involved in this transcription-dependent spatial restriction, truncation and deletion of the 5' terminal from the BmCatD promoter were conducted in vivo. The recombinant AcMNPV vector (Autographa californica multiple nucleopolyhedrovirus) with a dual-luciferase quantitative assay system was used as the transfer. A 289 bp DNA sequence (-1,214 to -925) upstream of the transcriptional start site is found to be responsible for promoting tissue-specific transcription. Further analysis of a series of deletion within the 289 bp region of overlapping deletion showed that a 33 bp region (-1,071 to -1,038) sequence suppresses the ectopic expression of the BmCatD promoter. These results suggest that this 33 bp region could function as a promoter element in the tissue-specificity expression.

[Microbial response mechanism for drying and rewetting effect on soil respiration in grassland ecosystem: a review].

As one of the most important and wide distribution community type among terrestrial ecosystems, grassland ecosystem plays a critical role in the global carbon cycles and climate regulation. China has extremely rich grassland resources, which have a huge carbon sequestration potential and are an important part of the global carbon cycle. Drying and rewetting is a common natural phenomenon in soil, which might accelerate soil carbon mineralization process, increase soil respiration and exert profound influence on microbial activity and community structure. Under the background of the global change, the changes in rainfall capacity, strength and frequency would inevitably affect soil drying and wetting cycles, and thus change the microbial activity and community structure as well as soil respiration, and then exert important influence on global carbon budget. In this paper, related references in recent ten years were reviewed. The source of soil released, the trend of soil respiration over time and the relationship between soil respiration and microbial biomass, microbial activity and microbial community structure during the processes of dry-rewetting cycle were analyzed and summarized, in order to better understand the microbial response mechanism for drying and rewetting effecting on soil respiration in grassland ecosystem, and provide a certain theoretical basis for more accurate evaluation and prediction of future global carbon balance of terrestrial ecosystems and climate change.

Identification of ecdysone response elements (EcREs) in the Bombyx mori cathepsin D promoter.

Bombyx mori Cathepsin D (BmCatD) is specifically expressed in the fat body, and plays a critical role for the programmed cell death of the larval fat body and pupal gut during metamorphosis. To better understand the transcriptional control of BmCatD expression, we conducted this study to identify the ecdysone response elements (EcREs) in the BmCatD promoter and clarify their regulational functions. We inserted EcREs into a recombinant AcMNPV (Autographa californica multiple nucleopolyhedrovirus) vector and performed luciferase assay with a dual-luciferase quantitative assay system. Three putative EcREs were located at positions -109 to -99, -836 to -826 and -856 to -846 relative to the transcription start site. Overlapping deletion studies of this EcRE region showed that the three EcREs could suppress the ectopic expression of the BmCatD promoter. EcRE mutations resulted in the loss of the fat body-specific expression of the BmCatD gene. These results suggest that the EcREs are vital for activation of the promoter by 20-hydroxyecdysone (20E) in the larval fat body and further support the crucial role of ecdysone signaling to control cathepsin D gene transcription. It may suggest that the heterodimeric complex EcR/USP mediates the activation of ecdysone-dependent BmCatD transcription in the larval fat body of B. mori.