Enhanced extracellular production of alpha-lactalbumin from Bacillus subtilis through signal peptide and promoter screening

Alpha-lactalbumin(α-LA)is a major whey protein found in breast milk and plays a crucial role in the growth and development of infants.In this study,Bacillus subtilis RIK1285 harboring AprE signal peptide(SP)was selected as the original strain for the production ofα-LA.It was found thatα-LA was identified in the pellet after ultrasonic disruption and centrifugation instead of in the fermentation supernatant.The original strain most likely only producedα-LA intracellular,but not extracellular.To improve the expression and secretion ofα-LA in RIK1285,a library of 173 homologous SPs from the B.subtilis 168 genome was fused with target LALBA gene in the pBE-S vector and expressed extracellularly in RIK1285.SP YjcN was determined to be the best signal peptide.Bands in supernatant were observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and purified by nickel column to calculate the highest yield signal peptide.In addition,different promoters(P_(aprE),P_(43),and P_(glv))were compared and applied.The results indicated that the strain RIK1285-pBE-P_(glv)-YjcN-LALBA had the highestα-LA yield,reaching 122.04μg/mL.This study demonstrates successful expression and secretion of humanα-LA in B.subtilis and establishes a foundation for simulating breast milk for infant formulas and developing bioengineered milk.

Understanding the catalytic performance and deactivation behaviour of second-promoter doped Pt/WO_(χ)/γ-Al_(2)O_(3) catalysts in the glycerol hydrogenolysis for selective and cleaner production of 1,3-propanediol

The selective aqueous-phase glycerol hydrogenolysis is a promising reaction to produce commercially useful 1,3-propanediol(1,3-PDO).The Pt-WOx bifunctional catalyst can catalyse the glycerol hydrogenol-ysis but the catalyst deactivation via sintering,metal leaching,and coking can predominantly occur in the aqueous phase reaction.In this work,the effect of reaction temperature,pressure and second promoter(Cu,Fe,Rh,Mn,Re,Ru,Ir,Sn,B,and P)on catalytic performance and deactivation behaviour of Pt/WOx/-Al2O3 was investigated.When doped with Rh,Mn,Re,Ru,Ir,B,and P,the second promoter boosts catalytic activity by promoting great dispersion of Pt on support and increasing Pt surface area.The increased Bronsted acid sites lead to selective synthesis of 1,3-PDO than 1,2-propanediol(1,2-PDO).The characterization studies of fresh and spent catalysts reveal that the main cause of catalyst deactivation is the Pt sintering,as interpreted based on XRD,CO chemisorption,and TEM analyses.The Pt sintering is affected depending on the second promoter that can either or reduce the interaction between Pt,WO_(χ)/γ and Al_(2)O_(3).As an electron acceptor of Pt in Pt/WO_(χ)/γ-Al_(2)O_(3),Re and Mn as second promoters resulted in increased Pt^(2+) on the catalytic surface,which strengthens the contact between Pt andγ-Al_(2)O_(3) and WO_(χ),resulting in a decrease in Pt sintering.The metal leaching and coking are not affected by the presence of second promoter.The catalyst modified with a second promoter possesses improved catalytic activity and 1,3-PDO production,however the stability continues to remain a challenge.The present work unrav-elled the determining parameters of catalytic activity and deactivation,thus providing a promising pro-tocol toward effective catalysts for glycerol hydrogenolysis.

Molecular cloning,characterization and promoter analysis of LbgCWIN1 and its expression profiles in response to exogenous sucrose during in vitro bulblet initiation in lily

Lily(Lilium spp.) is an important ornamental flower, which is mainly propagated by bulbs. Cell wall invertases(CWINs), which catalyze the irreversibly conversion of sucrose into glucose and fructose in the extracellular space, are key enzymes participating in sucrose allocation in higher plants. Previous studies have shown that CWINs play an essential role in bulblet initiation process in bulbous crops, but the underlying molecular mechanism remains unclear. Here, a CWIN gene of Lilium brownii var. giganteum(Lbg) was identified and amplified from genomic DNA. Quantitative RT-PCR assays revealed that the expression level of LbgCWIN1 was highly upregulated exactly when the endogenous starch degraded in non-sucrose medium during in vitro bulblet initiation in Lbg. Phylogenetic relationship, motif, and domain analysis of LbgCWIN1 protein and CWINs in other plant species showed that all sequences of these CWIN proteins were highly conserved. The promoter sequence of LbgCWIN1 possessed a number of alpha-amylase-, phytohormone-, light-and stress-responsive cis-elements. Meanwhile, β-glucuronidase(GUS) assay showed that the 459 bp upstream fragment from the translational start site displayed maximal promoter activity. These results revealed that LbgCWIN1 might function in the process of in vitro bulblet initiation and be in the response to degradation of endogenous starch.

Lineage-Specific Genome Architecture Links Enhancers and Non-coding Disease Variants to Target Gene Promoters

Long-range interactions between regulatory elements and gene promoters play key roles in transcriptional regulation.The vast majority of interactions are uncharted,constituting a major missing link in understanding genome control.Here,we use promoter capture Hi-C to identify interacting regions of 31,253 promoters in 17 human primary hematopoietic cell types.We show that promoter interactions are highly cell type specific and enriched for links between active promoters and epigenetically marked enhancers.Promoter interactomes reflect lineage relationships of the hematopoietic tree,consistent with dynamic remodeling of nuclear architecture during differentiation.Interacting regions are enriched in genetic variants linked with altered expression of genes they contact,highlighting their functional role.We exploit this rich resource to connect non-coding disease variants to putative target promoters,prioritizing thousands of disease-candidate genes and implicating disease pathways.Our results demonstrate the power of primary cell promoter interactomes to reveal insights into genomic regulatory mechanisms underlying common diseases.

Cloning of a flower-specific expression promoter from Arabidopsis thaliana and its plant expression vector construction

Given the sequence of Chs gene promoter from Arabidopsis thaliana reported in GenBank （AF248988）, a pair of specific PCR primers was designed with the Primer Premier 5.0 software. PCR products of about 0.5 kb were successfully amplified with the genome DNA of A. thaliana as a DNA template and Taq polymerase as DNA polymerase. The purified PCR products were ligated to the pMD18-T vector. The sequencing result showed that the Chs promoter from A. thaliana was 531 bp long. Sequence alignment analysis based on the DNAMAN software revealed that the sequence similarity between the cloned promoter and target promoter （AF248988） was up to 100%. Online PLACE analysis indicated that the Chs promoter contained cis-elements such as TATA-box, CAAT-box, pollen-box, G-box, ACGT-containing element, R response element, Myb recognition element and TACPyAT-box. At the same time, a plant expression vectorpAtChs：：GUS which fused the Chs promoter and the marker gene GUS was successfully constructed.

Promoter and coding sequence diversity of CsCCD1 may contribute to the differential accumulation of floralβ-ionone in fresh tea leaves

The carotenoid-derived volatileβ-ionone makes an important contribution to tea fragrance.Here,we qualitatively and quantitatively analysed 15 carotenoids in tea leaves of 13 cultivars by UHPLC-APCI-MS/MS.The 13 cultivars were divided into two groups by PCA(Principal Component Analysis)clustering analysis of their carotenoid content,and OPLS-DA(Orthogonal projections to latent structures)indicated that the levels ofβ-carotene(VIP=2.89)and lutein(VIP=2.30)were responsible for much of the variation between the two groups.Interestingly,theβ-carotene toβ-ionone conversion rates in Group 1 were higher than in Group 2,while theβ-carotene content was significantly lower in Group 1 than in Group 2.Theβ-ionone content was significantly higher in Group 1.Pearson Correlation Coefficient calculation between the transcription level of candidate genes(CsCCD1 and CsCCD4)and the accumulation ofβ-ionone indicated that CsCCD1 may involve in the formation ofβ-ionone in 13 cultivars.Prokaryotic expression and in vitro enzyme activity assays showed that‘Chuanhuang 1’had an amino acid mutation in carotenoid cleavage dioxygenases 1(CsCCD1)compared with‘Shuchazao’,resulting in a significantly higherβ-ionone content in‘Chuanhuang 1’.Sequence analysis showed that‘Chuanhuang 1’and‘Huangdan’had different CsCCD1 promoter sequences,leading to significantly higher CsCCD1 expression andβ-ionone accumulation in‘Chuanhuang 1’.These results indicated that the promoter and coding sequence diversity of CsCCD1 might contribute to the differential accumulation ofβ-ionone in different tea cultivars.

OsAMT1.1 Expression by Nitrate-Inducible Promoter of OsNAR2.1 Increases Nitrogen Use Efficiency and Rice Yield

Nitrate(NO_(3)^(-))and ammonium(NH_(4)^(+))are two main inorganic nitrogen(N)sources during crop growth.Here,we enhanced the expression of OsAMT1.1,which encodes a NH_(4)^(+)transporter,using the NO_(3)^(-)-inducible promoter of OsNAR2.1 and an ubiquitin promoter in transgenic rice plants.Under field condition of 120 kg/hm2 N,agronomic N use efficiency,N recovery efficiency and N transport efficiency,and grain yield of the pOsNAR2.1:OsAMT1.1 transgenic lines were increased compared with those of the wild type(WT)and the pUbi:OsAMT1.1 transgenic plants.Under 2.0 mmol/L NO_(3)^(-)+0.5 mmol/L NH_(4)^(+)and 0.5 mmol/L NO_(3)^(-)+2.0 mmol/L NH_(4)^(+)conditions of hydroponic culture,compared with the WT,both biomass and total N content were increased in the pOsNAR2.1:OsAMT1.1 transgenic lines.However,biomass was significantly reduced in pUbi:OsAMT1.1 transgenic plants under 0.5 mmol/L NO_(3)^(-)+2.0 mmol/L NH_(4)^(+)condition.The lines expressing pOsNAR2.1:OsAMT1.1 exhibited increased OsAMT1.1 expression and 15NH_(4)^(+)influx in roots under both 2.0 mmol/L NO_(3)^(-)+0.5 mmol/L NH_(4)^(+)and 0.5 mmol/L NO_(3)^(-)+2.0 mmol/L NH_(4)^(+)conditions.Our study showed that expression of OsAMT1.1 can be promoted when driven by the OsNAR2.1 promoter,especially under high-level nitrate condition,leading to enhancement of NH_(4)^(+)uptake,N use efficiency and grain yield.

Methyltransferase 3A-mediated promoter methylation represses retinoic acid receptor responder 3 expression in basal-like breast cancer

Retinoic acid receptor responder 3(RARRES3)has been characterized as a tumor suppressor in multiple types of cancer.This study aimed to examine the expression profile of RARRES3 across the PAM50 subtypes of breast cancer.The DNA methylation status of RARRES3 was checked in the basal-like subtype,and the underlying mechanisms of its dysregulation were explored.RNA-sequencing(seq)and methylation data from The Cancer Genome Atlas were used for in-silico analysis.Basal-like representative SUM149 and MDA-MB-468 cell lines were used for in vitro and in vivo studies.Compared to tumor-adjacent normal tissues,only the basal-like tumor tissues had significantly downregulated RARRES3 expression.The methylation level of four CpG sites in the promoter region showed a strong negative correlation with RARRES3 expression.The gene coding for DNA methyltransferase 3A(DNMT3A)had consistent positive correlations with the methylation of the CpG sites.Chromatin Immunoprecipitation-quantitative polymerase chain-reaction and bisulfite sequencing PCR showed that DNMT3A could bind to the promoter region of RARRES3 and promote methylation of the CpG sites within the region.DNMT3A knockdown significantly restored RARRES3 expression at the mRNA and protein level in the two cell lines.CCK-8,colony formation,and flow cytometric analysis showed that RARRES3 overexpression attenuated the growth-promoting effects of DNMT3A overexpression and also weakened the DNMT3A overexpression-induced activation of ERK1/2 and PI3K/AKT signaling.In summary,this study revealed that DNMT3A enhances promoter methylation of the RARRES3 gene and suppresses its transcription in basal-like breast cancer.The DNMT3A-RARRES3 signaling pathway might be a potential target for the treatment of this tumor subtype.

Theoretical study on the morphology of cobalt nanoparticles modulated by alkali metal promoters

Cobalt nanoparticles(NPs)catalysts are extensively used in heterogeneous catalytic reactions,and the addition of alkali metal promoters is a common method to modulate the catalytic performance because the catalyst's surface structures and morphologies are sensitive to the addition of promoters.However,the underlying modulation trend remains unclear.Herein,the adsorption of alkali metal promoters(Na and K)on the surfaces of face-centered-cubic(FCC)and hexagonal-closest packed(HCP)polymorphous cobalt was systematically investigated using density functional theory.Furthermore,the effect of alkali promoters on surface energies and nanoparticle morphologies was revealed on the basis of Wulff theory.For FCC-Co,the exposed area of the(111)facet in the nanoparticle increases with the adsorption coverage of alkali metal oxide.Meanwhile,the(311),(110),and(100)facets would disappear under the higher adsorption coverage of alkali metals.For HCPCo,the Wulff morphology is dominated by the(0001)and(1011)facets and is independent of the alkali metal adsorption coverage.This work provides insights into morphology modulation by alkali metal promoters for the rational design and synthesis of cobalt-based nanomaterials with desired facets and morphologies.

Measurement,associations,patterns,and promotion:Unveiling vital areas in physical activity epidemiology

Physical activity(PA)epidemiology first emerged as a new field of study around the 1950s with publication of seminal investigations on London civil servants1 and has continued to evolve as a scientific discipline to present time.This subdiscipline of epidemiology studies the frequencies,distributions,and dynamics of PA and its impact on human health,morbidity,and mortality.Research over the past 75 years has demonstrated numerous health benefits of PA.Based on these research findings,the World Health Organization recommends adults aged 18-64 years engage in at least 150-300 min of moderate-intensity aerobic PA per week,or at least 75-150 min of vigorous-intensity aerobic PA per week,or a combination of both intensities.

Shaping global health promotion:a comprehensive analysis of the 10 Global Conferences on Health Promotion Conferences(1986-2021)

Objective This study analyzed how the 10 Global Conferences on Health Promotion have played a significant role in shaping and promoting a worldwide consensus and actions on health promotion,effectively addressing diverse health challenges that evolved over different periods.Methods The textual analysis method was used in this study and text encoding was conducted to systematically examine the declarations and reports presented by the 10 Global Conferences on Health Promotion held during 1986-2021.We summarized the themes and key achievements,and key vocabulary in the conference declarations was extracted and analyzed to construct the global health promotion consensus and actions.Results The fundamental principles of the conferences are to foster consensus and initiate actions in the realm of health promotion on a global scale.The primary purpose and goal are to promote health from regional to global.Significantly,our findings highlight a transition in the primary actors driving health promotion.It underscores a shift in health promotion from being driven primarily by organizations like the World Health Organization,governments,and international bodies,to a more inclusive approach involving non-governmental organizations and the general public.This development implies that health promotion has evolved into a collective global endeavor,demanding the proactive involvement of various stakeholders,and forging new alliances in public health.Meanwhile,the coronavirus disease 2019(COVID-19)pandemic has further shaped the landscape of health promotion,underscoring the need for intensified focus on areas including disease prevention,health education,and the integration of digital health technologies,and emphasizing the importance of a multidimensional,responsive approach in public health initiatives.Conclusions Sustained collaboration and innovative strategies are pivotal to advancing health promotion globally.Countries,together with public and private entities,should intensify cooperation.Multisectoral collaboration among partners such as healthcare,education,social security,and the industry is vital for health promotion and achieving global health goals.

Research Progress on the Growth-Promoting Effect of Plant Biostimulants on Crops

A Plant Biostimulant is any substance or microorganism applied to plants to enhance nutrition efficiency,abioticstress tolerance,and/or crop quality traits,regardless of its nutrient content.The application of Plant biostimulants(PBs)in production can reduce the application of traditional pesticides and chemical fertilizers and improvethe quality and yield of crops,which is conducive to the sustainable development of agriculture.An in-depthunderstanding of the mechanism and effect of various PBs is very important for how to apply PBs reasonablyand effectively in the practice of crop production.This paper summarizes the main classification of PBs;Thegrowth promotion mechanism of PBs was analyzed from four aspects:improving soil physical and chemical properties,enhancing crop nutrient absorption capacity,photosynthesis capacity,and abiotic stress tolerance;At thesame time,the effects of PBs application on seed germination,seedling vigor,crop yield,and quality were summarized;Finally,how to continue to explore and study the use and mechanism of PBs in the future is analyzedand prospected,to better guide the application of PBs in crop production in the future.

Spatiotemporal variations,influencing factors,and configurational promotion paths of energy efficiency in China’s cities

The promotion of energy efficiency(EE)helps address energy constraints and promote environmental sustainability.This study comprehensively explores the spatiotemporal variations,influencing factors,and configuration promotion paths of EE in 284 Chinese cities during 2003‒2019 using the global super-efficiency minimum distance to strong efficient frontier(G-S-MinDS),exploratory spatial data analysis(ESDA),multiscale geographically weighted regression(MGWR),and fuzzy set qualitative comparative analysis(fsQCA)methods.The findings are:①China’s cities have an annual average EE of 0.658 with a growth rate of 0.53%,showing considerable promotion potential.②Industrial structure optimization,population agglomeration,economic development,and increased green coverage contribute positively,while government intervention and openness hinder China’s urban EE.③Four configurational promotion paths for enhancing China’s urban EE are identified,where among those paths population density is a core condition,while government intervention is not.This study provides valuable insights into substantially improving urban EE,emphasizing the need for targeted policies to address energy and environmental crises in China.

Promoting or Inhibiting?Influence of Railway Container Transportation on Regional Economic Development

In the context of building a country with a strong transportation network,railway container transportation(RCT)is an import-ant means of reducing costs,increasing efficiency,and adjusting transportation structures.Thus,its impact on regional economic devel-opment is important.Based on data from railway container-handling stations and spatial econometric models,this study discusses the differences in the development of RCT and their impact on regional economic development at different leves.This study has three main findings:first,there are significant regional differences in the development of the RCT.The intra-regional differences between the east-ern and central regions of China(which do not include Hong Kong,Macao and Taiwan)are gradually narrowing,while the regional dif-ferences in the western region are widening.Meanwhile,the intra-regional differences in important economic zones such as Pearl River Delta Economic Zone(PRDEZ),Chengdu-Chongqing Economic Zone(CYEZ),Bohai Rim Economic Zone(BHEZ),and Yangtze River Delta Economic Zone(YRDEZ)are narrowing daily.Second,the development differences of RCT in regional level and import-ant economic regions level show different trends.The unbalanced features of large regions are increasingly evident,whereas the differ-ences in economic regions are decreasing.However,the problem of overlapping RCT remains prominent.Third,the transformation of RCT development mode and fierce competition among transportation modes cause RCT to have a restraining effect on the regional eco-nomy at three levels.Rational allocation of resources and other means must be used to guide the transformation from inhibition to pro-motion,and by formulating targeted policies that will promote the development of RCT,which will improve the transportation structure and help construct a country with a strong transportation system.

The Promotion of China Tourism Day Held at the Bangladesh National Museum

On May 19,the China Tourism Day promotion co-sponsored by the Chinese Embassy in Bangladesh,the Bangladesh National Museum and the Bangladesh Travel Agents Association was successfully held at the Bangladesh National Museum.

Pueraria isoflavones inhibit XOD and GLUT9 to decrease uric acid production and promote uric acid excretion, respectively

Objective: To analyze the possible mechanism of Pueraria isoflavones inhibiting XOD and GLUT9 to reduce uric acid production and promote uric acid excretion. Methods: August 2021-April 2022, a total of forty SPF male Kunming mice were divided into the healthy group (carboxymethylcellulose sodium at a dose of 250 mg/kg), the model group (HUA mice were given carboxymethylcellulose sodium at a dose of 250 mg/kg), the low group (HUA mice were given pueraria isoflavone at a dose of 125 mg/kg), HUA mice were given pueraria isoflavones at a dose of 250 mg/kg once d frequency)and the high group (HUA mice were given pueraria isoflavones at a dose of 500 mg/kg once d frequency) dosage groups, with 8 mice in each group. The contents of uric acid (SUA), urea nitrogen (BUN) and creatinine (SCr) in serum and urine of each group were compared before and after intervention (30 d). Statistical differences of xanthine oxidase (XOD) and human glucose transporter 9(GLUT9), cy- clooxygenase- 2(COX-2), tumor necrosis factor (TNF-α) and interleukin-1 (IL-1β) contents in renal tissues of each group after intervention (30 d) were compared. Results: After intervention, kidney inflammatory factors (COX-2, TNF-α and IL-1β) in the model group were compared. Blood and urine indexes (SUA, BUN, SCr);The contents of XOD and GLUT9 were higher than those of healthy group(P<0.05). Renal inflammatory cytokines (COX-2, TNF-α and IL-1β) in low, medium and high dose groups;Blood and urine indexes (SUA, BUN, SCr);The contents of XOD and GLUT9 were lower than those of model group, and there were low > medium > high dose groups, the comparison between the two groups had statistical significance(P< 0.05). After intervention, the contents of 3 indicators in blood or urine(COX-2, TNF-α and IL-1β) all decreased compared with before intervention, and the differences in intra-group comparison were statistically significant (P<0.05). Conclusion: Pueraria isoflavones can treat HUA mice by inhibiting the expression of XOD and GLUT9, and then play a role in reducing uric acid pro- duction and promoting uric acid excretion, as well as alleviating the degree of disease inflammation.

From Inside Out:Internal Marketing as an Engine for Service Promotion-A Secondary Publication

Focusing on“from inside out”means understanding that the driving force behind a successful service promotion campaign lies in a deep connection with the internal team,turning it into the engine that propels a positive brand perception.The University of Panama faces challenges in the effective implementation of strategies involving its internal staff,impacting the coherence and strength of its promotional messages.The research aimed to determine internal marketing strategies as an engine for service promotion at the Central Campus and CRUSAM.The applied study was descriptive,non-experimental field research,and cross-sectional.The sample consisted of 80 individuals using a quota sampling method to measure and quantify the expression of a construct or variable that is not directly observable.A selfadministered questionnaire was used to collect information,measuring variables in their various dimensions.Content validity of the questionnaire was ensured through the evaluation of three experts,and the reliability of the Likert-type instrument was assessed using Cronbach’s α coefficient,which was found to be very high(0.918).The results led to the conclusion that establishing internal marketing strategies is necessary for the transformation and innovation process in the university through service promotion.

Study on the Promoting Role of Animating Zhuang Non-Heritage Folktales in the Ethnic Culture Industry

This study aims to explore the role of animating Zhuang non-heritage folktales in promoting the national culture industry.Using literature research,case studies,and comparative research methods,the positive impacts are analyzed in depth across four dimensions:cultural value inheritance,market demand,dissemination channels,and industrial innovation.The results show that animating Zhuang non-heritage folktales can help inherit national culture,meet market demand,broaden communication channels,and promote industrial innovation.Finally,this paper proposes corresponding policy suggestions and implementation paths to provide references for the sustainable development of the ethnic culture industry.

Study on the Promotion of Digital Trade to Northeast Asian Regional Economy

Society is currently transitioning from the era of informatization and digitization to the era of intelligence.Advanced technologies such as artificial intelligence,the Internet of Things,and big data are permeating every aspect of production and life.This penetration presents a rare opportunity for the development of digital trade and catalyzes innovation in traditional trade patterns.This paper first analyzes the inevitability of digital trade development in Northeast Asia.Secondly,it delves into the challenges faced by digital trade development and explores in-depth the promotion strategy for digital trade’s impact on the economic development of the Northeast Asia region,providing insights from various perspectives for reference.

A Multimodal Critical Discourse Analysis of Lingnan Cultural Promotional Videos on Social Media

In recent years,more and more directors of culture and tourism have taken part in the promotion of local cultural tourism by cross-dressing,talent shows,and pushing their limits on self-media platforms.This study investigates short videos of Lingnan culture promoted by directors general and deputy directors general of the Culture,Radio,Television,Tourism,and Sports Bureau of counties and cities in Guangdong Province on social media by the method of multimodal critical discourse analysis.The analysis of 33 videos shows that Lingnan culture is a domineering and confident culture,historical culture,graceful and elegant culture,and vibrant and active culture.Domineering and confident culture is embedded in the utterances and behaviors of the directors general or deputy directors general in the video.Historical culture is realized through the conversation with historical figures through time travel.Graceful and elegant culture is constructed in the depiction of sceneries and the depiction of characters’manners.Vibrant and active culture is represented in the depiction of the characters’actional process and analytical process.