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1.
Front Psychol ; 15: 1412596, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38860044

RESUMEN

Objectives: This study explores the significant impact of expressive ties (EI) between DanceSport couples on their competitive performance (CP). Utilizing a dyadic approach, we examined the performance achievement processes of DanceSport couples in relation to their EI. Methods: Participants comprised 67 dyads of Chinese elite dancers aged between 16 and 30 years. The dyadic analysis was carried out using a structural equation model based on the actor-partner interdependence mediation model. Results: With regard to actor effects, both male (ß = 0.292, p = 0.012) and female (ß = 0.443, p < 0.001) dancers' perceived quality of EI had a positive correlation with CP. The males' athlete engagement (AE) partially mediated the impact of EI on CP [indirect effect = 0.144, SE = 0.072, 95% confidence intervals (CI) = 0.020, 0.283]. Regarding partner effects, females' perceived EI quality positively influenced the male's CP (ß = 0.26, p = 0.023) and mediated this association through the male's AE [indirect effect = 0.086, SE = 0.041, 95% confidence intervals (CI) = 0.003, 0.149]. Similarly, the females' AE mediated the effect of males' perceived EI quality on the females' CP [indirect effect = 0.152, SE = 0.074, 95% confidence intervals (CI) =0.002, 0.256]. Conclusion: We not only validated the propositions of the self-determination theory but also provided valuable insights to further enrich it. Our findings underscore that self-determination theory must account for individual gender characteristics.

2.
Psychol Res Behav Manag ; 17: 2135-2147, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38826679

RESUMEN

Purpose: During the period of COVID-19 pandemic, the social restrictions and isolation exerted a significant impact on the sleep quality of Chinese college students. This study aims to delve into the influence of physical activity on the sleep quality of college students as well as the mediating roles of stress and smartphone addiction. Materials and Methods: A cohort of 274 eligible college students (146 males and 128 females) were selected for the investigation. The International Physical Activity Questionnaire Short Form, Stress Perception Scale, Smartphone Addiction Scale, and Pittsburgh Sleep Quality Index were employed to assess the levels of physical activity, stress, smartphone addiction, and sleep quality among college students. For data analysis, descriptive statistics, correlation analysis, and chained mediation effect tests were performed sequentially. Results: The findings revealed: (1) a significant negative correlation between physical activity and stress, smartphone addiction, and sleep quality among college students (r = -0.216, p < 0.001; r = -0.224, p < 0.001; r = -0.259, p < 0.001); (2) independent mediating roles of stress and smartphone addiction in the relationship between physical activity and sleep quality; and (3) chained mediating effects of stress and smartphone addiction in the association between physical activity and sleep quality. Conclusion: This study deepens our comprehension of how physical activity augments the quality of slumber, concurrently emphasizing that mitigating stress levels and alleviating smartphone addiction constitute effective strategies for preventing sleep issues among college students.

3.
Sci Rep ; 14(1): 11321, 2024 05 17.
Artículo en Inglés | MEDLINE | ID: mdl-38760471

RESUMEN

This study aimed to analyze anthropometric and physiological profiles of highly trained sailors and the differences between sailors regarding various training levels. Forty-two sailors (22 male, 22.4 ± 3.8 years; 20 females, 21.3 ± 3.6 years) were divided into helmsmen and crew groups, and the high- and low-level were distinguished. Sailors completed height, sitting height, legs length, weight, BMI, VO2max, 30 s all-out sprint, isometric mid-thigh pull (IMTP), countermovement jump, bench pull, core endurance tests. The results showed the crew had higher height, sitting height, weight, VO2max and lower trunk flexor endurance test times compared to the helmsmen (p < 0.05). The helmsmen had higher relative peak power/force in the 30 s all-out sprint and IMTP tests compared to the crew, whereas the crew had better absolute strength in bench pull, with significant differences between female sailors (p < 0.05). The high-level sailors showed more sailing experience than low-level sailors (p < 0.05). In conclusion, highly trained crew tend to be taller and heavier, while helmsmen have better trunk flexor endurance. For female sailors, helmsmen have better lower-body power and strength and crew have better upper-body strength. Sailing experience is a reliable variable to distinguish sailors' levels. The specific anthropometric and physiological profiles of sailors in various positions can assist sailing coaches in athlete selection and intervention training.


Asunto(s)
Antropometría , Resistencia Física , Humanos , Femenino , Masculino , Adulto , Resistencia Física/fisiología , Adulto Joven , Deportes Acuáticos/fisiología , Fuerza Muscular/fisiología , Navíos , Consumo de Oxígeno/fisiología , Personal Militar , Peso Corporal
4.
Front Physiol ; 15: 1321519, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38500539

RESUMEN

Objective: To assess the effects of 8 weeks of unilateral (UNI), bilateral (BI), and combined unilateral + bilateral (UNI + BI) resistance training on bench press and squat strength in adolescent boxers. Methods: Using the Gym Aware linear accelerometer, free-weight squat and bench press strength exercises were evaluated after an 8-week training intervention. Thirty adolescent boxers were randomly assigned to three groups: UNI, height: 1.73 ± 0.08 m, weight: 55.42 ± 5.85 kg; UNI + BI, height: 1.7 ± 0.06 m, weight: 54.73 ± 5.33 kg; and BI, height: 1.74 ± 0.06 m, weight: 59.67 ± 8.39 kg. Each group followed their designated UNI/BI/UNI + BI compound resistance training protocols, and the effects of 8 weeks of single-sided and bilateral intervention training on the performance of free-weight squat and bench press exercises at 30%, 50%, and 80% of 1-repetition maximum (1RM) were evaluated. Results: Significant improvements were observed in the 30% 1RM, 50% 1RM, and 80% 1RM outcomes for both squat and bench press exercises before and after the interventions (p < 0.05, p < 0.01). In the intergroup comparison, GymAware measurements revealed that the UNI and UNI + BI groups exhibited superior peak power values for squat and bench press exercises at 30% 1RM compared to the BI group. Discussion: UNI and UNI + BI training led to significantly higher output power values in bench press and squat exercises at 30% 1RM compared to the BI training group.

5.
Res Microbiol ; : 104188, 2024 Jan 28.
Artículo en Inglés | MEDLINE | ID: mdl-38286394

RESUMEN

The complete genome of Corynebacterium glutamicum contain a gene encoding murein endopeptidase MepA which maintain cell wall homeostasis by regulating peptidoglycan biosynthesis. In this study, we investigate the physiological function, localization and regulator of MepA. The result shows that mepA overexpression lead to peptidoglycan degradation and the defects in cell division. MepA-mCherry was shown to localizes exclusively at the cell cell septum. In addition, mepA overexpression increased cell permeability and reduced the resistance of cells to isoniazid, an antibiotic used to treat Mycobacterium tuberculosis infection. Furthermore, transcription analysis showed that mepA affected cell division and membrane transport pathways, and was coordinately regulated by the two-component systems MtrAB and MprAB(CgtS/R2).

6.
Biotechnol J ; 19(1): e2300187, 2024 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-38178735

RESUMEN

The ApxII toxin and the outer membrane lipoprotein (Oml) of Actinobacillus pleuropneumoniae are important vaccine antigens against porcine contagious pleuropneumonia (PCP), a prevalent infectious disease affecting the swine industry worldwide. Previous studies have reported the recombinant expression of ApxII and Oml in Escherichia coli; however, their yields were not satisfactory. Here, we aimed to enhance the production of ApxII and Oml by constructing a bicistronic expression system based on the widely used T7 promoter. To create efficient T7 bicistronic expression cassettes, 16 different fore-cistron sequences were introduced downstream of the T7 promoter. The expression of three vaccine antigens Oml1, Oml7, and ApxII in the four strongest bicistronic vectors were enhanced compared to the monocistronic control. Further optimization of the fermentation conditions in micro-well plates (MWP) led to improved production. Finally, the production yields reached unprecedented levels of 2.43 g L-1 of Oml1, 2.59 g L-1 of Oml7, and 1.21 g L-1 of ApxII, in a 5 L bioreactor. These three antigens also demonstrated well-protective immunity against A. pleuropneumoniae infection. In conclusion, this study establishes an efficient bicistronic T7 expression system that can be used to express recombinant proteins in E. coli and achieves the hyper-production of PCP vaccine proteins.


Asunto(s)
Infecciones por Actinobacillus , Pleuroneumonía Contagiosa , Porcinos , Animales , Proteínas Bacterianas , Escherichia coli/genética , Pleuroneumonía Contagiosa/prevención & control , Proteínas Recombinantes/genética , Infecciones por Actinobacillus/prevención & control , Vacunas de Subunidad/genética
7.
Cancer Biol Ther ; 24(1): 2279241, 2023 12 31.
Artículo en Inglés | MEDLINE | ID: mdl-38031910

RESUMEN

Cyclin-dependent Kinase 2 (CDK2) inhibition prevents supernumerary centrosome clustering. This causes multipolarity, anaphase catastrophe and apoptotic death of aneuploid cancers. This study elucidated how CDK2 antagonism affected centrosome stoichiometry. Focused ion beam scanning electron microscopy (FIB-SEM) and immunofluorescent imaging were used. Studies interrogated multipolar mitosis after pharmacologic or genetic repression of CDK2. CDK2/9 antagonism with CYC065 (Fadraciclib)-treatment disordered centrosome stoichiometry in aneuploid cancer cells, preventing centrosome clustering. This caused ring-like chromosomes or multipolar cancer cells to form before onset of cell death. Intriguingly, CDK2 inhibition caused a statistically significant increase in single centrioles rather than intact centrosomes with two centrioles in cancer cells having chromosome rings or multipolarity. Statistically significant alterations in centrosome stoichiometry were undetected in other mitotic cancer cells. To confirm this pharmacodynamic effect, CDK2 but not CDK9 siRNA-mediated knockdown augmented cancer cells with chromosome ring or multipolarity formation. Notably, engineered gain of CDK2, but not CDK9 expression, reversed emergence of cancer cells with chromosome rings or multipolarity, despite CYC065-treatment. In marked contrast, CDK2 inhibition of primary human alveolar epithelial cells did not confer statistically significant increases of cells with ring-like chromosomes or multipolarity. Hence, CDK2 antagonism caused differential effects in malignant versus normal alveolar epithelial cells. Translational relevance was confirmed by CYC065-treatment of syngeneic lung cancers in mice. Mitotic figures in tumors exhibited chromosome rings or multipolarity. Thus, CDK2 inhibition preferentially disorders centrosome stoichiometry in cancer cells. Engaging this disruption is a strategy to explore against aneuploid cancers in future clinical trials.


Asunto(s)
Centrosoma , Neoplasias , Humanos , Animales , Ratones , Quinasa 2 Dependiente de la Ciclina/genética , Quinasa 2 Dependiente de la Ciclina/metabolismo , Centrosoma/metabolismo , Anafase , Mitosis/genética , Aneuploidia , Neoplasias/genética , Neoplasias/metabolismo
8.
Front Psychol ; 14: 1278874, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-38034303

RESUMEN

Objectives: Although the positive association of partnership between DanceSport couples with competitive performance (CP) is documented, less is known about the mediating factors of this relationship. According to the related literature and self-determination theory (SDT), the present study finds and verifies that athlete engagement (AE) mediates the association between partnership and competitive performance. Methods: A total of 242 Chinese sports dancers were recruited using the purposive sampling method. The Partnership Scale-DanceSport Couples (PS-DSC), the Athlete Engagement Questionnaire (AEQ), and the Competitive Performance Questionnaire (CPQ) were adopted to collect data. Results: The obligatory instrumental ties, expressive ties, and interpersonal perception scores are all positively correlated with both athlete engagement and competitive performance, and athlete engagement scores are positively correlated with competitive performance. Athlete engagement completely mediates the association between obligatory instrumental ties and competitive performance, and it partially mediates the association between expressive ties, interpersonal perception, and competitive performance, with the mediating effect accounting for 25.29 and 24.40% of the total effect, respectively. Conclusion: Athlete engagement mediates the association between DanceSport couples' partnership and competitive performance. High levels of athlete engagement are needed to improve the chance of promoting obligatory instrumental ties, expressive ties, and interpersonal perception between DanceSport couples toward excellent competitive performance. Overall, the results represent an attempt to extend our understanding of the mechanisms by which the three partnership stereotype factors individually influence dancers' cognitive and psychological states.

9.
Chembiochem ; 24(24): e202300587, 2023 12 14.
Artículo en Inglés | MEDLINE | ID: mdl-37783667

RESUMEN

Most natural formate dehydrogenases (FDHs) exhibit NAD+ specificity, making it imperative to explore the engineering of FDH cofactor specificity for NADPH regeneration systems. The endogenous FDH of Komagataella phaffii (K. phaffii), termed KphFDH, is a typical NAD+ -specific FDH. However, investigations into engineering the cofactor specificity of KphFDH have yet to be conducted. To develop an NADP+ -specific variant of KphFDH, we selected D195, Y196, and Q197 as mutation sites and generated twenty site-directed variants. Through kinetic characterization, KphFDH/V19 (D195Q/Y196R/Q197H) was identified as the variant with the highest specificity towards NADP+ , with a ratio of catalytic efficiency (kcat /KM )NADP+ /(kcat /KM )NAD+ of 129.226. Studies of enzymatic properties revealed that the optimal temperature and pH for the reduction reaction of NADP+ catalyzed by KphFDH/V19 were 45 °C and 7.5, respectively. The molecular dynamics (MD) simulation was performed to elucidate the mechanism of high catalytic activity of KphFDH/V19 towards NADP+ . Finally, KphFDH/V19 was applied to an in vitro NADPH regeneration system with Meso-diaminopimelate dehydrogenase from Symbiobacterium thermophilum (StDAPDH/H227V). This study successfully created a KphFDH variant with high NADP+ specificity and demonstrated its practical applicability in an in vitro NADPH regeneration system.


Asunto(s)
NAD , Saccharomycetales , NADP/metabolismo , NAD/metabolismo , Formiato Deshidrogenasas/química , Saccharomycetales/metabolismo , Cinética
10.
Sci Rep ; 13(1): 14907, 2023 09 09.
Artículo en Inglés | MEDLINE | ID: mdl-37689790

RESUMEN

All-trans-retinoic acid (ATRA), the retinoic acid receptors (RARs) agonist, regulates cell growth, differentiation, immunity, and survival. We report that ATRA-treatment repressed cancer growth in syngeneic immunocompetent, but not immunodeficient mice. The tumor microenvironment was implicated: CD8+ T cell depletion antagonized ATRA's anti-tumorigenic effects in syngeneic mice. ATRA-treatment with checkpoint blockade did not cooperatively inhibit murine lung cancer growth. To augment ATRA's anti-tumorigenicity without promoting its pro-tumorigenic potential, an RARγ agonist (IRX4647) was used since it regulates T cell biology. Treating with IRX4647 in combination with an immune checkpoint (anti-PD-L1) inhibitor resulted in a statistically significant suppression of syngeneic 344SQ lung cancers in mice-a model known for its resistance to checkpoints and characterized by low basal T cell and PD-L1 expression. This combined treatment notably elevated CD4+ T-cell presence within the tumor microenvironment and increased IL-5 and IL-13 tumor levels, while simultaneously decreasing CD38 in the tumor stroma. IL-5 and/or IL-13 treatments increased CD4+ more than CD8+ T-cells in mice. IRX4647-treatment did not appreciably affect in vitro lung cancer growth, despite RARγ expression. Pharmacokinetic analysis found IRX4647 plasma half-life was 6 h in mice. Yet, RARα antagonist (IRX6696)-treatment with anti-PD-L1 did not repress syngeneic lung cancer growth. Together, these findings provide a rationale for a clinical trial investigating an RARγ agonist to augment check point blockade response in cancers.


Asunto(s)
Linfocitos T CD8-positivos , Neoplasias Pulmonares , Animales , Ratones , Interleucina-13 , Interleucina-5 , Microambiente Tumoral , Receptores de Ácido Retinoico , Neoplasias Pulmonares/tratamiento farmacológico , Tretinoina , Carcinogénesis
11.
Microb Cell Fact ; 22(1): 182, 2023 Sep 15.
Artículo en Inglés | MEDLINE | ID: mdl-37715258

RESUMEN

In the post-genomic era, the demand for faster and more efficient protein production has increased, both in public laboratories and industry. In addition, with the expansion of protein sequences in databases, the range of possible enzymes of interest for a given application is also increasing. Faced with peer competition, budgetary, and time constraints, companies and laboratories must find ways to develop a robust manufacturing process for recombinant protein production. In this review, we explore high-throughput technologies for recombinant protein expression and present a holistic high-throughput process development strategy that spans from genes to proteins. We discuss the challenges that come with this task, the limitations of previous studies, and future research directions.


Asunto(s)
Genómica , Laboratorios , Clonación Molecular , Secuencia de Aminoácidos , Proteínas Recombinantes/genética
12.
Reprod Domest Anim ; 58(11): 1559-1568, 2023 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-37712626

RESUMEN

Peroxisome proliferator-activated receptor γ (PPARγ) is highly expressed in trophoblast tissues in pregnancy during which the protein participates in diverse events, including embryo implantation and placental formation. However, little is known about the role of PPARγ in embryonic development. This study investigated the function of PPARγ in sheep trophoblast cells. The coding sequence of sheep PPARγ encoded 475 amino acids and included one synonymou mutation compared with the sheep reference sequence for PPARγ. The PPARγ protein was localized in the nucleus and cytoplasm of sheep trophoblasts. The relative expression of PPARγ was elevated in cells treated with rosiglitazone and reduced following administration of GW9662. Activation of PPARγ promoted cell proliferation and mobility, but inhibited apoptosis. In addition, stimulation of PPARγ promoted the expression of lipid metabolism-related genes FABP4 and PLIN2. The expression of prostaglandin metabolism-related genes PLA2G4A, PTGS2 and PTGES also was upregulated significantly in trophoblast cells when PPARγ was activated. In contrast, activation of PPARγ did not impact expression of the prostaglandin-related genes PGFS and SLCO2A1. At the same time, activation of PPARγ activity increased the ratio of PGE2 to PGF2α. Furthermore, fluorescence labelling showed that the numbers of cell lipid droplets increased after stimulation of PPARγ activity, but decreased when PPARγ was inhibited. In conclusion, PPARγ is critical for the regulation of lipid metabolism and prostaglandin synthesis and secretion in sheep trophoblast cells and also has a potent effect on cell proliferation and viability.


Asunto(s)
PPAR gamma , Trofoblastos , Embarazo , Femenino , Animales , Ovinos , PPAR gamma/genética , PPAR gamma/metabolismo , Placenta/metabolismo , Metabolismo de los Lípidos , Prostaglandinas
13.
Oncogene ; 42(44): 3260-3273, 2023 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-37740007

RESUMEN

Pancreatic cancer cells undergo intricate metabolic reprogramming to sustain their survival and proliferation. p53 exhibits a dual role in tumor cell ferroptosis. However, the precise role and mechanisms underlying wild-type p53 activation in promoting ferroptosis in pancreatic cancer cells remain obscure. In this study, we applied bioinformatics tools and performed an analysis of clinical tissue sample databases and observed a significantly upregulated expression of solute carrier family 35 member F2 (SLC35F2) in pancreatic cancer tissues. Our clinical investigations indicated that elevated SLC35F expression was related to adverse survival outcomes. Through multi-omics analyses, we discerned that SLC35F2 influences the transcriptome and inhibits ferroptosis in pancreatic cancer cells. Moreover, our findings reveal the pivotal involvement of p53 in mediating SLC35F2-mediated ferroptosis, both in vitro and in vivo. SLC35F2 inhibits ferroptosis by facilitating TRIM59-mediated p53 degradation. Further mechanistic investigations demonstrated that SLC35F2 competitively interacts with the E3 ubiquitin ligase SYVN1 of TRIM59, thereby stabilizing TRIM59 expression and consequentially promoting p53 degradation. Utilizing protein 3D structure analysis and drug screening, we identified irinotecan hydrochloride and lapatinib ditosylate as compounds targeting SLC35F2, augmenting the antitumor effect of imidazole ketone erastin (IKE) in a wild-type p53 patient-derived xenograft (PDX) model. However, in the p53 mutant PDX model, irinotecan hydrochloride and lapatinib ditosylate did not alter the sensitivity of the tumor xenograft model to IKE-triggered ferroptosis. In summary, our work establishes a novel mechanism wherein the SLC35F2-SYVN1-TRIM59 axis critically regulates ferroptosis of pancreatic cancer cells by inhibiting endogenous p53. Thus, SLC35F2 emerges as a promising therapeutic target for treating pancreatic cancer.


Asunto(s)
Ferroptosis , Neoplasias Pancreáticas , Humanos , Ferroptosis/genética , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Irinotecán/farmacología , Lapatinib/farmacología , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/genética , Línea Celular Tumoral , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas de Motivos Tripartitos/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Neoplasias Pancreáticas
14.
ACS Synth Biol ; 12(8): 2226-2235, 2023 08 18.
Artículo en Inglés | MEDLINE | ID: mdl-37463503

RESUMEN

Saccharomyces cerevisiae has been extensively used as a convenient synthetic biology chassis to reconstitute fungal polyketide biosynthetic pathways. Despite progress in refactoring these pathways for expression and optimization of the yeast production host by metabolic engineering, product yields often remain unsatisfactory. Such problems are especially acute when synthetic biological production is used for bioprospecting via genome mining or when chimeric fungal polyketide synthases (PKSs) are employed to produce novel bioactive compounds. In this work, we demonstrate that empirically balancing the expression levels of the two collaborating PKS subunits that afford benzenediol lactone (BDL)-type fungal polyketides is a facile strategy to improve the product yields. This is accomplished by systematically and independently altering the copy numbers of the two plasmids that express these PKS subunits. We applied this plasmid copy number engineering strategy to two orphan PKSs from genome mining where the yields of the presumed BDL products in S. cerevisiae were far too low for product isolation. This optimization resulted in product yield improvements of up to 10-fold, allowing for the successful isolation and structure elucidation of new BDL analogues. Heterocombinations of these PKS subunits from genome mining with those from previously identified BDL pathways led to the combinatorial biosynthesis of several additional novel BDL-type polyketides.


Asunto(s)
Policétidos , Policétidos/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Variaciones en el Número de Copia de ADN , Sintasas Poliquetidas/metabolismo , Lactonas/metabolismo , Plásmidos/genética
15.
Biotechnol Bioeng ; 120(10): 3039-3056, 2023 10.
Artículo en Inglés | MEDLINE | ID: mdl-37309999

RESUMEN

ß-elemene is one of the most commonly used antineoplastic drugs in cancer treatment. As a plant-derived natural chemical, biologically engineering microorganisms to produce germacrene A to be converted to ß-elemene harbors great expectations since chemical synthesis and plant isolation methods come with their production deficiencies. In this study, we report the design of an Escherichia coli cell factory for the de novo production of germacrene A to be converted to ß-elemene from a simple carbon source. A series of systematic approaches of engineering the isoprenoid and central carbon pathways, translational and protein engineering of the sesquiterpene synthase, and exporter engineering yielded high-efficient ß-elemene production. Specifically, deleting competing pathways in the central carbon pathway ensured the availability of acetyl-coA, pyruvate, and glyceraldehyde-3-phosphate for the isoprenoid pathways. Adopting lycopene color as a high throughput screening method, an optimized NSY305N was obtained via error-prone polymerase chain reaction mutagenesis. Further overexpression of key pathway enzymes, exporter genes, and translational engineering produced 1161.09 mg/L of ß-elemene in a shake flask. Finally, we detected the highest reported titer of 3.52 g/L of ß-elemene and 2.13 g/L germacrene A produced by an E. coli cell factory in a 4-L fed-batch fermentation. The systematic engineering reported here generally applies to microbial production of a broader range of chemicals. This illustrates that rewiring E. coli central metabolism is viable for producing acetyl-coA-derived and pyruvate-derived molecules cost-effectively.


Asunto(s)
Escherichia coli , Sesquiterpenos , Escherichia coli/genética , Escherichia coli/metabolismo , Ingeniería Metabólica/métodos , Acetilcoenzima A/metabolismo , Sesquiterpenos/metabolismo , Carbono/metabolismo
16.
ACS Synth Biol ; 12(7): 2157-2167, 2023 07 21.
Artículo en Inglés | MEDLINE | ID: mdl-37350137

RESUMEN

In synthetic biology, the precise control of gene expression is challenging due to the limited orthogonality of expression elements. Here, to address this issue and improve the reusability of genetic elements, we developed a bicistronic expression cassette in Corynebacterium glutamicum based on a leaderless promoter lacking a 5'UTR. The created leaderless bicistronic design (BCD) significantly improved the orthogonality of expression elements across different genes of interest. We also explored the importance of the fore-cistron and SD motif in maintaining the strength of leaderless BCDs. Additionally, we established a library containing 55,901 fore-cistrons and demonstrated that the regulatory range of gene expression in leaderless BCDs can be broader by modifying the fore-cistron sequence. This study provides a novel synthetic biology tool based on leaderless BCD for fine-tuning gene expression in C. glutamicum using fore-cistrons. Moreover, the strategy developed here can also be applied to improve the performance of other leaderless promoters in other bacteria.


Asunto(s)
Corynebacterium glutamicum , Corynebacterium glutamicum/genética , Corynebacterium glutamicum/metabolismo , Regiones Promotoras Genéticas/genética , Biblioteca de Genes , Expresión Génica , Regulación Bacteriana de la Expresión Génica/genética
17.
ACS Synth Biol ; 12(6): 1599-1607, 2023 06 16.
Artículo en Inglés | MEDLINE | ID: mdl-37172280

RESUMEN

Polyketides are a class of natural products with many applications but are mainly appealing as pharmaceuticals. Heterologous production of polyketides in the yeast Saccharomyces cerevisiae has been widely explored because of the many merits of this model eukaryotic microorganism. Although acetyl-CoA and malonyl-CoA, the precursors for polyketide synthesis, are distributed in several yeast subcellular organelles, only cytosolic synthesis of polyketides has been pursued in previous studies. In this study, we investigate polyketide synthesis by directly using acetyl-CoA in the peroxisomes of yeast strain CEN.PK2-1D. We first demonstrate that the polyketide flaviolin can be synthesized in this organelle upon peroxisomal colocalization of native acetyl-CoA carboxylase and 1,3,6,8-tetrahydroxynaphthalene synthase (a type III polyketide synthase). Next, using the synthesis of the polyketide triacetic acid lactone as an example, we show that (1) a new peroxisome targeting sequence, pPTS1, is more effective than the previously reported ePTS1 for peroxisomal polyketide synthesis; (2) engineering peroxisome proliferation is effective to boost polyketide production; and (3) peroxisomes provide an additional acetyl-CoA reservoir and extra space to accommodate enzymes so that utilizing the peroxisomal pathway plus the cytosolic pathway produces more polyketide than the cytosolic pathway alone. This research lays the groundwork for more efficient heterologous polyketide biosynthesis using acetyl-CoA pools in subcellular organelles.


Asunto(s)
Policétidos , Saccharomyces cerevisiae , Saccharomyces cerevisiae/metabolismo , Acetilcoenzima A/metabolismo , Policétidos/metabolismo , Malonil Coenzima A/metabolismo , Acetil-CoA Carboxilasa/genética , Acetil-CoA Carboxilasa/metabolismo
18.
Plant Biotechnol J ; 21(7): 1408-1425, 2023 07.
Artículo en Inglés | MEDLINE | ID: mdl-37031416

RESUMEN

Stone cells are often present in pear fruit, and they can seriously affect the fruit quality when present in large numbers. The plant growth regulator NAA, a synthetic auxin, is known to play an active role in fruit development regulation. However, the genetic mechanisms of NAA regulation of stone cell formation are still unclear. Here, we demonstrated that exogenous application of 200 µM NAA reduced stone cell content and also significantly decreased the expression level of PbrNSC encoding a transcriptional regulator. PbrNSC was shown to bind to an auxin response factor, PbrARF13. Overexpression of PbrARF13 decreased stone cell content in pear fruit and secondary cell wall (SCW) thickness in transgenic Arabidopsis plants. In contrast, knocking down PbrARF13 expression using virus-induced gene silencing had the opposite effect. PbrARF13 was subsequently shown to inhibit PbrNSC expression by directly binding to its promoter, and further to reduce stone cell content. Furthermore, PbrNSC was identified as a positive regulator of PbrMYB132 through analyses of co-expression network of stone cell formation-related genes. PbrMYB132 activated the expression of gene encoding cellulose synthase (PbrCESA4b/7a/8a) and lignin laccase (PbrLAC5) binding to their promotors. As expected, overexpression or knockdown of PbrMYB132 increased or decreased stone cell content in pear fruit and SCW thickness in Arabidopsis transgenic plants. In conclusion, our study shows that the 'PbrARF13-PbrNSC-PbrMYB132' regulatory cascade mediates the biosynthesis of lignin and cellulose in stone cells of pear fruit in response to auxin signals and also provides new insights into plant SCW formation.


Asunto(s)
Arabidopsis , Pyrus , Frutas/metabolismo , Lignina/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Ácidos Indolacéticos/metabolismo , Regulación de la Expresión Génica de las Plantas
19.
J Gen Appl Microbiol ; 69(1): 34-44, 2023 Jun 22.
Artículo en Inglés | MEDLINE | ID: mdl-36878578

RESUMEN

Protein synthesis in Corynebacterium glutamicum is critical for applications in biotechnology and medicine. However, the use of C. glutamicum for protein production is limited by its low expression and aggregation. To overcome these limitations, a molecular chaperone plasmid system was developed in this study to improve the efficiency of recombinant protein synthesis in C. glutamicum. The effect of molecular chaperones on target protein synthesis (Single-chain variable fragment, Scfv) under three different promoter strengths was tested. In addition, the plasmid containing the molecular chaperone and target protein was verified for growth stability and plasmid stability. This expression model was further validated using two recombinant proteins, human interferon-beta (Hifn) and hirudin variant III (Rhv3). Finally, the Rhv3 protein was purified, and analysis of Rhv3 activity confirmed that the use of a molecular chaperone led to an improvement in test protein synthesis. Thus, the use of molecular chaperones is believed to will improve recombinant proteins synthesis in C. glutamicum.


Asunto(s)
Corynebacterium glutamicum , Humanos , Corynebacterium glutamicum/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Plásmidos/genética , Regiones Promotoras Genéticas , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo
20.
Front Psychol ; 14: 1032767, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36910793

RESUMEN

Introduction: DanceSport is described as a dance involving a male-female partner. It is important to comprehend the partnership between dance couples so that their competitive performance can be effectively supported. However, only a few studies have verified the influence of partnership between DanceSport couples on competitive performance and explored its psychological mechanism to provide means to deal with the partnership. The reason was that there was a lack of appropriate assessment tools. Aims: This multi-study outlines the development, content, and construct validity of a novel, mixed-method tool to assess DanceSport partnership. Methods: The development of the Partnership Scale-DanceSport Couples (PS-DSC) included four studies and data from four samples of Chinese elite dancers (N = 914 total). In stage 1, outlined in study 1, PS-DSC items were generated and then refined using the feedback provided by academics, sports coaches, and elite dancers. In stage 2, outlined in studies 2 and 3, exploratory factor analysis and confirmatory factor analysis were used to examine the structure of the PS-DSC items. In stage 3, outlined in study 4, composite reliability, discriminant validity, and convergent validity were assessed. The result of this process was a 13-item three-factor instrument. Based on these initial findings, the PS-DSC provided the first valid and reliable way of measuring partnerships between DanceSport couples. Conclusion: This study has taken the promising first step in developing a tool to comprehensively measure partnerships between DanceSport couples.

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