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1.
Cancer Lett ; 483: 66-74, 2020 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-32142917

RESUMO

Endometrial cancer, a type of primary epithelial malignant tumor in the endometrium, is one of the three most common malignant tumors of the female reproductive system. While the incidence of endometrial cancer has been recently rising, its etiology remains unclear. In this study we found that EM2D9, an independently developed monoclonal antibody, specifically recognized endometrial cancer cells; we further determined that EM2D9 target protein was α5ß1. In vitro and in vivo experiments showed that EM2D9 inhibited the migration of endometrial cancer cells. Real-time quantitative PCR results showed that the expression of CD151 mRNA in endometrial carcinoma cells significantly decreased after EM2D9 treatment. We also found that EM2D9 affected the FAK signaling pathway. Collectively, these results shed light on a new mechanism for the development of endometrial carcinoma.

2.
Medicine (Baltimore) ; 99(12): e18930, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32195925

RESUMO

BACKGROUND: Lumbar disc herniation (LDH) is a common disease that seriously affects patients' quality of life. Although several articles have reported that acupuncture can improve the symptoms of LDH, different guidelines do not evaluate the efficacy of acupuncture consistently, new randomized controlled trials have been published in recent years.The purpose of this study is to evaluate the efficacy and safety of acupuncture for LDH. METHOD: Electronic resource databases, trial registration platform, and different types of grey literature will be systematically searched for eligible studies by 2 authors independently. The type of trial will be limited to randomized controlled trials on acupuncture treatment for LDH. Search strategy will be a combination of terms associated with LDH (eg, low back pain or sciatica) and study of design (eg, randomized controlled trials or clinical trial). Data from homogeneous studies will be combined in a fixed-effects model, and the evidence level will be measured by grading of recommendations assessment, development, and evaluation. RESULTS: This study will provide high-quality evidence to evaluate the relief of pain intensity and improvement of dysfunction of acupuncture in patients with LDH, and to evaluate the safety of acupuncture. CONCLUSION: This study will provide strong evidence for evaluating whether acupuncture therapy is effective and safe for LDH patients. PROSPERO REGISTRATION NUMBER: CRD 42019137399.


Assuntos
Terapia por Acupuntura/métodos , Deslocamento do Disco Intervertebral/terapia , Vértebras Lombares , Terapia por Acupuntura/efeitos adversos , Humanos , Qualidade de Vida , Ensaios Clínicos Controlados Aleatórios como Assunto , Projetos de Pesquisa
3.
Alzheimers Dement ; 16(3): 524-530, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32043803

RESUMO

INTRODUCTION: Clinical Alzheimer's disease (AD) and dementia with Lewy bodies often have mixed AD and Lewy pathology, making it difficult to delineate risk factors. METHODS: Six risk factors for earlier dementia onset due to autopsy-confirmed AD (n = 647), mixed AD and Lewy body disease (AD + LBD; n = 221), and LBD (n = 63) were entered into multiple linear regressions using data from the National Alzheimer's Coordinating Center. RESULTS: In AD and AD + LBD, male sex and apolipoprotein E (APOE) ɛ4 alleles each predicted a 2- to 3-year-earlier onset and depression predicted a 3-year-earlier onset. In LBD, higher education predicted earlier onset and depression predicted a 5.5-year-earlier onset. DISCUSSION: Male sex and APOE ɛ4 alleles increase risk for earlier dementia onset in AD but not LBD. Depression increases risk for earlier dementia onset in AD, LBD, and AD + LBD, but evaluating the course, treatment, and severity is needed in future studies.

4.
Oral Dis ; 26(4): 778-788, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31958204

RESUMO

OBJECTIVE: As an extracellular vesicle, exosomes can release from virus-infected cells containing various viral or host cellular elements and could stimulate recipient's cellular response. Enterovirus 71 (EV71), a single-strand positive-sense RNA virus, is known to cause hand, foot, and mouth disease (HFMD) in children and bring about severe clinical diseases. METHODS: Separated the human oral epithelial cells (OE cells) from normal buccal mucosa through enzyme digestion. Performed a comprehensive miRNA profiling in exosomes from EV71-infected OE cells through deep small RNA-seq. Using the Human Antiviral Response RT Profiler PCR Array profiles to explore the interactions of innate immune signaling networks with exosomal miR-30a. Knocked out the MyD88 gene in macrophages using CRISPR/Cas9-mediated genome editing method. RESULTS: Our study demonstrated that the miR-30a was preferentially enriched in exosomes that released from EV71-infected human oral epithelial cells through small RNA-seq. We found that the transfer of exosomal miR-30a to macrophages could suppress type Ⅰ interferon response through targeting myeloid differentiation factor 88 (MyD88) and subsequently facilitate the viral replication. CONCLUSIONS: Exosomes released from EV71-infected OE cells selectively packaged high level of miR-30a that can be functionally transferred to the recipient macrophages resulted in targeting MyD88 and subsequently inhibited type I interferon production in receipt cells, thus promoting the EV71 replication.

5.
Nat Prod Rep ; 37(2): 276-292, 2020 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-31515549

RESUMO

Covering: 2013-2018 Natural products bearing quaternary carbon stereocenters have attracted tremendous interest from the synthetic community due to their diverse biological activities and fascinating molecular architectures. However, the construction of these molecules in an enantioselective fashion remains a long-standing challenge because of the lack of efficient asymmetric catalytic methods for installing these motifs. The rapid progress in the development of new-generation efficient chiral catalysts has opened the door for several asymmetric reactions, such as Michael addition, dearomative cyclization, polyene cyclization, α-arylation, cycloaddition, allylation, for the construction of quaternary carbon stereocenters in a highly enantioselective fashion. These asymmetric catalytic methods have greatly facilitated the synthesis of complex natural products with improved output and overall efficiency. In this concise review, we highlight the progress in the last six years in complex natural product synthesis, in which at least one quaternary carbon stereocenter has been constructed via asymmetric catalytic technologies, with particular emphasis on the analysis of the stereochemical model of each enantioselective transformation.

6.
World J Microbiol Biotechnol ; 35(11): 171, 2019 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-31673786

RESUMO

Fungal endo-ß-1,4-xylanases (endo-xylanases) can hydrolyze xylan into xylooligosaccharides (XOS), and have potential biotechnological applications for the exploitation of natural renewable polysaccharides. In the current study, we aimed to screen and characterize an efficient fungal endo-xylanase from 100 natural humus-rich soil samples collected in Guizhou Province, China, using extracted sugarcane bagasse xylan (SBX) as the sole carbon source. Initially, 182 fungal isolates producing xylanases were selected, among which Trichoderma sp. strain TP3-36 was identified as showing the highest xylanase activity of 295 U/mL with xylobiose (X2) as the main product when beechwood xylan was used as substrate. Subsequently, a glycoside hydrolase family 11 endo-xylanase, TXyn11A, was purified from strain TP3-36, and its optimal pH and temperature for activity against beechwood xylan were identified to be 5.0 and 55 °C, respectively. TXyn11A was stable across a broad pH range (3.0-10.0), and exhibited strict substrate specificity, including xylan from beechwood, wheat, rye, and sugarcane bagasse, with Km and Vmax values of 5 mg/mL and 1250 µmol/mg min, respectively, toward beechwood xylan. Intriguingly, the main product obtained from hydrolysis of beechwood xylan by TXyn11A was xylobiose, whereas SBX hydrolysis resulted in both X2 and xylotriose. Overall, these characteristics of the endo-xylanase TXyn11A indicate several potential industrial applications.


Assuntos
Dissacarídeos/metabolismo , Endo-1,4-beta-Xilanases/química , Endo-1,4-beta-Xilanases/isolamento & purificação , Trichoderma/enzimologia , Xilanos/metabolismo , Celulose , China , Estabilidade Enzimática , Proteínas Fúngicas/isolamento & purificação , Concentração de Íons de Hidrogênio , Hidrólise , Cinética , Saccharum/metabolismo , Microbiologia do Solo , Especificidade por Substrato , Temperatura , Trichoderma/genética , Trichoderma/isolamento & purificação
7.
Appl Environ Microbiol ; 85(24)2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31604764

RESUMO

Transcriptional regulation of cellulolytic and xylolytic genes in ascomycete fungi is controlled by specific carbon sources in different external environments. Here, comparative transcriptomic analyses of Penicillium oxalicum grown on wheat bran (WB), WB plus rice straw (WR), or WB plus Avicel (WA) as the sole carbon source under solid-state fermentation (SSF) revealed that most of the differentially expressed genes (DEGs) were involved in metabolism, specifically, carbohydrate metabolism. Of the DEGs, the basic core carbohydrate-active enzyme-encoding genes which responded to the plant biomass resources were identified in P. oxalicum, and their transcriptional levels changed to various extents depending on the different carbon sources. Moreover, this study found that three deletion mutants of genes encoding putative transcription factors showed significant alterations in filter paper cellulase production compared with that of a parental P. oxalicum strain with a deletion of Ku70 (ΔPoxKu70 strain) when grown on WR under SSF. Importantly, the ΔPoxAtf1 mutant (with a deletion of P. oxalicum Atf1, also called POX03016) displayed 46.1 to 183.2% more cellulase and xylanase production than a ΔPoxKu70 mutant after 2 days of growth on WR. RNA sequencing and quantitative reverse transcription-PCR revealed that PoxAtf1 dynamically regulated the expression of major cellulase and xylanase genes under SSF. PoxAtf1 bound to the promoter regions of the key cellulase and xylanase genes in vitro This study provides novel insights into the regulatory mechanism of fungal cellulase and xylanase gene expression under SSF.IMPORTANCE The transition to a more environmentally friendly economy encourages studies involving the high-value-added utilization of lignocellulosic biomass. Solid-state fermentation (SSF), that simulates the natural habitat of soil microorganisms, is used for a variety of applications such as biomass biorefinery. Prior to the current study, our understanding of genome-wide gene expression and of the regulation of gene expression of lignocellulose-degrading enzymes in ascomycete fungi during SSF was limited. Here, we employed RNA sequencing and genetic analyses to investigate transcriptomes of Penicillium oxalicum strain EU2101 cultured on medium containing different carbon sources and to identify and characterize transcription factors for regulating the expression of cellulase and xylanase genes during SSF. The results generated will provide novel insights into genetic engineering of filamentous fungi to further increase enzyme production.

8.
Angew Chem Int Ed Engl ; 58(47): 17074-17080, 2019 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-31552701

RESUMO

Recently, CuH-catalyzed reductive coupling processes involving carbonyl compounds and imines have become attractive alternatives to traditional methods for stereoselective addition because of their ability to use readily accessible and stable olefins as surrogates for organometallic nucleophiles. However, the inability to use aldehydes, which usually reduce too rapidly in the presence of copper hydride complexes to be viable substrates, has been a major limitation. Shown here is that by exploiting relative concentration effects through kinetic control, this intrinsic reactivity can be inverted and the reductive coupling of 1,3-dienes with aldehydes achieved. Using this method, both aromatic and aliphatic aldehydes can be transformed into synthetically valuable homoallylic alcohols with high levels of diastereo- and enantioselectivities, and in the presence of many useful functional groups. Furthermore, using a combination of theoretical (DFT) and experimental methods, important mechanistic features of this reaction related to stereo- and chemoselectivities were uncovered.

9.
Angew Chem Int Ed Engl ; 58(38): 13573-13583, 2019 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-31343811

RESUMO

We herein report the development of a conformationally defined, electron-rich, C2 -symmetric, P-chiral bisphosphorus ligand, ArcPhos, by taking advantage of stereoelectronic effects in ligand design. With the Rh-ArcPhos catalyst, excellent enantioselectivities and unprecedentedly high turnovers (TON up to 10 000) were achieved in the asymmetric hydrogenation of aliphatic carbocyclic and heterocyclic tetrasubstituted enamides, to generate a series of chiral cis-2-alkyl-substituted carbocyclic and heterocyclic amine derivatives in excellent enantiomeric ratios. This method also enabled an efficient and practical synthesis of the Janus kinase inhibitor (R)-tofacitinib.

10.
Biotechnol Biofuels ; 12: 103, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31164922

RESUMO

Background: Solid-state fermentation (SSF) mimics the natural decay environment of soil fungi and can be employed to investigate the production of plant biomass-degrading enzymes. However, knowledge on the transcriptional regulation of fungal genes during SSF remains limited. Herein, transcriptional profiling was performed on the filamentous fungus Penicillium oxalicum strain HP7-1 cultivated in medium containing wheat bran plus rice straw (WR) under SSF (WR_SSF) and submerged fermentation (WR_SmF; control) conditions. Novel key transcription factors (TFs) regulating fungal cellulase and xylanase gene expression during SSF were identified via comparative transcriptomic and genetic analyses. Results: Expression of major cellulase genes was higher under WR_SSF condition than that under WR_SmF, but the expression of genes involved in the citric acid cycle was repressed under WR_SSF condition. Fifty-six candidate regulatory genes for cellulase production were screened out from transcriptomic profiling of P. oxalicum HP7-1 for knockout experiments in the parental strain ∆PoxKu70, resulting in 43 deletion mutants including 18 constructed in the previous studies. Enzyme activity assays revealed 14 novel regulatory genes involved in cellulase production in P. oxalicum during SSF. Remarkably, deletion of the essential regulatory gene PoxMBF1, encoding Multiprotein Bridging Factor 1, resulted in doubled cellulase and xylanase production at 2 days after induction during both SSF and SmF. PoxMBF1 dynamically and differentially regulated transcription of a subset of cellulase and xylanase genes during SSF and SmF, and conferred stress resistance. Importantly, PoxMBF1 bound specifically to the putative promoters of major cellulase and xylanase genes in vitro. Conclusions: We revealed differential transcriptional regulation of P. oxalicum during SSF and SmF, and identified PoxMBF1, a novel TF that directly regulates cellulase and xylanase gene expression during SSF and SmF. These findings expand our understanding of regulatory mechanisms of cellulase and xylanase gene expression during fungal fermentation.

11.
Biotechnol Biofuels ; 12: 105, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31073329

RESUMO

Background: Soil ascomycete fungi produce plant-biomass-degrading enzymes to facilitate nutrient and energy uptake in response to exogenous stress. This is controlled by a complex signal network, but the regulatory mechanisms are poorly understood. An essential Zn2Cys6 transcription factor (TF) PoxCxrA was identified to be required for cellulase and xylanase production in Penicillium oxalicum. The genome-wide regulon and DNA binding sequences of PoxCxrA were further identified through RNA-Sequencing, DNase I footprinting experiments and in vitro electrophoretic mobility shift assays. Moreover, a minimal DNA-binding domain in PoxCxrA was recognised. Results: A PoxCxrA regulon of 1970 members was identified in P. oxalicum, and it was displayed that PoxCxrA regulated the expression of genes encoding major plant cell wall-degrading enzymes, as well as important cellodextrin and/or glucose transporters. Interestingly, PoxCxrA positively regulated the expression of a known important TF PoxClrB. DNase I footprinting experiments and in vitro electrophoretic mobility shift assays further revealed that PoxCxrA directly bound the promoter regions of PoxClrB and a cellobiohydrolase gene cbh1 (POX05587/Cel7A-2) at different nucleic acid sequences. Remarkably, PoxCxrA autoregulated its own PoxCxrA gene expression. Additionally, a minimal 42-amino-acid PoxCxrA DNA-binding domain was identified. Conclusion: PoxCxrA could directly regulate the expression of cellulase genes and the regulatory gene PoxClrB via binding their promoters at different nucleic acid sequences. This work expands the diversity of DNA-binding motifs known to be recognised by Zn2Cys6 TFs, and demonstrates novel regulatory mechanisms of fungal cellulase gene expression.

12.
Cancer Med ; 8(4): 1806-1816, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30907072

RESUMO

Bladder cancer (BC) is a complex disease and could be classified into nonmuscle-invasive BC (NMIBC) or muscle-invasive BC (MIBC) subtypes according to the distinct genetic background and clinical prognosis. Until now, the golden standard and confirmed diagnosis of BC is cystoscopy and the major problems of BC are the high rate of recurrence and high costs in the clinic. Recent molecular and genetic studies have provided perspectives on the novel biomarkers and potential therapeutic targets of BC. In this article, we provided an overview of the traditional diagnostic approaches of BC, and introduced some new imaging, endoscopic, and immunological diagnostic technology in the accurate diagnosis of BC. Meanwhile, the minimally invasive precision treatment technique, immunotherapy, chemotherapy, gene therapy, and targeted therapy of BC were also included. Here, we will overview the diagnosis and therapy methods of BC used in clinical practice, focusing on their specificity, efficiency, and safety. On the basis of the discussion of the benefits of precision medicine in BC, we will also discuss the challenges and limitations facing the non-invasive methods of diagnosis and precision therapy of BC. The molecularly targeted and immunotherapeutic approaches, and gene therapy methods to BC treatment improved the prognosis and overall survival of BC patients.

13.
J Am Chem Soc ; 141(12): 5062-5070, 2019 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-30817137

RESUMO

Chiral tertiary alcohols are important building blocks for the synthesis of pharmaceutical agents and biologically active natural products. The addition of carbon nucleophiles to ketones is the most common approach to tertiary alcohol synthesis but traditionally relies on stoichiometric organometallic reagents that are difficult to prepare, sensitive, and uneconomical. We describe a mild and efficient method for the copper-catalyzed allylation of ketones using widely available 1,3-dienes as allylmetal surrogates. Homoallylic alcohols bearing a wide range of functional groups are obtained in high yield and with good regio-, diastereo-, and enantioselectivity. Mechanistic investigations using density functional theory (DFT) implicate the in situ formation of a rapidly equilibrating mixture of isomeric copper(I) allyl complexes, from which Curtin-Hammett kinetics determine the major isomer of the product. A stereochemical model is provided to explain the high diastereo- and enantioselectivity of this process. Finally, this method was applied to the preparation of an important drug, ( R)-procyclidine, and a key intermediate in the synthesis of several pharmaceuticals.

14.
Biotechnol Biofuels ; 12: 7, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30622649

RESUMO

Background: Raw-starch-digesting glucoamylases (RSDGs) from filamentous fungi have great commercial values in starch processing; however, the regulatory mechanisms associated with their production in filamentous fungi remain unknown. Penicillium oxalicum HP7-1 isolated by our laboratory secretes RSDG with suitable properties but at low production levels. Here, we screened and identified novel regulators of RSDG gene expression in P. oxalicum through transcriptional profiling and genetic analyses. Results: Penicillium oxalicum HP7-1 transcriptomes in the presence of glucose and starch, respectively, used as the sole carbon source were comparatively analyzed, resulting in screening of 23 candidate genes regulating the expression of RSDG genes. Following deletion of 15 of the candidate genes in the parental P. oxalicum strain ∆PoxKu70, enzymatic assays revealed five mutants exhibiting significant reduction in the production of raw-starch-digesting enzymes (RSDEs). The deleted genes (POX01907, POX03446, POX06509, POX07078, and POX09752), were the first report to regulate RSDE production of P. oxalicum. Further analysis revealed that ∆POX01907 lost the most RSDE production (83.4%), and that POX01907 regulated the expression of major amylase genes, including the RSDG gene POX01356/PoxGA15A, a glucoamylase gene POX02412, and the α-amylase gene POX09352/Amy13A, during the late-stage growth of P. oxalicum. Conclusion: Our results revealed a novel essential regulatory gene POX01907 encoding a transcription factor in controlling the production of RSDE, regulating the expression of an important RSDG gene POX01356/PoxGA15A, in P. oxalicum. These results provide insight into the regulatory mechanism of fungal amylolytic enzyme production.

15.
BMC Microbiol ; 18(1): 203, 2018 12 04.
Artigo em Inglês | MEDLINE | ID: mdl-30509171

RESUMO

BACKGROUND: Farnesol has potential antifungal activity against Candida albicans biofilms, but the molecular mechanism of this activity is still unclear. Farnesol inhibits hyphal growth by regulating the cyclic AMP (cAMP) signalling pathway in C. albicans, and CYR1 and PDE2 regulate a pair of enzymes that are directly responsible for cAMP synthesis and degradation. Here, we hypothesize that farnesol enhances the antifungal susceptibility of C. albicans biofilms by regulating CYR1 and PDE2. RESULTS: The resistance of the CYR1- and PDE2-overexpressing strains to caspofungin, itraconazole and terbinafine was increased in planktonic cells, and that to amphotericin B was increased in biofilms. Meanwhile, the biofilms of the CYR1- and PDE2-overexpressing strains were thicker (all p < 0.05) and consisted of more hyphae than that of the wild strain. The intracellular cAMP levels were higher in the biofilms of the CYR1-overexpressing strain than that in the biofilms of the wild strain (all p < 0.01), while no changes were found in the PDE2-overexpressing strain. Exogenous farnesol decreased the resistance of the CYR1- and PDE2-overexpressing strains to these four antifungals, repressed the hyphal and biofilm formation of the strains, and decreased the intracellular cAMP level in the biofilms (all p < 0.05) compared to the untreated controls. In addition, farnesol decreased the expression of the gene CYR1 and the protein CYR1 in biofilms of the CYR1-overexpressing strain (all p < 0.05) but increased the expression of the gene PDE2 and the protein PDE2 in biofilms of the PDE2-overexpressing strain (all p < 0.01). CONCLUSIONS: The results indicate that CYR1 and PDE2 regulate the resistance of C. albicans biofilms to antifungals. Farnesol suppresses the resistance of C. albicans biofilms to antifungals by regulating the expression of the gene CYR1 and PDE2, while PDE2 regulation was subordinate to CYR1 regulation.


Assuntos
Antifúngicos/farmacologia , Biofilmes/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Nucleotídeo Cíclico Fosfodiesterase do Tipo 2/metabolismo , Farmacorresistência Fúngica , Farneseno Álcool/farmacologia , Proteínas Fúngicas/metabolismo , Candida albicans/genética , Candida albicans/fisiologia , Caspofungina/farmacologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 2/genética , Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica , Hifas/efeitos dos fármacos , Hifas/genética , Hifas/metabolismo , Terbinafina/farmacologia
16.
Biotechnol Biofuels ; 11: 276, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30337955

RESUMO

Background: Perfect and low cost of fungal amylolytic and cellulolytic enzymes are prerequisite for the industrialization of plant biomass biorefinergy to biofuels. Genetic engineering of fungal strains based on regulatory network of transcriptional factors (TFs) and their targets is an efficient strategy to achieve the above described aim. Talaromyces pinophilus produces integrative amylolytic and cellulolytic enzymes; however, the regulatory mechanism associated with the expression of amylase and cellulase genes in T. pinophilus remains unclear. In this study, we screened for and identified novel TFs regulating amylase and/or cellulase gene expression in T. pinophilus 1-95 through comparative transcriptomic and genetic analyses. Results: Comparative analysis of the transcriptomes from T. pinophilus 1-95 grown on media in the presence and absence of glucose or soluble starch as the sole carbon source screened 33 candidate TF-encoding genes that regulate amylase gene expression. Thirty of the 33 genes were successfully knocked out in the parental strain T. pinophilus ∆TpKu70, with seven of the deletion mutants firstly displaying significant changes in amylase production as compared with the parental strain. Among these, ∆TpRfx1 (TpRfx1: Talaromyces pinophilus Rfx1) showed the most significant decrease (81.5%) in amylase production, as well as a 57.7% reduction in filter paper cellulase production. Real-time quantitative reverse transcription PCR showed that TpRfx1 dynamically regulated the expression of major amylase and cellulase genes during cell growth, and in vitro electrophoretic mobility shift assay revealed that TpRfx1 bound the promoter regions of genes encoding α-amylase (TP04014/Amy13A), glucoamylase (TP09267/Amy15A), cellobiohydrolase (TP09412/cbh1), ß-glucosidase (TP05820/bgl1), and endo-ß-1,4-glucanase (TP08514/eg1). TpRfx1 protein containing a regulatory factor X (RFX) DNA-binding domain belongs to RFX family. Conclusion: We identified a novel RFX protein TpRFX1 that directly regulates the expression of amylase and cellulase genes in T. pinophilus, which provides new insights into the regulatory mechanism of fungal amylase and cellulase gene expression.

17.
BMC Oral Health ; 18(1): 152, 2018 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-30157822

RESUMO

BACKGROUND: The root canal glide path has been recommended as the foundation for a safer root canal preparation. The aim of this study was to compare glide paths created with K-files, PathFiles, and the ProGlider file, and their effects on subsequent WaveOne preparation regarding canal transportation, canal volume increase, apical extruded debris, and working time in curved canals. METHODS: Sixty mesial canals of extracted human mandibular first molars were randomly assigned to the K-file (KF), PathFile (PF) and ProGlider file (PG) groups for glide path preparation. Then, canals were prepared using WaveOne files. Specimens were scanned (voxel size: 18 µm) three times using micro-computed tomography: pre-glide path, post-glide path, and post-root canal preparation. Canal transportations were measured at 1, 3, and 5 mm levels from the apical foramen, and canal volume increases were also accounted. Apical extruded debris during preparation was collected for measurement. Meanwhile,working time was recorded. Data were analyzed statistically using one-way analysis of variance and Tukey's post hoc tests (p < 0.05). RESULTS: After glide path preparation, the PG and PF groups showed significantly less canal transportation than the KF group at all levels (P < 0.05), while the PG group exhibited a significantly larger canal volume increase than the PF and KF groups (P < 0.05). After the subsequent canal preparation with WaveOne, the PG and PF groups showed significantly less canal transportation than the KF group at 3 and 5 mm levels, and the PG group showed significantly less canal transportation than the PF group at 5 mm level (P < 0.05). However, statistically similar canal volume increases occurred among the three groups. Additionally, the PG and PF groups produced less apical extruded debris compared to the KF group (P < 0.05). The working time of the PG group was the shortest, while that of the KF group was the longest. CONCLUSION: Compared with the PathFiles and K-files, the ProGlider file combined with the WaveOne file showed reduced canal transportation and working time.


Assuntos
Dente Molar/cirurgia , Preparo de Canal Radicular/instrumentação , Desenho de Equipamento , Humanos , Técnicas In Vitro , Mandíbula , Dente Molar/diagnóstico por imagem , Fatores de Tempo , Microtomografia por Raio-X
18.
Appl Microbiol Biotechnol ; 102(21): 9291-9301, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30155751

RESUMO

Raw starch-degrading enzymes (RSDEs) are capable of directly degrading raw starch granules below the gelatinization temperature of starch, which may significantly reduce the cost of starch-based biorefining. However, low yields of natural RSDEs from filamentous fungi limit their industrial application. In this study, transcriptomic and secretomic profiling was employed to screen strongest promoters and signal peptides for use in overexpression of a RSDE gene in Penicillium oxalicum. Top five strong promoters and three signal peptides were detected. Using a green fluorescent protein (GFP) as the reporter, the inducible promoter pPoxEgCel5B of an endoglucanase gene PoxEgCel5B and the signal peptide spPoxGA15A of a raw starch-degrading glucoamylase PoxGA15A were respectively identified as driving the highest GFP production in P. oxalicum. PoxGA15A-overexpressed P. oxalicum strain OXPoxGA15A, which was constructed based on both pPoxEgCel5B and spPoxGA15A, produced significantly higher amounts of recombinant PoxGA15A than the parental strain ∆PoxKu70. Furthermore, crude enzyme from the OXPoxGA15A strain exhibited high activities towards raw starch from cassava, potato, and uncooked soluble starch. Specifically, raw cassava starch-degrading enzyme activity reached 241.6 U/mL in the OXPoxGA15A, which was 3.4-fold higher than that of the ∆PoxKu70. This work provides a feasible method for hyperproduction of RSDEs in P. oxalicum.


Assuntos
Glucana 1,4-alfa-Glucosidase/genética , Penicillium/genética , Regiões Promotoras Genéticas/genética , Sinais Direcionadores de Proteínas/genética , Amido/genética , Fermentação/genética , Fungos/genética , Manihot/genética , Proteínas Recombinantes/genética , Solanum tuberosum/genética , Amido/metabolismo , Temperatura
19.
Appl Environ Microbiol ; 84(18)2018 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-29980558

RESUMO

Soil fungi produce a wide range of chemical compounds and enzymes with potential for applications in medicine and biotechnology. Cellular processes in soil fungi are highly dependent on the regulation under environmentally induced stress, but most of the underlying mechanisms remain unclear. Previous work identified a key GATA-type transcription factor, Penicillium oxalicum NsdD (PoxNsdD; also called POX08415), that regulates the expression of cellulase and xylanase genes in P. oxalicum PoxNsdD shares 57 to 64% identity with the key activator NsdD, involved in asexual development in Aspergillus In the present study, the regulatory roles of PoxNsdD in P. oxalicum were further explored. Comparative transcriptomic profiling revealed that PoxNsdD regulates major genes involved in starch, cellulose, and hemicellulose degradation, as well as conidiation and pigment biosynthesis. Subsequent experiments confirmed that a ΔPoxNsdD strain lost 43.9 to 78.8% of starch-digesting enzyme activity when grown on soluble corn starch, and it produced 54.9 to 146.0% more conidia than the ΔPoxKu70 parental strain. During cultivation, ΔPoxNsdD cultures changed color, from pale orange to brick red, while the ΔPoxKu70 cultures remained bluish white. Real-time quantitative reverse transcription-PCR showed that PoxNsdD dynamically regulated the expression of a glucoamylase gene (POX01356/Amy15A), an α-amylase gene (POX09352/Amy13A), and a regulatory gene (POX03890/amyR), as well as a polyketide synthase gene (POX01430/alb1/wA) for yellow pigment biosynthesis and a conidiation-regulated gene (POX06534/brlA). Moreover, in vitro binding experiments showed that PoxNsdD bound the promoter regions of the above-described genes. This work provides novel insights into the regulatory mechanisms of fungal cellular processes and may assist in genetic engineering of Poxalicum for potential industrial and medical applications.IMPORTANCE Most filamentous fungi produce a vast number of extracellular enzymes that are used commercially for biorefineries of plant biomass to produce biofuels and value-added chemicals, which might promote the transition to a more environmentally friendly economy. The expression of these extracellular enzyme genes is tightly controlled at the transcriptional level, which limits their yields. Hitherto our understanding of the regulation of expression of plant biomass-degrading enzyme genes in filamentous fungi has been rather limited. In the present study, regulatory roles of a key regulator, PoxNsdD, were further explored in the soil fungus Penicillium oxalicum, contributing to the understanding of gene regulation in filamentous fungi and revealing the biotechnological potential of Poxalicum via genetic engineering.


Assuntos
Proteínas Fúngicas/metabolismo , Regulação Fúngica da Expressão Gênica , Penicillium/metabolismo , Pigmentos Biológicos/biossíntese , Esporos Fúngicos/crescimento & desenvolvimento , Fatores de Transcrição/metabolismo , Biodegradação Ambiental , Celulase/genética , Celulase/metabolismo , Celulose/metabolismo , Proteínas Fúngicas/genética , Perfilação da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Glucana 1,4-alfa-Glucosidase/genética , Glucana 1,4-alfa-Glucosidase/metabolismo , Penicillium/enzimologia , Penicillium/genética , Penicillium/crescimento & desenvolvimento , Policetídeo Sintases/genética , Policetídeo Sintases/metabolismo , Esporos Fúngicos/genética , Esporos Fúngicos/metabolismo , Fatores de Transcrição/genética , alfa-Amilases/genética , alfa-Amilases/metabolismo
20.
Huan Jing Ke Xue ; 39(4): 1589-1597, 2018 Apr 08.
Artigo em Chinês | MEDLINE | ID: mdl-29964983

RESUMO

Dynamic variations and sources of nitrate during dry season in the Lijiang River were analyzed using the nitrate concentrations and 15 N and 18 O isotope techniques, from the samples obtained from 13 sections in the Lijiang River from September 28, 2016 to December 28, 2016. Results show that the nitrate concentrations range from 0.46 to 18.48 mg·L-1, with an average of 6.18 mg·L-1, and that the nitrate levels are low during the dry season. Nitrate concentrations in the Lijiang River increase slowly from September to December, mainly being influenced by rainfall, runoff, and human activity. Nitrate concentrations in the Lijiang River from upstream to downstream show a trend of "increase-decrease-increase." Nitrate in the Lijiang River during the dry season mainly originates from organic nitrogen in soil, human and animal feces, sewage (largely living sewage), human and animal waste, and tourism. In order to better protect the water quality of the Lijiang River, the urban sewage pipe network must be expanded, in addition to building small sewage treatment facilities and strengthening tourism management and environmental awareness.


Assuntos
Nitratos/análise , Rios/química , Estações do Ano , Poluentes Químicos da Água/análise , Animais , China , Monitoramento Ambiental , Humanos , Isótopos de Nitrogênio , Isótopos de Oxigênio , Esgotos , Solo/química , Qualidade da Água
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