Title: Improved Production of Recombinant Carboxylesterase FumDM by Co-Expressing Molecular Chaperones in Pichia pastoris Jiang L, Guan X, Liu H, Chang X, Sun J, Sun C, Zhao C Ref: Toxins (Basel), 15:, 2023 : PubMed
Fumonisins (FBs) are mycotoxins that threaten public health and food safety worldwide. Enzymatic degradation of Fumonisin B1 (FB(1)) through decarboxylation has attracted much attention, whereas application of FB(1) carboxylesterase in detoxification requires more effective expression of the recombinant carboxylesterase. In this study, the carboxylesterase FumDM from Sphingopyxis sp. ASAG22 was codon-optimized and co-expressed with five different molecular chaperones (PDI, CPR5, ERO1, HAC1, and Bip) in order to improve the expression level of FumDM in Pichia pastoris (also known as Komagataella phaffii) GS115. The co-expression of different chaperones caused varying degrees of improvement in FumDM activity for FB(1). The enzyme activities of recombinant strains over-expressing PDI and CPR5 reached the highest levels of 259.47 U/mL and 161.34 U/mL, 635% and 357% higher than the original enzyme activity, respectively. Transcriptomic analysis of the two recombinant strains in comparison with the control strain showed that the correct folding of proteins assisted by molecular chaperones played a key role in the improvement of FumDM expression and its enzyme activity. This study demonstrated that co-expression of carboxylesterase FumDM and folding chaperones was an efficient strategy and therefore might inspire new perspectives on the improvement of carboxylesterase for detoxification of FB(1).
Title: A smartphone-based fluorospectrophotometer and ratiometric fluorescence nanoprobe for on-site quantitation of pesticide residue Xiao M, Xu N, He A, Yu Z, Chen B, Jin B, Jiang L, Yi C Ref: iScience, 26:106553, 2023 : PubMed
Cost-effective and user-friendly quantitation at points-of-need plays an important role in food safety inspection, environmental monitoring, and biomedical analysis. This study reports a stand-alone smartphone-based fluorospectrophotometer (the SBS) installed with a custom-designed application (the SBS-App) for on-site quantitation of pesticide using a ratiometric sensing scheme. The SBS can collect fluorescence emission spectra in the wavelength range of 380-760 nm within 5 s. A ratiometric fluorescence probe is facilely prepared by directly mixing the blue-emissive carbon nanodots (the Fe(3+)-specific fluorometric indicator) and red-emissive quantum dots (the internal standard) at a ratio of 11.6 (w/w). Based on the acetylcholinesterase/choline oxidase dual enzyme-mediated cascade catalytic reactions of Fe(2+)/Fe(3+) transformation, a ratiometric fluorescence sensing scheme is developed. The practicability of the SBS is validated by on-site quantitation of chlorpyrifos in apple and cabbage with a comparable accuracy to the GC-MS method, offering a scalable solution to establish a cost-effective surveillance system for pesticide pollution.
Title: A Novel Nomogram for Predicting Risk Factors and Outcomes in Bloodstream Infections Caused by Klebsiella pneumoniae Chen Y, Ying S, Jiang L, Dong S, Dai J, Jin X, Yu W, Qiu Y Ref: Infect Drug Resist, 15:1317, 2022 : PubMed
BACKGROUND: Our study aimed to explore the risk factors in bloodstream infections Klebsiella pneumoniae (BSI-KP) patients and establish nomograms to predict the probability of BSI-CRKP and the prognosis of BSI-KP. METHODS: A total of 252 BSI-KP patients were enrolled from a tertiary teaching hospital between January 1, 2015, and May 31, 2020. Risk factors associated with BSI-CRKP and factors associated with the 30-day mortality were identified using LASSO analysis, univariate and multivariate analysis. RESULTS: There were 121 (48.0%) patients with carbapenem-resistant K. pneumoniae (CRKP) and 131 (52.0%) patients with carbapenem-susceptible K. pneumoniae (CSKP). The multivariate logistic regression analysis demonstrated that gastric tube indwelling before BSI (OR=2.442, P=0.043) and more types of antibiotics use before BSI (OR=1.305, P=0.009) were independent risk factors for BSI-CRKP. And previous transplantations, prior ICU stay, gastric tube indwelling before BSI, more types of antibiotics use before BSI, lower Hb and cholinesterase were associated with CRKP-BSI. The C-index of models indicated its good accuracy (C-index 0.816, 95% CI 0.763-0.868). In patients with BSI-CRKP, further logistic regression analysis revealed urinary catheterization (OR=0.298, P=0.017) was found to be an independent risk factor for 30-day mortality, while ceftazidime/avibactam use (OR=8.438, P=0.003) was an independent favorable prognostic factor. The nomogram predicated CRKP, ICU hospitalization, more types of antibiotics use, tigecycline, PLT, urinary catheterization were associated with 30-day mortality in patients with BSI-KP. The discriminative ability of the predictive model, as assessed by C-index, was 0.813 (95% CI: 0.780-0.867). CONCLUSION: Previous transplantations, prior ICU stay, gastric tube indwelling before BSI, more types of antibiotics use before BSI, lower Hb and cholinesterase represent significant risk factors for the development of BSI-CRKP. Our nomogram predicated thrombocytopenia was a sign for poor prognosis. Tigecycline resulted in higher mortality for patients with BSI-KP. Rational use of nomograms may help clinicians make better Clinical decisions when treating BSI-KP patients.
Terbuthylazine is an effective and widely used s-triazine herbicide. However, limited data exists on its toxicity and bioaccumulation in earthworms (Eisenia fetida). In this study, we investigated the bioaccumulation, antioxidant enzyme activity, detoxification enzyme activity, and DNA damage in earthworms when exposed to terbuthylazine. The results indicated that terbuthylazine in soil had low bioaccumulation in earthworms and the biota-soil accumulation factors of terbuthylazine declined with an increasing soil terbuthylazine concentration. In the enzyme activity assays, the superoxide dismutase (SOD), catalase (CAT), and glutathione-S-transferase (GST) activities showed upward trends when compared with the control. The carboxylesterase (CarE) activity increased on day 21. The 8-hydroxy-2-deoxyguanosine (8-OHdG) content, a DNA damage bioindicator, was higher than that of the control on day 21. Combined with the integrated biological response index version 2 analysis, these results can provide a comprehensive evaluation of the toxicological effects that terbuthylazine has on earthworms and soil ecosystems.
The stalling global progress in the fight against malaria prompts the urgent need to develop new intervention strategies. Whilst engineered symbiotic bacteria have been shown to confer mosquito resistance to parasite infection, a major challenge for field implementation is to address regulatory concerns. Here, we report the identification of a Plasmodium-blocking symbiotic bacterium, Serratia ureilytica Su_YN1, isolated from the midgut of wild Anopheles sinensis in China that inhibits malaria parasites via secretion of an antimalarial lipase. Analysis of Plasmodium vivax epidemic data indicates that local malaria cases in Tengchong (Yunnan province, China) are significantly lower than imported cases and importantly, that the local vector A. sinensis is more resistant to infection by P. vivax than A. sinensis from other regions. Analysis of the gut symbiotic bacteria of mosquitoes from Yunnan province led to the identification of S. ureilytica Su_YN1. This bacterium renders mosquitoes resistant to infection by the human parasite Plasmodium falciparum or the rodent parasite Plasmodium berghei via secretion of a lipase that selectively kills parasites at various stages. Importantly, Su_YN1 rapidly disseminates through mosquito populations by vertical and horizontal transmission, providing a potential tool for blocking malaria transmission in the field.
Title: The association between toxic pesticide environmental exposure and Alzheimer's disease: A scientometric and visualization analysis Li Y, Fang R, Liu Z, Jiang L, Zhang J, Li H, Liu C, Li F Ref: Chemosphere, 263:128238, 2021 : PubMed
Alzheimer's disease (AD) is one of the most common neurodegenerative diseases. The association between environmental factors (e.g., pesticide) and AD has attracted considerable attention. However, no systematic analysis has been performed and make it difficult to provide deeper insights of AD correlated with pesticide exposure. Hence, this study utilized a bibliometric and visual approach that included map collaborations, co-citations, and keywords, to identifying the knowledge structure, hot topics and the research trends based on 372 publications from the Web of Science Core Collection and PubMed databases. The results showed that 116 institutions from 52 countries published articles in this field. The United States and Israel played a leading role with numerous publications in related journals, as well as prolific institutions and authors, respectively. Three hot topics in pesticide-induced AD were recognized based on co-occurrence keywords detection, including acetylcholinesterase (AChE) inhibitor, oxidative stress, and AChE. Moreover, analysis of keywords burst suggests that some potential molecular mechanisms and therapy targets of pesticide-induced AD, especially for mitochondrial dysfunction and monoamine oxidase-B (MAO-B) that catalyzes the oxidative deamination and causes oxidative stress, are emerging trends. In addition, the study of various pesticides and the assessment method of pesticide exposure will step forward as well. To the best of our knowledge, this study is the first to specifically visualize the relationship between AD and pesticide exposure and to predict potential future research directions.
Title: Degradation and toxicity of the antidepressant fluoxetine in an aqueous system by UV irradiation Pan C, Zhu F, Wu M, Jiang L, Zhao X, Yang M Ref: Chemosphere, :132434, 2021 : PubMed
Fluoxetine (FLU), a selective serotonin reuptake inhibitor, is commonly found in aquatic environments. Ultraviolet (UV) photolysis is widely used to remove certain pharmaceuticals from water and wastewater. The present study aimed to investigate the toxicity of FLU and its transformed products formed during UV photolysis by using zebrafish embryos (Danio rerio) as a model. The degradation rates of FLU for five days were approximately 63.6% +/- 2.14%, 84.6% +/- 0.99%, and 97.5% +/- 0.25% after 15, 30, and 60 min of UV irradiation, respectively. Furthermore, the degradation mechanism was explored using LC-MS measurements and density flooding theory (DFT) theoretical calculations. Comprehensive toxicity preassessment of FLU and its degradation products was carried out using the T.E.S.T. software. The effects of physiological and biochemical parameters and neuron- and apoptosis-related gene expression were examined in zebrafish embryos exposed to non-irradiated (0-min) and irradiated (15, 30- and 60-min) solutions from 4 h post-fertilization (hpf) to 120 hpf. The hatching time of zebrafish embryos exposed to the non-irradiated solution (0-min) and irradiated solution (60-min) was delayed, their heart rate at 48 and 72 hpf increased, and their body length at 120 hpf decreased. Significant differences were found between the non-irradiated (0-min) and UV-irradiated (15- or 30-min) groups. A dynamic response involving acetylcholinesterase (AChE) and superoxide dismutase (SOD) activity was also observed in the non-irradiated and UV-irradiated groups. During the UV treatment experiments, the expression levels of neuron-related and apoptosis-related genes were significantly reduced over time alongside the formation of FLU degradation products. Overall, this study provides new concepts to remove and assess the toxicity of emerging contaminants in aquatic environments and highlights the need to consider the formation and persistence of toxic transformation products.
Title: Enhancing Therapeutic Efficacy of Donepezil by Combined Therapy: A Comprehensive Review Rong X, Jiang L, Qu M, Hassan SSU, Liu Z Ref: Curr Pharm Des, 27:332, 2021 : PubMed
Combination therapy involving different therapeutic strategies mostly provides more rapid and effective results as compared to monotherapy in diverse areas of clinical practice. The most worldwide famous acetylcholinesterase inhibitor (AChEIs) donepezil for its dominant role in Alzheimer's disease (AD) has also attracted the attention of many pharmaceuticals due to its promising pharmacological potencies such as neuroprotective, muscle relaxant, and sleep inducer. Recently, a combination of donepezil with other agents has displayed better desirable results in managing several disorders, including the most common Alzheimer's disease (AD). This study involves all the data regarding the therapeutic effect of donepezil in its combination with other agents and explains its therapeutic targets and mode of action. Furthermore, this review also puts light on the current status of donepezil with other agents in clinical trials. The combination therapy of donepezil with symptomatic relief drugs and disease-modifying agents opens a new road for treating multiple pathological disorders. To the best of our knowledge, this is the first report encircling all the pharmacologic effects of donepezil in its combination therapy with other agents and their current status in clinical trials.
Title: Lipase catalysis of alpha-linolenic acid-rich medium- and long-chain triacylglycerols from perilla oil and medium-chain triacylglycerols with reduced by-products Huang Z, Cao Z, Guo Z, Chen L, Wang Z, Sui X, Jiang L Ref: J Sci Food Agric, 100:4565, 2020 : PubMed
BACKGROUND: Medium- and long- chain triacylglycerols (MLCTs) are functional structural lipids that can provide the human body with essential fatty acids and a faster energy supply. This study aimed to prepare MLCTs rich in alpha-linolenic by enzymatic interesterification of perilla oil and medium-chain triacylglycerols (MCTs), catalyzed by Lipozyme RM IM, Lipozyme TL IM, Lipozyme 435, and Novozyme 435 respectively. RESULTS: The effects of lipase loading, concentration of MCTs, reaction temperature, and reaction time on the yield of MLCTs were investigated. It was found that the reaction achieved more than a 70% yield of MLCTs in triacylglycerols under the conditions of 400 g kg(-1) MCTs and 60 g kg(-1) lipase loading after equilibrium. A novel two-stage deodorization was also applied to purify the interesterification products. The triacylglycerols reach over 97% purity in the products with significant removal (P < 0.05) of the free fatty acids, and the trans fatty acids were strictly controlled at below 1%. There was more than 40% alpha-linolenic in the purified products, with long-chain fatty acids mostly occupying the desired sn-2 position in acylglycerols, which are more active in hydrolysis. CONCLUSION: A series of novel alpha-linolenic acid-rich medium- and long-chain triacylglycerols was prepared. Under appropriate reaction conditions, the yield of MLCTs in triacylglycerols was above 70%. A novel two-stage deodorization can be used to promote the elimination of free fatty acids and limit the generation of trans fatty acids.
Title: Rhizomucor miehei lipase-catalysed synthesis of cocoa butter equivalent from palm mid-fraction and stearic acid: Characteristics and feasibility as cocoa butter alternative Huang Z, Guo Z, Xie D, Cao Z, Chen L, Wang H, Jiang L, Shen Q Ref: Food Chem, :128407, 2020 : PubMed
In this study, cocoa butter equivalents (CBEs) were prepared through enzymatic interesterification of palm mid-fraction (PMF) with stearic acid (SA). The reaction process parameters were experimented and the performance of the product was analysed. PMF and stearic acid (at a mass ratio of 1:2) were catalysed by 80 g kg(-1) enzyme loading of Lipozyme RM IM fromRhizomucor mieheiat 60 degreeC for 120 min. The yield of the CBE product was more than 92%, and the CBE resembled cocoa butter (CB) in terms of its triacylglycerol composition. The hardness of the CBE product was higher than that of CB at different storage temperatures, but this difference was not obvious at 25 degreeC. The polymorphic structures and SFC curve of the CBE were similar to those of the CB. In addition, the CBE could be mixed with CB in any ratio without an obvious eutectic phenomena. Up to 40% CBE could be added to CB without significantly affecting the thermodynamic properties of CB. Thus, replacing CB with the CBE product is feasible.
Title: Immobilized lipase in bio-based metal-organic frameworks constructed by biomimetic mineralization: A sustainable biocatalyst for biodiesel synthesis Li Q, Chen Y, Bai S, Shao X, Jiang L Ref: Colloids Surf B Biointerfaces, 188:110812, 2020 : PubMed
Herein, thermophilic lipase QLM from Alcaligenes sp. has been successfully immobilized in bio-based metal-organic frameworks (MOFs) through biomimetic mineralization, using zinc acetate and adenine as metal ion and organic ligand, respectively. The morphology and structure of lipase@Bio-MOF was systematically characterized by scanning electron microcopy (SEM), transmission electron microcopy (TEM), powder X-ray diffraction (PXRD) and Fourier transform infrared spectra (FT-IR). The enzyme loading in immobilized enzyme was measured to be 15.9 % by thermogravimetric analysis (TGA). Further, it was demonstrated to possess favorable catalytic activity and stability under high temperature and alkaline conditions and in the presence of metal ions, using the hydrolysis of p-nitrophenyl caprylate as a model. Finally, the immobilized enzyme was successfully applied in the preparation of biodiesel through the trans-esterification of sunflower oil with methanol, obtaining a conversion of >60 % at a high oil/methanol ratio of 8:1. Meanwhile, it showed excellent recyclability during the biodiesel production, and no changes of morphology and crystal structure were observed after being used for 3 cycles. Overall, the immobilized lipase in bio-based MOFs provided an economically and environmentally viable biocatalyst for the synthesis of biodiesel.
Title: The influence mechanism of temperature on solid phase denitrification based on denitrification performance, carbon balance, and microbial analysis Shen Q, Ji F, Wei J, Fang D, Zhang Q, Jiang L, Cai A, Kuang L Ref: Sci Total Environ, 732:139333, 2020 : PubMed
In this work, the influence mechanism of temperature on solid phase denitrification (SPD) was investigated using a pilot-scale reactor supported with polycaprolactone (PCL). The results showed that under nitrate loads of ~31.5 mg N/(L.h), as temperature decreased from 30 degrees C to 13 degrees C, the nitrate removal efficiency declined from 94% to 57%. Furthermore, denitrification rate constants were input into Arrhenius equation and the resulting temperature coefficient was 1.04. Significantly nitrite accumulation and less effluent COD residue occurred at low-temperatures. Via stoichiometry, the sludge yield coefficient and COD demand for nitrate removal both increased as a function of increasing temperature; and were calculated at 20 degrees C as 0.069 g MLVSS/(g COD.d) and 3.265 g COD/g N, respectively. Carbon balance analysis indicated that the COD release rate (upsilon) at 30 degrees C was twice that at 13 degrees C. LEfSe analysis demonstrated that Desulfomicrobium, Desulfovibrio, and Meganema were abundant at low-temperature, while Simplicispira, Aquabacterium, and Acidovorax were enriched at high-temperature. Besides, carboxylesterase (PCL depolymerase) was more abundant at high-temperature, implying an association with a fast upsilon. Moreover, nar was enriched at low-temperature, while nir was depleted, which led to nitrite accumulation. These results provide reference for SPD design parameter estimation and/or optimal operation strategy.
Title: An enzyme inhibition-based lab-in-a-syringe device for point-of-need determination of pesticides Yang L, Wang J, Qu L, Liu Z, Jiang L Ref: Analyst, :, 2020 : PubMed
An enzyme inhibition-based lab-in-a-syringe (EI-LIS) device was developed by integrating a 1-naphthol-linked bi-enzymatic reaction (sensor core) into the LIS (sensor device) for point-of-need monitoring of pesticide residues. The integration relies on the rational design of two reaction pads. The conjugate pad is a polyester fiber membrane loaded with plant-esterase, an alternative to acetylcholinesterase. Besides pesticide capture, plant-esterase also mediates the hydrolysis of 1-naphthyl acetate, generating 1-naphthol. The detection pad is an agarose gel entrapping oxidized 3,3',5,5'-tetramethylbenzidine (oxTMB) from Fe(iii) meso-tetra(N-methyl-4-pyridyl) porphyrin (FeTMPyP4)-catalyzed TMB oxidation. Both pads were embedded into their cartridges and then connected to a syringe. Under syringe pumping, 1-naphthol vertically flowed from the conjugate to the detection cartridge, linking the two pads. If plant-esterase was intact, 1-naphthol would reduce oxTMB, causing a color change of the detection pad from blue to colorless. If the plant-esterase activity was inhibited by pesticides, less 1-naphthol was produced, and the blue color of the detection pad would be partially or wholly retained. The deeper the blue color, the greater the pesticide concentration. This chromogenic pattern is responsible for a highly sensitive readout (detection limits of dichlorvos: 0.1 nM with the naked eye and 0.07 nM with a spectrometer).
Humans and rodents with Comparative Gene Identification-58 (CGI-58) mutations manifest nonalcoholic fatty liver disease (NAFLD). Here we show that liver CGI-58 knockout (LivKO) mice fed a Western diet rapidly develop advanced NAFLD, including nonalcoholic steatohepatitis (NASH) and hepatic fibrosis. After 14 weeks of diet challenge, starting at 6 weeks of age, LivKO mice showed increased inflammatory cell infiltration and proinflammatory gene expression in the liver, which was associated with elevated plasma levels of aminotransferases. Hepatic ductular reactions, pericellular fibrosis, and bridging fibrosis were observed only in the LivKO mice. Consistently, the KO mice had a significant increase in hepatic mRNAs for fibrogenic genes. In addition, LivKO mice displayed massive accumulation of lipid droplets (LDs) in hepatocytes. LDs were also observed in the cholangiocytes of the LivKO mice, but not the floxed controls. Four of the five LD coat proteins, including perilipins 2, 3, 4, and 5, were increased in the CGI-58 KO liver. CRISPR/Cas9-mediated knockout of CGI-58 in Huh7 human hepatoma cells induced LD deposition and perilipin expression, suggesting a cell autonomous effect. Our findings establish the Western diet-fed LivKO mice as an animal model of NASH and hepatic fibrosis. These animals may facilitate preclinical screening of therapeutic agents that counter against NAFLD progression.
Title: Effects of Lecanicillium lecanii strain JMC-01 on the physiology, biochemistry, and mortality of Bemisia tabaci Q-biotype nymphs Xie T, Jiang L, Li J, Hong B, Wang X, Jia Y Ref: PeerJ, 7:e7690, 2019 : PubMed
Background: Lecanicillium lecanii is an entomopathogenic fungi, which was isolated from insects suffering from disease. Now, it is an effective bio-control resource that can control agricultural pests such as whitefly and aphids. There are many studies on the control of various agricultural pests by L. lecanii, but no report on its control of Bemisia tabaci biotype-Q exists. In this work, we studied the susceptibility of B. tabaci Q-biotype (from Ningxia, China) to L. lecanii JMC-01 in terms of nymph mortality and the changes in detoxifying protective enzymes activities. Methods: B. tabaci nymphs were exposed to L. lecanii JMC-01 conidia by immersion with the host culture. Mortality was assessed daily for all nymph stages. The detoxifying and protective enzyme activity changes, weight changes, and fat, and water contents of the nymphs were determined spectrophotometrically. Results: All instars of B. tabaci died after being infested with 1 x 10(8) conidia/mL. The 2nd-instar nymphs were the most susceptible, followed by the 3rd-instar nymphs. The corrected cumulative mortality of the 2nd- and 3rd-instar nymphs was 82.22% and 75.55%, respectively. The levels of detoxifying and protective enzymes initially increased and then decreased. The highest activities of carboxylesterase, acetylcholinesterase, peroxidase, and catalase occurred on the 3rd day, reaching 10.5, 0.32, 20, and 6.3 U/mg prot, respectively. These levels were 2.2-, 4.3-, 2.4-, and 1.4-fold the control levels, respectively. The highest activities of glutathione-S transferase and superoxide dismutase on the 2nd day were, respectively, 64 and 43.5 U/mg prot. These levels were, respectively, 2.7 and 1.1-fold that of the control level. The water and fat content in the infected B. tabaci nymphs decreased and differed significantly from the control levels. The weight increased continuously in the first 24 h, decreasing thereafter. At 72 h, the infestation level was about 0.78-fold that of the control level. Conclusions: The studied L. lecanii JMC-01 strain is pathogenic to the B. tabaci Q-biotype. This strain interferes with the normal functioning of detoxifying and protective enzymes, and is also involved in the disruption of normal physiological metabolism in B. tabaci.
Title: Tissue bioconcentration and effects of fluoxetine in zebrafish (Danio rerio) and red crucian cap (Carassius auratus) after short-term and long-term exposure Pan C, Yang M, Xu H, Xu B, Jiang L, Wu M Ref: Chemosphere, 205:8, 2018 : PubMed
Fluoxetion (FLU) is an antidepressant pharmaceutical most commonly detected in the aquatic environment. The present study aims to elucidate the tissue accumulation and effects of FLU using two different fish models. First, the multiple effects and the FLU levels in fish, were examined in zebrafish (Danio rerio) embryos exposed to FLU concentrations (0, 0.1, 1, 10, 100, 1000mug/L) from 4h post-fertilization (hpf) until 120 hpf. Exposure to FLU accelerated heart rates, postponed hatching time, and increased swimming speed of fish. A dynamic response of acetylcholinesterase (AChE) activity was also displayed in the fish. Second, a 30-day exposure experiment using red crucian carp (Carassius auratus) was performed, and it found that the concentration of FLU in fish organs increased with increasing water concentrations, but the highest FLU bioconcentration was present in the lowest FLU exposure group (0.1mug/L). Finally, 6 days of exposure to 0.1mug/L of FLU followed by a 6-day clearance experiment was performed with both adult zebrafish and red crucian carp. The FLU levels in different fish organs increased as exposure time increased, but they sharply declined following the 6-day clearance. Correspondingly, the changes in brain AChE activity and in antioxidant parameters in the liver were consistent with the FLU levels in the fish organs. Our study provides fundamental data on the tissue accumulation and concentration-dependent effects in fish exposed to fluoxetine.
OBJECTIVES: Familial hypercholesterolemia (FH) is an autosomal dominant disorder of cholesterol metabolism. Three recognized genes (LDLR, APOB and PCSK9) present in only 20-30% of patients with possible FH cases. Additional FH-causing genes need to be explored. The present study found an isolated gene change, sEH R287Q, in a core family of FH. In this study, we aimed to investigate the roles of R287Q on sEH expression and on LDLR expression, LDL binding to LDLR and LDL internalization. MATERIALS AND METHODS: 167 lipid-related genes of a core FH family were sequenced using a gene-capture chip. Through carrier dependent protein expression, the expression level (western blot), hydrolase activity (fluorescent chemistry) and intracellular localization (immunofluorescence and Confocal Laser Scanning Microscope) of recombinant sEH R287Q in cultured BEL-7402 cells were conducted. The effect of wild type and R287Q of sEH on LDLR expression, LDL binding to LDLR and LDL internalization were also conducted through Flow Cytometry. RESULTS: sEH R287Q was the only gene changes among 167 lipid-related genes in the FH core family. Both expression level and hydrolase activity of recombinant sEH R287Q in cultured cells were significantly declined compared with that of the wild type sEH. sEH R287Q also decreased the binding of LDL to LDLR and LDL internalization and had no effect on cell-surface LDLR protein level. CONCLUSION: Our results suggest that sEH R287Q may have a role in the elevation of blood LDL in FH. The exactly role of sEH R287Q on FH deserves further study.
Title: Preparation of Margarine Stock Rich in Naturally Bioactive Components by Enzymatic Interesterification Yu D, Qi X, Jiang Y, Zou D, Wang L, Jiang L, Qin L Ref: J Oleo Sci, 67:29, 2018 : PubMed
Fully hydrogenated expanded press soybean oil (FHEPSO) rich in naturally bioactive components was prepared using Palladium on Carbon (Pd/C) catalyst. Interesterified fat was prepared from binary blends of FHEPSO and cold press corn oil (CPCO) with FHEPSO/CPCO mass ratios of 50:50, 40:60 and 30:70. Lipozyme RM IM (6 wt% of total substrate) was used in a supercritical CO(2) system to catalyze the transesterification. The fatty acid compositions had no significant changes in the fats before and after interesterification, and trans-fatty acid (TFA) was not detected. The fatty acid compositions within triacylglycerol (TAG) were rearranged, and the amounts of trisaturated and triunsaturated TAG decreased, whereas that of mixed TAG increased as a result of interesterification. The enzymatic interesterified fats (EIEF) had a lower solid fat content (SFC), broader melting and plasticity ranges compared to the noninteresterified blend (NIB). According to X-ray diffraction (XRD), the predominant crystal form had changed from beta to beta'. EIEF contained 0.33-0.51 g/100 g phospholipids, 88.6-105.6 mg/100 g total tocopherols, and 916-1053 mg/100 g total phytosterols, which could confer health benefits. The results indicated that EIEF may have a potential use in trans-free margarine stock preparation.
The major mechanism of antibody-mediated neutralization of the Middle East respiratory syndrome coronavirus (MERS-CoV) involves competition with the cellular receptor dipeptidyl peptidase 4 (DPP4) for binding to the receptor-binding domain (RBD) of the spike (S) glycoprotein. Here, we report a unique epitope and unusual neutralizing mechanism of the isolated human antibody MERS-4. Structurally, MERS-4 approached the RBD from the outside of the RBD-DPP4 binding interface. Such binding resulted in the folding of the beta5-beta6 loop toward a shallow groove on the RBD interface critical for accommodating DPP4. The key residues for binding are identified through site-directed mutagenesis. Structural modeling revealed that MERS-4 binds to RBD only in the "up" position in the S trimer. Furthermore, MERS-4 demonstrated synergy with several reported antibodies. These results indicate that MERS-4 neutralizes MERS-CoV by indirect rather than direct competition with DPP4. This mechanism provides a valuable addition for the combined use of antibodies against MERS-CoV infection.
Strigolactones (SLs) are the latest confirmed phytohormones that regulate shoot branching by inhibiting bud outgrowth in higher plants. Perception of SLs depends on a novel mechanism employing an enzyme-receptor DWARF14 (D14) that hydrolyzes SLs and becomes covalently modified. This stimulates the interaction between D14 and D3, leading to the ubiquitination and degradation of the transcriptional repressor protein D53. However, the regulation of SL perception in rice remains elusive. In this study, we provide evidences that D14 is ubiquitinated after SL treatment and degraded through the 26S proteasome system. The Lys280 site of the D14 amino acid sequence was important for SL-induced D14 degradation, but did not change the subcellular localization of D14 nor disturbed the interaction between D14 and D3, nor D53 degradation. Biochemical and genetic analysis indicated that the key amino acids in the catalytic center of D14 were essential for D14 degradation. We further showed that D14 degradation is dependent on D3 and is tightly correlated with protein levels of D53. These findings revealed that D14 degradation takes place following D53 degradation and functions as an important feedback regulation mechanism of SL perception in rice.
Recent developments in the generation of neuronal population-specific, genetically modified mouse lines have allowed precise identification and selective stimulation of cholinergic neurons in vivo. Although considerably less laborious than studies conducted with post hoc identification of cholinergic neurons by immunostaining, it is not known whether the genetically based labeling procedures that permit in vivo identification are electrophysiologically benign. In this study, we use mice carrying a bacterial artificial chromosome transgene that drives expression of a tau-green fluorescent fusion protein specifically in cholinergic neurons. This allowed us to visualize basal forebrain cholinergic neurons in acute slice preparations. Using whole cell, patch clamp electrophysiological recording in acute brain slices, here we present original data about the basic electrical properties of these genetically tagged cholinergic neurons including firing rate, resting membrane potential, rheobase, and various characteristics of their action potentials and after-hyperpolarization potentials. The basic electrical properties are compared (i) with non-cholinergic neurons in the same brain regions; (ii) in cholinergic neurons between immature animals and young adults; and (iii) with cholinergic neurons that are expressing light-sensitive channels. Our conclusions based on these data are (i) cholinergic neurons are less excitable then their non-cholinergic neighbors, (ii) the basic properties of cholinergic neurons do not significantly change between adolescence and young adulthood and (iii) these properties are not significantly affected by chronic expression of the excitatory opsin, oChIEF. This is an article for the special issue XVth International Symposium on Cholinergic Mechanisms.
Being a neurodegenerative disorder, Alzheimer's disease (AD) is the one of the most terrible diseases. And acetylcholinesterase (AChE) is considered as an important target for treating AD. Acetylcholinesterase inhibitors (AChEI) are considered to be one of the effective drugs for the treatment of AD. The aim of this study is to find a novel potential AChEI as a drug for the treatment of AD. In this study, instead of using the synthetic compounds, we used those extracted from plants to investigate the interaction between floribundiquinone B (FB) and AChE by means of both the experimental approach such as fluorescence spectra, ultraviolet-visible (UV-vis) absorption spectrometry, circular dichroism (CD) and the theoretical approaches such as molecular docking. The findings reported here have provided many useful clues and hints for designing more effective and less toxic drugs against Alzheimer's disease.
We examined the contribution of endogenous cholinergic signaling to the acquisition and extinction of fear- related memory by optogenetic regulation of cholinergic input to the basal lateral amygdala (BLA). Stimulation of cholinergic terminal fields within the BLA in awake-behaving mice during training in a cued fear-conditioning paradigm slowed the extinction of learned fear as assayed by multi-day retention of extinction learning. Inhibition of cholinergic activity during training reduced the acquisition of learned fear behaviors. Circuit mechanisms underlying the behavioral effects of cholinergic signaling in the BLA were assessed by in vivo and ex vivo electrophysiological recording. Photostimulation of endogenous cholinergic input (1) enhances firing of putative BLA principal neurons through activation of acetylcholine receptors (AChRs), (2) enhances glutamatergic synaptic transmission in the BLA, and (3) induces LTP of cortical-amygdala circuits. These studies support an essential role of cholinergic modulation of BLA circuits in the inscription and retention of fear memories.
Strigolactones are important rhizosphere signals that act as phytohormones and have multiple functions, including modulation of lateral root (LR) development. Here, we show that treatment with the strigolactone analog GR24 did not affect LR initiation, but negatively influenced LR priming and emergence, the latter especially near the root-shoot junction. The cytokinin module ARABIDOPSIS HISTIDINE KINASE3 (AHK3)/ARABIDOPSIS RESPONSE REGULATOR1 (ARR1)/ARR12 was found to interact with the GR24-dependent reduction in LR development, because mutants in this pathway rendered LR development insensitive to GR24. Additionally, pharmacological analyses, mutant analyses, and gene expression analyses indicated that the affected polar auxin transport stream in mutants of the AHK3/ARR1/ARR12 module could be the underlying cause. Altogether, the data reveal that the GR24 effect on LR development depends on the hormonal landscape that results from the intimate connection with auxins and cytokinins, two main players in LR development.
Title: Comment on A bacterium that degrades and assimilates poly(ethylene terephthalate) Yang Y, Yang J, Jiang L Ref: Science, 353:759, 2016 : PubMed
Yoshida et al (Report, 11 March 2016, p. 1196) reported that the bacterium Ideonella sakaiensis 201-F6 can degrade and assimilate poly(ethylene terephthalate) (PET). However, the authors exaggerated degradation efficiency using a low-crystallinity PET and presented no straightforward experiments to verify depolymerization and assimilation of PET. Thus, the authors' conclusions are rather misleading.
The large yellow croaker Larimichthys crocea (L. crocea) is one of the most economically important marine fish in China and East Asian countries. It also exhibits peculiar behavioral and physiological characteristics, especially sensitive to various environmental stresses, such as hypoxia and air exposure. These traits may render L. crocea a good model for investigating the response mechanisms to environmental stress. To understand the molecular and genetic mechanisms underlying the adaptation and response of L. crocea to environmental stress, we sequenced and assembled the genome of L. crocea using a bacterial artificial chromosome and whole-genome shotgun hierarchical strategy. The final genome assembly was 679 Mb, with a contig N50 of 63.11 kb and a scaffold N50 of 1.03 Mb, containing 25,401 protein-coding genes. Gene families underlying adaptive behaviours, such as vision-related crystallins, olfactory receptors, and auditory sense-related genes, were significantly expanded in the genome of L. crocea relative to those of other vertebrates. Transcriptome analyses of the hypoxia-exposed L. crocea brain revealed new aspects of neuro-endocrine-immune/metabolism regulatory networks that may help the fish to avoid cerebral inflammatory injury and maintain energy balance under hypoxia. Proteomics data demonstrate that skin mucus of the air-exposed L. crocea had a complex composition, with an unexpectedly high number of proteins (3,209), suggesting its multiple protective mechanisms involved in antioxidant functions, oxygen transport, immune defence, and osmotic and ionic regulation. Our results reveal the molecular and genetic basis of fish adaptation and response to hypoxia and air exposure. The data generated by this study will provide valuable resources for the genetic improvement of stress resistance and yield potential in L. crocea.
Title: Strigolactone Signaling in Arabidopsis Regulates Shoot Development by Targeting D53-Like SMXL Repressor Proteins for Ubiquitination and Degradation Wang L, Wang B, Jiang L, Liu X, Li X, Lu Z, Meng X, Wang Y, Smith SM, Li J Ref: Plant Cell, 27:3128, 2015 : PubMed
Strigolactones (SLs) are carotenoid-derived phytohormones that control many aspects of plant development, including shoot branching, leaf shape, stem secondary thickening, and lateral root growth. In rice (Oryza sativa), SL signaling requires the degradation of DWARF53 (D53), mediated by a complex including D14 and D3, but in Arabidopsis thaliana, the components and mechanism of SL signaling involving the D3 ortholog MORE AXILLARY GROWTH2 (MAX2) are unknown. Here, we show that SL-dependent regulation of shoot branching in Arabidopsis requires three D53-like proteins, SUPPRESSOR OF MORE AXILLARY GROWTH2-LIKE6 (SMXL6), SMXL7, and SMXL8. The smxl6 smxl7 smxl8 triple mutant suppresses the highly branched phenotypes of max2 and the SL-deficient mutant max3. Overexpression of a mutant form of SMXL6 that is resistant to SL-induced ubiquitination and degradation enhances shoot branching. Exogenous application of the SL analog rac-GR24 causes ubiquitination and degradation of SMXL6, 7, and 8; this requires D14 and MAX2. D53-like SMXLs form complexes with MAX2 and TOPLESS-RELATED PROTEIN2 (TPR2) and interact with D14 in a GR24-responsive manner. Furthermore, D53-like SMXLs exhibit TPR2-dependent transcriptional repression activity and repress the expression of BRANCHED1. Our findings reveal that in Arabidopsis, D53-like SMXLs act with TPR2 to repress transcription and so allow lateral bud outgrowth but that SL-induced degradation of D53-like proteins activates transcription to inhibit outgrowth.
Title: Conversion of inhibition biosensing to substrate-like biosensing for quinalphos selective detection Yang L, Han J, Liu W, Li J, Jiang L Ref: Analytical Chemistry, 87:5270, 2015 : PubMed
Since all of the organophosphorus pesticides (OPP) inhibit the cholinesterases with a common mechanism, it is still challenging to detect OPP selectively with inhibition-based biosensors. This study focuses on the conversion of a typical inhibition biosensing to a selective substrate-like biosensing. The interaction of quinalphos with plant-esterase involves not only a decrease in enzyme activity but also a heterolytic bond cleavage of quinalphos. The leaving group eliminated from quinalphos is an ideal biomarker due to its specificity in most OPP. Thus, using 2-hydroxyquinoxaline (HQO), the leaving group of quinalphos, as the biomarker and meso-tetra (4-sulfonatophenyl) porphine (TPPS4) as an optical probe, quinalphos can be selectively detected. The molecular recognition between TPPS4 and HQO leads to a considerable sensitivity of the detection. The spectral responses of TPPS4 show a linear dependence on quinalphos concentration in the presence of plant-esterase within the 0.01-1 mg kg(-1) range. The detection limit is 0.01 mg kg(-1), well below the maximum residue limits (MRLs) defined by European Union (0.05 mg kg(-1)) and China (0.2 mg kg(-1)).
Title: Biodegradation and Mineralization of Polystyrene by Plastic-Eating Mealworms: Part 1. Chemical and Physical Characterization and Isotopic Tests Yang Y, Yang J, Wu WM, Zhao J, Song Y, Gao L, Yang R, Jiang L Ref: Environ Sci Technol, 49:12080, 2015 : PubMed
Polystyrene (PS) is generally considered to be durable and resistant to biodegradation. Mealworms (the larvae of Tenebrio molitor Linnaeus) from different sources chew and eat Styrofoam, a common PS product. The Styrofoam was efficiently degraded in the larval gut within a retention time of less than 24 h. Fed with Styrofoam as the sole diet, the larvae lived as well as those fed with a normal diet (bran) over a period of 1 month. The analysis of fecula egested from Styrofoam-feeding larvae, using gel permeation chromatography (GPC), solid-state (13)C cross-polarization/magic angle spinning nuclear magnetic resonance (CP/MAS NMR) spectroscopy, and thermogravimetric Fourier transform infrared (TG-FTIR) spectroscopy, substantiated that cleavage/depolymerization of long-chain PS molecules and the formation of depolymerized metabolites occurred in the larval gut. Within a 16 day test period, 47.7% of the ingested Styrofoam carbon was converted into CO2 and the residue (ca. 49.2%) was egested as fecula with a limited fraction incorporated into biomass (ca. 0.5%). Tests with alpha (13)C- or beta (13)C-labeled PS confirmed that the (13)C-labeled PS was mineralized to (13)CO2 and incorporated into lipids. The discovery of the rapid biodegradation of PS in the larval gut reveals a new fate for plastic waste in the environment.
Title: Identification of differentially expressed proteins and phosphorylated proteins in rice seedlings in response to strigolactone treatment Chen F, Jiang L, Zheng J, Huang R, Wang H, Hong Z, Huang Y Ref: PLoS ONE, 9:e93947, 2014 : PubMed
Strigolactones (SLs) are recently identified plant hormones that inhibit shoot branching and control various aspects of plant growth, development and interaction with parasites. Previous studies have shown that plant D10 protein is a carotenoid cleavage dioxygenase that functions in SL biosynthesis. In this work, we used an allelic SL-deficient d10 mutant XJC of rice (Oryza sativa L. spp. indica) to investigate proteins that were responsive to SL treatment. When grown in darkness, d10 mutant seedlings exhibited elongated mesocotyl that could be rescued by exogenous application of SLs. Soluble protein extracts were prepared from d10 mutant seedlings grown in darkness in the presence of GR24, a synthetic SL analog. Soluble proteins were separated on two-dimensional gels and subjected to proteomic analysis. Proteins that were expressed differentially and phosphoproteins whose phosphorylation status changed in response to GR24 treatment were identified. Eight proteins were found to be induced or down-regulated by GR24, and a different set of 8 phosphoproteins were shown to change their phosphorylation intensities in the dark-grown d10 seedlings in response to GR24 treatment. Analysis of these proteins revealed that they are important enzymes of the carbohydrate and amino acid metabolic pathways and key components of the cellular energy generation machinery. These proteins may represent potential targets of the SL signaling pathway. This study provides new insight into the complex and negative regulatory mechanism by which SLs control shoot branching and plant development.
Title: Tacrine induces apoptosis through lysosome- and mitochondria-dependent pathway in HepG2 cells Gao C, Ding Y, Zhong L, Jiang L, Geng C, Yao X, Cao J Ref: Toxicol In Vitro, 28:667, 2014 : PubMed
Tacrine (THA) is a competitive inhibitor of cholinesterase. Administration of THA for the treatment of Alzheimer's disease results in a reversible hepatotoxicity in 30-50% of patients, as indicated by elevated alanine aminotransferase levels. However, the intracellular mechanisms have not yet been elucidated. In our previous study, we found that THA induced cytotoxicity and mitochondria dysfunction by ROS generation and 8-OHdG formation in mitochondrial DNA in HepG2 cells. In this study, the mechanism underlying was further investigated. Our results demonstrated that THA induced dose-dependent apoptosis with cytochrome c release and activation of caspase-3. THA-induced apoptosis was inhibited by treating cells with a ROS inhibitor, YCG063. In addition, we observed that THA led to an early lysosomal membrane permeabilization and release of cathepsin B. Pretreatment with CA-074Me, a specific cathepsin B inhibitor resulted in a significant but not complete decrease in tacrine-induced apoptosis. These data suggest that tacrine-induced cell apoptosis involves both mitochondrial damage and lysosomal membrane destabilization, and ROS is the critical factor that integrates tacrine-induced mitochondrial and lysosomal death pathways.
Title: Draft Genome Sequence of Defluviimonas indica Strain 20V17T, Isolated from a Deep-Sea Hydrothermal Vent Environment in the Southwest Indian Ocean Jiang L, Long M, Shao Z Ref: Genome Announc, 2:, 2014 : PubMed
Here, we present the draft genome sequence of Defluviimonas indica 20V17(T), which was isolated from a deep-sea hydrothermal vent chimney sample in the southwest Indian Ocean. The draft genome sequence contains 4,268,338 bp, with a G+C content of 66.33%.
Molecular manipulations and targeted pharmacological studies provide a compelling picture of which nicotinic receptor subtypes are where in the central nervous system (CNS) and what happens if one activates or deletes them. However, understanding the physiological contribution of nicotinic receptors to endogenous acetylcholine (ACh) signaling in the CNS has proven a more difficult problem to solve. In this review, we provide a synopsis of the literature on the use of optogenetic approaches to control the excitability of cholinergic neurons and to examine the role of CNS nicotinic ACh receptors (nAChRs). As is often the case, this relatively new technology has answered some questions and raised others. Overall, we believe that optogenetic manipulation of cholinergic excitability in combination with some rigorous pharmacology will ultimately advance our understanding of the many functions of nAChRs in the brain.
Title: An example of enzymatic promiscuity: the Baylis-Hillman reaction catalyzed by a biotin esterase (BioH) from Escherichia coli Jiang L, Yu HW Ref: Biotechnol Lett, 36:99, 2014 : PubMed
Ten lipases and esterases have been examined to catalyse the reaction between p-nitrobenzaldehyde and methyl vinyl ketone, the Baylis-Hillman reaction, to form 3-[hydroxyl-(4-nitrophenyl)-methyl]-but-3-en-2-one. Among these enzymes, Escherichia coli BioH esterase had the best activity. Optimal conditions for this reaction were: 0.1 mmol aldehyde, 0.1 mmol activated alkene, 30 mg E. coli BioH, 1 ml acetonitrile at 30 degrees C for 96 h. In addition to the named substrates, four other aldehydes and three activated alkenes were also investigated to determine the substrate range of the enzyme. The structures of nine products were confirmed by NMR and yields of the corresponding products ranged from 21 to 46 %.
In the wake of recent progress of high throughput transcriptome profiling technologies, extensive housekeeping gene mining has been conducted in humans. However, very few studies have been reported in maize (Zea mays L.), an important crop plant, and none were conducted on a genome -wide level. In this study, we surveyed housekeeping genes throughout the maize transcriptome using RNA-seq and microarray techniques, and validated the housekeeping profile with quantitative polymerase chain reaction (qPCR) under a series of conditions including different genotypes and nitrogen supplies. Seven microarray datasets and two RNA-seq libraries representing 40 genotypes at more than 20 developmental stages were selected to screen for commonly expressed genes. A total of 1,661 genes showed constitutive expression in both microarray and RNA-seq datasets, serving as our starting housekeeping gene candidates. To determine for stably expressed housekeeping genes, NormFinder was used to select the top 20 % invariable genes to be the more likely candidates, which resulted in 48 and 489 entries from microarray and RNA-seq data, respectively. Among them, nine genes (2OG-Fe, CDK, DPP9, DUF, NAC, RPN, SGT1, UPF1 and a hypothetical protein coding gene) were expressed in all 40 maize diverse genotypes tested covering 16 tissues at more than 20 developmental stages under normal and stress conditions, implying these as being the most reliable reference genes. qPCR analysis confirmed the stable expression of selected reference gene candidates compared to two widely used housekeeping genes. All the reference gene candidates showed higher invariability than ACT and GAPDH. The hypothetical protein coding gene exhibited the most stable expression across 26 maize lines with different nitrogen treatments with qPCR, followed by CDK encoding the cyclin-dependent kinase. As the first study to systematically screen for housekeeping genes in maize, we identified candidates by examining the transcriptome atlas generated from RNA-seq and microarray technologies. The nine top-ranked qPCR-validated novel housekeeping genes provide a valuable resource of reference genes for maize gene expression analysis.
Title: A novel cephalosporin deacetylating acetyl xylan esterase from Bacillus subtilis with high activity toward cephalosporin C and 7-aminocephalosporanic acid Tian Q, Song P, Jiang L, Li S, Huang H Ref: Applied Microbiology & Biotechnology, 98:2081, 2014 : PubMed
A cephalosporin deacetylating acetyl xylan esterase was cloned from the genomic DNA of Bacillus subtilis CICC 20034 and functionally expressed in Escherichia coli. Its gene contained an open reading frame of 957 bp encoding 318 amino acids with a calculated mass of 35,607 Da, and it displayed significant identity to acetyl xylan esterases from Bacillus sp. 916, B. subtilis 168, and Bacillus pumilus Cect5072. The enzyme was a native homohexamer but a trimer under the condition of 1% sodium dodecyl sulfate (SDS); both forms were active and could transit to each other by incubating in or removing SDS. The enzyme belongs to carbohydrate esterase family 7 and had a double specificity on both the acetylated oligosaccharide and cephalosporin C (CPC) and 7-aminocephalosporanic acid (7-ACA). The activity of this purified enzyme toward CPC and 7-ACA was highest among all the acetyl xylan esterase from CE family 7, which were 484 and 888 U/mg, respectively, and endowed itself with great industrial interest on semi-synthetic beta-lactam antibiotics. The optimum pH of the purified enzyme was 8.0, and the optimum temperature was 50 degrees C, and the enzyme had high thermal stability, broad range of pH tolerance, and extremely organic solvent tolerance.
Title: Evidence of polyethylene biodegradation by bacterial strains from the guts of plastic-eating waxworms Yang J, Yang Y, Wu WM, Zhao J, Jiang L Ref: Environ Sci Technol, 48:13776, 2014 : PubMed
Polyethylene (PE) has been considered nonbiodegradable for decades. Although the biodegradation of PE by bacterial cultures has been occasionally described, valid evidence of PE biodegradation has remained limited in the literature. We found that waxworms, or Indian mealmoths (the larvae of Plodia interpunctella), were capable of chewing and eating PE films. Two bacterial strains capable of degrading PE were isolated from this worm's gut, Enterobacter asburiae YT1 and Bacillus sp. YP1. Over a 28-day incubation period of the two strains on PE films, viable biofilms formed, and the PE films' hydrophobicity decreased. Obvious damage, including pits and cavities (0.3-0.4 mum in depth), was observed on the surfaces of the PE films using scanning electron microscopy (SEM) and atomic force microscopy (AFM). The formation of carbonyl groups was verified using X-ray photoelectron spectroscopy (XPS) and microattenuated total reflectance/Fourier transform infrared (micro-ATR/FTIR) imaging microscope. Suspension cultures of YT1 and YP1 (10(8) cells/mL) were able to degrade approximately 6.1 +/- 0.3% and 10.7 +/- 0.2% of the PE films (100 mg), respectively, over a 60-day incubation period. The molecular weights of the residual PE films were lower, and the release of 12 water-soluble daughter products was also detected. The results demonstrated the presence of PE-degrading bacteria in the guts of waxworms and provided promising evidence for the biodegradation of PE in the environment.
Title: Enzymatic resolution of ibuprofen in an organic solvent under ultrasound irradiation Zhao D, Yue H, Chen G, Jiang L, Zhang H, Wang Z, Liu G Ref: Biotechnol Appl Biochem, 61:655, 2014 : PubMed
Ultrasound has been successfully adopted to improve the biocatalytic properties of APE1547 (a novel esterase from the archaeon Aeropyrum pernix K1) in the resolution of ibuprofen. After optimizing the conditions (ultrasound power, 200 W; temperature, 35 degrees C), the best biocatalytic performance of APE1547 (enzyme activity, 5.39 micromol/H/mg; E value, 130.8) was obtained. Compared with the conventional reaction in an orbital shaker, the enzyme activity was significantly enhanced about 90-fold, and the enantioselectivity was enhanced about fourfold after an ultrasound. The results of scanning electron microscopy clearly indicated that the activation effect of ultrasound on APE1547 originated mainly in the morphological change of the enzyme powder. Both lower particle size and conformational change of APE1547 under ultrasound might be helpful to enhance the enantioselectivity. In addition, APE1547 kept its best performance under the low-power ultrasound for at least five reaction cycles.
Title: Type III neuregulin 1 is required for multiple forms of excitatory synaptic plasticity of mouse cortico-amygdala circuits Jiang L, Emmetsberger J, Talmage DA, Role LW Ref: Journal of Neuroscience, 33:9655, 2013 : PubMed
The amygdala plays an important role in the formation and storage of memories associated with emotional events. The cortical glutamatergic inputs onto pyramidal neurons in the basolateral nucleus of the amygdala (BLA) contribute to this process. As the interaction between neuregulin 1 (Nrg1) and its ErbB receptors has been implicated in the pathological mechanisms of schizophrenia, loss of Nrg1 may disrupt cortical-amygdala neural circuits, resulting in altered processing of salient memories. Here we show that Nrg1 is critical in multiple forms of plasticity of cortical projections to pyramidal neurons of the BLA. The miniature EPSCs in Nrg1 heterozygous animals have a faster time constant of decay and evoked synaptic currents have a smaller NMDA/AMPA ratio than those recorded in wild-type (WT) littermates. Both high-frequency electrical stimulation of cortical inputs and theta burst stimulation combined with nicotine exposure results in long-lasting potentiation in WT animals. However, the same manipulations have little to no effect on glutamatergic synaptic plasticity in the BLA from Nrg1 heterozygous mice. Comparison of WT, Nrg1 heterozygous animals and alpha7 nicotinic receptor heterozygous mice reveals that the sustained phase of potentiation of glutamatergic transmission after theta burst stimulation with or without nicotine only occurs in the WT mice. Together, these findings support the idea that type III Nrg1 is essential to multiple aspects of the modulation of excitatory plasticity at cortical-BLA synapses.
Strigolactones (SLs) are a group of newly identified plant hormones that control plant shoot branching. SL signalling requires the hormone-dependent interaction of DWARF 14 (D14), a probable candidate SL receptor, with DWARF 3 (D3), an F-box component of the Skp-Cullin-F-box (SCF) E3 ubiquitin ligase complex. Here we report the characterization of a dominant SL-insensitive rice (Oryza sativa) mutant dwarf 53 (d53) and the cloning of D53, which encodes a substrate of the SCF(D3) ubiquitination complex and functions as a repressor of SL signalling. Treatments with GR24, a synthetic SL analogue, cause D53 degradation via the proteasome in a manner that requires D14 and the SCF(D3) ubiquitin ligase, whereas the dominant form of D53 is resistant to SL-mediated degradation. Moreover, D53 can interact with transcriptional co-repressors known as TOPLESS-RELATED PROTEINS. Our results suggest a model of SL signalling that involves SL-dependent degradation of the D53 repressor mediated by the D14-D3 complex.
Title: Enhanced enantioselectivity of a carboxyl esterase from Rhodobacter sphaeroides by directed evolution Ma J, Wu L, Guo F, Gu J, Tang X, Jiang L, Liu J, Zhou J, Yu H Ref: Applied Microbiology & Biotechnology, 97:4897, 2013 : PubMed
The present work created an esterase variant from Rhodobacter sphaeroides (RspE) with enhanced selectivity in hydrolytic kinetic resolutions by directed evolution. A "model" substrate, methyl mandelate, was introduced in the high-throughput screening procedure. E values of a variant CH (Asn62Cys/Leu145His) for six different esters were 10-83, which were a relative improvement compared to 2-20 for the wild type. Our subsequent crystal structure interpretation and molecular dynamics simulations helped shed light on the source of enantioselectivity modified by directed evolution. Though mutations displayed no "direct" interaction with the substrate, they were hypothesized to strengthen the intramolecular interaction in the catalytic cavity of variant. Conformation analysis revealed that the enhanced enantioselectivity of variant CH for the seven substrates applied in this study was derived from the decrease in size of the substrate binding pocket.
Human monoglyceride lipase (MGL) is a recently identified lipase and very little is known about its regulation and function in cellular regulatory processes, particularly in context to human malignancy. In this study, we investigated the regulation and function of MGL in human cancer(s) and report that MGL expression was either absent or reduced in the majority of primary colorectal cancers. Immunohistochemical studies showed that reduction of MGL expression in the colorectal tumor tissues predominantly occurred in the cancerous epithelial cells. MGL was found to reside in the core surface of a cellular organelle named 'lipid body'. Furthermore, it was found to interact selectively with a number of phospholipids, including phosphatidic acid and phosphoinositol(3,4,5)P3, phosphoinositol(3,5)P2, phosphoinositol(3,4)P2 and several other phosphoinositides, and among all phosphoinositides analyzed, its interaction with PI(3,4,5)P3 was found to be the strongest. In addition, overexpression of MGL suppressed colony formation in tumor cell lines and knockdown of MGL resulted in increased Akt phosphorylation. Taken together, our results suggest that MGL plays a negative regulatory role in phosphatidylinositol-3 kinase/Akt signaling and tumor cell growth.
The spike glycoprotein (S) of recently identified Middle East respiratory syndrome coronavirus (MERS-CoV) targets the cellular receptor, dipeptidyl peptidase 4 (DPP4). Sequence comparison and modeling analysis have revealed a putative receptor-binding domain (RBD) on the viral spike, which mediates this interaction. We report the 3.0 A-resolution crystal structure of MERS-CoV RBD bound to the extracellular domain of human DPP4. Our results show that MERS-CoV RBD consists of a core and a receptor-binding subdomain. The receptor-binding subdomain interacts with DPP4 beta-propeller but not its intrinsic hydrolase domain. MERS-CoV RBD and related SARS-CoV RBD share a high degree of structural similarity in their core subdomains, but are notably divergent in the receptor-binding subdomain. Mutagenesis studies have identified several key residues in the receptor-binding subdomain that are critical for viral binding to DPP4 and entry into the target cell. The atomic details at the interface between MERS-CoV RBD and DPP4 provide structural understanding of the virus and receptor interaction, which can guide development of therapeutics and vaccines against MERS-CoV infection.
Strigolactones (SLs), a newly discovered class of carotenoid-derived phytohormones, are essential for developmental processes that shape plant architecture and interactions with parasitic weeds and symbiotic arbuscular mycorrhizal fungi. Despite the rapid progress in elucidating the SL biosynthetic pathway, the perception and signalling mechanisms of SL remain poorly understood. Here we show that DWARF 53 (D53) acts as a repressor of SL signalling and that SLs induce its degradation. We find that the rice (Oryza sativa) d53 mutant, which produces an exaggerated number of tillers compared to wild-type plants, is caused by a gain-of-function mutation and is insensitive to exogenous SL treatment. The D53 gene product shares predicted features with the class I Clp ATPase proteins and can form a complex with the alpha/beta hydrolase protein DWARF 14 (D14) and the F-box protein DWARF 3 (D3), two previously identified signalling components potentially responsible for SL perception. We demonstrate that, in a D14- and D3-dependent manner, SLs induce D53 degradation by the proteasome and abrogate its activity in promoting axillary bud outgrowth. Our combined genetic and biochemical data reveal that D53 acts as a repressor of the SL signalling pathway, whose hormone-induced degradation represents a key molecular link between SL perception and responses.
Title: Enhanced pesticide sensitivity of novel housefly acetylcholinesterases: a new tool for the detection of residual pesticide contamination Tan F, Wang L, Wang J, Wu X, Zhu H, Jiang L, Tao S, Zhao K, Yang Y, Tang X Ref: Bioprocess Biosyst Eng, 34:305, 2011 : PubMed
The full-length cDNA encoding an acetylcholinesterase (AChE) was cloned and sequenced from the housefly, Musca domestica, by reverse transcriptase-polymerase chain reaction (RT-PCR). Sequence analysis revealed that this 2,076 bp sequence encodes a mature protein of 612 amino acids (67 kDa) and a 79 residue signal peptide. The amino acid sequence shared 52.8-81.4% identity with the AChE proteins of other insects. The cDNA sequence, which lacked the signal peptide was inserted into the vector pPIC9K and then introduced into strain GS115 of the yeast Pichia pastoris. The recombinant AChE protein was then expressed in P. pastoris strain GS115 by methanol induction. Site-directed mutagenesis of the A262G, Y327F, Y327D and I374D residues, either singly or in combination, was performed by reverse PCR. These mutants improved the catalytic activity and sensitivity to the organophosphate and carbamate insecticides. Although the sensitivity of other mutants was slightly increased, the results still showed that the sensitivity of triple mutant, GDD (A262G/Y327D/I374D), enhanced remarkably as much as 16 times for methomyl, 14 times for both carbofuran and chlorpyrifos, and ten times for parathion-methyl, compared to that of the wild-type. The results strongly suggested that these residues are the key structural elements controlling AChE enzyme catalytic activity and sensitivity to inhibition by insecticides. The AChE enzyme obtained by this method could be used to detect the organophosphate and carbamate insecticide residues in fruits and vegetables, a characteristic of great potential research and industrial application.
Title: Functionalized ionic liquid modified mesoporous silica SBA-15: a novel, designable and efficient carrier for porcine pancreas lipase Zou B, Hu Y, Yu D, Jiang L, Liu W, Song P Ref: Colloids Surf B Biointerfaces, 88:93, 2011 : PubMed
A series of functionalized ionic liquid modified mesoporous silicas SBA-15 (FIL-SBA) were synthesized by modulating the loading and cation/anion ratio of the functionalized ionic liquid (FIL). The prepared materials FIL-SBA were used as a novel carrier system to immobilize porcine pancreas lipase (PPL). Enzymatic activity and reusability of the immobilized enzyme were investigated using the triacetin hydrolysis reaction. The combined advantages of the nano-sized pore diameter, large surface area and high pore volume of SBA-15, and the tunable properties of the FIL for enzymes immobilized in FIL-SBA gave a maximum improvement of 570% in relative activity, with 63% retention of initial activity after five cycles of use. Carriers and immobilized enzymes were characterized using nitrogen adsorption, small-angle X-ray diffraction (SXRD), Fourier transform infrared (FT-IR), elemental analysis, nuclear magnetic resonance (NMR), scanning and transmission electron microscopy (SEM and TEM). It was shown that the introduction of FIL influenced the catalytic behavior of PPL significantly by changing the structure and surface properties of the carriers.
Streptomyces bingchenggensis is a soil-dwelling bacterium producing the commercially important anthelmintic macrolide milbemycins. Besides milbemycins, the insecticidal polyether antibiotic nanchangmycin and some other antibiotics have also been isolated from this strain. Here we report the complete genome sequence of S. bingchenggensis. The availability of the genome sequence of S. bingchenggensis should enable us to understand the biosynthesis of these structurally intricate antibiotics better and facilitate rational improvement of this strain to increase their titers.
The pathogenic bacterium Bacillus anthracis has become the subject of intense study as a result of its use in a bioterrorism attack in the United States in September and October 2001. Previous studies suggested that B. anthracis Ames Ancestor, the original Ames fully virulent plasmid-containing isolate, was the ideal reference. This study describes the complete genome sequence of that original isolate, derived from a sample kept in cold storage since 1981.
Title: A new microdialysis-electrochemical device for in vivo simultaneous determination of acetylcholine and choline in rat brain treated with N-methyl-(R)-salsolinol Zhu W, An Y, Zheng J, Tang L, Zhang W, Jin L, Jiang L Ref: Biosensors & Bioelectronics, 24:3594, 2009 : PubMed
Acetylcholine (ACh) and choline (Ch) play a critical role in cholinergic neurotransmission and the abnormalities in their concentrations are related to several neural diseases. Therefore, the in vivo determination of ACh and Ch is important to the research on neurodegenerative disorders. In this work, electrochemical biosensors based on poly(m-(1,3)-phenylenediamine) (pmPD) and polytyramine (PTy) modified enzyme electrodes were fabricated. The electropolymerized pmPD polymer was used to exclude interfering substances and the PTy layer facilitated the immobilization of acetylcholinesterase (AChE) and choline oxidase (ChOx). Then, ACh/Ch sensor and Ch sensor were coupled with microdialysis to produce a novel device, which provides a sensitive and selective method for simultaneous determination of ACh and Ch. This method has detection limits of 63.0+/-3.4 nM for ACh and 25.0+/-1.2 nM for Ch. The integrated device was successfully applied to assessing the impact of endogenous neurotoxin N-methyl-(R)-salsolinol [1(R),2-dimethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, (R)-NMSal] on ACh and Ch concentration, which is of great benefit to understand the pathogenesis of Parkinson's disease.
Title: Facilitation of cortico-amygdala synapses by nicotine: activity-dependent modulation of glutamatergic transmission Jiang L, Role LW Ref: Journal of Neurophysiology, 99:1988, 2008 : PubMed
The basolateral nucleus of the amygdala (BLA) receives cholinergic innervation from the basal forebrain and nicotine, via activation of neuronal nicotinic acetylcholine receptors (nAChRs), can improve performance in amygdala-based learning tasks. We tested the hypothesis that acute and prenatal nicotine exposure modulates cortico-amygdala synaptic transmission. We found that low-dose, single-trial exposures to nicotine can elicit lasting facilitation, the extent of which is dependent on the level of stimulation of the cortical inputs to the BLA. In addition, sustained facilitation is ablated by prenatal exposure to nicotine. This study examined synaptic transmission in 238 patch-clamp recordings from BLA neurons in acute slice from mouse brain. Pharmacological studies in wild-type and nAChR subunit knock-out mice reveal that activation of presynaptic alpha 7, containing (alpha 7*) and non-alpha 7* nAChRs, facilitates glutamatergic transmission in an activity-dependent manner. Without prior stimulation, application of nicotine elicits modest and transient facilitation of glutamatergic postsynaptic currents (PSCs) in about 40% of BLA neurons. With low-frequency stimulation of cortical inputs nicotine elicits robust facilitation of transmission at about 60% of cortico-BLA synapses and synaptic strength remains elevated at about 40% of these connections for >15 min after nicotine washout. Following paired-pulse stimulation nicotine elicits long-lasting facilitation of glutamatergic transmission at about 70% of cortico-BLA connections. Nicotine reduces the threshold for activation of long-term potentiation of cortico-BLA synapses evoked by patterned stimulation. Prenatal exposure to nicotine reduced subsequent modulatory responses to acute nicotine application.
Title: Molecular mechanism of age-specific hepatic lipid accumulation in PPARalpha (+/-):LDLR (+/-) mice, an obese mouse model Li Y, Sugiyama E, Yokoyama S, Jiang L, Tanaka N, Aoyama T Ref: Lipids, 43:301, 2008 : PubMed
This study aimed to clarify the molecular mechanisms of age-specific hepatic lipid accumulation accompanying hyperinsulinemia in a peroxisome proliferator-activated receptor alpha (PPARalpha) (+/-):low-density lipoprotein receptor (LDLR) (+/-) mouse line. The hepatic fat content, protein amounts, and mRNA levels of genes involved in hepatic lipid metabolism were analyzed in 25-, 50-, 75- and 100-week-old mice. Severe fatty liver was confirmed only in 50- and 75-week-old mice. The hepatic expression of proteins that function in lipid transport and catabolism did not differ among the groups. In contrast, the mRNA levels and protein amounts of lipogenic enzymes, including acetyl-coenzyme A carboxylase-1, fatty acid synthase, and glycerol-3-phosphate acyltransferase, enhanced in the mice with fatty liver. Elevated mRNA and protein levels of lipoprotein lipase and fatty acid translocase, which are involved in hepatic lipid uptake, were also detected in mice with fatty liver. Moreover, both protein and mRNA levels of sterol regulatory element-binding protein-1 (SREBP-1), a transcription factor regulating lipid synthesis, had age-specific patterns similar to those of the proteins described above. Therefore, the age-specific fatty liver found in the PPARalpha (+/-):LDLR (+/-) mouse line is probably caused by age-specific expression of SREBP-1 and its downstream lipogenic genes, coordinated by the increased uptake of lipids. All of these factors might be affected by age-specific changes in serum insulin concentration.
Title: Lipase-catalyzed esterification of conjugated linoleic acid with L-carnitine in solvent-free system and acetonitrile Li Z, Yang D, Jiang L, Ji J, Ji H, Zeng X Ref: Bioprocess Biosyst Eng, 30:331, 2007 : PubMed
Lipase-catalyzed esterification of conjugated linoleic acid (CLA) with L-carnitine in solvent-free system and acetonitrile was studied. Three lipases (Novzym 435, Amamo AY30 and Amano AYS) have been assayed as suitable biocatalysts in the reaction. It was found that Amano AY30 was the most effective biocatalyst in both solvent-free system and acetonitrile. The conversion rate varied from 8.05 to 60.9% in terms of reaction conditions such as the amount of lipase, the presence of water, the amount of molecular sieves and reaction time. The conversions of substrate in solvent-free system were higher than that in acetonitrile. When the substrates were 1 mmol CLA and 1 mmol L-carnitine, the maximum conversion (60.9%) was obtained in solvent-free system with 150 mg lipase AY30, 50% water content and 150 mg molecular sieves at the reaction time of 24 h. A novel CLA ester product was successfully isolated and characterized by ESI-MS and (1)H NMR.
Title: Huperzine A regulates amyloid precursor protein processing via protein kinase C and mitogen-activated protein kinase pathways in neuroblastoma SK-N-SH cells over-expressing wild type human amyloid precursor protein 695 Peng Y, Lee DY, Jiang L, Ma Z, Schachter SC, Lemere CA Ref: Neuroscience, 150:386, 2007 : PubMed
Alpha-secretase (alpha-secretase), cleaves the amyloid precursor protein (APP) within the amyloid-beta (Abeta) sequence, resulting in the release of a secreted fragment of APP (alphaAPPs) and precluding Abeta generation. We investigated the effects of the acetylcholinesterase inhibitor, huperzine A (Hup A), on APP processing and Abeta generation in human neuroblastoma SK-N-SH cells overexpressing wild-type human APP695. Hup A dose-dependently (0-10 microM) increased alphaAPPs release. Therefore, we evaluated two alpha-secretase candidates, a disintegrin and metalloprotease (ADAM) 10 and ADAM17 in Hup A-induced non-amyloidogenic APP metabolism. Hup A enhanced the level of ADAM10, and the inhibitor of tumor necrosis factor-alpha converting enzyme (TACE)/ADAM17 inhibited the Hup A-induced rise in alphaAPPs levels, further suggesting Hup A directed APP metabolism toward the non-amyloidogenic alpha-secretase pathway. Hup A had no effect on Abeta generation in this cell line. The steady-state levels of full-length APP and cell viability were unaffected by Hup A. Alpha-APPs release induced by Hup A treatment was significantly reduced by muscarinic acetylcholine receptor antagonists (particularly by an M1 antagonist), protein kinase C (PKC) inhibitors, GF109203X and calphostin C, and the mitogen-activated kinase kinase (MEK) inhibitors, U0126 and PD98059. Furthermore, Hup A markedly increased the phosphorylation of p44/p42 mitogen-activated protein (MAP) kinase, which was blocked by treatment with U0126 and PD98059. In addition, Hup A inhibited acetylcholinesterase activity by 20% in neuroblastoma cells. Our results indicate that the activation of muscarinic acetylcholine receptors, PKC and MAP kinase may be involved in Hup A-induced alphaAPPs secretion in neuroblastoma cells and suggest multiple pharmacological mechanisms of Hup A regarding the treatment of Alzheimer's disease (AD).
Title: Complete sequence analysis of novel plasmids from emetic and periodontal Bacillus cereus isolates reveals a common evolutionary history among the B. cereus-group plasmids, including Bacillus anthracis pXO1 Rasko DA, Rosovitz MJ, Okstad OA, Fouts DE, Jiang L, Cer RZ, Kolsto AB, Gill SR, Ravel J Ref: Journal of Bacteriology, 189:52, 2007 : PubMed
The plasmids of the members of the Bacillus cereus sensu lato group of organisms are essential in defining the phenotypic traits associated with pathogenesis and ecology. For example, Bacillus anthracis contains two plasmids, pXO1 and pXO2, encoding toxin production and encapsulation, respectively, that define this species pathogenic potential, whereas the presence of a Bt toxin-encoding plasmid defines Bacillus thuringiensis isolates. In this study the plasmids from B. cereus isolates that produce emetic toxin or are linked to periodontal disease were sequenced and analyzed. Two periodontal isolates examined contained almost identical approximately 272-kb plasmids, named pPER272. The emetic toxin-producing isolate contained one approximately 270-kb plasmid, named pCER270, encoding the cereulide biosynthesis gene cluster. Comparative sequence analyses of these B. cereus plasmids revealed a high degree of sequence similarity to the B. anthracis pXO1 plasmid, especially in a putative replication region. These plasmids form a newly defined group of pXO1-like plasmids. However, these novel plasmids do not contain the pXO1 pathogenicity island, which in each instance is replaced by plasmid specific DNA. Plasmids pCER270 and pPER272 share regions that are not found in any other pXO1-like plasmids. Evolutionary studies suggest that these plasmids are more closely related to each other than to other identified B. cereus plasmids. Screening of a population of B. cereus group isolates revealed that pXO1-like plasmids are more often found in association with clinical isolates. This study demonstrates that the pXO1-like plasmids may define pathogenic B. cereus isolates in the same way that pXO1 and pXO2 define the B. anthracis species.
Title: Effects of huperzine A on amyloid precursor protein processing and beta-amyloid generation in human embryonic kidney 293 APP Swedish mutant cells Peng Y, Jiang L, Lee DY, Schachter SC, Ma Z, Lemere CA Ref: Journal of Neuroscience Research, 84:903, 2006 : PubMed
The amyloid precursor protein (APP) is cleaved enzymatically by nonamyloidogenic and amyloidogenic pathways. alpha-Secretase (alpha-secretase), cleaves APP within the beta-amyloid (Abeta) sequence, resulting in the release of a secreted fragment of APP (alphaAPPs) and precluding Abeta generation. In this study, we investigated the effects of an acetylcholinesterase inhibitor, huperzine A (Hup A), on APP processing and Abeta generation in human embryonic kidney 293 cells transfected with human APP bearing the Swedish mutation (HEK293 APPsw). Hup A dose dependently (0-10 microM) increased alphaAPPs release and membrane-coupled APP CTF-C83, suggesting increased APP metabolism toward the nonamyloidogenic alpha-secretase pathway. The metalloprotease inhibitor TAPI-2 inhibited the Hup A-induced increase in alphaAPPs release, further suggesting a modulatory effect of Hup A on alpha-secretase activity. The synthesis of full-length APP and cell viability were unchanged after Hup A incubation, whereas the level of Abeta(Total) was significantly decreased, suggesting an inhibitory effect of Hup A on Abeta production. Hup A-induced alphaAPPs release was significantly reduced by the protein kinase C (PKC) inhibitors GF109203X and Calphostin C. These data, together with the finding that the PKCalpha level was enhanced prior to the increase of alphaAPPs secretion, indicate that PKC may be involved in Hup A-induced alphaAPPs secretion by HEK293 APPsw cells. Our data suggest alternative pharmacological mechanisms of Hup A relevant to the treatment of Alzheimer's disease.
We report the genome sequence of Theileria parva, an apicomplexan pathogen causing economic losses to smallholder farmers in Africa. The parasite chromosomes exhibit limited conservation of gene synteny with Plasmodium falciparum, and its plastid-like genome represents the first example where all apicoplast genes are encoded on one DNA strand. We tentatively identify proteins that facilitate parasite segregation during host cell cytokinesis and contribute to persistent infection of transformed host cells. Several biosynthetic pathways are incomplete or absent, suggesting substantial metabolic dependence on the host cell. One protein family that may generate parasite antigenic diversity is not telomere-associated.
Staphylococcus aureus is an opportunistic pathogen and the major causative agent of numerous hospital- and community-acquired infections. Staphylococcus epidermidis has emerged as a causative agent of infections often associated with implanted medical devices. We have sequenced the approximately 2.8-Mb genome of S. aureus COL, an early methicillin-resistant isolate, and the approximately 2.6-Mb genome of S. epidermidis RP62a, a methicillin-resistant biofilm isolate. Comparative analysis of these and other staphylococcal genomes was used to explore the evolution of virulence and resistance between these two species. The S. aureus and S. epidermidis genomes are syntenic throughout their lengths and share a core set of 1,681 open reading frames. Genome islands in nonsyntenic regions are the primary source of variations in pathogenicity and resistance. Gene transfer between staphylococci and low-GC-content gram-positive bacteria appears to have shaped their virulence and resistance profiles. Integrated plasmids in S. epidermidis carry genes encoding resistance to cadmium and species-specific LPXTG surface proteins. A novel genome island encodes multiple phenol-soluble modulins, a potential S. epidermidis virulence factor. S. epidermidis contains the cap operon, encoding the polyglutamate capsule, a major virulence factor in Bacillus anthracis. Additional phenotypic differences are likely the result of single nucleotide polymorphisms, which are most numerous in cell envelope proteins. Overall differences in pathogenicity can be attributed to genome islands in S. aureus which encode enterotoxins, exotoxins, leukocidins, and leukotoxins not found in S. epidermidis.
Bacillus anthracis is the etiologic agent of anthrax, an acute fatal disease among mammals. It was thought to differ from Bacillus cereus, an opportunistic pathogen and cause of food poisoning, by the presence of plasmids pXO1 and pXO2, which encode the lethal toxin complex and the poly-gamma-d-glutamic acid capsule, respectively. This work describes a non-B. anthracis isolate that possesses the anthrax toxin genes and is capable of causing a severe inhalation anthrax-like illness. Although initial phenotypic and 16S rRNA analysis identified this isolate as B. cereus, the rapid generation and analysis of a high-coverage draft genome sequence revealed the presence of a circular plasmid, named pBCXO1, with 99.6% similarity with the B. anthracis toxin-encoding plasmid, pXO1. Although homologues of the pXO2 encoded capsule genes were not found, a polysaccharide capsule cluster is encoded on a second, previously unidentified plasmid, pBC218. A/J mice challenged with B. cereus G9241 confirmed the virulence of this strain. These findings represent an example of how genomics could rapidly assist public health experts responding not only to clearly identified select agents but also to novel agents with similar pathogenic potentials. In this study, we combined a public health approach with genome analysis to provide insight into the correlation of phenotypic characteristics and their genetic basis.
We sequenced the complete genome of Bacillus cereus ATCC 10987, a non-lethal dairy isolate in the same genetic subgroup as Bacillus anthracis. Comparison of the chromosomes demonstrated that B.cereus ATCC 10987 was more similar to B.anthracis Ames than B.cereus ATCC 14579, while containing a number of unique metabolic capabilities such as urease and xylose utilization and lacking the ability to utilize nitrate and nitrite. Additionally, genetic mechanisms for variation of capsule carbohydrate and flagella surface structures were identified. Bacillus cereus ATCC 10987 contains a single large plasmid (pBc10987), of approximately 208 kb, that is similar in gene content and organization to B.anthracis pXO1 but is lacking the pathogenicity-associated island containing the anthrax lethal and edema toxin complex genes. The chromosomal similarity of B.cereus ATCC 10987 to B.anthracis Ames, as well as the fact that it contains a large pXO1-like plasmid, may make it a possible model for studying B.anthracis plasmid biology and regulatory cross-talk.
Methanotrophs are ubiquitous bacteria that can use the greenhouse gas methane as a sole carbon and energy source for growth, thus playing major roles in global carbon cycles, and in particular, substantially reducing emissions of biologically generated methane to the atmosphere. Despite their importance, and in contrast to organisms that play roles in other major parts of the carbon cycle such as photosynthesis, no genome-level studies have been published on the biology of methanotrophs. We report the first complete genome sequence to our knowledge from an obligate methanotroph, Methylococcus capsulatus (Bath), obtained by the shotgun sequencing approach. Analysis revealed a 3.3-Mb genome highly specialized for a methanotrophic lifestyle, including redundant pathways predicted to be involved in methanotrophy and duplicated genes for essential enzymes such as the methane monooxygenases. We used phylogenomic analysis, gene order information, and comparative analysis with the partially sequenced methylotroph Methylobacterium extorquens to detect genes of unknown function likely to be involved in methanotrophy and methylotrophy. Genome analysis suggests the ability of M. capsulatus to scavenge copper (including a previously unreported nonribosomal peptide synthetase) and to use copper in regulation of methanotrophy, but the exact regulatory mechanisms remain unclear. One of the most surprising outcomes of the project is evidence suggesting the existence of previously unsuspected metabolic flexibility in M. capsulatus, including an ability to grow on sugars, oxidize chemolithotrophic hydrogen and sulfur, and live under reduced oxygen tension, all of which have implications for methanotroph ecology. The availability of the complete genome of M. capsulatus (Bath) deepens our understanding of methanotroph biology and its relationship to global carbon cycles. We have gained evidence for greater metabolic flexibility than was previously known, and for genetic components that may have biotechnological potential.
Bacillus anthracis is an endospore-forming bacterium that causes inhalational anthrax. Key virulence genes are found on plasmids (extra-chromosomal, circular, double-stranded DNA molecules) pXO1 (ref. 2) and pXO2 (ref. 3). To identify additional genes that might contribute to virulence, we analysed the complete sequence of the chromosome of B. anthracis Ames (about 5.23 megabases). We found several chromosomally encoded proteins that may contribute to pathogenicity--including haemolysins, phospholipases and iron acquisition functions--and identified numerous surface proteins that might be important targets for vaccines and drugs. Almost all these putative chromosomal virulence and surface proteins have homologues in Bacillus cereus, highlighting the similarity of B. anthracis to near-neighbours that are not associated with anthrax. By performing a comparative genome hybridization of 19 B. cereus and Bacillus thuringiensis strains against a B. anthracis DNA microarray, we confirmed the general similarity of chromosomal genes among this group of close relatives. However, we found that the gene sequences of pXO1 and pXO2 were more variable between strains, suggesting plasmid mobility in the group. The complete sequence of B. anthracis is a step towards a better understanding of anthrax pathogenesis.
Comparison of the whole-genome sequence of Bacillus anthracis isolated from a victim of a recent bioterrorist anthrax attack with a reference reveals 60 new markers that include single nucleotide polymorphisms (SNPs), inserted or deleted sequences, and tandem repeats. Genome comparison detected four high-quality SNPs between the two sequenced B. anthracis chromosomes and seven differences among different preparations of the reference genome. These markers have been tested on a collection of anthrax isolates and were found to divide these samples into distinct families. These results demonstrate that genome-based analysis of microbial pathogens will provide a powerful new tool for investigation of infectious disease outbreaks.
The complete genome sequence of the radiation-resistant bacterium Deinococcus radiodurans R1 is composed of two chromosomes (2,648,638 and 412,348 base pairs), a megaplasmid (177,466 base pairs), and a small plasmid (45,704 base pairs), yielding a total genome of 3,284, 156 base pairs. Multiple components distributed on the chromosomes and megaplasmid that contribute to the ability of D. radiodurans to survive under conditions of starvation, oxidative stress, and high amounts of DNA damage were identified. Deinococcus radiodurans represents an organism in which all systems for DNA repair, DNA damage export, desiccation and starvation recovery, and genetic redundancy are present in one cell.
