Direct observation of chemical short-range order in a medium-entropy alloy.

Complex concentrated solutions of multiple principal elements are being widely investigated as high- or medium-entropy alloys (HEAs or MEAs)1-11, often assuming that these materials have the high configurational entropy of an ideal solution. However, enthalpic interactions among constituent elements are also expected at normal temperatures, resulting in various degrees of local chemical order12-22. Of the local chemical orders that can develop, chemical short-range order (CSRO) is arguably the most difficult to decipher and firm evidence of CSRO in these materials has been missing thus far16,22. Here we discover that, using an appropriate zone axis, micro/nanobeam diffraction, together with atomic-resolution imaging and chemical mapping via transmission electron microscopy, can explicitly reveal CSRO in a face-centred-cubic VCoNi concentrated solution. Our complementary suite of tools provides concrete information about the degree/extent of CSRO, atomic packing configuration and preferential occupancy of neighbouring lattice planes/sites by chemical species. Modelling of the CSRO order parameters and pair correlations over the nearest atomic shells indicates that the CSRO originates from the nearest-neighbour preference towards unlike (V-Co and V-Ni) pairs and avoidance of V-V pairs. Our findings offer a way of identifying CSRO in concentrated solution alloys. We also use atomic strain mapping to demonstrate the dislocation interactions enhanced by the CSROs, clarifying the effects of these CSROs on plasticity mechanisms and mechanical properties upon deformation.

Harnessing instability for work hardening in multi-principal element alloys.

The strength-ductility trade-off has long been a Gordian knot in conventional metallic structural materials and it is no exception in multi-principal element alloys. In particular, at ultrahigh yield strengths, plastic instability, that is, necking, happens prematurely, because of which ductility almost entirely disappears. This is due to the growing difficulty in the production and accumulation of dislocations from the very beginning of tensile deformation that renders the conventional dislocation hardening insufficient. Here we propose that premature necking can be harnessed for work hardening in a VCoNi multi-principal element alloy. Lüders banding as an initial tensile response induces the ongoing localized necking at the band front to produce both triaxial stress and strain gradient, which enables the rapid multiplication of dislocations. This leads to forest dislocation hardening, plus extra work hardening due to the interaction of dislocations with the local-chemical-order regions. The dual work hardening combines to restrain and stabilize the premature necking in reverse as well as to facilitate uniform deformation. Consequently, a superior strength-and-ductility synergy is achieved with a ductility of ~20% and yield strength of 2 GPa during room-temperature and cryogenic deformation. These findings offer an instability-control paradigm for synergistic work hardening to conquer the strength-ductility paradox at ultrahigh yield strengths.

Effector Cs02526 from Ciboria shiraiana induces cell death and modulates plant immunity.

Sclerotinia disease is one of the most devastating fungal diseases worldwide, as it reduces the yields of many economically important crops. Pathogen-secreted effectors play crucial roles in infection processes. However, key effectors of Ciboria shiraiana, the pathogen primarily responsible for sclerotinia disease in mulberry (Morus spp.), remain poorly understood. In this study, we identified and functionally characterized the effector Cs02526 in C. shiraiana and found that Cs02526 could induce cell deathin a variety of plants. Moreover, Cs02526-induced cell death was mediated by the central immune regulator BRASSINOSTEROID INSENSITIVE 1-associated receptor kinase 1 (BAK1), dependent on a 67-amino acid fragment. Notably, Cs02526 homologues were widely distributed in hemibiotrophic and necrotrophic phytopathogenic fungi, but the homologues failed to induce cell death in plants. Pre-treatment of plants with recombinant Cs02526 protein enhanced resistance against both C. shiraiana and Sclerotinia sclerotiorum. Furthermore, the pathogenicity of C. shiraiana was diminished upon spraying plants with synthetic dsRNA-Cs02526. In conclusion, our findings highlight the cell death-inducing effector Cs02526 as a potential target for future biological control strategies against plant diseases.

High-intensity interval training induces lactylation of fatty acid synthase to inhibit lipid synthesis.

BACKGROUND: The aim of study was to observe the effect of increased lactate levels during high-intensity interval training (HIIT) on protein lactylation, identify the target protein, and investigate the regulatory effect of lactylation on the function of the protein. METHODS: C57B/L6 mice were divided into 3 groups: the control group, HIIT group, and dichloroacetate injection + HIIT group (DCA + HIIT). The HIIT and DCA + HIIT groups underwent 8 weeks of HIIT treatment, and the DCA + HIIT group was injected DCA before HIIT treatment. The expression of lipid metabolism-related genes was determined. Protein lactylation in subcutaneous adipose tissue was identified and analyzed using 4D label-free lactylation quantitative proteomics and bioinformatics analyses. The fatty acid synthase (FASN) lactylation and activity was determined. RESULTS: HIIT had a significant effect on fat loss; this effect was weakened when lactate production was inhibited. HIIT significantly upregulated the protein lactylation while lactate inhibition downregulated in iWAT. FASN had the most modification sites. Lactate treatment increased FASN lactylation levels, inhibited FASN activity, and reduced palmitate and triglyceride synthesis in 3T3-L1 cells. CONCLUSIONS: This investigation revealed that lactate produced by HIIT increased protein pan-lactylation levels in iWAT. FASN lactylation inhibited de novo lipogenesis, which may be an important mechanism in HIIT-induced fat loss.

Crosstalk between ABA and ethylene in regulating stomatal behavior in tomato under high CO2 and progressive soil drying.

Increasing atmospheric CO2 concentrations accompanied by intensifying drought markedly impact plant growth and physiology. This study aimed to explore the role of abscisic acid (ABA) in mediating the response of stomata to elevated CO2 (e[CO2]) and drought. Tomato plants with different endogenous ABA concentrations [Ailsa Craig (AC), the ABA-deficient mutant flacca, and ABA-overproducing transgenic tomato SP5] were grown in ambient (a[CO2], 400 µmol mol-1) and elevated (e[CO2],800 µmol mol-1) CO2 environments and subjected to progressive soil drying. Compared with a[CO2] plants, e[CO2] plants had significantly lower stomatal conductance in AC and SP5 but not in flacca. Under drought, e[CO2] plants had better water status and higher water use efficiency. e[CO2] promoted the accumulation of ABA in leaves of plants subjected to drought, which coincided with the up-regulation of ABA biosynthetic genes and down-regulation of ABA metabolic genes. Although the increase of ABA induced by drought in flacca was much less than in AC and SP5, flacca accumulated large amounts of ethylene, suggesting that in plants with ABA deficiency, ethylene might play a compensatory role in inducing stomatal closure during soil drying. Collectively, these findings improve our understanding of plant performance in a future drier and higher-CO2 environment.

Relationship between social network and individual performance of core members from aged care services social organizations: cross-sectional study.

BACKGROUND: The social network of core members can affect the performance of the organization, while there is a lack of research on the relationship between the social network of core members of social organizations and individual performance in the field of aged care services. This study aimed to explore the relationship between social network and individual performance of core members from social organizations engaged in aged care services and explore measures to promote the development of aged care services. METHODS: We used a multi-stage stratified sampling method to conduct a cross-sectional study and collected the required data in six cities in Anhui Province, China. Univariate analysis and binary logistic regression were used to estimate the relationship between social network and individual performance. RESULTS: Our results indicated that core members with higher social network scores were more likely to yield better individual performance, including receiving awards or recognitions related to aged care services (AOR=2.534; 95% CI: 1.397-4.596). Moreover, teams led by the core members were more likely to receive awards or recognitions related to aged care services (AOR=2.930; 95% CI: 1.740-4.933). The core members or the teams led by them were more likely to be reported by the media (AOR=1.748; 95% CI: 1.030-2.966) and participate in the drafting or discussion of local aged care service standards or service specifications (AOR=2.088; 95% CI: 1.093-3.911). In addition, demographic variables such as gender, marital status, and education of core members were significantly related to their performance (P<0.05). CONCLUSIONS: The social network of core members of aged care service social organizations has an impact on their individual performance. To improve the performance of the core members of senior citizens services and organizations, relevant measures should be taken from the government, social organizations and core members to strengthen the social network construction of core members.

Lessons from assembling a microbial natural product and pre-fractionated extract library in an academic laboratory.

Microbial natural products are specialized metabolites that are sources of many bioactive compounds including antibiotics, antifungals, antiparasitics, anticancer agents, and probes of biology. The assembly of libraries of producers of natural products has traditionally been the province of the pharmaceutical industry. This sector has gathered significant historical collections of bacteria and fungi to identify new drug leads with outstanding outcomes-upwards of 60% of drug scaffolds originate from such libraries. Despite this success, the repeated rediscovery of known compounds and the resultant diminishing chemical novelty contributed to a pivot from this source of bioactive compounds toward more tractable synthetic compounds in the drug industry. The advent of advanced mass spectrometry tools, along with rapid whole genome sequencing and in silico identification of biosynthetic gene clusters that encode the machinery necessary for the synthesis of specialized metabolites, offers the opportunity to revisit microbial natural product libraries with renewed vigor. Assembling a suitable library of microbes and extracts for screening requires the investment of resources and the development of methods that have customarily been the proprietary purview of large pharmaceutical companies. Here, we report a perspective on our efforts to assemble a library of natural product-producing microbes and the establishment of methods to extract and fractionate bioactive compounds using resources available to most academic labs. We validate the library and approach through a series of screens for antimicrobial and cytotoxic agents. This work serves as a blueprint for establishing libraries of microbial natural product producers and bioactive extract fractions suitable for screens of bioactive compounds. ONE-SENTENCE SUMMARY: Natural products are key to discovery of novel antimicrobial agents: Here, we describe our experience and lessons learned in constructing a microbial natural product and pre-fractionated extract library.

Development of HPLC-MS/MS assay for quantitation of ensartinib in human plasma and its application to a pharmacokinetics study in Chinese patients.

Ensartinib is a novel anaplastic lymphoma kinase (ALK) inhibitor with potent activity against a broad range of known crizotinib-resistant ALK mutations and is developed to treat patients with non-small-cell lung cancer. This study was the first to develop and validate a rapid and sensitive HPLC-MS/MS method for the determination of ensartinib in human plasma. The plasma samples were extracted using liquid extraction, and chromatographic separation was performed using a Phenomenex, Luna phenyl-hexyl column (50 × 2.0 mm, 5 µm). Electrospray ionization in positive-ion mode and multiple reaction monitoring were used to monitor ion transitions at m/z 561.3 → 257.1 (ensartinib) and 565.2 → 261.2 (internal standard: X-396-d4), respectively. The method yielded excellent linearity in the range of 0.5-500 ng/ml with the lowest quantification of 0.5 ng/ml. Both intra- and inter-run precisions (relative standard deviation %) were less than 15%, with accuracy (relative error %) between ±15%. Extraction recovery, matrix effect, selectivity, and stability were also validated and found to be satisfactory. Finally, the validated method was successfully applied in a phase I clinical study of ensartinib in Chinese subjects with advanced ALK-positive non-small-cell lung cancer.

Two Different Isocitrate Dehydrogenases from Pseudomonas aeruginosa: Enzymology and Coenzyme-Evolutionary Implications.

Pseudomonas aeruginosa PAO1, as an experimental model for Gram-negative bacteria, harbors two NADP+-dependent isocitrate dehydrogenases (NADP-IDHs) that were evolved from its ancient counterpart NAD-IDHs. For a better understanding of PaIDH1 and PaIDH2, we cloned the genes, overexpressed them in Escherichia coli and purified them to homogeneity. PaIDH1 displayed higher affinity to NADP+ and isocitrate, with lower Km values when compared to PaIDH2. Moreover, PaIDH1 possessed higher temperature tolerance (50 °C) and wider pH range tolerance (7.2-8.5) and could be phosphorylated. After treatment with the bifunctional PaIDH kinase/phosphatase (PaIDH K/P), PaIDH1 lost 80% of its enzymatic activity in one hour due to the phosphorylation of Ser115. Small-molecule compounds like glyoxylic acid and oxaloacetate can effectively inhibit the activity of PaIDHs. The mutant PaIDH1-D346I347A353K393 exhibited enhanced affinity for NAD+ while it lost activity towards NADP+, and the Km value (7770.67 µM) of the mutant PaIDH2-L589 I600 for NADP+ was higher than that observed for NAD+ (5824.33 µM), indicating a shift in coenzyme specificity from NADP+ to NAD+ for both PaIDHs. The experiments demonstrated that the mutation did not alter the oligomeric state of either protein. This study provides a foundation for the elucidation of the evolution and function of two NADP-IDHs in the pathogenic bacterium P. aeruginosa.

Enzymatic Characterization of the Isocitrate Dehydrogenase with Dual Coenzyme Specificity from the Marine Bacterium Umbonibacter marinipuiceus.

Isocitrate dehydrogenase (IDH) can be divided into NAD+-dependent and NADP+-dependent types based on the coenzyme specificity. It is worth noting that some IDHs exhibit dual coenzyme specificity characteristics. Herein, a dual coenzyme-dependent IDH from Umbonibacter Marinipuiceus (UmIDH) was expressed, purified, and identified in detail for the first time. SDS-PAGE and Gel filtration chromatography analyses showed that UmIDH is an 84.7 kDa homodimer in solution. The Km values for NAD+ and NADP+ are 1800.0 ± 64.4 µM and 1167.7 ± 113.0 µM in the presence of Mn2+, respectively. Meanwhile, the catalytic efficiency (kcat/Km) of UmIDH is only 2.3-fold greater for NADP+ than NAD+. The maximal activity for UmIDH occurred at pH 8.5 (with Mn2+) or pH 8.7 (with Mg2+) and at 35 °C (with Mn2+ or Mg2+). Heat inactivation assay revealed that UmIDH sustained 50% of maximal activity after incubation at 57 °C for 20 min with either Mn2+ or Mg2+. Moreover, three putative core coenzyme binding residues (R345, L346, and V352) of UmIDH were evaluated by site-directed mutagenesis. This recent work identified a unique dual coenzyme-dependent IDH and achieved the groundbreaking bidirectional modification of this specific IDH's coenzyme dependence for the first time. This provides not only a reference for the study of dual coenzyme-dependent IDH, but also a basis for the investigation of the coenzyme-specific evolutionary mechanisms of IDH.

Effects of floor- and net-rearing systems on intestinal growth and microbial diversity in the ceca of ducks.

BACKGROUND: Rearing systems can affect livestock production directly, but whether they have effects on intestinal growth states and ceca microorganisms in ducks is largely unclear. The current study used Nonghua ducks to estimate the effects of rearing systems on the intestines by evaluating differences in intestinal growth indices and cecal microorganisms between ducks in the floor-rearing system (FRS) and net-rearing system (NRS). RESULTS: The values of relative weight (RW), relative length (RL) and RW/RL of the duodenum, jejunum, ileum and ceca in the FRS were significantly higher than those in the NRS during weeks 4, 8 and 13 (p < 0.05). A total of 157 genera were identified from ducks under the two systems, and the dominant microorganisms in both treatments were Firmicutes, Bacteroidetes, Actinobacteria and Proteobacteria at the phylum level. The distribution of microorganisms in the ceca of the two treatments showed significant separation during the three time periods, and the value of the Simpson index in the FRS was significantly higher than that in the NRS at 13 weeks (p < 0.05). Five differential microorganisms and 25 differential metabolic pathways were found in the ceca at week 4, seven differential microorganisms and 25 differential metabolic pathways were found in the ceca at week 8, and four differential microorganisms and two differential metabolic pathways were found in the ceca at week 13. CONCLUSIONS: The rearing system influences duck intestinal development and microorganisms. The FRS group had higher intestinal RL, RW and RW/RL and obviously separated ceca microorganisms compared to those of the NRS group. The differential metabolic pathways of cecal microorganisms decreased with increasing age, and the abundance of translation pathways was higher in the NRS group at week 13, while cofactor and vitamin metabolism were more abundant in the FRS group.

An activatable near-infrared hemicyanine-based probe for selective detection and imaging of Hg2+ in living cells and animals.

Fluorescence imaging in the near-infrared (NIR) window has great potential for clinical diagnosis and treatment because of its deep penetration and high contrast. Here, a NIR hemicyanine-based probe (CyP) was synthesized for selective detection and imaging of Hg2+ in living cells and animals. Using the rigid xanthene structure as the electron donor, trimethylindolenine as the electron acceptor and diphenylphosphinothioic chloride as the recognition element, the probe CyP could specifically respond to Hg2+ and transform into an activated donor-π-acceptor (D-π-A) motif with a NIR emission maximum at 710 nm. Cell and animal imaging experiments showed that the probe CyP could be activated by Hg2+ and had good concentration dependence in imaging. Meanwhile, animal imaging experiments showed that the activated CyP probe exhibited a higher tissue penetration depth and spatiotemporal resolution. Thus, the probe CyP could be a useful tool for monitoring and visually evaluating Hg2+ in living systems.

Oestrogen promotes lipids transportation through oestrogen receptor α in hepatic steatosis of geese in vitro.

Evidence has shown that oestrogen suppresses lipids deposition in the liver of mammals. However, the molecular mechanism of oestrogen action in hepatic steatosis of geese liver has yet to be determined. This study aimed to investigate the effect of oestrogen on lipid homeostasis at different states of geese hepatocytes in vitro. The results showed that an in vitro model of hepatic steatosis was induced by 1.5 mM sodium oleate via detecting the viability of hepatocytes and content of lipids. When the normal hepatocytes were administrated with different concentrations of oestrogen (E2 ), the expression levels of diacylglycerol acyltransferase 2 (DGAT2), microsomal triglyceride transfer protein (MTTP) and oestrogen receptors (ERs, alpha and beta) were up-regulated only at high concentrations of E2 , whereas the lipid content was not a significant difference. In goose hepatocytes of hepatic steatosis, however, the expression levels of MTTP, apolipoprotein B (apoB) and ERα/ß significantly increased at 10-7 or 10-6  M E2 . Meanwhile, the lipids content significantly increased at 10-9 and 10-8  M E2 and decreased at 80 µM E2 . Further heatmap analysis showed that ERα was clustered with apoB and MTTP in either normal hepatocytes or that of hepatic steatosis. Taken together, E2  might bind to ERα to up-regulate the expression levels of apoB and MTTP, promoting the transportation of lipids and alleviating lipids overload in hepatic steatosis of geese in vitro.

Factors related to turnover intention among staff of elderly caring social organizations in Anhui Province, China: a cross-sectional study.

OBJECTIVE: Turnover intention of employees in elderly caring social organizations has a significant impact on elderly care service delivery. This study investigated the associated factors of turnover intention among employees of elderly caring social organizations in Anhui Province, China. METHODS: A total of 605 participants were selected using a multi-stage stratified random sampling method. A self-administered questionnaire was used to collect information on socio-demographic, social support, and turnover intention from the participants. The data was analyzed through descriptive statistical analysis, one-way variance analysis, Spearman correlation analysis, and multiple linear regression were used to analyze the factors related to turnover intention. RESULTS: Results of our study showed that the total score of turnover intention, turnover intention I (possibility of quitting a current job),turnover intention II (motivation to find other jobs) and turnover intention III (obtaining the external possibility of work) were 8.84, 2.32, 2.38, and 4.14, respectively. Social support negatively correlated with turnover intention I and turnover intention II. However, it showed positive correlation with turnover intention III and total turnover intention scores; turnover intentionI (coefficient: - 0.082), turnover intention II (coefficient: - 0.071), turnover intention III (coefficient: 0.19), Total score of turnover intention (coefficient: 0.093). Ethnic group, age, education level, and job satisfaction were associated with turnover intention. CONCLUSION: Improvement of social support play an important role in reducing the turnover intention of employees in elderly caring social organizations. It is important to increase organizational commitment and strengthen psychological empowerment, combined with decreasing job burnout for stability.

RedH and PigC Catalyze the Biosynthesis of Hybrubins via Phosphorylation of 4'-Methoxy-2,2'-Bipyrrole-5'-Carbaldehyde.

Hybrubins are "unnatural" alkaloids with the same 4'-methoxy-2,2'-bipyrrole-5'-methine moiety found in prodiginines and a different ring derived from tetramic acids. Here, we demonstrated that RedH, a homologue of prodigiosin synthetase PigC, was responsible for the biosynthesis of hybrubins A and B in Streptomyces lividansIn vitro reactions indicated that RedH and PigC catalyzed the intermolecular condensation between 4'-methoxy-2,2'-bipyrrole-5'-carbaldehyde (MBC) and (Z)-5-ethylidenetetramic acid (ETA) to produce hybrubin B. Moreover, we demonstrated that RedH and PigC activated MBC via phosphorylation of the aldehyde group to form an intermediate Pi-MBC and that the subsequent condensation between Pi-MBC and (Z)-5-ethylidenetetramic acid occurs in a nonenzymatic way.IMPORTANCE Hybrubins are an emerging class of prodiginines possessing a new C ring derived from 5'-substituted tetramic acids and the methylene bridge connecting the C ring at a different position. We have supposed that condensation between 4'-methoxy-2,2'-bipyrrole-5'-carbaldehyde (MBC) and 5-ethylidenetetramic acid (ETA) yields the hybrid natural products hybrubins, which was proposed to be catalyzed by the undecylprodigiosin synthetase RedH. However, it is doubted whether RedH is able to catalyze another type of condensation between MBC and tetramic acids. In this study, we have demonstrated that the MBC-ETA condensation proceeds through RedH/PigC-catalyzed enzymatic activation of MBC via phosphorylation and a nonenzymatic condensation of Pi-MBC with ETA. Since MBC analogues have been shown to be accepted by PigC, more hybrubin analogues might be produced by using combinations of MBC analogues and other tetramic acids in future studies.

Functional Genome Mining Reveals a Class V Lanthipeptide Containing a d-Amino Acid Introduced by an F420 H2 -Dependent Reductase.

Lantibiotics are a type of ribosomally synthesized and post-translationally modified peptides (termed lanthipeptides) with often potent antimicrobial activity. Herein, we report the discovery of a new lantibiotic, lexapeptide, using the library expression analysis system (LEXAS) approach. Lexapeptide has rare structural modifications, including N-terminal (N,N)-dimethyl phenylalanine, C-terminal (2-aminovinyl)-3-methyl-cysteine, and d-Ala. The characteristic lanthionine moiety in lexapeptide is formed by three proteins (LxmK, LxmX, and LxmY), which are distinct from enzymes known to be involved in lanthipeptide biosynthesis. Furthermore, a novel F420 H2 -dependent reductase (LxmJ) from the lexapeptide biosynthetic gene cluster (BGC) is identified to catalyze the reduction of dehydroalanine to install d-Ala. Our findings suggest that lexapeptide is the founding member of a new class of lanthipeptides that we designate as class V. We also identified further class V lanthipeptide BGCs in actinomycetes and cyanobacteria genomes, implying that other class V lantibiotics await discovery.

Vascular endothelial growth factor B inhibits lipid accumulation in C2C12 myotubes incubated with fatty acids.

To investigate (1) the effect of vascular endothelial growth factor B (VEGFB) on lipid accumulation and the alteration of fatty acids and fatty acid-related enzymes in C2C12 myotubes incubated with fatty acids and (2) the regulatory effect of VEGFB on skeletal muscle lipid metabolism. Mouse C2C12 myotubes were incubated with oleic acid (OA) and palmitic acid (PA), and differentiated mature C2C12 myotubes were treated with VEGFB. Oil-red O staining, BODIPY staining and cell triglycerides (TG) content were examined. Total RNA was isolated, and real-time PCR analysis was performed. Treatment with 100 µM OA and 50 µM PA induced lipid droplet accumulation and increased TG content (p < .01), and 100 ng/mL VEGFB reduced lipid droplet accumulation and decreased TG content (p < .01). Treatment with 100 ng/mL VEGFB significantly induced the mRNA expression of fatty acid transport protein 1 (FATP1) (p < .01) and FATP4 (p < .01). Treatment with 100 ng/mL VEGFB significantly induced the mRNA expression of adipose TG lipase and hormone-sensitive lipase (p < .01) as well as carnitine palmitoyltransferase I (p < .01), peroxisome proliferator-activated receptor-γ coactivator-1α (p < .01), acyl-coa dehydrogenase very long chain (p < .05), acyl-coa synthetase long-chain family member 1 (p < .01), peroxisomal acyl-coenzyme A oxidase 1 (p < .05), and mitochondrial uncoupling protein 3 (p < .01). VEGFB enhanced FATP1and FATP4 expression, promoted C2C12 myotube fatty acid oxidation and TG decomposition, and inhibited C2C12 myotube fatty acid re-esterification, thus inhibiting lipid accumulation in C2C12 myotubes incubated with fatty acids.

Biosynthesis of Tropolones in Streptomyces spp.: Interweaving Biosynthesis and Degradation of Phenylacetic Acid and Hydroxylations on the Tropone Ring.

Tropolonoids are important natural products that contain a unique seven-membered aromatic tropolone core and exhibit remarkable biological activities. 3,7-Dihydroxytropolone (DHT) isolated from Streptomyces species is a multiply hydroxylated tropolone exhibiting antimicrobial, anticancer, and antiviral activities. In this study, we determined the DHT biosynthetic pathway by heterologous expression, gene deletion, and biotransformation. Nine trl genes and some of the aerobic phenylacetic acid degradation pathway genes (paa) located outside the trl biosynthetic gene cluster are required for the heterologous production of DHT. The trlA gene encodes a single-domain protein homologous to the C-terminal enoyl coenzyme A (enoyl-CoA) hydratase domain of PaaZ. TrlA truncates the phenylacetic acid catabolic pathway and redirects it toward the formation of heptacyclic intermediates. TrlB is a 3-deoxy-d-arabino-heptulosonic acid-7-phosphate (DAHP) synthase homolog. TrlH is an unusual bifunctional protein bearing an N-terminal prephenate dehydratase domain and a C-terminal chorismate mutase domain. TrlB and TrlH enhanced de novo biosynthesis of phenylpyruvate, thereby providing abundant precursor for the prolific production of DHT in Streptomyces spp. Six seven-membered carbocyclic compounds were identified from the trlC, trlD, trlE, and trlF deletion mutants. Four of these chemicals, including 1,4,6-cycloheptatriene-1-carboxylic acid, tropone, tropolone, and 7-hydroxytropolone, were verified as key biosynthetic intermediates. TrlF is required for the conversion of 1,4,6-cycloheptatriene-1-carboxylic acid into tropone. The monooxygenases TrlE and TrlCD catalyze the regioselective hydroxylations of tropone to produce DHT. This study reveals a natural association of anabolism of chorismate and phenylpyruvate, catabolism of phenylacetic acid, and biosynthesis of tropolones in Streptomyces spp.IMPORTANCE Tropolonoids are promising drug lead compounds because of the versatile bioactivities attributed to their highly oxidized seven-membered aromatic ring scaffolds. Our present study provides clear insight into the biosynthesis of 3,7-dihydroxytropolone (DHT) through the identification of key genes responsible for the formation and modification of the seven-membered aromatic core. We also reveal the intrinsic mechanism of elevated production of DHT and related tropolonoids in Streptomyces spp. The study on DHT biosynthesis in Streptomyces exhibits a good example of antibiotic production in which both anabolic and catabolic pathways of primary metabolism are interwoven into the biosynthesis of secondary metabolites. Furthermore, our study sets the stage for metabolic engineering of the biosynthetic pathway for natural tropolonoid products and provides alternative synthetic biology tools for engineering novel tropolonoids.

[Efficacy of endoscopic transcaruncular approach to reconstruct isolated medial orbital fracture].

OBJECTIVE: To study the efficacy of endoscopic transcaruncular approach in the repair of isolated medial orbital fracture. METHODS: It was a retrospective case series study.Retrospective chart was reviewed in 21 patients (21 eyes) receiving endoscopic transcaruncular approach to reconstruct isolated medial orbital fracture at Quzhou People's Hospital from May 2011 to May 2012. RESULTS: of visual acuity, diplopia, protrusion degree of both eyes and the movement of eye balls was recorded for analysis. RESULTS: All patients were followed up for 8-20 months. There was no extrusion, rejection, infection or other complications of Medpor surgical implant during follow-up. There was no instance of decreased visual acuity at post-operation. The post-operative protrusion degree of both eyes was almost identical at less than 2 mm. The movement of eye balls was satisfactory in all directions. Diplopia disappeared in 13 cases, 1 case improved. CONCLUSION: Endoscopic transcaruncular approach is a safe and effective treatment in the repair of isolated medial orbital fracture.

Further delineation of phenotype and genotype of Kenny-Caffey syndrome type 2 (phenotype and genotype of KCS type 2).

BACKGROUND: Kenny-Caffey syndrome type 2 (KCS2) is an extremely rare inherited disorder characterized by proportionate short stature, skeletal defects, ocular and dental abnormalities, and transient hypocalcemia. It is caused by variants in FAM111A gene. Diagnosis of KCS2 can be challenging because of its similarities to other syndromes, the absence of clear hallmarks and the deficient number of genetically confirmed cases. Here, we aimed to further delineate and summarize the genotype and phenotype of KCS2, in order to get a better understanding of this rare disorder, and promote early diagnosis and intervention. METHODS: We present clinical and genetic characteristics of eight newly affected individuals with KCS2 from six families, including one family with three individuals found to be a father-to-daughter transmission, adding to the limited literature. Furthermore, we performed a review of genetically confirmed KCS2 cases in PubMed, MEDLINE and CNKI databases. RESULTS: There were six females and two males in our cohort. All the patients presented with short stature (100.0%). Clinical manifestations included ocular defects such as hypermetropia (5/8), dental problems such as defective dentition (3/8) and dental caries (3/8), skeletal and brain anomalies such as delayed closure of anterior fontanelle (6/8), cerebral calcification (3/8), cortical thickening (3/8) and medullary stenosis (4/8) of tubular bones. Endocrinologic abnormalities included hypoparathyroidism (5/8) and hypocalcemia (3/8). One male patient had micropenis and microorchidism. All cases harboured missense variants of FAM111A, and nucleotides c.1706 arose as a mutational hotspot, with seven individuals harbouring a c.1706G>A (p.Arg569His) variant, and one child harbouring a c.1531T>C (p.Tyr511His) variant. Literature review yielded a total of 46 patients from 20 papers. Data analysis showed that short stature, hypoparathyroidism and hypocalcemia, ocular and dental defects, skeletal features including cortical thickening and medullary stenosis of tubular bones, and seizures/spasms were present in more than 70% of the reported KCS2 cases. CONCLUSION: We provide detailed characteristics of the largest KCS2 group in China and present the first genetically confirmed instance of father-to-daughter transmission of KCS2. Our study confirms that Arg569His is the hot spot variant and summarizes the typical phenotypes of KCS2, which would help early diagnosis and intervention.