竹节参鲨烯环氧酶基因的克隆与生物信息学分析

根据已报道的植物鲨烯环氧酶(SE)基因特异性引物克隆竹节参SE基因.结果表明,克隆得到竹节参SE全长为1 632 bp,编码539个氨基酸,命名为PjSE.生物信息学分析指出,PjSE基因的氨基酸序列与人参属植物人参、三七、越南人参的同源性依次为99％、98％和98％.推测PjSE基因定位于线粒体(mTP)、叶绿体(cTP)和分泌途径(SP).PjSE基因存在保守结构域FAD结合位点,含4个跨膜结构域和7个motif结构位点.PjSE蛋白二级结构以无规则卷曲(Random coli)和α-螺旋(Alpha helix)为主要结构元件,延伸链(Extended strand)和β转角(Beta turn)分散于整个蛋白质中,百分比依次占40.82％、37.48％、15.77％、5.94％;该蛋白能折叠形成典型的三维结构.     

Impact of manipulation of glycerol/diol dehydratase activity on intestinal microbiota ecology and metabolism

Glycerol/diol dehydratases (GDH) are enzymes that catalyse the production of propionate from 1,2-propanediol, and acrolein from glycerol. Acrolein reacts with dietary carcinogenic heterocyclic amines (HCA), reducing HCA mutagenicity, but is itself also an antimicrobial agent and toxicant. Gut microbial GDH activity has been suggested as an endogenous acrolein source; however, there is limited information on the potential of the intestinal microbiota to have GDH activity, and what impact it can have on the intestinal ecosystem and host health. We hypothesized that GDH activity of gut microbiota is determined by the abundance and distribution of GDH-active taxa and can be enhanced by supplementation of the GDH active Anaerobutyricum hallii, and tested this hypothesis combining quantitative profiling of gdh, model batch fermentations, microbiota manipulation, and kinetic modelling of acrolein formation. Our results suggest that GDH activity is a common trait of intestinal microbiota shared by a few taxa, which was dependent on overall gdh abundance. Anaerobutyricum hallii was identified as a key taxon in GDH metabolism, and its supplementation increased the rate of GDH activity and acrolein release, which enhanced the transformation of HCA and reduced fermentation activity. The findings of this first systematic study on acrolein release by intestinal microbiota indicate that dietary and microbial modulation might impact GDH activity, which may influence host health.     

A new Illumina MiSeq high-throughput sequencing-based method for evaluating the composition of the Bacteroides community in the intestine using the rpsD gene sequence

Bacteroides is a bacterial genus that is known to closely interact with the host. The potential role of this genus is associated with its ecological status and distribution in the intestine. However, the current 16S V3–V4 region sequencing method can only detect the abundance of this genus, revealing a need for a novel sequencing method that can elucidate the composition of Bacteroides in the human gut microbiota. In this study, a core gene, rpsD, was selected as a template for the design of a Bacteroides-specific primer set. We used this primer set to develop a novel assay based on the Illumina MiSeq sequencing platform that enabled an accurate assessment of the Bacteroides compositions in complex samples. Known amounts of genomic DNA from 10 Bacteroides species were mixed with a complex sample and used to evaluate the performance and detection limit of our assay. The results were highly consistent with those of direct sequencing with a low Bacteroides DNA detection threshold (0.01 ng), supporting the reliability of our assay. In addition, the assay could detect all the known Bacteroides species within the faecal sample. In summary, we provide a sensitive and specific approach to determining the Bacteroides species in complex samples.     

Circulating exosomes repair endothelial cell damage by delivering miR-193a-5p

Circulating exosomes delivering microRNAs are involved in the occurrence and development of cardiovascular diseases. How are the circulating exosomes involved in the repair of endothelial injury in acute myocardial infarction (AMI) convalescence (3-7 days) was still not clear. In this study, circulating exosomes from AMI patients (AMI-Exo) and healthy controls (Normal-Exo) were extracted. In vitro and in vivo, our study showed that circulating exosomes protected endothelial cells (HUVECs) from oxidative stress damage; meanwhile, Normal-Exo showed better protective effects. Through the application of related inhibitors, we found that circulating exosomes shuttled between HUVECs via dynamin. Microarry analysis and qRT-PCR of circulating exosomes showed higher expression of miR-193a-5p in Normal-Exo. Our study showed that miR-193a-5p was the key factor on protecting endothelial cells in vitro and in vivo. Bioinformatics analyses found that activin A receptor type I (ACVR1) was the potential downstream target of miR-193a-5p, which was confirmed by ACVR1 expression and dual-luciferase report. Inhibitor of ACVR1 showed similar protective effects as miR-193a-5p. While overexpression of ACVR1 could attenuate protective effects of miR-193a-5p. To sum up, these findings suggest that circulating exosomes could shuttle between cells through dynamin and deliver miR-193a-5p to protect endothelial cells from oxidative stress damage via ACVR1.     

Inhibition of cyclooxygenase-2 enhanced intestinal epithelial homeostasis via suppressing β-catenin signalling pathway in experimental liver fibrosis

The intestinal barrier dysfunction is crucial for the development of liver fibrosis but can be disturbed by intestinal chronic inflammation characterized with cyclooxygenase-2 (COX-2) expression. This study focused on the unknown mechanism by which COX-2 regulates intestinal epithelial homeostasis in liver fibrosis. The animal models of liver fibrosis induced with TAA were established in rats and in intestinal epithelial–specific COX-2 knockout mice. The impacts of COX-2 on intestinal epithelial homeostasis via suppressing β-catenin signalling pathway were verified pharmacologically and genetically in vivo. A similar assumption was tested in Ls174T cells with goblet cell phenotype in vitro. Firstly, disruption of intestinal epithelial homeostasis in cirrhotic rats was ameliorated by celecoxib, a selective COX-2 inhibitor. Then, β-catenin signalling pathway in cirrhotic rats was associated with the activation of COX-2. Furthermore, intestinal epithelial–specific COX-2 knockout could suppress β-catenin signalling pathway and restore the disruption of ileal epithelial homeostasis in cirrhotic mice. Moreover, the effect of COX-2/PGE2 was dependent on the β-catenin signalling pathway in Ls174T cells. Therefore, inhibition of COX-2 may enhance intestinal epithelial homeostasis via suppression of the β-catenin signalling pathway in liver fibrosis.     

Physical interactions facilitate sex change in the protogynous orange-spotted grouper,Epinephelus coioides

Sex change in teleost fishes is commonly regulated by social factors. In species that exhibit protogynous sex change, such as the orange-spotted grouper Epinephelus coioides, when the dominant males are removed from the social group, the most dominant female initiates sex change. The aim of this study was to determine the regulatory mechanisms of socially controlled sex change in E. coioides. We investigated the seasonal variation in social behaviours and sex change throughout the reproductive cycle of E. coioides, and defined the behaviour pattern of this fish during the establishment of a dominance hierarchy. The social behaviours and sex change in this fish were affected by season, and only occurred during the prebreeding season and breeding season. Therefore, a series of sensory isolation experiments was conducted during the breeding season to determine the role of physical, visual and olfactory cues in mediating socially controlled sex change. The results demonstrated that physical interactions between individuals in the social groups were crucial for the initiation and completion of sex change, whereas visual and olfactory cues alone were insufficient in stimulating sex change in dominant females. In addition, we propose that the steroid hormones 11-ketotestosterone and cortisol are involved in regulating the initiation of socially controlled sex change.     

Microtubule-assisted mechanism for toxisome assembly in Fusarium graminearum

In Fusarium graminearum, a trichothecene biosynthetic complex known as the toxisome forms ovoid and spherical structures in the remodelled endoplasmic reticulum (ER) under mycotoxin-inducing conditions. Previous studies also demonstrated that disruption of actin and tubulin results in a significant decrease in deoxynivalenol (DON) biosynthesis in F. graminearum. However, the functional association between the toxisome and microtubule components has not been clearly defined. In this study we tested the hypothesis that the microtubule network provides key support for toxisome assembly and thus facilitates DON biosynthesis. Through fluorescent live cell imaging, knockout mutant generation, and protein–protein interaction assays, we determined that two of the four F. graminearum tubulins, α₁ and β₂ tubulins, are indispensable for DON production. We also showed that these two tubulins are directly associated. When the α₁–β₂ tubulin heterodimer is disrupted, the metabolic activity of the toxisome is significantly suppressed, which leads to significant DON biosynthesis impairment. Similar phenotypic outcomes were shown when F. graminearum wild type was treated with carbendazim, a fungicide that binds to microtubules and disrupts spindle formation. Based on our results, we propose a model where α₁–β₂ tubulin heterodimer serves as the scaffold for functional toxisome assembly in F. graminearum.     

SHORT-ROOT 1 is critical to cell division and tracheary element development in rice roots

The exocyst is a key factor in vesicle transport and is involved in cell secretion, cell growth, cell division and other cytological processes in eukaryotes. EXO70 is the key exocyst subunit. We obtained a gene, SHORT-ROOT 1 (SR1), through map-based cloning and genetic complementation. SR1 is a conserved protein with an EXO70 domain in plants. SR1 mutation affected the whole root-development process: producing shorter radicles, adventitious roots and lateral roots, and demonstrating abnormal xylem development, resulting in dwarfing and reduced water potential and moisture content. SR1 was largely expressed in the roots, but only in developing root meristems and tracheary elements. The shortness of the sr1 mutant roots was caused by the presence of fewer meristem cells. The in situ histone H4 expression patterns confirmed that cell proliferation during root development was impaired. Tracheary element dysplasia was caused by marked decreases in the inner diameters of and distances between the perforations of adjacent tracheary elements. The membrane transport of sr1 mutants was blocked, affecting cell division in the root apical region and the development of root tracheary elements. The study of SR1 will deepen our understanding of the function of EXO70 genes in Oryza sativa (rice) and guide future studies on the molecular mechanisms involved in plant root development.