荒漠甲虫小胸鳖甲抗冻蛋白的酵母表达及应用

昆虫抗冻蛋白(Antifreeze protein,AFP)的抗冻活性很高,可应用于生物组织和细胞的低温保存。为了在酵母中表达荒漠甲虫小胸鳖甲Microdera punctipennis抗冻蛋白Mp AFP698,并确定其在低温下的保护作用,本文通过构建真核表达载体p PIC9K-Mpafp698,转化巴斯德毕赤酵母GS115,诱导表达小胸鳖甲抗冻蛋白Mp AFP698。利用免疫印迹(Western blotting)分析Mp AFP698蛋白的特异性表达,结果显示Mpafp698基因可整合到酵母基因组中并分泌表达,且酵母自身蛋白很少分泌表达。检测抗冻蛋白的低温保护作用,结果发现,小胸鳖甲抗冻蛋白可显著改善冷冻小鼠肝脏等器官的细胞形态,降低血细胞在4℃的溶血率,提高SF9细胞冻融后的存活率。本研究表明,小胸鳖甲AFP可以在毕赤酵母中分泌表达,便于纯化,有良好的低温保护效果。     

Epigenetic silencing of microRNA-149 in cancer-associated fibroblasts mediates prostaglandin E2/interleukin-6 signaling in the tumor microenvironment

Tumor initiation and growth depend on its microenvironment in which cancer-associated fibroblasts (CAFs) in tumor stroma play an important role. Prostaglandin E2 (PGE2) and interleukin (IL)-6 signal pathways are involved in the crosstalk between tumor and stromal cells. However, how PGE2-mediated signaling modulates this crosstalk remains unclear. Here, we show that microRNA (miR)-149 links PGE2 and IL-6 signaling in mediating the crosstalk between tumor cells and CAFs in gastric cancer (GC). miR-149 inhibited fibroblast activation by targeting IL-6 and miR-149 expression was substantially suppressed in the CAFs of GC. miR-149 negatively regulated CAFs and their effect on GC development both in vitro and in vivo. CAFs enhanced epithelial-to-mesenchymal transition (EMT) and the stem-like properties of GC cells in a miR-149-IL-6-dependent manner. In addition to IL-6, PGE2 receptor 2 (PTGER2/EP2) was revealed as another potential target of miR-149 in fibroblasts. Furthermore, H. pylori infection, a leading cause of human GC, was able to induce cyclooxygenase-2 (COX-2)/PGE2 signaling and to enhance PGE2 production, resulting in the hypermethylation of miR-149 in CAFs and increased IL-6 secretion. Our findings indicate that miR-149 mediates the crosstalk between tumor cells and CAFs in GC and highlight the potential of interfering miRNAs in stromal cells to improve cancer therapy.     

Arabidopsis SEIPIN Proteins Modulate Triacylglycerol Accumulation and Influence Lipid Droplet Proliferation

The lipodystrophy protein SEIPIN is important for lipid droplet (LD) biogenesis in human and yeast cells. In contrast with the single SEIPIN genes in humans and yeast, there are three SEIPIN homologs in Arabidopsis thaliana, designated SEIPIN1, SEIPIN2, and SEIPIN3. Essentially nothing is known about the functions of SEIPIN homologs in plants. Here, a yeast (Saccharomyces cerevisiae) SEIPIN deletion mutant strain and a plant (Nicotiana benthamiana) transient expression system were used to test the ability of Arabidopsis SEIPINs to influence LD morphology. In both species, expression of SEIPIN1 promoted accumulation of large-sized lipid droplets, while expression of SEIPIN2 and especially SEIPIN3 promoted small LDs. Arabidopsis SEIPINs increased triacylglycerol levels and altered composition. In tobacco, endoplasmic reticulum (ER)-localized SEIPINs reorganized the normal, reticulated ER structure into discrete ER domains that colocalized with LDs. N-terminal deletions and swapping experiments of SEIPIN1 and 3 revealed that this region of SEIPIN determines LD size. Ectopic overexpression of SEIPIN1 in Arabidopsis resulted in increased numbers of large LDs in leaves, as well as in seeds, and increased seed oil content by up to 10% over wild-type seeds. By contrast, RNAi suppression of SEIPIN1 resulted in smaller seeds and, as a consequence, a reduction in the amount of oil per seed compared with the wild type. Overall, our results indicate that Arabidopsis SEIPINs are part of a conserved LD biogenesis machinery in eukaryotes and that in plants these proteins may have evolved specialized roles in the storage of neutral lipids by differentially modulating the number and sizes of lipid droplets.     

Valsartan Attenuates Atherosclerosis via Upregulating the Th2 Immune Response in Prolonged Angiotensin II–Treated ApoE?/? Mice

Valsartan has a protective effect against hypertension and atherosclerosis in humans and experimental animal models. This study aimed to determine the effect of prolonged treatment with angiotensin II (Ang II) on atherosclerosis and the effect of valsartan on the activity of CD4⁺ T lymphocyte subsets. The results showed that prolonged treatment (8 wks) with exogenous Ang II resulted in an increased atherosclerotic plaque size and a switch of stable-to-unstable plaque via modulating on CD4⁺ T lymphocyte activity, including an increase in the T helper cell type 1 (Th1) and Th17 cells and a decrease in Th2 and regulatory T (Treg) cells. In contrast, valsartan treatment efficiently reversed the imbalance in CD4⁺ T lymphocyte activity, ameliorated atherosclerosis and elicited a stable plaque phenotype in addition to controlling blood pressure. In addition, treatment with anti-interleukin (IL)-5 monoclonal antibodies weakened the antiatherosclerotic effects of valsartan without affecting blood pressure.     

Enterocytozoon bieneusi Genotypes in Yaks (Bos grunniens) and Their Public Health Potential

Enterocytozoon bieneusi, the most frequently diagnosed microsporidian species in humans, is also identified in a wide range of animals. To date, few data are available on E. bieneusi in yaks (Bos grunniens). In this study, we examined the occurrence and genotype identity of E. bieneusi in yaks in four counties in Qinghai Province of China. Of 327 fecal specimens examined by nested PCR analysis of the ribosomal internal transcribed spacer, 23 (7.0%) were E. bieneusi‐positive. DNA sequence analysis of the PCR products revealed the presence of five distinct genotypes: three Group 2 genotypes previously reported in cattle as well as humans (BEB4, I and J) and two novel genotypes (CHN11 and CHN12) belonging to the large zoonotic group (Group 1). Data of the study suggest that these animals could be potential reservoirs for human E. bieneusi infection.     

Exosomes in Mesenchymal Stem Cells,a New Therapeutic Strategy for Cardiovascular Diseases?

Cardiovascular diseases (CVDs) are still a major cause of people deaths worldwide, and mesenchymal stem cells (MSCs) transplantation holds great promise due to its capacity to differentiate into cardiovascular cells and secrete protective cytokines, which presents an important mechanism of MSCs therapy for CVDs. Although the capability of MSCs to differentiate into cardiomyocytes (CMCs), endothelial cells (ECs) and vascular smooth muscle cells (VSMCs) has been well recognized in massive previous experiments both in vitro and in vivo, low survival rate of transplanted MSCs in recipient hearts suggests that therapeutic effects of MSCs transplantation might be also correlated with other underlying mechanisms. Notably, recent studies uncovered that MSCs were able to secret cholesterol-rich, phospholipid exosomes which were enriched with microRNAs (miRNAs). The released exosomes from MSCs acted on hearts and vessels, and then exerted anti-apoptosis, cardiac regeneration, anti-cardiac remodeling, anti-inflammatory effects, neovascularization and anti-vascular remodeling, which are considered as novel molecular mechanisms of therapeutic potential of MSCs transplantation. Here we summarized recent advances about the role of exosomes in MSCs therapy for CVDs, and discussed exosomes as a novel approach in the treatment of CVDs in the future.     

Avianbase: a community resource for bird genomics

Giving access to sequence and annotation data for genome assemblies is important because, while facilitating research, it places both assembly and annotation quality under scrutiny, resulting in improvements to both. Therefore we announce Avianbase, a resource for bird genomics, which provides access to data released by the Avian Phylogenomics Consortium.Access to complete genome sequences provides the first step towards the understanding of the biology of organisms. It is the template that underpins the phenotypic characteristics of individuals and ultimately separates species due to the accumulation and fixation of mutations over evolutionary timescales. In terms of the available genomic datasets for species, birds, as our more distant relatives, have been historically underrepresented. The high cost of sequencing and annotation in the past led to a bias towards accumulating data for species that are either established model organisms or economically significant (that is, chicken, turkey and duck, representing two sister orders within the Galloanseriformes clade from the large and diverse phylogeny of birds). The recent release of genome assemblies and initial predictions of protein-coding genes [1-4] for 44 bird species, including representatives from all major branches of the bird phylogeny, is, therefore, highly significant.One of the major challenges with the release of this number of newly sequenced genomes and the many more to come [5] is how to make these available to the various research communities in a way that supports basic research. Providing access to the sequences and initial annotations in the format of text files will limit the potential usage of the data as they require significant resources, including bioinformatics personnel and computer infrastructure in place to access and mine - for example, searching for genes belonging to certain protein families or searching for orthologous genes. These overheads pose a serious bottleneck that can hinder research and requires concerted action by the relevant research communities.Once genomes are submitted to public databases, genome-wide annotations are frequently generated and released either via the Ensembl project [6] or by the National Center for Biotechnology Information [7] and sequence and annotation are then made visually available online in integrated views via the Ensembl or the University of California Santa Cruz (UCSC) genome browsers [8]. These systems provide search facilities, sequence alignment tools like BLAT/BLAST and various analysis tools to facilitate subsetting and computational retrieval of the data, including UCSC’s Table Browser or Ensembl’s Perl and REST APIs and BioMart system.While these systems have become almost indispensable for research, not all sequenced genomes are annotated and displayed in genome browsers. Full genome annotation remains time consuming and resource intensive: a full evidence-based Ensembl genebuild takes approximately 4 months. Thus, the list of species represented is currently limited and depends on various factors, including the completeness of the assembled genome sequence and the overall demand in the scientific community for the resources, including whether the species is a model organism (for example, human or mouse), economically important (for example, farmed animals) or of specific phylogenetic interest. Many of the recently sequenced bird genomes do not obviously fall within these categories.     

Tissue‐specific and pathogen‐inducible expression of a fusion protein containing a Fusarium‐specific antibody and a fungal chitinase protects wheat against Fusarium pathogens and mycotoxins

Fusarium head blight (FHB) in wheat and other small grain cereals is a globally devastating disease caused by toxigenic Fusarium pathogens. Controlling FHB is a challenge because germplasm that is naturally resistant against these pathogens is inadequate. Current control measures rely on fungicides. Here, an antibody fusion comprised of the Fusarium spp.‐specific recombinant antibody gene CWP2 derived from chicken, and the endochitinase gene Ech42 from the biocontrol fungus Trichoderma atroviride was introduced into the elite wheat cultivar Zhengmai9023 by particle bombardment. Expression of this fusion gene was regulated by the lemma/palea‐specific promoter Lem2 derived from barley; its expression was confirmed as lemma/palea‐specific in transgenic wheat. Single‐floret inoculation of independent transgenic wheat lines of the T₃ to T₆ generations revealed significant resistance (type II) to fungal spreading, and natural infection assays in the field showed significant resistance (type I) to initial infection. Gas chromatography–mass spectrometry analysis revealed marked reduction of mycotoxins in the grains of the transgenic wheat lines. Progenies of crosses between the transgenic lines and the FHB‐susceptible cultivar Huamai13 also showed significantly enhanced FHB resistance. Quantitative real‐time PCR analysis revealed that the tissue‐specific expression of the antibody fusion was induced by salicylic acid drenching and induced to a greater extent by F. graminearum infection. Histochemical analysis showed substantial restriction of mycelial growth in the lemma tissues of the transgenic plants. Thus, the combined tissue‐specific and pathogen‐inducible expression of this Fusarium‐specific antibody fusion can effectively protect wheat against Fusarium pathogens and reduce mycotoxin content in grain.