Increase of anti-HIV activity of C-peptide fusion inhibitors using a bivalent drug design approach

We reported the design of fusion inhibitors with improved activity using a multivalent inhibitor design strategy. First, we chose C29 as the template sequence, which is a 29-mer peptide derived from HIV-1 gp41 CHR domain and has anti-HIV activity of IC₅₀ 118 nM in a cell–cell fusion assay. We optimized the crosslink sites and linkers of the template peptide. We found that N-terminal crosslink caused activity improvement based on the multivalent co-operative effect. Especially, the IC₅₀ of peptide (CAcaC29)₂ was improved from 49.02 (monomeric form) to 5.71 nM. Compared with long peptides, short peptides may be more suitable to analyze the co-operative effect. So we selected a shorter peptide C22 to synthesize the bivalent inhibitors. Due its weak helicity, no co-operative effect appeared. Therefore, we chose SC22EK, which were introduced salt bridges to consolidate the helicity based on the natural sequence C22. The cross-linked (CAcaSC22EK)₂ was four times more potent than the monomer SC22EK in anti-HIV activity, with an IC₅₀ value of 4.92 nM close to the high active peptide fusion inhibitor C34. The strategy used in this study may be used to design new fusion inhibitors to interfere similar processes.     

Impact of GC content on gene expression pattern in chicken

Background

GC content varies greatly between different genomic regions in many eukaryotes. In order to determine whether this organization named isochore organization influences gene expression patterns, the relationship between GC content and gene expression has been investigated in man and mouse. However, to date, this question is still a matter for debate. Among the avian species, chicken (Gallus gallus) is the best studied representative with a complete genome sequence. The distinctive features and organization of its sequence make it a good model to explore important issues in genome structure and evolution.

Methods

Only nuclear genes with complete information on protein-coding sequence with no evidence of multiple-splicing forms were included in this study. Chicken protein coding sequences, complete mRNA sequences (or full length cDNA sequences), and 5^′ untranslated region sequences (5^′ UTR) were downloaded from Ensembl and chicken expression data originated from a previous work. Three indices i.e. expression level, expression breadth and maximum expression level were used to measure the expression pattern of a given gene. CpG islands were identified using hgTables of the UCSC Genome Browser. Correlation analysis between variables was performed by SAS Proprietary Software Release 8.1.

Results

In chicken, the GC content of 5^′ UTR is significantly and positively correlated with expression level, expression breadth, and maximum expression level, whereas that of coding sequences and introns and at the third coding position are negatively correlated with expression level and expression breadth, and not correlated with maximum expression level. These significant trends are independent of recombination rate, chromosome size and gene density. Furthermore, multiple linear regression analysis indicated that GC content in genes could explain approximately 10% of the variation in gene expression.

Conclusions

GC content is significantly associated with gene expression pattern and could be one of the important regulation factors in the chicken genome.     

马哈利樱桃大孢子发生和雌配子体的发育

采用石蜡切片法对马哈利樱桃大孢子发生和雌配子体发育过程进行观察研究。结果表明:(1)马哈利樱桃雌配子体发育早期,在单室子房内可以看到2个倒生胚珠,但在后期其中一个退化,另一个发育为种子;其胚珠具双珠被,为厚珠心。(2)大孢子母细胞减数分裂形成直线型四分体,功能大孢子位于合点端;胚囊发育为蓼型,成熟胚囊为七细胞八核。(3)根据不同时间花的外部形态特征与内部解剖学对比的观察结果,在陕西关中地区,三月下旬是马哈利樱桃雌性生殖细胞分化和发育的重要时期,果园在此期间应加强肥水管理。     

XBP-1u suppresses autophagy by promoting the degradation of FoxO1 in cancer cells

Autophagy is activated to maintain cellular energy homeostasis in response to nutrient starvation. However, autophagy is not persistently activated, which is poorly understood at a mechanistic level. Here, we report that turnover of FoxO1 is involved in the dynamic autophagic process caused by glutamine starvation. X-box-binding protein-1u (XBP-1u) has a critical role in FoxO1 degradation by recruiting FoxO1 to the 20S proteasome. In addition, the phosphorylation of XBP-1u by extracellular regulated protein kinases1/2 (ERK1/2) on Ser61 and Ser176 was found to be critical for the increased interaction between XBP-1u and FoxO1 upon glutamine starvation. Furthermore, knockdown of XBP-1u caused the sustained level of FoxO1 and the persistent activation of autophagy, leading to a significant decrease in cell viability. Finally, the inverse correlation between XBP-1u and FoxO1 expression agrees well with the expression profiles observed in many human cancer tissues. Thus, our findings link the dynamic process of autophagy to XBP-1u-induced FoxO1 degradation.     

Immunosensor based on magnetic relaxation switch and biotin-streptavidin system for the detection of Kanamycin in milk

A rapid, sensitive, and simple immunosensor was developed for the detection of Kanamycin (KM) in milk. This immunosensor is based on magnetic relaxation switch (MRS) assay and biotin-streptavidin system (B-SA system). The target analyte (KM) competed with those on the surface of the superparamagnetic iron oxide (SPIO) nanoparticles and hence affected the formation of SPIO aggregates. The dispersed and aggregated states of SPIO can modulate the spin-spin relaxation time (T(2)) of the neighboring water molecule. T(2) was then changed as an effect of the target analyte. The B-SA system was used to amplify the SPIO binding, thus enhance the sensitivity. The detection working was 1.5 to 25.2ngmL(-1) and limit of detection (LOD) was determined to be 0.1ngmL(-1). The LOD of the immunosensor decreased tenfold, and its analysis time (45min) was much shorter than that of enzyme-linked immunosorbent assay (6h to 8h). The average recoveries of the KM at various spiking levels ranged from 80.2% to 85.6% with a relative standard deviation (RSD) below 4.0%. The results showed that the MRS immunosensor was a promising platform for the determination of small molecular residues because of its high sensitivity, specificity, homogeneity, and speed.     

Purification and characterization of a dehalogenase from Pseudomonas stutzeri DEH130 isolated from the marine sponge Hymeniacidon perlevis

2-haloacid dehalogenases are enzymes that are capable of degrading 2-haloacid compounds. These enzymes are produced by bacteria, but so far they have only been purified and characterized from terrestrial bacteria. The present study describes the purification and characterization of 2-haloacid dehalogenase from the marine bacterium Pseudomonas stutzeri DEH130. P. Stutzeri DEH130 contained two kinds of 2-haloacid dehalogenase (designated as Dehalogenase I and Dehalogenase II) as detected in the crude cell extract after ammonium sulfate fractionation. Both enzymes appeared to exhibit stereo-specificity with respect to substrate. Dehalogenase I was a 109.9-kDa enzyme that preferentially utilized D-2-chloropropropionate and had optimum activity at pH 7.5. Dehalogenase II, which preferentially utilized L-2-chloropropionate, was further purified by ion-exchange chromatography and gel filtration. Purified Dehalogenase II appeared to be a dimeric enzyme with a subunit of 26.0-kDa. It had maximum activity at pH 10.0 and a temperature of 40 °C. Its activity was not inhibited by DTT and EDTA, but strongly inhibited by Cu²⁺, Zn²⁺, and Co²⁺. The K _m and V _max for L-2-chloropropionate were 0.3 mM and 23.8 μmol/min/mg, respectively. Its substrate specificity was limited to short chain mono-substituted 2-halocarboxylic acids, with no activity detected toward fluoropropionate and monoiodoacetate. This is the first report on the purification and characterization of 2-haloacid dehalogenase from a marine bacterium.     

Screening tomato-associated bacteria for biological control of grey mold on tomato

Aiming at discovering effective biocontrol agents (BCAs) against grey mold on tomato caused by Botrytis cinerea Pers., we selected 819 bacterial isolates from the surface as well as the interior of the roots, stems, and leaves of tomato plants grown in B. cinerea-infested fields. In a dual-culture assay, 116 isolates (14.16%) showed antagonism against B. cinerea and fewer ones against five additional tomato-associated fungal pathogens – Pythium ultimum, Phytophthora capsici, Fusarium oxysporum f. sp. lycopersici, Sclerotinia sclerotiorum and Ralstonia solanacearum. Thirty-one isolates with antagonism to B. cinerea and at least one of the five additional pathogens were assessed for their efficacy in controlling grey mold on tomato in a greenhouse test. Thirteen of them attained the efficacy over 50% and were subjected to the second greenhouse test, in which 12 isolates consistently accomplished the biocontrol efficacy over 50%, with isolates ABc28 and ABc22 achieving the efficacy of 66.71% and 64.90%, respectively. Under greenhouse conditions, the above two as well as isolates ABc2, ABc11 and ABc17 increased tomato biomass by more than 20% in comparison with the control. The 12 antagonistic isolates accomplishing the biocontrol efficacy over 50% in both greenhouse tests were considered potential BCAs against grey mold, which were identified as Pseudomonas spp., Pantoea spp., Bacillus spp. and Chryseobacterium spp. Ten of them were found to produce at least one of the three hydrolytic enzymes (protease, cellulase and chitinase) and/or siderophore, which might be involved in their mechanisms of suppressing the disease. Based on the origin of these 12 strains, the leaf tissue, especially the leaf interior, of tomato plants grown in a B. cinerea-infested field appears to be a good source of potential BCAs against grey mold.     

Control efficacy of Bacillus thuringiensis and a new granulovirus isolate against Cydia pomonella in orchards

Abstract

Codling moth, Cydia pomonella, is one of the most serious pests of apple and pear worldwide. This study evaluates the efficacies of a granulovirus, a Bacillus thuringiensis (Bt) strain and their combination in the control of C. pomonella in China. A Cydia pomonella granulovirus (CpGV) was isolated from C. pomonella cadavers in an orchard in Gansu, China. Droplet-feeding bioassays showed the median lethal concentration (LC₅₀) of this CpGV isolate (CpGV-C1) against the third instar C. pomonella larvae was 770 OBs µl^?1. The LC₅₀ values of Bt C-33 and kurstaki HD-1 against the third instar larvae were 26.3 µg ml^?1 and 15.7 µg ml^?1, respectively. Field tests indicated the control efficacies of CpGV-C1 and the combination of CpGV-C1 and Bt against C. pomonella larvae in apple orchards were similar to that of beta-cypermethrin. Our data demonstrated that the combination of CpGV and Bt might effectively protect apple fruits from the damage of C. pomonella larvae and had the potential to be developed as a low-cost, highly effective insecticide.     

A Method for the Enhancement of Environmental Stress Resistance of Endoparasitic Fungus Esteya vermicola

Esteya vermicola is the first recorded endoparasitic fungus of the pinewood nematode, Bursaphelenchus xylophilus, which is the causal agent for the pine wilt disease. Culture on modified agar media with herbal extraction (0.5%) was found to be able to induce resistance to UV radiation, heat and drought conditions in Esteya vermicola. Herba Houttuyniae, Tatraxacum officinale and Scutellaria baicalensis Georgi exhibited the highest improvement on environmental competence of Esteya vermicola at all the tested time points under the stress conditions. In addition, improved quality and effective viability of Esteya vermicola were observed amended with the three herbal extractions in culture media. Enhanced stress resistance was associated with herbal metabolites. These findings provided a green, feasible, economical method for developing an open‐field spay application of fungal biocontrol agents against pine wilt disease.     

Tim-1分子在约氏疟原虫感染模型中的表达分析

Tim-1分子表达在T细胞和调节性B细胞表面,具有促进Th2应答的作用。为探讨Tim-1分子在抗疟疾免疫应答中的作用,利用致死型约氏疟原虫(Plasmodium yoelii)感染鼠疟模型研究了Tim-1分子表达与小鼠感染结局和免疫应答模式的关系。结果显示,BALB/c小鼠对P.yoelii易感,在感染后6～7 d全部死亡,感染后第3、5天脾内细胞因子IFN-γmRNA水平明显低于对P.yoelii抵抗的DBA2小鼠,而脾IL-10 mRNA水平显著高于DBA2小鼠。BALB/c小鼠脾内Tim-1+T、B细胞百分比在感染后第3、5天显著升高,而DBA2小鼠感染前后脾内Tim-1+T、B细胞百分比没有显著变化。结果提示,Tim-1分子参与抗P.yoelii免疫应答的调节,可能与易感鼠产生高水平Th2型细胞因子有关。