Frequency of infection with A and B supergroup <Emphasis Type="Italic">Wolbachia</Emphasis> in insects and pests associated with mulberry and silkworm

Wolbachia is a ubiquitous, Gram-negative, vertically transmitted, alpha-proteobacterium that causes an array of reproductive abnormalities including cytoplasmic incompatibility, feminization of genetic males, parthenogenesis in a number of insect species, among others. Wolbachia is now being exploited as an agent for pest and vector control. Previous surveys indicated that it is commonly seen in 16–76% of arthropods. In this paper, using polymerase chain reaction assay based on specific amplification of the ftsZ-A and-B supergroup Wolbachia gene fragments, we found that 30% of insects and pests screened were positive for Wolbachia. Among them 66.7% harbour double Wolbachia infection, while 33.3% harbour single Wolbachia infection. These results indicate widespread infection with both double and single Wolbachia, and provide a wealth of information to exploit this endobacterium for the management of pests and vectors.     

与成纤维细胞共培养的肺泡II型上皮细胞的生物学特性

建立胎鼠肺泡II型上皮细胞(AECII)与肺成纤维细胞(LF)共培养模型,观察与LF共培养下AECII的生物学特性。倒置相差显微镜观察AECII形态和基本生长情况;RT-PCR和流式细胞术分别检测肺泡表面活性蛋白-C(SP-C)、水通道蛋白5(AQP5)mRNA及蛋白质表达;流式细胞术检测细胞周期及Ki67表达。结果显示,与LF共培养时,AECII能较好地保留其细胞形态,SP-CmRNA及其蛋白质表达明显增加,而AQP5mRNA及其蛋白质表达则明显减少;LF促进AECII增殖,使G2/M、S期细胞及表达Ki67 细胞的比率明显增多。结果提示,AECII与LF共培养时,能更好地保留其细胞形态、分化及增殖特性。     

金鱼Prox1基因cDNA的克隆及其在眼睛发育中的表达分析

脊椎动物的Prox1基因,与果蝇的转录因子prospero同源。为了探讨Prox1基因在金鱼眼睛发生过程中的表达图式,我们从金鱼眼睛SMART库中克隆了Prox1cDNA。它全长共2851bp,编码739个氨基酸。组织分布研究表明,Prox1主要分布于眼、脑、心、肝、脾和肾中。整体原位杂交显示,Prox1mRNA首先是在晶体期的晶体原基中有转录,心跳期则在未成熟晶体的细胞中和视网膜的幼芽区可以检测到。晶体纤维形成后,它主要定位于视纤维层和内网织细胞层。免疫组化显示,心跳期Prox1蛋白的定位与mRNA相同,晶体纤维形成以后,Prox1蛋白主要定位在晶体上皮细胞内侧的晶体纤维上一个环状区域,与Prox1mRNA的定位不同。这说明,Prox1基因在晶体发生过程中有重要作用,且在晶体的不同发育时期起的作用可能有所不同。另外,Prox1在晶体发育过程中有一个从内向外的变化过程。     

Structural basis of the collagen-binding mode of discoidin domain receptor 2

Discoidin domain receptor (DDR) is a cell-surface receptor tyrosine kinase activated by the binding of its discoidin (DS) domain to fibrillar collagen. Here, we have determined the NMR structure of the DS domain in DDR2 (DDR2-DS domain), and identified the binding site to fibrillar collagen by transferred cross-saturation experiments. The DDR2-DS domain structure adopts a distorted jellyroll fold, consisting of eight beta-strands. The collagen-binding site is formed at the interloop trench, consisting of charged residues surrounded by hydrophobic residues. The surface profile of the collagen-binding site suggests that the DDR2-DS domain recognizes specific sites on fibrillar collagen. This study provides a molecular basis for the collagen-binding mode of the DDR2-DS domain.     

Genetic relationships among Chinese pigs and other pig populations from Hunan Province, China

In this study, protein-level polymorphisms of transferrin, pre-albumin, hemopexin, ceruloplasmin and amylase were investigated in Hunan native pigs and Large Yorkshire pigs collected from Hunan (a province of China), allowing calculations of allele frequencies, average heterozygosities, inbreeding coefficients and genetic distances. The genetic relationship between Southeast Asian native pigs and American pigs was more distant than those among Southeast Asian native pig breeds. The genetic relationship between Southeast Asian native pig breeds and Hampshire pigs was the most distant.     

Phenotypical characteristics of Shiga-like toxin Escherichia coli isolated from sheep dairy products

AIMS: To analyse phenotypical characteristics of Shiga toxin-producing Escherichia coli (STEC) strains from ovine origin. METHODS AND RESULTS: A total of 13 STEC strains (eight O157 and five non-O157) isolated from sheep dairy products were used in this study. Biochemical traits, motility, haemolytic activity, resistance to tellurite-cefixime, maximum growth temperature and antibiotic resistance were determined. The STEC strains were grouped into nine biochemical and physiological biotypes (five for the O157 and four for the non-O157 strains). All STEC strains showed resistance to bacitracin, cloxacilin, penicillin and tylosin. CONCLUSIONS: Different biotypes and antibiotic resistance patterns of STEC isolated from sheep dairy products were observed. SIGNIFICANCE AND IMPACT OF THE STUDY: This work will be a contribution to the better characterization of STEC isolated from sheep dairy products, which have, to date, been scarcely studied, and to the better understanding of the risks associated with its consumption.     

Gas exchange and water relations of three <Emphasis Type="Italic">Vitis vinifera</Emphasis> L. cultivars growing under Mediterranean climate

Optical characteristics, contents of photosynthetic pigments, total soluble sugars, and starch, rates of gas exchange, chlorophyll (Chl) a fluorescence, and leaf water relations were analysed in three Vitis vinifera L. cultivars, Tinto Cão (TC), Touriga Nacional (TN), and Tinta Roriz (TR), grown in Mediterranean climate. Chl content was significantly lower in TC than in TN and TR leaves, while the Chl a/b ratio was higher. TR had the lowest net photosynthetic rate, stomatal conductance, and contents of soluble sugars and starch than TN and TC. In spite of low Chl content, TC showed the lowest photon absorbance and the highest photochemical efficiency of photosystem 2. TC had the lowest predawn and midday leaf water potential. The capability for osmotic adjustment was similar among cultivars and the calculated modulus of elasticity was higher in TC leaves. The typical lighter green leaves of TC seemed to be an adaptive strategy to high irradiance and air temperature associated to water stress.     

FTIR studies of the redox partner interaction in cytochrome P450: The Pdx–P450cam couple

Recently we have developed a new approach to study protein–protein interactions using Fourier transform infrared spectroscopy in combination with titration experiments and principal component analysis (FTIR-TPCA). In the present paper we review the FTIR-TPCA results obtained for the interaction between cytochrome P450 and the redox partner protein in two P450 systems, the Pseudomonas putida P450cam (CYP101) with putidaredoxin (P450cam–Pdx), and the Bacillus megaterium P450BM-3 (CYP102) heme domain with the FMN domain (P450BMP–FMND). Both P450 systems reveal similarities in the structural changes that occur upon redox partner complex formation. These involve an increase in β-sheets and α-helix content, a decrease in the population of random coil/3₁₀-helix structure, a redistribution of turn structures within the interacting proteins and changes in the protonation states or hydrogen-bonding of amino acid carboxylic side chains. We discuss in detail the P450cam–Pdx interaction in comparison with literature data and conclusions drawn from experiments obtained by other spectroscopic techniques. The results are also interpreted in the context of a 3D structural model of the Pdx–P450cam complex.     

Characterization and development of a peptide substrate-based phosphate transfer assay for the human vascular endothelial growth factor receptor-2 tyrosine kinase

Vascular endothelial growth factor (VEGF), a critical regulator in angiogenesis, exerts its angiogenic effect via binding to its receptor, VEGF receptor-2 tyrosine kinase (VEGFR2) or kinase insert domain-containing receptor (Kdr), on the surface of endothelial cells. Kdr-mediated signaling plays an important role in the proliferation, migration, differentiation, and survival of endothelial cells. Therefore, the inhibition of this signaling pathway represents a promising therapeutic approach for the discovery of novel anticancer agents by destabilizing the progression of solid tumors via abrogating tumor-induced angiogenesis. To explore Kdr as an anticancer target and further characterize the enzyme, we purified a cytoplasmic domain of human Kdr (Kdr-CD) and characterized its autophosphorylation activity. We also designed and synthesized peptides containing amino acid sequences corresponding to the autophosphorylation sites of Kdr and developed a simple, robust, high-throughput assay for measuring the phosphate transfer activity of the enzyme. This assay was validated by the experiments showing that the phosphate transfer activity of the purified Kdr-CD required Mg2+ or Mn2+ and preactivation by adenosine 5'-triphosphate (ATP) and was inhibited by known Kdr inhibitors. Using this assay, we examined effects of Mg2+ and Mn2+ on the enzyme activity; optimized the concentrations of Kdr-CD, peptide and ATP substrates, and metal ions in the assay; and determined the kinetic properties of the enzyme for the peptide and ATP as well as IC50 values of two known Kdr inhibitors. Thus, the results of these studies have validated the utilities of this assay for biochemical characterizations of the enzyme and its inhibitors. This approach of designing peptides corresponding to the autophosphorylation sites of Kdr as substrates for the enzyme has general practical implications to other kinases.