Embryonic expression of juvenile hormone binding protein and its relationship to the toxic effects of juvenile hormone in Manduca sexta

The juvenile hormones (JHs) regulate a diverse array of insect developmental and reproductive processes. One molecular target of JH action is its transporter, hemolymph JH binding protein (hJHBP); in the larva of the tobacco hornworm, Manduca sexta, low doses of JH can immediately increase hJHBP gene expression. Less explored are the effects of JH on embryological development, where early hormonal treatment has been shown to affect embryonic development and pupation. This study examines the egg form of JHBP and its gene expression during embryogenesis of M. sexta, as well as the phenotypic effect JH treatment has on embryos and on JHBP gene expression. We here demonstrate that the preponderance of JHBP found in the egg is maternally derived and that the embryonic gene and protein appear identical to those found in the larva. Expression of the JHBP gene begins in both the embryo itself and extra-embryonic tissues 15 h after fertilization, long before emergence of a functional fat body and circulatory system. Topical application of low JH doses to early embryos resulted in larval abnormalities while high doses of the hormone induced embryonic mortality. These effects are not mediated through regulation of the JHBP gene, since embryonic expression appears invariant in response to JH challenge. The toxicity of JH is tightly correlated with the concentration of unbound hormone.     

Expression and characterization of an epoxide hydrolase from Anopheles gambiae with high activity on epoxy fatty acids

In insects, epoxide hydrolases (EHs) play critical roles in the metabolism of xenobiotic epoxides from the food resources and in the regulation of endogenous chemical mediators, such as juvenile hormones. Using the baculovirus expression system, we expressed and characterized an epoxide hydrolase from Anopheles gambiae (AgEH) that is distinct in evolutionary history from insect juvenile hormone epoxide hydrolases (JHEHs). We partially purified the enzyme by ion exchange chromatography and isoelectric focusing. The experimentally determined molecular weight and pI were estimated to be 35 kD and 6.3 respectively, different than the theoretical ones. The AgEH had the greatest activity on long chain epoxy fatty acids such as 14,15-epoxyeicosatrienoic acids (14,15-EET) and 9,10-epoxy-12Z-octadecenoic acids (9,10-EpOME or leukotoxin) among the substrates evaluated. Juvenile hormone III, a terpenoid insect growth regulator, was the next best substrate tested. The AgEH showed kinetics comparable to the mammalian soluble epoxide hydrolases, and the activity could be inhibited by AUDA [12-(3-adamantan-1-yl-ureido) dodecanoic acid], a urea-based inhibitor designed to inhibit the mammalian soluble epoxide hydrolases. The rabbit serum generated against the soluble epoxide hydrolase of Mus musculus can both cross-react with natural and denatured forms of the AgEH, suggesting immunologically they are similar. The study suggests there are mammalian sEH homologs in insects, and epoxy fatty acids may be important chemical mediators in insects.     

A novel alkyne cholesterol to trace cellular cholesterol metabolism and localization

Cholesterol is an important lipid of mammalian cells and plays a fundamental role in many biological processes. Its concentration in the various cellular membranes differs and is tightly regulated. Here, we present a novel alkyne cholesterol analog suitable for tracing both cholesterol metabolism and localization. This probe can be detected by click chemistry employing various reporter azides. Alkyne cholesterol is accepted by cellular enzymes from different biological species (Brevibacterium, yeast, rat, human) and these enzymes include cholesterol oxidases, hydroxylases, and acyl transferases that generate the expected metabolites in in vitro and in vivo assays. Using fluorescence microscopy, we studied the distribution of cholesterol at subcellular resolution, detecting the lipid in the Golgi and at the plasma membrane, but also in the endoplasmic reticulum and mitochondria. In summary, alkyne cholesterol represents a versatile, sensitive, and easy-to-use tool for tracking cellular cholesterol metabolism and localization as it allows for manifold detection methods including mass spectrometry, thin-layer chromatography/fluorography, and fluorescence microscopy.     

中枢神经系统内神经元信号处理(英文)

一种新颖的轴突断端(axon bleb)膜片钳记录方法大力促进了中枢神经系统轴突功能的研究。我们的工作应用这一方法揭示了大脑皮层锥体神经元的数码信号(具全或无特性的动作电位)的爆发和传播机制。在轴突始段(axon initial segment,AIS)远端高密度聚集的低阈值Na+通道亚型Nav1.6决定动作电位的爆发;而在AIS近端高密度聚集的高阈值Na+通道亚型Nav1.2促进动作电位向胞体和树突的反向传播。应用胞体和轴突的同时记录,我们发现胞体阈下膜电位的变化可以在轴突上传播较长的距离并可到达那些离胞体较近的突触前终末。进一步的研究证明了胞体膜电位的变化调控动作电位触发的突触传递,该膜电位依赖的突触传递是一种模拟式的信号传递。轴突上一类特殊K+通道(Kv1)的活动调制动作电位的波形,特别是其波宽,从而调控各种突触前膜电位水平下突触强度的变化。突触前终末的背景Ca2+浓度也可能参与模拟信号的传递。这些发现深化了我们对中枢神经系统内神经信号处理基本原理的认识,进而帮助我们理解脑如何工作。     

Cell cycle arrest induced by the vitamin D(3) analog EB1089 in NCI-H929 myeloma cells is associated with induction of the cyclin-dependent kinase inhibitor p27

EB1089, a 1,25-dihydroxyvitamin D(3) analog, has been known to have potent antiproliferative properties in a variety of malignant cells in vitro and in vivo. In the present study, we analyzed the effect of EB1089 on human myeloma cell lines. EB1089 inhibited the proliferation of NCI-H929 cells and RPMI8226 cells in a dose-dependent manner among three myeloma cell lines tested. The antiproliferative effect of EB1089 on myeloma cells was related to the expression level of vitamin D receptor. To investigate the mechanism of the antiproliferative effect of EB1089, cell cycle analysis was attempted in EB1089-sensitive NCI-H929 cells. EB1089 (1 x 10(-8) M) efficiently induced G(1) arrest of the cell cycle. Analysis of G(1) regulatory proteins demonstrated that protein levels of CDK2, CDK4, cyclin D1, and cyclin A were decreased in a time-dependent manner, but not those of CDK6 and cyclin E, by EB1089. In addition, EB1089 (1 x 10(-8) M, 72 h) increased the protein level of the CDKI p27 and markedly enhanced the binding of p27 with CDK2 compared to EB1089-untreated cells. Furthermore, the activity of CDK2-associated cyclin kinase was decreased, which was accompanied by the reduction of cyclin-D1-, cyclin-E-, and cyclin-A-associated kinase activities, resulting in the hypophosphorylation of Rb protein. These results suggest that EB1089 can inhibit the proliferation of human myeloma cells, especially NCI-H929 cells, via a G(1) block in association with the induction of p27 and the reduction of CDK2 activity.     

Effect of various metal ions on growth of two wheat lines known to differ in aluminium tolerance

The effects of aluminium (Al), manganese (Mn), zinc (Zn), copper (Cu), boron (B), iron (Fe), gallium (Ga), scandium (Sc) and lanthanum (La) on growth of an Al-tolerant and an Al-sensitive line of wheat (Triticum aestivum L.) were measured in solution culture. The concentrations of nutrients in the basal nutrient solution were (M) 500 Ca, 100 Mg, 300 K, 600 N (150 NH₄, 450 NO₃), 600 SO₄, 2.5 P, 3 B, 2.5 Fe, 0.5 Zn, 0.5 Mn, 0.1 Cu at a pH of 4.7. The major solution nutrient concentrations were maintained at the nominal concentration with monitoring, frequent additions and weekly renewal. Differentiation in yield between the Al-tolerant and Al-sensitive line only occurred in the presence of Al indicating that, in the long term, none of the other metals tested could be used as an analog for Al. The visual symptoms in the roots of Cu toxicity (in both lines) and Al toxicity (in the sensitive line) were similar. The solution concentration (M) at which yield of the roots of the tolerant line was reduced by 50% was, in order of increasing tolerance, Cu 0.5, Sc 1.1, La 7.1, Ga 8.6, Al 15, Zn 19, Fe 84, B 490 and Mn 600.     

昆虫卵黄发生研究进展

昆虫卵黄发生研究进展李乾君,龚和,管致和（中国科学院动物研究所北京１０００８０）（北京农业大学植保系北京１０００９４）昆虫卵的成熟一般分为三个时期－－卵黄发生前期（Ｐｒｅｖｉｔｅｌｌｏｇｅｎｉｃｓｔａｇｅ）、卵黄发生期（ｖｉｔｅｌｌｏｇｅｎｉｃｓｔａ...     

小粒野生稻STK类抗病基因同源序列的克隆与分析

根据丝氨酸/苏氨酸蛋白激酶类(serine-threonine kinase,STK)抗病基因结构中保守结构域,设计引物,以小粒野生稻基因组DNA为模板,扩增获得10条STK类抗病基因同源序列.同源性分析表明其都具有STK保守结构域,与已克隆的STK类抗病基因有不同程度的相似性,为进一步克隆小粒野生稻中的STK类抗病基因提供依据.     

抗肿瘤药物帕玛度胺类似物的合成

目的：设计并合成抗肿瘤药物帕玛度胺的3位N取代的新型类似物。方法：从3-硝基邻苯二甲酸（2）和N-（叔丁氧羰基）．L-谷氨酰胺（4）出发,经过六步反应得到目标化合物。3-硝基邻苯二甲酸（2）经脱水制得3-硝基邻苯二甲酸酐（3）。用N-（叔丁氧羰基）-L-谷氨酰胺（4）经闭环、脱保护制得3-氨基-2,6-哌啶二酮三氟乙酸盐（6）。4与6经缩合、钯碳催化氢化制得免疫调节剂Pomalidomide（8）,（8）经过酰化得到3-乙酰氨基-N-（2,6-二氧代-3-哌啶基）邻苯二甲酰亚胺（1）。以（4）计总收率约34．9％。结果：得到3-乙酰氨基-N-（2,6-二氧代-3-哌啶基）-邻苯二甲酰亚胺（1）,应用于细胞活性测试。结论：改进了帕玛度胺的合成工艺,得到了3位N乙酰化的新型帕玛度胺类似物（1）,初步研究显示（1）的生物活性与帕玛度胺接近。