Identifying the basal angiosperm node in chloroplast genome phylogenies: sampling one's way out of the Felsenstein zone

While there has been strong support for Amborella and Nymphaeales (water lilies) as branching from basal-most nodes in the angiosperm phylogeny, this hypothesis has recently been challenged by phylogenetic analyses of 61 protein-coding genes extracted from the chloroplast genome sequences of Amborella, Nymphaea, and 12 other available land plant chloroplast genomes. These character-rich analyses placed the monocots, represented by three grasses (Poaceae), as sister to all other extant angiosperm lineages. We have extracted protein-coding regions from draft sequences for six additional chloroplast genomes to test whether this surprising result could be an artifact of long-branch attraction due to limited taxon sampling. The added taxa include three monocots (Acorus, Yucca, and Typha), a water lily (Nuphar), a ranunculid (Ranunculus), and a gymnosperm (Ginkgo). Phylogenetic analyses of the expanded DNA and protein data sets together with microstructural characters (indels) provided unambiguous support for Amborella and the Nymphaeales as branching from the basal-most nodes in the angiosperm phylogeny. However, their relative positions proved to be dependent on the method of analysis, with parsimony favoring Amborella as sister to all other angiosperms and maximum likelihood (ML) and neighbor-joining methods favoring an Amborella + Nymphaeales clade as sister. The ML phylogeny supported the later hypothesis, but the likelihood for the former hypothesis was not significantly different. Parametric bootstrap analysis, single-gene phylogenies, estimated divergence dates, and conflicting indel characters all help to illuminate the nature of the conflict in resolution of the most basal nodes in the angiosperm phylogeny. Molecular dating analyses provided median age estimates of 161 MYA for the most recent common ancestor (MRCA) of all extant angiosperms and 145 MYA for the MRCA of monocots, magnoliids, and eudicots. Whereas long sequences reduce variance in branch lengths and molecular dating estimates, the impact of improved taxon sampling on the rooting of the angiosperm phylogeny together with the results of parametric bootstrap analyses demonstrate how long-branch attraction might mislead genome-scale phylogenetic analyses.     

The effects of phenytoin and its metabolite 5-(4-hydroxyphenyl)-5-phenylhydantoin on cellular glucose transport

Glucose is the principal fuel for brain metabolism and its movement across the blood-brain barrier depends on Glut1. Impaired glucose transport to the brain may have deleterious consequences. For example, Glut1 deficiency syndrome (Glut1DS) is the result of heterozygous loss of function Glut1 mutation leading to energy failure of the brain and subsequently, epileptic encephalopathy. To preserve the integrity of the energy supply to the brain in patients with compromised glucose transport function, consumption of compounds with glucose transport inhibiting properties should be avoided. Phenytoin is a widely used anticonvulsant that affects carbohydrate metabolism. In this study, the hypothesis that phenytoin and its metabolite 5-(4-hydroxyphenyl)-5-phenylhydantoin (HPPH) affect cellular glucose transport was tested. With a focus on Glut1, the effects of phenytoin and HPPH on cellular glucose transport were studied. Glucose uptake assay measuring the zero-trans influx of radioactive-labeled glucose analogues showed that phenytoin and HPPH did not exert immediate effects on erythrocyte Glut1 activity or glucose transport in Hs68 control fibroblasts, Glut1DS primary fibroblasts isolated from two patients, or in rat primary astrocytes. Prolonged exposure to the two compounds could stimulate glucose transport by up to 30-60% over the control level (p <0.05) in Hs68 and Glut1DS fibroblasts as well as in rat astrocytes. The stimulation of glucose transport by HPPH was dose-dependent and accompanied by an up-regulation of GLUT1 mRNA expression (p <0.05). In conclusion, phenytoin and HPPH do not compromise cellular glucose transport. Prolonged exposure to these compounds can modify carbohydrate homeostasis by up-regulating glucose transport in both normal and Glut1DS conditions in vitro.     

Syntheses of quinolone hydrochloride enantiomers from synthons (R)- and (S)-2-methylpiperazine

A series of R and S enantiomers of 7-(3-methylpiperazin-1-yl) quinolone derivatives were synthesized from (R)- and (S)-tert-butyl 2-methylpiperazine-1-carboxylate and tested for their antibacterial activities on 14 kinds of bacteria. Although no distinct difference in in vitro antibacterial activities was observed, 2-64-fold difference between R and S enantiomers was observed in approximately 52% of cases.     

噪声习服对听觉损伤的保护作用机制探讨

目的:探讨噪声习服对听觉损伤的保护作用机制.方法:建立噪声习服实验动物模型.采用免疫组织化学、激光扫描共聚焦显微镜(LSCM)及图像分析等技术,定量研究噪声习服后毛细胞内纤维状肌动蛋白(F-actin)、钙调蛋白(CaM)、热休克蛋白70(HSP70)的表达及游离Ca2 浓度的变化.结果:噪声暴露后毛细胞中F-actin、CaM及HSP70的表达均呈增加趋势.与噪声损伤暴露组(H组)比较,噪声习服后损伤暴露组(CH组)中F-actin和HSP70的表达均明显增多,CaM的表达具有增加趋势.声暴露后毛细胞内游离Ca2 浓度升高,噪声损伤暴露组毛细胞内游离Ca2 浓度明显高于噪声习服组(C组)和习服后损伤暴露组.结论:噪声习服使毛细胞对于其后声刺激的保护性反应增强,毛细胞内细胞骨架系统的加强及胞内钙稳态的维持在噪声习服的保护机制中具有重要意义.     

醛固酮对新生大鼠心肌成纤维细胞分泌ET、NO功能的影响

目的: 探讨醛固酮对新生大鼠心肌成纤维细胞(CFs)分泌内皮素(ET)、一氧化氮(NO)的影响.方法: 采用胰酶消化法和差速贴壁分离法获取CFs,应用放射免疫分析法、硝酸还原酶法分别测定不同条件下培养的CFs培养液中的ET、NO水平.结果: 一定浓度范围内的醛固酮可按剂量依赖方式促进CFs分泌ET,抑制CFs分泌NO,使ET/NO比值上升;螺内酯可阻断醛固酮的上述作用(P<0.01).结论: 醛固酮可能通过影响CFs分泌ET、NO,从而改变生物活性物质网络平衡关系,发挥其促心肌纤维化作用.     

Protein subunit interfaces: heterodimers versus homodimers

Protein dimers are either homodimers (complexation of identical monomers) or heterodimers (complexation of non-identical monomers). These dimers are common in catalysis and regulation. However, the molecular principles of protein dimer interactions are difficult to understand mainly due to the geometrical and chemical characteristics of proteins. Nonetheless, the principles of protein dimer interactions are often studied using a dataset of 3D structural complexes determined by X-ray crystallography. A number of physical and chemical properties govern protein dimer interactions. Yet, a handful of such properties are known to dominate protein dimer interfaces. Here, we discuss the differences between homodimer and heterodimer interfaces using a selected set of interface properties.     

Mapping of B cell epitopes on the zona pellucida 2 protein of a marsupial, the brushtail possum (Trichosurus vulpecula)

Immunocontraceptive vaccines against zona pellucida (ZP) proteins are being developed for brushtail possum (Trichosurus vulpecula) management in New Zealand. Mapping of B cell epitopes on the ZP2 protein of possums was undertaken in this study to define the antigenic regions that may be crucial to sperm-egg binding. The amino acid sequence of the full-length possum ZP2 protein (712 amino acids) was used to synthesize a complete set of 71 (15-mer) biotinylated peptides with an offset of five amino acids. The peptides were used in a modified enzyme-linked immunosorbent assay (ELISA) to identify continuous epitopes recognized by antibodies in the sera of possums immunized with recombinant possum ZP2 (rZP2) constructs. Seventeen continuous epitopes were located on possum ZP2 protein. Comparisons of the peptide binding pattern of antibodies in individual sera with the fertility status of the same immunized possums revealed three significant infertility-relevant peptide epitopes (amino acids 111-125, 301-315, and 431-445). One of these (amino acids 431-445) bound to possum spermatozoa from the caudal epididymis. The implications of these findings for developing immunocontraceptive vaccines for possum control are discussed.     

Drosophila Nod protein binds preferentially to the plus ends of microtubules and promotes microtubule polymerization in vitro

Nod, a nonmotile kinesin-like protein, plays a critical role in segregating achiasmate chromosomes during female meiosis. In addition to localizing to oocyte chromosomes, we show that functional full-length Nod-GFP (Nod(FL)-GFP) localizes to the posterior pole of the oocyte at stages 9-10A, as does kinesin heavy chain (KHC), a plus end-directed motor. This posterior localization is abolished in grk mutants that no longer maintain the microtubule (MT) gradient in the oocyte. To test the hypothesis that Nod binds to the plus ends of MTs, we expressed and purified both full-length Nod (Nod(FL)) and a truncated form of Nod containing only the motor-like domain (Nod318) from Escherichia coli and assessed their interactions with MTs in vitro. Both Nod(FL) and Nod318 demonstrate preferential binding to the ends of the MTs, displaying a strong preference for binding to the plus ends. When Nod318-GFP:MT collision complexes were trapped by glutaraldehyde fixation, the preference for binding to plus ends versus minus ends was 17:1. Nod(FL) and Nod318 also promote MT polymerization in vitro in a time-dependent manner. The observation that Nod is preferentially localized to the plus ends of MTs and stimulates MT polymerization suggests a mechanism for its function.