Cloning and expression of thermophilic catechol 1,2-dioxygenase gene (catA) from Streptomyces setonii

Streptomyces setonii (ATCC 39116) degrades various single aromatic compounds such as phenol or benzoate via an ortho-cleavage pathway using catechol 1,2-dioxygenase (C12O). A PCR using degenerate primers based on the conserved regions of known C12O-encoding genes amplified a 0.45-kbp DNA fragment from S. setonii total DNA. A Southern hybridization analysis and size-selected DNA library screening using the 0.45-kbp PCR product as a probe led to the isolation of a 6.4-kbp S. setonii DNA fragment, from which the C12O-encoding genetic locus was found to be located within a 1.4-kbp DNA fragment. A complete nucleotide sequencing analysis of the 1.4-kbp DNA fragment revealed a 0.84-kbp open reading frame, which showed a strong overall amino acid similarity to the known high-G+C Gram-positive (but significantly less to the Gram-negative) bacterial mesophilic C12Os. The heterologous expression of the cloned 1.4-kbp DNA fragment in Escherichia coli demonstrated that this C12O possessed a thermophilic activity within a broad temperature range (up to 65 degrees C) and showed a higher activity against 3-methylcatechol than catechol or 4-methylcatechol, but no activity against protocatechuate.     

Molecular phylogeny and species delimitation of Stachyuraceae: Advocating a herbarium specimen‐based phylogenomic approach in resolving species boundaries

Species concept and delimitation are fundamental to taxonomic and evolutionary studies. Both inadequate informative sites in the molecular data and limited taxon sampling have often led to poor phylogenetic resolution and incorrect species delineation. Recently, the whole chloroplast genome sequences from extensive herbarium specimen samples have been shown to be effective to amend the problem. Stachyuraceae are a small family consisting of only one genus Stachyurus of six to 16 species. However, species delimitation in Stachyurus has been highly controversial because of few and generally unstable morphological characters used for classification. In this study, we sampled 69 individuals of seven species (each with at least three individuals) covering the entire taxonomic diversity, geographic range, and morphological variation of Stachyurus from herbarium specimens for genome‐wide plastid gene sequencing to address species delineation in the genus. We obtained high‐quality DNAs from specimens using a recently developed DNA reconstruction technique. We first assembled four whole chloroplast genome sequences. Based on the chloroplast genome and one nuclear ribosomal DNA sequence of Stachyurus, we designed primers for multiplex polymerase chain reaction and high throughput sequencing of 44 plastid loci for species of Stachyurus. Data of these chloroplast DNA and nuclear ribosomal DNA internal transcribed spacer sequences were used for phylogenetic analyses. The phylogenetic results showed that the Japanese species Stachyurus praecox Siebold & Zucc. was sister to the rest in mainland China, which indicated a typical Sino‐Japanese distribution pattern. Based on diagnostic morphological characters, distinct distributional range, and monophyly of each clade, we redefined seven species for Stachyurus following an integrative species concept, and revised the taxonomy of the family based on previous reports and specimens, in particular the type specimens. Furthermore, our divergence time estimation results suggested that Stachyuraceae split from its sister group Crossosomataceae from the New World at ca. 54.29 Mya, but extant species of Stachyuraceae started their diversification only recently at ca. 6.85 Mya. Diversification time of Stachyurus in mainland China was estimated to be ca. 4.45 Mya. This research has provided an example of using the herbarium specimen‐based phylogenomic approach in resolving species boundaries in a taxonomically difficult genus.     

Two effects of combined nitrogen on the adhesion of Rhizobium etli to bean roots

Combined forms of nitrogen negatively influence rhizobia-legume symbiosis. The effects of combined nitrogen are known for nodulation and dinitrogen (N₂) fixation, but little is known about the effect on preinfection events. Here, we studied the effects of combined nitrogen on the adhesion of Rhizobium etli to common bean (Phaseolus vulgaris L.) roots. When potassium nitrate (KNO₃) or sodium glutamate was added to an incubation mixture of rhizobia and plants that were previously grown in nitrogen-free solution, rhizobial adhesion to roots was stimulated. However, the rhizobial adhesion to bean roots that were previously grown with 10 mM KNO₃ was reduced by half. A fraction of the bean root exudates, which is thermolabile and has molecular mass larger than 12 kDa stimulated rhizobial adhesion, but this stimulatory activity was lost in root exudates obtained with 10 mM KNO₃. Thus, the inhibition of symbiosis in response to combined nitrogen may be controlled by the plant at the preinfection stage as well.     

中国沙棘幼苗对土壤镉生理耐性和富集转运特征的性别差异

为探究中国沙棘对土壤镉(Cd)胁迫的性别响应差异,该研究以中国沙棘2年生幼苗为材料,利用盆栽试验研究在不同浓度Cd处理下(0(CK)、25、50、100和200 mg·kg^-1)雌、雄株幼苗的生长、叶片生理特性以及Cd富集特征的差异。结果表明：(1)Cd处理下中国沙棘幼苗雌、雄株的株高和基径生长以及各器官生物量均表现出低浓度(<50 mg·kg^-1)促进,高浓度(> 100 mg·kg^-1)抑制的现象;低浓度Cd处理下雌株的株高、基径增长率和生物量的增幅均高于雄株;高浓度Cd处理下(200 mg·kg^-1)雄株株高增长率、叶生物量和总生物量分别较CK显著降低,而雌株均未显著下降。(2)随着Cd浓度升高,雌、雄株叶片光合色素含量和抗氧化酶活性呈先升后降的变化趋势,丙二醛(MDA)和渗透调节物质含量呈上升趋势;Cd浓度为50～200 mg·kg^-1时,雌株叶片的光合色素含量、抗氧化酶活性和渗透调节物质含量均高于雄株,而MDA含量始终低于雄株。(3)随着Cd浓度升高,雌、...     

中国晚期旧石器的碳-14断代和问题

根据已公布的47个碳-14数据,进而分析10处中国晚期遗址的年代和问题,并提出不同的看法。文中强调:露天遗址中碳-14数据的异常现象,往往与各种原因形成的再次堆积有关。因此必须注意样品的采集和避免引用孤零的碳-14数据,同时还要结合地层和文化性质的分析,才可能保证断代的准确性。     

The Expression of Connexins and SOX2 Reflects the Plasticity of Glioma Stem-Like Cells

Glioblastoma (GBM) is the most malignant primary brain tumor, with an average survival rate of 15 months. GBM is highly refractory to therapy, and such unresponsiveness is due, primarily, but not exclusively, to the glioma stem-like cells (GSCs). This subpopulation express stem-like cell markers and is responsible for the heterogeneity of GBM, generating multiple differentiated cell phenotypes. However, how GBMs maintain the balance between stem and non-stem populations is still poorly understood. We investigated the GBM ability to interconvert between stem and non-stem states through the evaluation of the expression of specific stem cell markers as well as cell communication proteins. We evaluated the molecular and phenotypic characteristics of GSCs derived from differentiated GBM cell lines by comparing their stem-like cell properties and expression of connexins. We showed that non-GSCs as well as GSCs can undergo successive cycles of gain and loss of stem properties, demonstrating a bidirectional cellular plasticity model that is accompanied by changes on connexins expression. Our findings indicate that the interconversion between non-GSCs and GSCs can be modulated by extracellular factors culminating on differential expression of stem-like cell markers and cell-cell communication proteins. Ultimately, we observed that stem markers are mostly expressed on GBMs rather than on low-grade astrocytomas, suggesting that the presence of GSCs is a feature of high-grade gliomas. Together, our data demonstrate the utmost importance of the understanding of stem cell plasticity properties in a way to a step closer to new strategic approaches to potentially eliminate GSCs and, hopefully, prevent tumor recurrence.     

Morphological alterations by ectopic expression of the rice OsMADS4 gene in tobacco plants

OsMADS4, a rice MADS-box gene, is a member of the GLO/PI family that specifies the identity of petals and stamens in combination with other MADS-box genes. We report here the ectopic expression of OsMADS4 fused to the CaMV 35S promoter in tobacco plants. Transgenic plants carrying the CaMV 35S promoter::OsMADS4 construct generated mutant flowers with a mosaic carpel, in which the tissue around the nectary was elongated and the styles reduced. The fruits were distorted, but viable seeds did develop. These phenotypes mimicked those of transgenic tobacco plants that ectopically express Antirrhinum GLO. However, unlike GLO, OsMADS4 did not cause any homeotic change in the first whorl of the transgenic flowers. These results suggest that the functional role of OsMADS4 in the outer whorls has diverged from that of its dicot counterparts.     

Catalpol protects dopaminergic neurons from LPS-induced neurotoxicity in mesencephalic neuron-glia cultures

Inflammation plays an important role in the pathogenesis of Parkinson's disease (PD). Microglia, the resident immune cells in the central nervous system, are pivotal in the inflammatory reaction. Activated microglia can induce expression of inducible nitric-oxide synthase (iNOS) and release significant amounts of nitric oxide (NO) and TNF-alpha, which can damage the dopaminergic neurons. Catalpol, an iridoid glycoside, contained richly in the roots of Rehmannia glutinosa, was found to be neuroprotective in gerbils subjected to transient global cerebral ischemia. But the effect of catalpol on inflammation-mediated neurodegeneration has not been examined. In this study, microglia in mesencephalic neuron-glia cultures were activated with lipopolysaccharide (LPS) and the aim of the study was to examine whether catalpol could protect dopaminergic neurons from LPS-induced neurotoxicity. The results showed that catalpol significantly reduced the release of reactive oxygen species (ROS), TNF-alpha and NO after LPS-induced microglial activation. Further, catalpol attenuated LPS-induced the expression of iNOS. As determined by immunocytochemical analysis, pretreatment by catalpol dose-dependently protected dopaminergic neurons against LPS-induced neurotoxicity. These results suggest that catalpol exerts its protective effect on dopaminergic neurons by inhibiting microglial activation and reducing the production of proinflammatory factors. Thus, catalpol may possess therapeutic potential against inflammation-related neurodegenerative diseases.     

Synonymous Mutations Reduce Genome Compactness in Icosahedral ssRNA Viruses

Recent studies have shown that single-stranded (ss) viral RNAs fold into more compact structures than random RNA sequences with similar chemical composition and identical length. Based on this comparison, it has been suggested that wild-type viral RNA may have evolved to be atypically compact so as to aid its encapsidation and assist the viral assembly process. To further explore the compactness selection hypothesis, we systematically compare the predicted sizes of >100 wild-type viral sequences with those of their mutants, which are evolved in silico and subject to a number of known evolutionary constraints. In particular, we enforce mutation synonynimity, preserve the codon-bias, and leave untranslated regions intact. It is found that progressive accumulation of these restricted mutations still suffices to completely erase the characteristic compactness imprint of the viral RNA genomes, making them in this respect physically indistinguishable from randomly shuffled RNAs. This shows that maintaining the physical compactness of the genome is indeed a primary factor among ssRNA viruses’ evolutionary constraints, contributing also to the evidence that synonymous mutations in viral ssRNA genomes are not strictly neutral.