Analysis of genetic diversity in Glyptosternum maculatum (Sisoridae, Siluriformes) populations with AFLP markers

We determined the genetic diversity of geographic populations from three spawning grounds (Nyang River, Lhasa River, Shetongmon Reach of Yarlung Zangbo River) of Glyptosternum maculatum with amplified fragment length polymorphism (AFLP) markers. Five primer combinations detected 332 products, 51 of them (15.4%) were polymorphic in at least one population. The Shetongmon population was found to be the richest in genetic diversity as was indicated by the percentage of polymorphic loci and heterozygosity, followed by the Nyang population and the Lhasa population. The pair-wise genetic distance between populations were all very close, ranging from 0.0015 to 0.0042 with an average of 0.0024. The genetic distance was not proportional to the geographic distance. The analysis of molecular variance demonstrated that all variation occurred within populations. The average estimated fixation index (F _st) of three populations across all polymorphic loci was −0.0184, indicating the absence of genetic differences among the three sampled populations. The differentiation among populations was not significant, and population structure was weak. Our observations will help identify the genetic relationship among populations as the first approach to understand the genetic diversity of Glyptosternum maculatum.     

The impact of the International Atomic Energy Agency (IAEA) program on radiation and tissue banking in Indonesia

In 1986, the National Nuclear Energy Agency (Batan) in Jakarta started the research and development for the setting up of a tissue bank (Batan Research Tissue Bank/BRTB) by preserving fresh amnion or fetal membranes by lyophilisation and then sterilising by gamma irradiation. During the period of 1990 and 2000, three more tissue banks were set up, i.e., Biomaterial Centre in Surabaya, Jamil Tissue Bank in Padang, and Sitanala Tissue Bank in Tangerang. In 1994, BRTB produced bone allografts. The banks established under the IAEA program concentrated its work on the production of amnion, bone and soft tissues allografts, as well as bone xenografts. These tissues (allografts and xenografts) were sterilised using gamma irradiation (about 90%) and the rest were sterilized by ETO and those products have been used in the treatment of patients at more than 50 hospitals in Indonesia. In 2004, those tissue banks produced 8,500 grafts and 5,000 of them were amnion grafts for eye treatment and wound dressing. All of those grafts were used for patients as well as for research. In 2006, the production increased to 9,000 grafts. Although the capacity of those banks can produce more grafts, we are facing problems on getting raw materials from suitable donors. To fulfill the demand of bone grafts we also produced bone xenografts. The impact of the IAEA program in tissue banking activities in Indonesia can be summarised as follows: to support the national program on importing substitutes for medical devices. The price of imported tissues are between US$ 50 and US$ 6,000 per graft. Local tissue bank can produce tissues with the same quality with the price for about 10–30% of the imported tissues.     

Biochemical Analysis of a β-d-Xylosidase and a Bifunctional Xylanase-Ferulic Acid Esterase from a Xylanolytic Gene Cluster in Prevotella ruminicola 23

Prevotella ruminicola 23 is an obligate anaerobic bacterium in the phylum Bacteroidetes that contributes to hemicellulose utilization within the bovine rumen. To gain insight into the cellular machinery that this organism elaborates to degrade the hemicellulosic polymer xylan, we identified and cloned a gene predicted to encode a bifunctional xylanase-ferulic acid esterase (xyn10D-fae1A) and expressed the recombinant protein in Escherichia coli. Biochemical analysis of purified Xyn10D-Fae1A revealed that this protein possesses both endo-β-1,4-xylanase and ferulic acid esterase activities. A putative glycoside hydrolase (GH) family 3 β-d-glucosidase gene, with a novel PA14-like insertion sequence, was identified two genes downstream of xyn10D-fae1A. Biochemical analyses of the purified recombinant protein revealed that the putative β-d-glucosidase has activity for pNP-β-d-xylopyranoside, pNP-α-l-arabinofuranoside, and xylo-oligosaccharides; thus, the gene was designated xyl3A. When incubated in combination with Xyn10D-Fae1A, Xyl3A improved the release of xylose monomers from a hemicellulosic xylan substrate, suggesting that these two enzymes function synergistically to depolymerize xylan. Directed mutagenesis studies of Xyn10D-Fae1A mapped the catalytic sites for the two enzymatic functionalities to distinct regions within the polypeptide sequence. When a mutation was introduced into the putative catalytic site for the xylanase domain (E280S), the ferulic acid esterase activity increased threefold, which suggests that the two catalytic domains for Xyn10D-Fae1A are functionally coupled. Directed mutagenesis of conserved residues for Xyl3A resulted in attenuation of activity, which supports the assignment of Xyl3A as a GH family 3 β-d-xylosidase.β-1,4-linked xylopyranose is the principal component of plant cell wall hemicellulose, which represents the second largest reservoir of fixed carbon in the biosphere (11, 30, 34, 38, 42, 72). The catabolic breakdown of hemicellulose thus represents a critical step in the recycling of carbon in nature and has been targeted as a subject of intense research with respect to renewable energy resources. Two enzymes of principal importance for recycling hemicellulosic material are endo-1,4-β-xylanases (EC 3.2.1.8), which cleave the xylan backbone, and β-d-xylosidases (EC 3.2.1.37), which cleave xylose monomers from the nonreducing end of xylo-oligosaccharides (17).Prevotella ruminicola 23 is an important member of the anaerobic rumen microbiota (60) that contributes to the utilization of noncellulosic polysaccharides, such as starch and xylan (15, 25, 35, 55). Despite its documented importance in rumen physiology, relatively little is known about the cellular machinery that P. ruminicola 23 employs to harvest energy from hemicellulosic substrates. Several studies have explored the carbohydrate active enzymes from the related taxon Prevotella bryantii B₁4 (previously classified as P. ruminicola B₁4) (25, 27, 28, 50, 52, 66, 73); however, less is known about the xylanolytic system that P. ruminicola 23 elaborates. These two species share 88.9% nucleotide identity in their 16S rRNA genes (1,473 nucleotides aligned); however, there is evidence that these two species exhibit differences in their xylanolytic systems (2). Two xylanases from P. ruminicola 23 have been characterized: a 66-kDa xylanase with significant sequence identity to glycoside hydrolase (GH) family 5 endoglucanases (69, 70) and an 80-kDa xylanase related to GH family 10 endoxylanases which possesses a 260-amino-acid insertion within the putative catalytic domain (25). Recently, the draft genome sequence for P. ruminicola 23 has been obtained (http://www.tigr.org/tdb/rumenomics/), and a large number of GHs have been identified. Consistent with the predicted role of P. ruminicola 23 in hemicellulose degradation, many of these genes appear to encode enzymes important for hemicellulose depolymerization and utilization.In this study, we report the biochemical properties for two GHs with unique domain architectures from P. ruminicola 23. These two enzymes function synergistically to release monomeric sugars from a complex hemicellulosic substrate, xylan. This study, therefore, provides important insight into the molecular strategies for xylan depolymerization by P. ruminicola 23.     

GTP induces S-phase cell-cycle arrest and inhibits DNA synthesis in K562 cells but not in normal human peripheral lymphocytes

Since differentiation therapy is one of the promising strategies for treatment of leukemia, universal efforts have been focused on finding new differentiating agents. In that respect, we used guanosine 5'-triphosphate (GTP) to study its effects on K562 cell line. GTP, at concentrations between 25-200 microM, inhibited proliferation (3-90%) and induced 5-78% increase in benzidine-positive cells after 6-days of treatments of K562 cells. Flow cytometric analyses of glycophorine A (GPA) showed that GTP can induce expression of this marker in more mature erythroid cells in a time- and dose-dependent manner. These effects of GTP were also accompanied with inhibition of DNA synthesis (measured by [3H]-thymidine incorporation) and early S-phase cell cycle arrest by 96 h of exposure. In contrast, no detectable effects were observed when GTP administered to unstimulated human peripheral blood lymphocytes (PBL). However, GTP induced an increase in proliferation, DNA synthesis and viability of mitogen-stimulated PBL cells. In addition, growth inhibition and differentiating effects of GTP were also induced by its corresponding nucleotides GDP, GMP and guanosine (Guo). In heat-inactivated medium, where rapid degradation of GTP via extracellular nucleotidases is slow, the anti-proliferative and differentiating effects of all type of guanine nucleotides (except Guo) were significantly decreased. Moreover, adenosine, as an inhibitor of Guo transporter system, markedly reduced the GTP effects in K562 cells, suggesting that the extracellular degradation of GTP or its final conversion to Guo may account for the mechanism of GTP effects. This view is further supported by the fact that GTP and Guo are both capable of impeding the effects of mycophenolic acid. In conclusion, our data will hopefully have important impact on pharmaceutical evaluation of guanine nucleotides for leukemia treatments.     

新疆阿勒泰山两河源自然保护区地面生地衣的物种多样性

阿勒泰山地是我国著名水系额尔齐斯河和乌伦古河的发源地,该山地地衣植物的研究在中国乃至中亚都占有非常重要的科学地位。新疆阿勒泰山两河源自然保护区位于阿勒泰山东段,气候在新疆最为潮湿,其地衣植物种类十分丰富。作者在该保护区选择6个植被带类型,即山地荒漠带、山地草原带、针阔混交林带、针叶林带、亚高山草甸带、高山草甸带,研究了其地面生地衣植物的物种多样性特征。结果表明:阿勒泰山两河源自然保护区地衣植物区系成分丰富而且复杂,共有地面生地衣植物5科6属46种,以石蕊科种类最为丰富,约32种。该地区不同植被带类型下地面生地衣植物物种的S?renson相似性系数在0.200–0.739之间,以针阔混交林和针叶林带的相似性为最高(0.739),针阔混交林和高山草甸带地衣植物物种相似性最低(0.200)。各植被带地衣优势种中白边岛衣(Cetrarialaeuigata)、林鹿蕊(Cladoniaarbuscula)、佐木氏珊瑚枝(Stereocaulonsasakii)、鳞地卷(Peltigeralepido-phora)、喇叭粉石蕊(Cladoniachlorophaea)、东方鹿蕊(Cladinagrisea)等的重要值在0.5以上,其余优势种的重要值均在0.5以下;山地森林带地衣植物物种多样性最为丰富,在整个阿勒泰山两河源自然保护区地面生地衣植物群落中占据优势地位,为该地区地衣植物多样性的分布中心,是地衣植物多样性保护的关键地区。     

新疆榅桲中的鞣质类化合物及其生物活性

榅桲是传统的中药之一,富含具有药用价值的化学成分,如鞣质类、有机酸、氨基酸、挥发油和黄酮等。鞣质类物质主要包括没食子酸鞣质、矢车菊苷配质、儿茶精、表儿茶精等,具有抗菌、抗病毒、抗突变、抗肿瘤、抗脂质过氧化、抑制酶活性,以及抗溃疡、止血、收敛、解毒等功效,对心脏和心血管也有一定的效应。     

Cutting edge: mechanisms of IL-2-dependent maintenance of functional regulatory T cells

IL-2 controls the survival of regulatory T cells (Tregs), but it is unclear whether IL-2 also directly affects Treg suppressive capacity in vivo. We have found that eliminating Bim-dependent apoptosis in IL-2- and CD25-deficient mice restored Treg numbers but failed to cure their lethal autoimmune disease, demonstrating that IL-2-dependent survival and suppressive activity can be uncoupled in Tregs. Treatment with IL-2-anti-IL-2-Ab complexes enhanced the numbers and suppressive capacity of IL-2-deprived Tregs with striking increases in CD25, CTLA-4, and CD39/CD73 expression. Although cytokine treatment induced these suppressive mechanisms in both IL-2(-/-) and IL-2(-/-)Bim(-/-) mice, it only reversed autoimmune disease in the latter. Our results suggest that successful IL-2 therapy of established autoimmune diseases will require a threshold quantity of Tregs present at the start of treatment and show that the suppressive capacity of Tregs critically depends on IL-2 even when Treg survival is independent of this cytokine.