氨基糖苷抗生素庆大霉素： 基础研究的新进展及应用研究的新潜力

庆大霉素是临床上重要的氨基糖苷类抗生素。近年来,伴随现代生物技术水平的发展,人们对该抗生素有了更深入的了解。本文综述了近年来国内外关于庆大霉素的作用机制和耐药机制、生物合成途径和结构改造,及其新活性等方面研究成果,并对庆大霉素的应用与发展前景进行展望。     

2‐photon excitation fluorescence microscopy enables deeper high‐resolution imaging of voltage and Ca2+ in intact mice,rat, and rabbit hearts

We describe a novel two‐photon (2P) laser scanning microscopy (2PLSM) protocol that provides ratiometric transmural measurements of membrane voltage (V_m) via Di‐4‐ANEPPS in intact mouse, rat and rabbit hearts with subcellular resolution. The same cells were then imaged with Fura‐2/AM for intracellular Ca²⁺ recordings. Action potentials (APs) were accurately characterized by 2PLSM vs. microelectrodes, albeit fast events (<1 ms) were sub‐optimally acquired by 2PLSM due to limited sampling frequencies (2.6 kHz). The slower Ca²⁺ transient (CaT) time course (>1ms) could be accurately described by 2PLSM. In conclusion, V_m ‐ and Ca²⁺‐sensitive dyes can be 2P excited within the cardiac muscle wall to provide AP and Ca²⁺ signals to ～400 µm. (© 2013 WILEY‐VCH Verlag GmbH &Co. KGaA, Weinheim)     

Bt工程菌WG-001与化学杀虫剂复配最佳配方筛选及田间防效研究

为了延缓小菜蛾Plutella xylostella(L.)抗药性的产生和发展,筛选Bt工程菌WG-001与化学杀虫剂的最佳复配配方,本研究采用浸叶法分别测定了Bt工程菌WG-001与茚虫威、丁醚脲和巴丹复配对小菜蛾的增效作用。结果表明,当Bt工程菌WG-001与茚虫威以有效成分之比为100.10∶0.72复配时,增效作用最强,共毒系数为178.05。而Bt工程菌WG-001分别与丁醚脲和巴丹复配表现相加或拮抗作用。田间试验结果表明,Bt工程菌WG-001与茚虫威以100.10∶0.72的有效成分之比进行复配后,药后7d,500倍液对小菜蛾的平均防效为84.88%,显著优于Bt工程菌WG-001和茚虫威单用的平均防效。     

N-乙酰氨基葡萄糖转移酶（OGT）研究进展

O-GlcNAc是一种广泛存在于蛋白质丝/苏氨酸残基上的动态、可逆的蛋白翻译后修饰,它广泛分布在细胞浆和细胞核中,参与调节多种细胞途径。研究表明蛋白的O-GlcNAc糖基化与神经退行性疾病、糖尿病和癌症等疾病相关。在体内,O-GlcNAc动态修饰由N-乙酰氨基葡萄糖转移酶（OGT）和N-乙酰氨基葡萄糖苷酶（OGA）协同完成。近年来,OGT逐渐成为糖生物学领域的研究热点,在其结构、作用机制及晶体学方面取得了快速发展。     

1,25-Dihydroxyvitamin D3 Controls a Cohort of Vitamin D Receptor Target Genes in the Proximal Intestine That Is Enriched for Calcium-regulating Components

1,25-Dihydroxyvitamin D₃ (1,25(OH)₂D₃) plays an integral role in calcium homeostasis in higher organisms through its actions in the intestine, kidney, and skeleton. Interestingly, although several intestinal genes are known to play a contributory role in calcium homeostasis, the entire caste of key components remains to be identified. To examine this issue, Cyp27b1 null mice on either a normal or a high calcium/phosphate-containing rescue diet were treated with vehicle or 1,25(OH)₂D₃ and evaluated 6 h later. RNA samples from the duodena were then subjected to RNA sequence analysis, and the data were analyzed bioinformatically. 1,25(OH)₂D₃ altered expression of large collections of genes in animals under either dietary condition. 45 genes were found common to both 1,25(OH)₂D₃-treated groups and were composed of genes previously linked to intestinal calcium uptake, including S100g, Trpv6, Atp2b1, and Cldn2 as well as others. An additional distinct network of 56 genes was regulated exclusively by diet. We then conducted a ChIP sequence analysis of binding sites for the vitamin D receptor (VDR) across the proximal intestine in vitamin D-sufficient normal mice treated with vehicle or 1,25(OH)₂D₃. The residual VDR cistrome was composed of 4617 sites, which was increased almost 4-fold following hormone treatment. Interestingly, the majority of the genes regulated by 1,25(OH)₂D₃ in each diet group as well as those found in common in both groups contained frequent VDR sites that likely regulated their expression. This study revealed a global network of genes in the intestine that both represent direct targets of vitamin D action in mice and are involved in calcium absorption.     

Bortezomib and sphingosine kinase inhibitor interact synergistically to induces apoptosis in BCR/ABl cells sensitive and resistant to STI571 through down-regulation Mcl-1

Interactions between the proteasome inhibitor, bortezomib, and the sphingosine kinase (SPK1) inhibitor, SKI, were examined in BCR/ABL human leukemia cells. Coexposure of K562 or chronic myeloid leukemia (CML) cells from patients to subtoxic concentrations of SKI (10 μM) and bortezomib (100 nM) resulted in a synergistic increase in caspase-3 cleavage and apoptosis. These events were associated with the downregulation of BCR–ABL and Mcl-1, and a marked reduction in SPK1 expression. In imatinib mesylate-resistant K562 cells that displayed decreased BCR–ABL expression, bortezomib/SKI treatment markedly increased apoptosis and inhibited colony-formation in association with the downregulation of Mcl-1. Finally, the bortezomib/SKI regimen also potently induced the downregulation of BCR/ABL and Mcl-1 in human leukemia cells. Collectively, these findings suggest that combining SKI and bortezomib may represent a novel strategy in leukemia, including apoptosis-resistant BCR–ABL⁺ hematologic malignancies.     

Management of threatened insect species in Australia, with particular reference to butterflies

Abstract   Advances in determination of conservation status of insects through evaluation of threats (rather than simple 'rarity' alone) help to set priorities for allocation of conservation resources, and formulate effective legislative treatment and field management for the most needy species. Summaries of several recent conservation studies on butterflies exemplify the variety of practical needs, amongst which habitat protection and enhancement is paramount. However, the paucity of biological and distributional information on most Australian insects necessitates considerable flexibility in approach, so that perceptions of conservation need may be highly subjective. Listing species for protection and conservation priority must be viewed as a dynamic process, with opportunity for rapid action, either to list or de-list taxa as research and recovery actions lead to change in perceived or real status and conservation need.     

Effects of pressure overload-induced hypertrophy on TTX-sensitive inward currents in guinea pig left ventricle

We investigated the effects of pressure overload hypertrophy on inward sodium (I _Na) and calcium currents (I _Ca) in single left ventricular myocytes to determine whether changes in these current systems could account for the observed prolongation of the action potential. Hypertrophy was induced by pressure overload caused by banding of the abdominal aorta. Whole-cell patch clamp experiments were used to measure tetrodotoxin (TTX)-sensitive inward currents. The main findings were that I _Ca density was unchanged whereas I _Na density after stepping from –80 to –30 mV was decreased by 30% (–9.0 ± 1.16 pA pF^–1 in control and –6.31 ± 0.67 pA pF^–1 in hypertrophy, p < 0.05, n= 6). Steady-state activation/inactivation variables of I _Na, determined by using double-pulse protocols, were similar in control and hypertrophied myocytes, whereas the time course of fast inactivation of I _Na was slowed (p < 0.05) in hypertrophied myocytes. In addition, action potential clamp experiments were carried out in the absence and presence of TTX under conditions where only Ca²⁺ was likely to enter the cell via TTX-sensitive channels. We show for the first time that a TTX-sensitive inward current was present during the plateau phase of the action potential in hypertrophied but not control myocytes. The observed decrease in I _Na density is likely to abbreviate rather than prolong the action potential. Delayed fast inactivation of Na⁺ channels was not sustained throughout the voltage pulse and may therefore merely counteract the effect of decreased I _Na density so that net Na⁺ influx remains unaltered. Changes in the fast I _Na do not therefore appear to contribute to lengthening of the action potential in this model of hypertrophy. However, the presence of a TTX-sensitive current during the plateau could potentially contribute to the prolongation of the action potential in hypertrophied cardiac muscle. (Mol Cell Biochem 261: 217–226, 2004)     

The Role of Weak Specific and Nonspecific Interactions in Enzymatic Recognition and Conversion of Long DNAs

According to the currently accepted model, enzymes searching for specific recognition sequences or structural elements (modified nucleotides, breaks, single-stranded DNA fragments, etc.) slide at a high rate along DNA. Such sliding is possible only if the enzymes possess sufficiently high affinity for all DNA, sequence notwithstanding. Therefore, significant differences in their affinity for specific and nonspecific DNA sequences are unlikely, and the formation of a complex between an enzyme and its target DNA is not a basic factor of enzyme specificity. To elucidate such factors, we have analyzed many DNA replication, DNA repair, topoisomerization, integration, and recombination enzymes using a number of physicochemical methods, including the method of stepwise increase in ligand complexity developed in our laboratory. It has been shown that high affinity of all studied enzymes for long DNAs is provided by the formation of many weak contacts of the enzyme with all nucleotide units covered by the protein globule. The main role lies in the contact between positively charged amino acid residues and internucleoside phosphate groups; however, the contribution of each contact is very small, and the full contact interface usually resembles that characteristic of interactions between oppositely charged biopolymer surfaces. In some cases, a significant contribution to the affinity is made through hydrophobic and/or van der Waals interactions of the enzymes with nucleotide bases. On the whole, such nonspecific interactions provide for five to eight orders of enzyme affinity for DNA, depending on the enzyme. Specific interactions of enzymes with long DNAs, in contrast to their contacts with small ligands, are usually weak and comparable in efficiency with weak nonspecific contacts. The sum of specific interactions most often provides for approximately one or, rarely, two orders of affinity. According to structural data, DNA binding to any of the investigated enzymes is followed by a stage of DNA conformation adjustment, which includes partial or complete DNA melting, deformation of its backbone, stretching, compression, bending or kinking, eversion of nucleotides from the DNA helix, etc. The full set of such changes is specific for each individual enzyme. The fact that all enzyme-dependent changes in DNA are effected through weak specific (rather than strong) interactions is very important. Enzyme-specific changes in DNA conformation are required for effective adjustment of reacting orbitals to an accuracy of 10°–15°, which is possible only in the case of specific DNAs. A transition from nonspecific to specific DNA leads to an increase in the reaction rate (k _cat) by four to eight orders of magnitude. Thus, the stages of DNA conformation adjustment and catalysis proper provide for the high specificity of enzyme action.