Factors influencing swimming in bay scallops, Argopectenirradians (Lamarck, 1819)

Factors influencing the probability, distance, and direction of swimming in bay scallops (Argopectenirradions Lamarck, 1819) were studied through a series of experimental releases in the field and in a 3-m tank. The probability of a scallop swimming was significantly influenced by the type of substratum on which it was released (sand vs. grassbed), by contact with two natural gastropod predators (Murex, Fasciolaria), and by the amount of rest allowed after a previous swim. The horizontal distance traveled by a swimming scallop was significantly influenced by artificial weight of a magnitude equivalent to a normal load of shell-encrusting organisms, by the amount of rest allowed after a previous swim, by the height attained in the water column, and by the scallop's size. The direction of scallop swimming was significantly influenced by the location along the mantle edge where a predator was contacted, and by factors probably related to the asymmetrical water flow pattern through the mantle cavity. Swimming in bay scallops apparently serves to maintain position in grassbeds and to avoid predators.     

植物群落的冗余结构——对生态系统稳定性的一种解释

应用冗余理论探讨植物群落的稳定性机理。植物群落的冗余是由植物体的器官冗余、种群内遗传结构冗余、物种冗余和层次冗余组成的。植物群落的抵抗力主要来自物种冗余和种群内遗传结构冗余,而恢复力来自层次冗余和器官冗余。植物群落的冗余结构决定了其稳定性是抵抗的,或是恢复的,或即是抵抗的又是恢复的。研究表明,冗余理论比多样性导致稳定性更能合理地解释植物群落的稳定性。植物群落冗余按其组成万分的性质可分为两种：数量冗     

Hydrodynamics-based transfection of the liver: entrance into hepatocytes of DNA that causes expression takes place very early after injection

BACKGROUND: The mechanism of gene transfer into hepatocytes by the hydrodynamics-based transfection procedure is not clearly understood. It has been shown that, after a hydrodynamic injection, a large proportion of plasmid DNA remains intact in the liver where it is bound to plasma membrane and suggested that this DNA could be responsible for the efficiency of the transfection. METHODS: We have investigated the problem by giving mice a hydrodynamic injection of isotonic NaCl, followed at different time intervals by a conventional injection of DNA, cold or labelled with (35)S, with cDNA of luciferase as a reporter gene. Then, we determined the consequences of that dual injection on luciferase expression and on DNA uptake by the liver and its intracellular fate. By such experiments, it is possible to establish the time dependency of the induction of liver changes caused by a hydrodynamic injection on the one hand and the expression and DNA uptake and fate on the other. Moreover, some experiments have been performed on primary cultures of hepatocytes isolated after a hydrodynamic injection of DNA. RESULTS: When DNA is given to mice by a conventional injection a few seconds after an hydrodynamic injection of isotonic NaCl, luciferase expression in the liver is considerably lower than that observed after a single hydrodynamic injection of the plasmid. On the other hand, as assessed by the rate of DNA degradation and by centrifugation results obtained after injection of (35)S-DNA, the uptake and the intracellular fate of the bulk of DNA are similar whether DNA is administered by a single hydrodynamic injection or by a conventional injection given up to at least 2 h after a hydrodynamic injection of isotonic NaCl. Hepatocytes isolated a few minutes after a hydrodynamic injection exhibit a maximal expression that does not depend on the large amount of DNA that remains bound to the plasma membrane for a relatively long time. CONCLUSIONS: Our results show that the efficiency of hydrodynamics-based transfection depends on a process that takes place very quickly after injection and is not linked to a delay of DNA degradation and the persistence of a large proportion of DNA bound to hepatocytes of the plasma membrane, strongly suggesting that expression after a hydrodynamic injection is caused by a small proportion of DNA molecules that rapidly enter the cytosol probably by plasma membrane pores generated by the hydrodynamic pressure.     

Effects of tributyltin on barrier functions in human intestinal Caco-2 cells

The effect of tributyltin (TBT) on human intestinal epithelial cell functions was investigated by using human intestinal Caco-2 cell monolayers. We paid particular attention to the effect of TBT on two barrier functions: the tight junction as a physical barrier and MDR1/P-glycoprotein as a biological barrier. A loss of monolayer integrity was apparent from the TBT treatment and the paracellular permeability was increased by TBT. On the other hand, the activity of P-glycoprotein, which was examined by measuring the accumulation of Rhodamine-123 and daunomycin, was increased by prolonged TBT treatment in a concentration-dependent manner (1-100 nM). Furthermore, it was clarified by Western and Northern blots that this increase was accompanied by the increased expression of MDR1 mRNA and protein. The activation of a multidrug resistance transporter P-glycoprotein by TBT would cause a disorder of the human intestines by changing the drug pharmacokinetics.     

Mechanisms underlying burst generation of the pyloric muscle in the mantis,shrimp, Squilla oratoria

The pyloric constrictor muscles of the stomach in Squilla can generate spikes by synaptic activation via the motor nerve from the stomatogastric ganglion. Spikes are followed by slow depolarizing afterpotentials (DAPs) which lead to sustained depolarization during a burst of spikes. 1. The frequency of rhythmic bursts induced by continuous depolarization is membrane voltage-dependent. A brief depolarizing or hyperpolarizing pulse can trigger or terminate bursts, respectively, in a threshold-dependent manner. 2. The conductance increases during the DAP response. The amplitude of DAP decreases by imposed depolarization, whereas it increases by hyperpolarization. DAPs from successive spikes sum to produce a sustained depolarizing potential capable of firing a burst. 3. The spike and DAP are reduced in amplitude by decreasing [Ca]o, enhanced by Sr2+ or Ba2+ substituted for Ca2+, and blocked by Co2+ or Mn2+. DAPs are selectively blocked by Ni2+, and the spike is followed by a hyperpolarizing afterpotential. 4. The spike and DAP are prolonged by intracellular injection of the Ca2+ chelator EGTA. A hyperpolarizing afterpotential is abolished by EGTA and enhanced by increasing [Ca]o. The DAP is diminished in Na(+)-free saline and reduced by tetrodotoxin. 5. It is concluded that the muscle fiber is endowed with endogenous oscillatory properties and that the oscillatory membrane events result from changes of a voltage- and time-dependent conductance to Ca2+ and Na+ and a Ca2+ activated conductance to K+.     

Conformational changes of beta-lactoglobulin induced by anionic phospholipid

Conformational changes of beta-lactoglobulin (beta-LG) induced by anionic phospholipid (dimyristoylphosphatidylglycerol, DMPG) at physiological conditions (pH 7.0) have been investigated by UV-VIS, circular dichroism (CD) and fluorescence spectra. The experimental results suggest that beta-LG-DMPG interactions cause beta-LG a structural reorganization of the secondary structure elements accompanied by an increase in alpha-helical content, and a loosening of the protein tertiary structure. The interaction forces between beta-LG and DMPG are further evaluated by fluorescence spectra. The fluorescence spectral data show that conformational changes in the protein are driven by electrostatic interaction at first, then by hydrophobic interaction between a protein with a negative net charge and a negatively charged phospholipid.     

miR-29a通过调控PTEN表达对脂多糖诱导人肺微血管内皮细胞损伤的保护作用研究*

目的:探讨miR-29a在脂多糖(LPS)诱导人肺微血管内皮细胞(HPMVECs)损伤中的作用及机制。方法:构建LPS损伤HPMVECs模型。RT-qPCR检测miR-29a表达变化;试剂盒测乳酸脱氢酶(LDH)释放量;MTT和流式细胞术分别检测细胞存活率及凋亡率;Western blot测蛋白质表达水平;Microcosm、starBase、Pictar、TargetScan软件预测 miR-29a的可能靶基因,双萤光素酶实验验证miR-29a和PTEN的靶向关系。结果:使用LPS处理HPMVECs,显著降低细胞中miR-29a的表达和细胞存活率,诱导LDH释放量和HPMVECs凋亡率增加,上调细胞中PTEN、Bim蛋白表达,下调p-Akt/Akt、p-FOXO3a/FOXO3a表达 (P<0.05);过表达miR-29a逆转LPS对HPMVECs的损伤作用。萤光素酶报告基因实验证实miR-29a 靶向PTEN,转染miR-29a mimics显著下调PTEN蛋白表达,转染miR-29a inhibitors明显上调PTEN蛋白表达 (P<0.05),但PTEN mRNA表达水平差异均无统计学意义(P>0.05)。结论:过表达miR-29a可能通过抑制PTEN蛋白的表达水平、激活Akt/FOXO3a/Bim信号通路对LPS致HUVECs的损伤发挥保护作用。     

Surveying a local fitness landscape of a protein with epistatic sites for the study of directed evolution

We present a method for analysis of a fitness landscape of a biopolymer with significantly epistatic sites. The analysis is based on a quasi-additive fitness model. The fitness model is constructed with additive terms conducted by "site-fitness" and epistatic terms conducted by "pair-fitness," where the site-fitness is a fitness contribution from an independent residue and the pair-fitness is a fitness contribution from a pair of epistatic residues. As a case study, we analyzed the sequence-fitness data for 45 clones of thermostable prolyl endopeptidase mutants. They were generated by a mutation scrambling method, which can accumulate advantageous mutations. The fitness contributions from 14 single-point mutations including E67Q and Q656R were identified by the analysis. As a result, we found that the fitness model with a significant epistatic term by a pair of the 67th site and 656th site was in good agreement with the experimental data and that the explored landscape in the binary 14-dimensional sequence space is still a mountainous landscape with twin peaks. The validity was supported by the analysis of mutant fitness distributions derived from another mutation scrambling experiment and by (3D) structural data.     

Chemical synthesis of 4,5-dioxovaleric acid and its nonenzymatic transamination to 5-aminolevulinic acid

4,5-Dioxovaleric acid (DOVA) was synthesized from 5-bromolevulinic acid via formation of the pyridinium bromide of 5-bromolevulinic acid, followed by nitrone formation with p-nitrosodimethylaniline, and hydrolysis of the nitrone to yield DOVA. Partial purification of DOVA was obtained by passage of the reaction mixture through a cation exchange column. DOVA was identified by paper electrophoresis and by a specific fluorometric assay. DOVA was nonenzymatically transaminated to 5-aminolevulinic acid (ALA) with glycine serving as the amino donor. Other compounds tested were less effective amino donors. Glyoxylic acid was identified as a reaction product by paper electrophoresis and a specific calorimetric test. ALA was identified by paper electrophoresis, paper chromatography of a pyrrole derivative, reaction with Ehrlich reagent, and by its enzymatic conversion by a barley extract to porphobilinogen and uroporphyrin. The nonenzymatic transamination was inhibited by Tris and was stimulated by high pH. The existence of this nonenzymatic activity is discussed in relation to previous reports of dova transaminase activity in cell extracts.     

Inhibition of nuclear transport of caspase-7 by its prodomain

Apoptosis is a major form of cell death, characterized by a series of morphological changes induced by cleaving cytoplasmic and nuclear proteins via active caspases. The data presented here show, by fluorescence microscopic and immunoblotting analyses, that a prodomain of caspase-7 inhibits its nuclear translocation and apoptosis-inducing activity. This nuclear localization is dependent on the presence of a basic tetrapeptide that is conserved in mammalian and Xenopus caspase-7 and that is located downstream of a cleavage site between a prodomain and a catalytic protease domain. Furthermore, an attachment of the caspase-7 prodomain (31 amino acids) represses the nuclear transport of a fusion protein of a heterologous protein and the caspase-7 nuclear localization signal (19 amino acids), suggesting that the inhibition of nuclear localization by the prodomain is mediated by the interaction of these short peptides.     

Measurements of intra-and extracellular pH in the liverwort Conocephalum conicum during action potentials

During action potentials triggered by electricity and light, measurements of intra- and extracellular pH in the liverwort Conocephalum conicum L. were carried out by the use of antimony-filled H⁺-sensitive microelectrodes. Intracellular pH increased transiently by about 0.05 unit in the course of an action potential, while extracellular pH, measured at the surface of the thallus, remained unchanged. Switching the light off caused a transient increase in intracellular pH by less than 0.1 unit. Turning the light on produced a fast pH decrease by about 0.15 unit followed by a slow increase. When the light was intensive enough, the action potential thus triggered caused a slight increase in intracellular pH superimposed on the phase of a slow pH increment.
The magnitude and time course of the intracellular pH changes seem insufficient for a role as messenger between action potential and the up to 100% increase in the rate of respiration that has been registered in Conocephalum conicum as a consequence of excitation.     

胃癌相关mir-148a靶向调控胃泌素受体CCKBR

目的：研究胃癌相关miR-148a与胃泌素受体CCKBR的调控关系,并分析其调控结合位点。方法生物信息学预测人CCKBR 3’ UTR上miR-148a 的结合位点;利用 PCR扩增 miR-148a 前体构建真核表达载体;Northern Blot检测miR-148a真核表达载体的表达;构建CCKBR 3’UTR野生型和突变型荧光素酶报告载体,并利用双荧光素酶活性分析检测分析miR-148a对CCKBR基因表达的调控和结合位点;Western Blot检测miR-148a过表达对CCKBR蛋白表达的作用。结果在人CCKBR 3’UTR上找到3个miR-148a的潜在结合位点;miR-148a真核表达载体构建成功,转染胃癌细胞后可显著过表达;miR-148a通过人CCKBR 3’UTR上423bp处的结合位点抑制CCKBR的基因表达;miR-148a过表达显著抑制胃癌细胞中CCKBR的蛋白表达。结论 CCKBR是胃癌相关miR-148a的靶基因,miR-148a通过其3’UTR上的结合位点抑制CCKBR的基因表达和蛋白合成,提示miR-148a可能通过调控CCKBR参与胃癌的发生发展。     

Robust development as a consequence of generated positional information

The origin and robustness of morphogenesis are studied by dynamical system modeling of a cell society, in which cells possessing internal chemical reaction dynamics interact with each other through their mutual interaction with diffusive chemicals in a two-dimensional medium. It is found that stem-type cells differentiate into various cell types (where a cell 'type' is defined by a type of intra-cellular dynamics) due to a dynamic instability caused by cell-cell interactions in a manner described by the isologous diversification theory. The differentiations are spatially regulated by the concentration of chemicals in the medium, while the chemical concentrations are locally influenced by the intra-cell dynamics. Through this reciprocal relationship, chemical concentrations come to exhibit spatial variation as differentiated cell types begin to emerge, and as a result the regulation exercised by the chemical concentrations become spatially inhomogeneous. This reinforces the process of differentiation, through which spatial patterns of differentiated cells appear. Within this reciprocal relationship, the concentration gradients are read and interpreted by the cell as positional information. A spatial order of cells realized in this process represents a stable state of the system governed by this reciprocal relationship, and that the developmental process through which this state is realized is robust with respect to perturbations. The dependence of the morphogenesis on history and the community effect in cell differentiation are also discussed.     

Microbial metabolism of methanesulfonic acid

Methanesulfonic acid is a very stable strong acid and a key intermediate in the biogeochemical cycling of sulfur. It is formed in megatonne quantities in the atmosphere from the chemical oxidation of atmospheric dimethyl sulfide (most of which is of biogenic origin) and deposited on the Earth in rain and snow, and by dry deposition. Methanesulfonate is used by diverse aerobic bacteria as a source of sulfur for growth, but is not known to be used by anaerobes either as a sulfur source, a fermentation substrate, an electron acceptor, or as a methanogenic substrate. Some specialized methylotrophs (including Methylosulfonomonas, Marinosulfonomonas, and strains of paragraph signHyphomicrobium and Methylobacterium) can use it as a carbon and energy substrate to support growth. Methanesulfonate oxidation is initiated by cleavage catalysed by methanesulfonate monooxygenase, the properties and molecular biology of which are discussed.     

A new preparation of S-100 protein from rat and bovine brains

The S-100 nervous system protein was purified from bovine and rat brains by a modification of the original procedure. The main modification consisted in substituting a step of calciumdependent binding of S-100 to a phenyl-Sepharose column for the original step of chromatography on G-200 Sephadex. The proteins were pure as determined by SDS gel electrophoresis. HPLC on a reversed phase and on a size-separation column, and by immunological criteria. The bovine S-100 behaved as previously described, during calcium binding, by displaying a conformational change as evidenced by increase in native fluorescence.     

Partial Characterization of Cell-bound Lipase of Candida paralipolytica

In accordance with the regulation by aspartate of phosphoenolpyrubate (PEP*) carboxylase, glutamate formation in Brevibacterium flavum, a glutamate-producing bacterium, was inhibited by the addition of aspartate. Furthermore, an increase in aspartate formation caused by a mutational decrease in citrate synthase specific activity was accompanied by a decrease in the total amount of glutamate and aspartate formed. However, a mutational decrease in glutamate dehydrogenase activity caused a decrease in the total amount without increasing the asparate formation but with accumulation of 2-oxoglutarate, suggesting that the feedback inhibition by the aspartate of PEP carboxylase was enhanced by 2-oxoglutarate. In fact, partially purified PEP carboxylase from this organism was found to be synergistically inhibited by aspartate and 2-oxoglutarate, citrate, cis-aconitase, or isocitrate. Among them, the effects of tricarboxylic acids were attributed to their non-specific chelating action with Mn²⁺, an activator of the enzyme. The synergistic action of 2-oxoglutarate was accompanied by a decrease in Hill coefficient for the aspartate of the enzyme.     

Shell disintegration and taphonomic loss in rudist biostromes

Radiolitid biostromes in the Upper Cretaceous of Austria and Italy record a marked taphonomic loss controlled mainly by the composition of the biocoenosis, by the density of rudist colonization, by the style of radiolitid shell disintegration and by early diagenetic processes. Radiolitid shells consisted of a calcitic ostracum and an originally aragonitic hypostracum. The attached valve of most radiolitids was built of (1) an outermost ostracal layer of delicate calcite lamellae, (2) a thick layer of ‘boxwork ostracum’ built of radial funnel plates and cell walls, (3) a thin, inner ‘ostracal layer 3’ of thick-walled boxwork, and (4) the hypostracum that formed the innermost shell layer. The attached valve disintegrated by spalling of radial funnel plates of layer 2, and by selective removal of the boxwork ostracum. In the free valve, the ostracum consisted of two layers: (a) an inner, lid-shaped layer of dense calcite, and (b) an outer layer composed of calcite lamellae. The free valve disintegrated by spalling into ostracal and hypostracal portions, by spalling of the ostracum into layers a and b, and by disintegration of layer b into packages of calcite lamellae and individual lamellae. The specific style of disintegration of the radiolitids was aided or induced by discontinuities in shell structure. Lamellar fragments from the ostracum of the upper valve and from the radial funnel plates of the lower valve locally are abundant in free-valve-funnel-plate floatstones that comprise the matrix of or occur in lenses within radiolitid biostromes. In biostromes with an open parautochthonous fabric, selective removal of the boxwork ostracum of the attached valve occurred by mechanical spalling and, most probably, by early diagenetic dissolution. Complete removal of the boxwork ostracum yielded thin, relict shells composed of the ‘ostracal layer 3’ and the hypostracum. During early diagenesis, the hypostracum was replaced by blocky calcite spar, or was dissolved and became filled by internal sediments. The combination of both selective removal of boxwork ostracum and early diagenetic dissolution of aragonite locally resulted in the formation of ghost biostromes that entirely or largely consist of faint relics of radiolitids. The syndepositional formation of radiolitid shell relics and the presence of radiolitid ghost biostromes produced by bios-tratinomic and early diagenetic processes show that rudist biostromes can undergo marked taphonomic loss during fossilization. The presence of ghost biostromes with a burrowed, open parautochthonous rudist fabric indicates that the final preservation of a rudist biostrome was directly influenced by the characteristics of the biocoenosis, including unpreserved burrowing taxa. Rudist biostromes may be of markedly different taphonomy as a result of the taxonomic composition of the entire assemblage and the density of colonization by the rudists.