Uptake, transport and accumulation of nicotine by the Golden Potho (Epipremnum aureum): the central role of root pressure

The roots of Epipremnum aureum, though not synthesizing nicotine themselves, take up exogenously fed nicotine as a xenobiotic. The alkaloid is subsequently translocated to the leaves, via the xylem path, where it accumulates in the mesophyll up to levels comparable with nicotine-rich Nicotiana species. The Epipremnum plants accept nicotine only up to a distinct level; saturation is reached after about 10 days. All mature, non-senescent leaves accumulate the same amount of nicotine. By different experimental approaches, unequivocal evidence could be provided that root pressure is the 'translocative force' for nicotine transport in E. aureum. Xylem sap exudates, collected from shoot stumps that were connected to an intact root system immersed in nicotine solution were analyzed for nicotine content. Nicotine uptake from the medium by the root and its subsequent transfer into the xylem of the shoot persisted for more than 10h without measurable decline of the transport rate, provided the nicotine concentrations applied were < or =0.05%. In intact plants, where both components of water transport in the xylem--root pressure and transpirative water flow--are in operation, no surplus transport of nicotine from the roots into the leaves took place beyond the level observed in amputated plants. Under the influence of inhibitors of root respiration, nicotine uptake was halted slowly in case of oxygen deprivation and in case of cyanide, or it stopped very rapidly when CCCP, an uncoupler of mitochondrial ATP formation, was applied to the roots. This threshold of toxicity against the xenobiotic was established by dose effect curves for nicotine sensitivity of the roots for root respiration and by transpiration measurements. Leaves, bearing a heavy 'nicotine load', showed symptoms of senescence only after 3-6 weeks, as indicated by a decline in the chlorophyll content, the chl a/b ratio, and the maximal quantum yield efficiency (Fv/Fm), and by an increase in catalase activity. Our results provide insight into the mechanisms of uptake, transport and storage of nicotine as a xenobiotic.     

Dose response study of human exposure to 60 Hz electric and magnetic fields

This human exposure study examined the relationship between field strength and biological response and tested whether the exposure levels at which the greatest effects occur differ for different endpoints. Three matched groups of 18 men each participated in two 6 h exposure test sessions. All subjects were sham exposed in one session. In the other session, each group of subjects was exposed at a different level of combined electric and magnetic field strength (low group: 6 kV/m, 10 μT; medium group: 9 kV/m, 20 μT; and high group: 12 kV/m, 30 μT). The study was performed double blind, with exposure order counterbalanced. Significant slowing of heart rate, as well as alterations in the latency and amplitude of event-related brain potential measures derived from the electro encephalogram (EEG), occurred in the group exposed to the 9 kV/m, 20 μT combined field (medium group). Exposure at the other field strength levels had no influence on cardiac measures and differential effects on EEG activity. Significant decrements in reaction time and in performance accuracy on a time estimation task were observed only in the low group. These results provide support for the hypothesis that humans may be more responsive to some combinations or levels of field strength than to others and that such differences in responsivity may depend, in part, on the endpoint of interest. © 1994 Wiley-Liss, Inc.     

Changes in levels of pancreatic endoplasmic reticulum proteins that function in translocation and maturation of secretory proteins in response to cholecystokinin

Two pathways operate to target newly-synthesised proteins to the endoplasmic reticulum. In one, the signal recognition particle attaches to the signal sequences of nascent chains on ribosomes and slows or stops translation until contact is made with the docking protein at the membrane. The second operates via molecular chaperons. The pathways converge at the level of a 43 kDa signal binding protein integrated into the membrane, where translocation through a proteinaceous pore is initiated. In the lumen, proteins fold and disulphide formation is catalysed by the enzyme protein disulphide isomerase. The heavy chain binding protein may attach to unassembled or unfolded proteins and prevent their exit from the ER to the Golgi. Cholecystokinin (CCK) treatment increases the biosynthesis and secretion of pancreatic proteins, increases the levels of PDI and the 43 kDa binding protein, and reduces levels of BiP. These proteins may be possible targets for genetic manipulation to improve processing of heterologous proteins from cultured mammalian cells.     

Molecular flexibility and discontinuous translocation of a non-templated polymerase

Little is known regarding the translocation of non-templated nucleic acid polymerases with respect to single-stranded primers. VP55, the vaccinia virus poly(A) polymerase, translocates as it processively adds a approximately 3-7 adenylate tail to primers possessing only three ribouridylate residues (as an (rU)(2)-N(15)-rU motif), and a approximately 25-30 adenylate tail to primers that are more U-rich. Here, three models were addressed for the translocation of VP55 with respect to its primer, namely: (a) rigid protein/rigid nucleic acid; (b) flexible protein/rigid nucleic acid; (c) rigid protein/flexible nucleic acid. Analysis of free and covalently VP55-attached primers favored either (b) or a version of (c) incorporating a passive steric block, and suggested two regions of relative motion between polymerase and primer. Inclusion of a 6nt uridylate-rich patch at the primer 3' end switched the polymerase from approximately 3-7 nt to approximately 25-30 nt tail addition without affecting initial binding affinity. By synthesizing this patch as a (rU/dC) pool, discontinuous polymerase movements could be detected.     

The dose response relationship obtained at constant irradiation times for the induction of chromosome aberrations in human lymphocytes by cobalt-60 gamma rays

Summary The induction of unstable chromosome aberrations in human peripheral blood lymphocytes exposed in vitro to protracted doses of cobalt-60 radiation is presented. Four dose response curves have been produced using constant exposure times of 1, 3, 6, and 12 h. The data fit well to the linear quadratic model and the yield coefficients have been compared with those obtained for acute (< 10 min) exposure. The quadratic coefficient is dependent on irradiation time and decreases approximately as predicted by Lea and Catcheside'sG-function hypothesis. The possibility of a small proportion of much longer lived breaks is discussed. For purposes of biological dosimetry it is sufficient to assume a mean time of 2 h and a single exponential function for the repair of lesions when relating the effects of brief and protracted exposure.     

Trans-cis isomerization of retinal and a mechanism for ion translocation in halorhodopsin

The chromophore in halorhodopsin (HR) which acts as a light-driven chloride pump in halobacteria shares many properties with its counterpart in bacteriorhodopsin (BR): (i) a similar retinal protein interaction, (ii) trans to cis isomerization and (iii) similar intermediates of its photocycle. One major difference between the two chromoproteins is that the HR chromophore does not become deprotonated during its photocycle. A mechanism for the photocycle of HR is presented, which, in close analogy to an earlier proposed mechanism for BR, involves the sequence of all-trans 13-cis, 14s-cis 13-cis all-trans isomerizations of the chromophore, a Schiff base of retinal. In contrast to the situation in BR the 13-cis, 14s-cis13-cis isomerization is induced not by deprotonation of the retinal Schiff base chromophore but rather by the movement of an anion (Cl^-) towards the protonated nitrogen of the Schiff's base. The suggested mechanism involves the Schiff base directly in the chloride translocation in halorhodopsin.     

Conformational change of 23S RNA in 50S ribosome is responsible for translocation in protein synthesis

Since the recognition of the ‘translocation’ phenomenon during protein synthesis several theories have been proposed, without much success, to explain the translocation of peptidyl tRNA from the aminoacyl site to the peptidyl site. The involvement of L7/L12 proteins and therefore the L7/L12 stalk region of 50S ribosomes in the translocation process has been widely accepted. The mobility of the stalk region, as recognised by many workers, must be of physiological significance. It has recently been shown in this laboratory that 50S ribosomes derived from tight and loose couple 70S ribosomes differ markedly in quite a few physical and biological properties and it appears that these differences are due to the different conformations of 23S RNAs. It has also been possible to interconvert tight and loose couple 50S ribosomes with the help of the agents, elongation factor -G, GTP (and its analogues) which are responsible for translocation. Thus loose couple 70S ribosomes so long thought to be inactive ribosomes are actually products of translocation. Further, the conformational change of 23S RNA appears to be responsible for the interconversion of tight and loose couple 50S ribosomes and thus the process of translocation. A model has been proposed for translocation on the basis of the direct experimental evidences obtained in this laboratory.     

Non-bridging phosphate oxygen atoms within the tRNA anticodon stem-loop are essential for ribosomal A site binding and translocation

The conformation of the anticodon stem-loop of tRNAs required for correct decoding by the ribosome depends on intramolecular and intermolecular interactions that are independent of the tRNA nucleotide sequence. Non-bridging phosphate oxygen atoms have been shown to be critical for the structure and function of several RNAs. However, little is known about the role they play in ribosomal A site binding and translocation of tRNA to the P site. Here, we show that non-bridging phosphate oxygen atoms within the tRNA anticodon stem-loop at positions 33, 35, and 37 are important for A site binding. Those at positions 34 and 36 are not necessary for binding, but are essential for translocation. Our results correlate with structural data, indicating that position 34 interacts with the highly conserved 16S rRNA base G966 and position 36 interacts with the universally conserved tRNA base U33 during translocation to the P site.     

Evidence for the simulaneous translocation of muscarinic acetylcholine receptor and G protein by carbachol

Interactions between guanine nucleotide regulatory proteins (G proteins) and muscarinic acetylcholine receptors (mAChRs) were studied in vivo following carbachol treatment. Rat brain homogenates were separated by high speed ultracentrigation into heavy and light membrane and 300,000 g supernate franctions. The G proteins were partially purified by Sephadex-G200 and heptylamine-Sepharose and the mAChRs by (3,2′-aminobenzhydryloxy)-tropane-(ABT)-affinity chromatographies. Radioligand binding assays showed that acute carbachol induced a biphasic translocation of the mAChRs and G proteins into the light membrane fraction with an initial release at 5–10 min and a second phase at 60 min. Portions of the released mAChRs and the G proteins, were found in the 300,000 g supernates and light membranes and were eluted in the same peak fractions from a Sephadex G-200 column. This dually labelled peak dissociated in the presence of digitonin, suggesting close association between the mAChR and G protein. ABT-affinity chromatography yielded dually labelled mAChR-G protein fractions which eluated as a single radioactive peak on a second ABT column. the partially purified G proteins from these fractions were photoaffinity labelled with 8-azidoguanosine-5′-triphosphate, [γ-³²P]. SDS-PAGE autoradiography revealed the presence of G_0ξ and G_i which may be released simultaneously with the mAChRs from the plasma membrane. In addition, a 110,000 molecular weight polypeptide was dually labelled by [³H]-PrBCM and [γ-³²P]-8-azido-GTP suggesting the presence of a “mAChR-G protein complex.” These findings provide direct evidence for the release of mAChRs and G proteins and a mAChR-G protein complex by agonist occupation of the mAChRs.     

小麦籽粒蛋白质组分的生态变异及与植株氮转化的关系研究

小麦籽粒蛋白质和蛋白质组分含量决定了小麦面团流变学特性和加工品质、籽粒蛋白质组分的形成与植株氮素吸收利用密切相关,且蛋白质组分的合成有一定的顺序性。对不同生态区的小麦籽粒蛋白质组分合成进行了研究。结果表明,蛋白质及组分含量存在明显的基因型差异,徐州点蛋白质和谷蛋白含量显著大于南京点;花期氮素积累量和花后氮素转运量徐州点显著大于南京点,而花后氮同化量南京点显著大于徐州点;清蛋白、醇溶蛋白和谷蛋白含量与籽粒蛋白质含量呈显著正相关,与花后植株氮素转运量呈显著正相关。不同生态点小麦花后发育速率模式基本一致,但南京点发育进程迟于徐州点;在籽粒形成期和灌浆后期,南京点的发育速率较快,降低了清蛋白和谷蛋白的合成,导致南京点籽粒蛋白质含量降低。     

Bioaccumulation and Physiological Response of Five Willows to Toxic Levels of Cadmium and Zinc

We evaluated the phytoremediation potential of Salix spp. exposed to high cadmium (Cd) and zinc (Zn) concentrations to select feasible plant materials for restoration and revegetation of mining soil contaminated by heavy metals on the basis of their Cd and Zn accumulation, Cd-Zn interaction on bioaccumulation, and the changes of photosynthetic parameters. The Cd and Zn concentrations were in the order of root > leaf > stem, regardless of the species. In the combined Cd and Zn treatment, the leaf and stem Cd concentration in all species were higher relative to Cd-alone treatment. In contrast, the Zn concentration in plant tissues when exposed to the combined Cd + Zn treatment decreased relative to the Zn-alone treatment. The translocation factor (TF) of Cd and Zn from root to leaf was generally higher compared to TF from root to stem than those in the single treatment. The Cd + Zn treatments resulted in enhanced translocation of Cd from root to aboveground tissue (synergistic), while the same treatment suppressed the Zn translocation from root to leaf and stem (antagonistic). The reduction of photosynthetic parameters in Zn alone and Cd + Zn treatments was generally higher than that of Cd-alone treatment. Among the different species, S. caprea and P. alba×glandulosa have the lowest photosynthetic reduction relative to the control. Overall, S. caprea could be a potential candidate for phytoremediation of Cd- and Zn-contaminated sites.     

Reciprocal chromosome translocation associated with TDNA-insertion mutation in Arabidopsis: genetic and cytological analyses of consequences for gametophyte development and for construction of doubly mutant lines

Chromosomal rearrangements may complicate construction of Arabidopsis with multiple TDNA-insertion mutations. Here, crossing two lines homozygous for insertions in AtREV3 and AtPOLH (chromosomes I and V, respectively) and selfing F1 plants yielded non-Mendelian F2 genotype distributions: frequencies of +/++/+ and 1/1 2/2 progeny were only 0.42 and 0.25%. However, the normal development and fertility of double mutants showed AtPOLH-1 and AtREV3-2 gametes and 1/1 2/2 embryos to be fully viable. F2 distributions could be quantitatively predicted by assuming that F1 selfing produced inviable (1,2) and (+,+) gametophytes 86% of the time. Some defect intrinsic to the F1 selfing process itself thus appeared responsible. In selfing AtREV3 ^+/2 single mutants, imaging of ovules and pollen showed arrest or abortion, respectively, of half of gametophytes; however, gametogenesis was normal in AtREV3 ^2/2 homozygotes. These findings, taken together, suggested that T-DNA insertion at AtREV3 on chromosome I had caused a reciprocal I–V translocation. Spreads of meiosis I chromosomes in selfing AtREV3 ^+/2 heterozygotes revealed the predicted cruciform four-chromosome structures, which fluorescence in situ hybridization showed to invariably include both translocated and normal chromosomes I and V. Sequencing of the two junctions of T-DNA with AtREV3 DNA and the two with gene At5g59920 suggested translocation via homologous recombination between independent inverted-repeat T-DNA insertions. Thus, when crosses between TDNA-insertion mutants yield anomalous progeny distributions, TDNA-linked translocations should be considered. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Early effects of the strategies of creating a genetic refuge and direct translocation for conserving and restoring populations of native brown trout

1. The conservation of salmonid inter‐ and intra‐specific diversity is a well‐known challenge, and general management guidelines and conservation processes are available. However, research demonstrating the outcomes of practical conservation actions is largely lacking. 2. We monitored the spatiotemporal genetic and demographic evolution of a native Mediterranean brown trout population in a river in the French Alps to assess the efficacy and early effects of genetic refuge (i.e. cessation of stocking) and wild trout translocation strategies. We also studied the use of angling as a tool to limit the introgression of the wild standing population. 3. We found that the rate of non‐native alleles in wild populations was age dependent, underpinning the importance of using age profiles in the design of genetic conservation studies. 4. Genetic refuge and direct translocation of wild trout resulted in a rapid and significant decrease in the percentages of non‐native alleles. Moreover, the genetic refuge strategy resulted in a significant reduction in the number of pure non‐native individuals, without changing trout densities, whilst direct translocations resulted in the establishment of dense, self‐sustaining native trout populations. Direct translocations changed the distribution of genotype categories and increased densities up to 55‐fold in 3 years. Our results also showed that angling resulted in a selective pressure on non‐native trout introduced at fry stage, whereas non‐native trout issued from natural recruitment were not affected. 5. Our study provides insights for improving the efficacy of practical conservation policies and can be used in other native freshwater fish conservation plans. Proactive measures such as direct translocation need to be implemented together with passive approaches such as genetic refuge policies. Before implementing such actions, accurate genetic and demographic studies at small geographical scales are essential to ensure that no self‐sustaining population of non‐native fish is present. To obtain rapid colonisation, we recommend introducing fish along whole river sections rather than concentrating on a few river stretches. Angling pressure can be used as an additional tool to improve restoration.     

Early development in an algal gametophyte: role of the cytoskeleton in germination and nuclear translocation

Summary Biflagellate zoospores from the giant kelpMacrocystis pyrifera underwent germination after adhering to a substrate and produced germ tubes that were approximately 13–15 m in length. Coincident with the germ tube elongation was organelle (other than the nucleus) translocation along the tube. Shortly after formation of the germ tube, the zoospore nucleus divided and one daughter nucleus translocated along the germ tube. The nucleus did not appear to undergo chromosomal condensation prior to division. The nuclear division and/or translocation of the daughter nucleus did not begin until well after germ tube elongation was complete, demonstrating that these are temporally distinct developmental events. The translocation of one daughter nucleus coincided with differentiation of the distal end of the germ tube into a bulbous structure. Following this, the first gametophytic cross wall was formed and, subsequently, the daughter nucleus remaining in the original zoospore body underwent repositioning, assuming a position in the germ tube near the cross wall. Cytochalasin D inhibited germ tube elongation suggesting that actin microfilaments are probably involved in this developmental process. In addition, when cytochalasin D was added to the culture after the germ tube elongation was complete, it did not affect either nuclear division or translocation, indicating that microfilaments were not directly involved in these nuclear events. Colchicine and the plant specific microtubule disrupting agent, amiprophos methyl blocked nuclear division and translocation without affecting germination or germ tube elongation. These data suggest that actin microfilaments are primarily responsible for complete germination, specifically germ tube elongation, while microtubules are involved in nuclear division and translocation. The present study demonstrates that germination (and germ tube elongation) and nuclear translocation inM. pyrifera gametophytes are temporally and mechanistically distinct developmental events.     

Effect of exogenous amino acids on Cu uptake and translocation in maize seedlings

This study aimed to assess the effects of four contrasting proteinogenic amino acids on copper (Cu) uptake and translocation in maize (Zea mays L.) seedlings grown in a modified Hoagland solution. Glycine, aspartic acid and lysine at three concentrations (10, 25 and 100 μM) did not have any significant effect on Cu uptake and translocation in maize seedlings over a two-day experimental period. However, cysteine (a reductive amino acid) at the three concentrations increased very significantly (P < 0.01) Cu accumulations in the root symplast and the shoots of maize seedlings in comparison to the control. Cu uptake in the whole plant and Cu translocation from root to shoot were also increased in the cysteine treatments. In the 25 μM cysteine treatment, where cysteine was in moderate excess, the Cu uptake in the whole plant and Cu translocation from root to shoot were significantly (P < 0.01) higher than those of the 10 or 100 μM cysteine treatments, where the concentration of cysteine was equivalent to that of Cu(II) or in great excess according to the stoichiometry of the redox reaction of cysteine with Cu(II). It is hypothesized that the cysteine-induced oxidation state alteration from Cu(II) to Cu(I) could be responsible for the increased Cu uptake and Cu translocation, on the ground that Cu(I), as free cuprous ion or cysteine cuprous complex, may be more available to maize roots than Cu(II).     

Cyclin-dependent protein kinase 2 activity is required for mitochondrial translocation of Bax and disruption of mitochondrial transmembrane potential during etoposide-induced apoptosis

Previous studies have suggested that upregulation of Cyclin A-dependent protein kinase 2 (Cdk2) activity is an essential event in apoptotic progression and the mitochondrial permeability transition in human cancer cells. Here, we show that upregulated Cyclin A/Cdk2 activity precedes the proteolytic cleavage of PARP and is correlated with the mitochondrial translocation of Bax and the loss of mitochondrial transmembrane potential (Δψm) during etoposide-induced apoptosis in human cervical adenocarcinoma (HeLa) cells. Etoposide-induced apoptotic cell death is efficiently prevented in cells that overexpress a dominant negative mutant of Cdk2 (Cdk2-dn) or p21^WAF1/CIP1, a specific Cdk inhibitor. Conversely, apoptotic cell death is promoted in Cyclin A-expressing cells. Disruption of the mitochondrial transmembrane potential in etoposide-induced cells is prevented in cells that overexpress Cdk2-dn or p21^WAF1/CIP1, while this transition is prominently promoted in Cyclin A-expressing cells. We screened for mitochondrial Cdk2 targets in the etoposide-induced cells and found that the mitochondrial level of Bax is elevated by more than three fold in etoposide-treated cells and this elevation is effectively prevented in cells expressing Cdk2-dn under the same conditions. Thus, we suggest that Cdk2 activity is involved in the mitochondrial translocation of Bax, which plays an important role in the mitochondrial membrane permeability transition during apoptotic progression.     

Physiological performance of largemouth bass related to local adaptation and interstock hybridization: implications for conservation and management

Four genetically distinct stocks of age 2+ years largemouth bass Micropterus salmoides were produced using adults collected from two regions in the upper midwest (central Illinois, IL and south-eastern Wisconsin, WI, U.S.A.). Two pure stocks (IL × IL and WI × WI), as well as both of their reciprocal F1 interstock hybrids (IL × WI and WI × IL) were produced in research ponds in Champaign, IL. In general, swimming performance, routine oxygen consumption and activity were highest at 18 × C, intermediate at 12 × C, and lowest at 6. C for all stocks. However, performance indicators varied among stocks at each of the temperatures. The pure Illinois stock (IL × IL) had the lowest activity: cost ratio at 18 × C and the highest at 6_ C (based upon swimming strength, routine activity rates and routine metabolic rates). The opposite pattern was observed for the other pure stock (WI × WI). Although differences were less distinct at lower temperatures, the two pure stocks (IL × IL and WI × WI) outperformed both interstock hybrids. These results indicate that not only do non-native stocks appear to have reduced performance relative to locally adapted stocks, but also that interstock hybrids exhibit performance impairments, not hybrid vigour.     

Physiological and morphological study of encapsulated Saccharomyces cerevisiae

The impact of encapsulation on the anaerobic growth pattern of S. cerevisiae CBS 8066 in a defined synthetic medium over 20 consecutive batch cultivations was investigated. In this period, the ethanol yield increased from 0.43 to 0.46 g/g, while the biomass and glycerol yields decreased by 58 and 23%, respectively. The growth rate of the encapsulated cells in the first batch was 0.13 h⁻¹, but decreased gradually to 0.01 h⁻¹ within the 20 sequential batch cultivations. Total RNA content of these yeast cells decreased by 39% from 90.3 to 55 mg/g, while the total protein content decreased by 24% from 460 to 350 mg/g. On the other hand, the stored carbohydrates, that is, glycogen and trehalose content, increased by factors of 4.5 and 4 within 20 batch cultivations, respectively. Higher biomass concentrations inside capsules led to a lower glucose diffusion rate through the membrane, and volumetric mass transfer coefficient for glucose was drastically decreased from 6.28 to 1.24 (cm³/min) by continuing the experiments. Most of the encapsulated yeast existed in the form of single and non-budding cells after long-term application.