Expression in Yeast and Purification of Functional Recombinant Human Poly(ADP-RIBOSE)Polymerase(PARP). Comparative Pharmacological Profile with that of the Rat Enzyme

Abstract

Human poly(ADP-ribose)polymerase (PARP) was expressed in the yeast line JELl under the control of a GAL promoter. Proteins were extracted and human recombinant PARP purified to apparent homogeneity. The pharmacological profile of this human enzyme was characterised in terms of the effects of known inhibitors of PARP belonging to various chemical families and this was compared with that of the rat enzyme purified from rat testes. using the same purification protocol. The rat and the human enzymes appeared very similar in terms of their sensitivities to those selected inhibitors.     

Improved sensitivity and coherence selection for [15N,1H]-TROSY elements in triple resonance experiments

In experiments with proteins of molecular weights around 100 kDa the implementation of [¹⁵N,¹H]-TROSY-elements in [¹⁵N]-constant-time triple resonance experiments yields sensitivity enhancements of one to two orders of magnitude. An additional gain of 10 to 20% may be obtained with the use of sensitivity enhancement elements. This paper describes a novel sensitivity enhancement scheme which is based on concatenation of the ¹³ C ¹⁵N magnetization transfer with the ST2-PT element, and which enables proper TROSY selection of the ¹⁵N multiplet components.     

Developmental patterns of a large set of barley (Hordeum vulgare) cultivars in response to ambient temperature

Ambient temperature plays an important role in plant development. In cereals, little is known about the exact effects of ambient temperature in the range between it being a vernalising agent and an abiotic stress factor; thus the genetic determinants involved in the registering and response to ambient temperature, and their natural variation has not been dissected either. Principally, we wished to establish the level of natural variation in response to ambient temperature in barley via studying plant phenological development. The responses to temperature of 168 barley genotypes of different provenances and seasonal growth habit groups were observed in controlled environments. The effects of four temperature regimes (13°C, 16.5°C, 18°C and 23°C) on the duration of plant phenophases were examined. The plant development was characterised in a series of consecutive phenophases that span the plant life cycle from germination through flowering to attainment of maximum plant height. Ambient temperature affected significantly plant development, with substantial variation in responses among the genotypes. Six major types of responses were identified, which depended strongly on seasonal growth habit, with only a small degree of overlap. Although the differences in the timing of development among clusters were significant under each temperature regime, the 23°C treatment resulted in the largest diversity of responses, with significant changes in the ranking of the six clusters compared to other treatments. Two clusters showed particularly unusual responses to 23°C: the development of one winter barley cluster was extremely accelerated by the 23°C treatment, whereas the development of one spring barley cluster was significantly delayed. Ambient temperature assumes importance as a regulatory cue in the intricate and complex temporal and spatial regulation network of plant development in cereals and acts mostly through its regulatory effect on certain developmental phases such as the onset and duration of the intensive stem elongation.     

Role of c-Jun in cellular sensitivity to the microtubule inhibitor vinblastine

The role of c-Jun in the apoptotic response of cells to the microtubule inhibitor vinblastine was investigated using fibroblasts lacking or overexpressing c-Jun. c-Jun null cells were found to be more sensitive than wild-type cells at low (1-3 nM) concentrations of vinblastine, but showed essentially identical apoptotic responses as wild-type cells at a higher concentration of 10nM. In contrast, c-Jun overexpressing cells were highly vinblastine-resistant, with an IC50 of 12-fold greater than wild-type cells. The fate of cells exposed to lethal concentrations of vinblastine was examined by propidium iodide staining and flow cytometry. All cell types appeared to undergo mitotic arrest prior to apoptosis. Apoptosis of wild-type cells was associated with significant DNA re-replication. In contrast, DNA re-replication was much less prominent in vinblastine-treated c-Jun null cells and absent during apoptosis of c-Jun overexpressing cells. These results suggest that c-Jun plays a key role in the cellular sensitivity to vinblastine. In addition, c-Jun appears to regulate the pathway to cell death following mitotic arrest.     

Chromatic and achromatic stimulus discrimination of long wavelength (red) visual stimuli by the honeybee Apis mellifera

It has long been assumed that bees cannot see red. However, bees visit red flowers, and the visual spectral sensitivity of bees extends into wavelengths to provide sensitivity to such flowers. We thus investigated whether bees can discriminate stimuli reflecting wavelengths above 560 nm, i.e., which appear orange and red to a human observer. Flowers do not reflect monochromatic (single wavelength) light; specifically orange and red flowers have reflectance patterns which are step functions, we thus used colored stimuli with such reflectance patterns. We first conditioned honey bees Apis mellifera to detect six stimuli reflecting light mostly above 560 nm and found that bees learned to detect only stimuli which were perceptually very different from a bee achromatic background. In a second experiment we conditioned bees to discriminate stimuli from a salient, negative (un-rewarded) yellow stimulus. In subsequent unrewarded tests we presented the bees with the trained situation and with five other tests in which the trained stimulus was presented against a novel one. We found that bees learned to discriminate the positive from the negative stimulus, and could unambiguously discriminate eight out of fifteen stimulus pairs. The performance of bees was positively correlated with differences between the trained and the novel stimulus in the receptor contrast for the long-wavelength bee photoreceptor and in the color distance (calculated using two models of the honeybee colors space). We found that the differential conditioning resulted in a concurrent inhibitory conditioning of the negative stimulus, which might have improved discrimination of stimuli which are perceptually similar. These results show that bees can detect long wavelength stimuli which appear reddish to a human observer. The mechanisms underlying discrimination of these stimuli are discussed. Handling Editor: Lars Chittka.     

Structural organisation of prolamellar bodies (PLB) isolated from Zea mays. Parallel TEM, SAXS and absorption spectra measurements on samples subjected to freeze-thaw, reduced pH and high-salt perturbation

Well-organised PLB gives rise to a X-ray diffraction pattern overlaid by a scattering pattern arising from individual tubules within less well-organised regions of the lattice. TEM and SAXS measurements were used to characterise the structural changes in PLB subjected to perturbation by freeze-thaw, exposure to pH 6.5, or resuspension in high-salt media. Comparison of SAXS patterns measured, before and after structural perturbation allows the separation of the contributions from ordered and disordered PLB. The diffraction pattern is shown to be based on a diamond cubic (Fd3m) lattice of unit cell a = 78 nm. Freeze-thaw and high-salt disruption lead to the breakdown of ordered PLB into disordered tubules of similar dimensions to those making up the original PLB lattice. Their scattering patterns suggest that they are approximately 26 nm in diameter with a central lumen about 16 nm in diameter. The tubules formed at pH 6.5 are appreciably narrower, probably reflecting changes in the pattern of ionisation of charged groups at the membrane surface. Absorption spectra of PLB in media containing different concentrations of salts indicated that the structural and spectral changes are related. NADPH, have a significant role in the protection of POR-PChlide₆₅₀ but to have only a relatively small effect on the preservation of PLB organisation indicating that the retention of POR-PChlide₆₅₀ in isolated PLB preparations is a poor guide to their structural integrity.     

天山中西段不同地区雪岭云杉径向生长对气候变暖的响应差异

为研究乌苏和石河子地区雪岭云杉（Picea schrenkiana）径向生长对气候变暖的响应差异状况,利用响应函数及滑动相关分析等树轮气候学方法,研究了2个地区森林下限雪岭云杉径向生长与气候因子的响应关系。结果表明：乌苏和石河子地区雪岭云杉生长均与生长季的气候因子呈显著相关关系。但乌苏和石河子地区雪岭云杉生长对当地气候因子的响应也存在差异,乌苏地区雪岭云杉径向生长与上一年7、9月平均气温呈显著负相关,与当年8月降水、上一年9月及当年8月平均相对湿度和上一年7~10月的scPDSI呈显著正相关。石河子地区雪岭云杉径向生长与当年1月平均湿度和当年1、2月的scPDSI呈显著正相关。此外,乌苏地区树轮宽度指数具有与温度升高而下降的“分离现象”,而石河子地区树轮宽度指数具有与温度升高相一致的趋势。升温导致的水分胁迫是造成乌苏地区雪岭云杉径向生长与温度变化趋势相反的重要因素。生长季的温度和降水的增加对石河子地区雪岭云杉径向生长有促进作用。乌苏地区雪岭云杉径向生长对5~7月平均气温和降水量变化敏感性上升;石河子地区雪岭云杉径向生长对5~7月降水量变化敏感性上升,而对5~7月平均气温变化敏感性下降。结果显示,气候变化的区域差异是造成乌苏和石河子地区雪岭云杉径向生长趋势不同以及各年表序列对温度和降水变化敏感性变化的主要气候因素。     

Is climate warming more consequential towards poles? The phenology of Lepidoptera in Finland

The magnitude and direction of phenological shifts from climate warming could be predictably variable across the planet depending upon the nature of physiological controls on phenology, the thermal sensitivity of the developmental processes and global patterns in the climate warming. We tested this with respect to the flight phenology of adult nocturnal moths (3.33 million captures of 334 species) that were sampled at sites in southern and northern Finland during 1993–2012 (with years 2005–2012 treated as an independent model validation data set). We compared eight competing models of physiological controls on flight phenology to each species and found strong support for thermal controls of phenology in 66% of the species generations. Among species with strong thermal control of phenology in both the south and north, the average development rate was higher in northern vs. southern populations at 10 °C, but about the same at 15 and 20 °C. With a 3 °C increase in temperature (approximating A2 scenario of IPPC for 2090–2099 relative to 1980–1999) these species were predicted to advance their phenology on average by 17 (SE ± 0.3) days in the south vs. 13 (±0.4) days in the north. The higher development rates at low temperatures of poleward populations makes them less sensitive to climate warming, which opposes the tendency for stronger phenological advances in the north from greater increases in temperature.     

Modelling the spatiotemporal dynamics of chemovirotherapy cancer treatment

Chemovirotherapy is a combination therapy with chemotherapy and oncolytic viruses. It is gaining more interest and attracting more attention in the clinical setting due to its effective therapy and potential synergistic interactions against cancer. In this paper, we develop and analyse a mathematical model in the form of parabolic non-linear partial differential equations to investigate the spatiotemporal dynamics of tumour cells under chemovirotherapy treatment. The proposed model consists of uninfected and infected tumour cells, a free virus, and a chemotherapeutic drug. The analysis of the model is carried out for both the temporal and spatiotemporal cases. Travelling wave solutions to the spatiotemporal model are used to determine the minimum wave speed of tumour invasion. A sensitivity analysis is performed on the model parameters to establish the key parameters that promote cancer remission during chemovirotherapy treatment. Model analysis of the temporal model suggests that virus burst size and virus infection rate determine the success of the virotherapy treatment, whereas travelling wave solutions to the spatiotemporal model show that tumour diffusivity and growth rate are critical during chemovirotherapy. Simulation results reveal that chemovirotherapy is more effective and a good alternative to either chemotherapy or virotherapy, which is in agreement with the recent experimental studies.