Three sea urchin actin genes show different patterns of expression: muscle specific, embryo specific, and constitutive.

The expression of three different actin genes in the sea urchin, Strongylocentrotus purpuratus, was monitored in embryos and adult tissues by using untranslated mRNA sequences as specific hybridization probes. Three distinct patterns of expression were found: muscle specific, embryo specific, and constitutive (i.e., present in all tissues examined). The actin genes encoding the muscle-specific and constitutively expressed genes were each found to be present once in the haploid genome. The embryo-specific probe could derive from either a single gene or a small subset of actin genes. These data demonstrate that at least three members of the sea urchin actin gene family are expressed in distinct ways and thus are probably associated with different regulatory programs of gene expression necessary for development of this metazoan.     

Comparison of methods for extracting intracellular pools of inorganic nitrogen from marine phytoplankton

Marine phytoplankton can accumulate large intracellular poolsof nitrate and ammonium under some growth conditions. A numberof different methods have been used to extract these pools whichdiffer primarily in the way the cells are broken to releasethe pools. In this study, seven methods of extracting nitrateand ammonium pools were examined in four species of marine phytoplanktongrown so that extraction of both large and small pools couldbe tested. The methods of breaking cells examined included variouscombinations of osmotic shock, heat treatment, grinding, sonication,and freeze–thawing. Of these, two methods which use osmoticshock or osmotic shock combined with heating, yielded the highestpool concentrations, the best percent recoveries, and the leastvariability. Osmotic shock combined with heat is the recommendedextraction procedure since it may be more effective with cellswhich are difficult to break. ¹Current address: Bigelow Laboratory for Ocean Sciences, MckownPoint, West Boothbay, Maine 04575, USA     

Requirement for different Ca2+ pools in the activation of rabbit platelets: II. Phospholipase activity

The effects of extracellular Ca²⁺ concentration and the putative antagonist of intracellular Ca²⁺ movement, 8-(N,N-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8) on platelet phospholipase activity and thromboxane B₂ synthesis were examined in rabbit platelets stimulated by platelet activating factor, thrombin and ionophore A23187. TMB-8 markedly inhibited the platelet activating factor-induced decrease in [¹⁴C]arachidonate content in platelet phsophatidylacholine and phosphatidylinositol, while showing minimal effects on thrombin-induced phospholipase activation. A23187 stimulation of these processes was inhibited to an intermediated degree by TMB-8. In contrast, extracellular Ca²⁺ removal inhibited phospholipase activity to a similar degree with all three stimuli. Moreover, the threshold concentration of extracelullar Ca²⁺ for phospholiphase activation, as measured by thromboxane B₂ synthesis, was similar for platelet activating factor- and thrombin-stimulated platelets. The data provide evidence that, while platelet activating factor and thrombin may, to some extent, have similar requirements for extracellular Ca²⁺, they utilize a TMB-8 sensitive step to different degrees during activation of platelet phospholipase.     

Diel patterns of phytoplankton productivity in Lake Vechten

Summary A review of the literature on phytoplankton primary production studies in lakes indicates that in most cases the daily primary production rates have been derived by multiplying the short-term (usually 3 or 4 hours) production measurement with a light factor. This factor is a ratio between the total irradiance during the day and during the period of incubation. The aim of the present study was to verify this method of calculation. Since the primary production rate is of great importance for the carbon cycle of lakes, this study can be regarded as a fundamental contribution to the integrated research of the ecosystem of lake Vechten (the Netherlands) (BESTet al., 1978).Photosynthetic carbon fixation was determined using the radiocarbon method of STEEMANN NIELSEN (1952). Thein situ measurements were carried out for 2 hours incubation periods starting at 15–30 minutes before sunrise and ending 15–30 minutes after sunset. Also measured were: water temperature, light penetration, oxygen and chlorophyll. Total incident solar radiation data were obtained from the Royal Netherlands Meteorological Institute (K.N.M.I.) at de Bilt (situated at 5 km from lake Vechten). The production during two consecutive 2-hourly periods,e.g. period 1 and period 2, were summed up and using the irradiance data during the 4 hours, daily production was calculated. These calculations were repeated for period 2 and 3, and so on. The sum of all the short-term measurements during a day,i.e. the total daily primary production, was taken for comparison with the production calculated by using the sum of the production during two successive periods and the light factor.Daily production rates obtained by extrapolating from short-term incubation periods both in the morning and in the late afternoon appeared to overestimate the real daily production (Fig. 1) up to 100%. It is likely, that in the morning the lack of nutrient limitation and relatively better physiological state of algae, and in the afternoon increase of phytoplankton biomass during the day can be considered as the factors causing the discrepancy between the calculated and real production. On the other hand, the underestimation of daily production rate based on midday time intervals on sunny days, was about 25%. It is attributed to photoinhibition, especially in the present study where the measurements were limited to the upper water strata (0–4m).Is is concluded that short-termin situ exposures between 10.00 and 14.00 hr appeared to be a good indicator of total primary production, although underestimates up to 25% on sunny days are to be taken into account.     

Efflux of cholesterol from different cellular pools

Free cholesterol is very efficiently removed from cells by 2-hydroxypropyl-beta-cyclodextrins. The efflux of cholesterol occurs from two distinct kinetic pools: the half-times (t(1/2)) for the two pools in CHO-K1 cells are 15 +/- 5 s and 21 +/- 6 min and they represent 25% +/- 5% and 75% +/- 5% of the readily exchangeable cell cholesterol, respectively. In this study we have determined that the fast pool and the majority of the slow kinetic pool for cholesterol efflux are apparently present in the plasma membrane. Numerous agents that inhibit intracellular cholesterol trafficking are unable to affect either the size or the t(1/2) for efflux of either kinetic pool. In contrast, treatment of the cells with N-ethylmaleimide (NEM), exogenous lipases such as sphingomyelinase and phospholipase C, calcium ionophore A23187, or heat resulted in the dramatic increase in the size of the fast kinetic pool of cholesterol. These changes in the kinetics of cholesterol efflux are not specific to the nature of the extracellular acceptor indicating that they are a consequence of changes in the cell plasma membrane. The above treatments disrupt the normal organization of the lipids in the plasma membrane via either hydrolysis or randomization. The phosphatidylcholine and sphingomyelin present in the plasma membrane are critical for maintaining the two kinetic pools of cholesterol; any alteration in the amount or the location of these phospholipids results in an enhancement of efflux by redistributing cholesterol into the fast kinetic pool.     

Biodiversity correlates with regional patterns of forest litter pools

We compared litter pools of more than 1,000 forests differing in tree species diversity over a large scale in Catalonia (NE Spain). Monospecific forests always had smaller litter pools than mixed (from 2 to 5 tree species) forests. Whether there was a positive effect beyond two species mixtures depended on the species and functional identity of the dominant tree species. In sclerophyllous forests the positive effect of diversity was a step-function from one to more species. However, in conifers, litter pools increased constantly with tree diversity. The identity of the dominant tree species and functional type had also a significant effect on litter pools. For instance, forests dominated by sclerophyllous tree species had larger litter pools than forests dominated by deciduous and conifer tree species. When other forest structure parameters (i.e. tree basal area, wood production, successional stage, shrub cover and leaf area index) and environmental factors (i.e. mean annual temperature, mean annual precipitation, annual evapotranspiration and hillside position) where included in the analysis only leaf area index, basal area, wood production and mean temperature influenced litter pools positively. Our analysis emphasizes that at the regional scale, the litter compartment can be as influenced by biodiversity components as by other forest structure and climate components. In mixed forests, species and functional identity of the trees determine whether litter pools increase with tree diversity.     

Changing patterns of carbon allocation in lake phytoplankton: an FTIR analysis

Fourier transform infra-red (FTIR) spectroscopy was used to assess carbon allocation in selected phytoplankton (Pediastrum duplex, Ceratium hirundinella, Anabaena flos-aquae) in a eutrophic lake. In contrast to bulk-analysis techniques, FTIR spectroscopy can provide information on carbon allocation at the species level within natural mixed populations. Carbon allocation was determined using lipid/protein (L/P) and carbohydrate/protein (C/P) ratios and multivariate analysis of complete spectra and showed considerable intraspecific heterogeneity within samples dates, and clear seasonal changes in the spectra of Pediastrum and Anabaena, with both alga showing increased L/P and C/P ratios on selected sampling dates. Although increased ratios can indicate nutrient deficiency, the alga were not nutrient deficient during these periods, as indicated by continued population growth and high (>0.1%) internal phosphorus concentrations. FTIR spectra from Ceratium did not show any clear seasonal change. In all three algae, the transition from population growth to decline was not accompanied by any significant change in ratios, nor were low epilimnetic nutrients correlated with increased ratios, marking a significant contrast to laboratory batch cultures (previous studies) where low nutrients and the transition to stationary phase led to marked spectral changes.     

The effect of different zooplankton grazing patterns resulting from diel vertical migration on phytoplankton growth and composition: a laboratory experiment

Diel vertical migration (DVM) of herbivorous zooplankton is a widespread behavioural phenomenon in freshwater ecosystems. So far only little attention has been paid to the impact of DVM on the phytoplankton community in the epilimnion. Some theoretical models predict that algal population growth in the epilimnion should depend on the herbivores migration and grazing patterns: even if migrating zooplankton consume the same total amount of algae per day in the epilimnion as non-migrating zooplankton, nocturnal grazing should result in enhanced algal growth and favour algal species with high intrinsic growth rates over species with lower intrinsic growth rates. To test these hypotheses we performed experiments in which several algal species were confronted with different feeding regimes of Daphnia. In the experiments algal growth did not only depend on the absolute time of grazing but was comparatively higher when grazing took place only during the night, even when the grazing pressure was the same. Furthermore, algal species with higher intrinsic growth rates had higher advantages when being grazed upon only discontinuously during the night than algal species with a smaller intrinsic growth rate. The grazing pattern itself was an important factor for relative algal performance.     

Aspects of aerobic mineralization during spring in Lake Vechten with special reference to the 14C-labelling technique

Aerobic mineralization, i.e. seston respiration, microbial breakdown of detritus and microbial assimilation-dissimilation of photosynthetically derived D(issolved) O(rganic) C(arbon) was measured in concentrated samples from the pelagic zone of Lake Vechten. The samples were described by cell numbers of dominant algae prior to concentrating by centrifuge with continuous rotor. The concentrated samples were incubated in the laboratory at in situ temperature and a light intensity of 30 W · m^–2 for measuring primary production and photosynthetically derived DOC. After filtration the particulate fraction was incubated in unlabelled lake water far measuring respiratory production of DI¹⁴C. Portions of the same particulate fraction were sterilized and incubated in unlabelled sample concentrate for measuring microbial breakdown of detritus. The seston respiration amounted to 19–30% loss of the particulate fraction over 15–45 h. Microbial breakdown of detritus amounted to 28–40% loss of the particulate fraction over 24–168 h. In both cases P(articulate) O(rganic) ¹⁴C(arbon) was transformed to mainly DI¹⁴C. Microbial assimilation-dissimilatioh of photosynthetically derived DOC could not be measured reliably.     

The influence of different glycosylation patterns on factor VII biological activity

In this study the bioactivity of three differently glycosylated blood coagulation factor VII (FVII) variants (human plasma FVII, recombinant human FVII produced in CHO and BHK cell cultures) were analyzed and compared. Surface plasmon resonance studies of FVII interaction with soluble and full length TF together with FVII autoactivation assays revealed that BHK-derived FVII has the highest bioactivity, while human plasma and CHO-derived FVII showed very similar bioactivity. The affinity of FVII variants to TF correlates with FVII autoactivation rates – the higher the affinity, the faster the autoactivation rate.     

Optimization of 14C-lysine concentration and specific activity for the radiometric detection of microorganisms

Summary The sensitivity of the radiometric detection of microbial contamination based on the labeling of cells by ¹⁴C-lysine was studied as a function of the lysineconcentration and its specific activity for a strain of E. coli and a strain of S. cerevisiae. It was found that best conditions of detection were given by a labelled lysine specific activity of 200 mC_i/mmole and a medium radioactivity of 0.2 C_i/ml.     

Invertebrate communities in dry-season pools of a large subtropical river: patterns and processes

In order to develop an optical model to map the extent of coastal waters, the authors analyzed variations in bio-optical constituents and submarine optical properties along a transect from the nutrient-enriched coastal bay, Himmerfjärden, out into the open Baltic Sea. The model is a simple implementation of the “ecosystem approach,” because the optical constituents are proxies for important components of ecosystem state. Yellow substance or colored dissolved organic matter (CDOM) is often a marker for terrestrial freshwater or decay processes in the littoral zone. Phytoplankton pigments, especially chlorophyll a, are used as a proxy for phytoplankton biomass that may be stimulated by fluvial or coastal inputs of anthropogenic nutrients. Suspended particulate matter (SPM) is placed in suspension by tidal or wind-wave stirring of shallow seabeds, and is therefore an indicator for physical forcing. It is the thesis of this article that such constituents, and the optical properties that they control, can be used to provide an ecological definition of the extent of the coastal zone. The spatial distribution of the observations was analyzed using a steady-state model that assumes diffusional transport of bio-optical variables along an axis perpendicular to the coast. According to the model, the resulting distribution along this axis can be described as a low-order polynomial (of order 1–3) when moving from a “source” associated with land to the open-sea “sink.” Order 1 implies conservative mixing, and the higher orders imply significant biological or chemical processes within the gradient. The analysis of the transect data confirmed that the trend of each optical component could be described well using a low-order polynomial. Multiple regression analysis was then used to weigh the contribution of each optical component to the spectral attenuation coefficient K _d(490) along the transect. The results showed that in this Swedish Baltic case study, the inorganic fraction of the SPM may be used to distinguish between coastal and open-sea waters, as it showed a clear break between coastal and open-sea waters. Alternative models may be needed for coastal waters in which fronts interrupt the continuity of mixing.     

Temporal patterns of carbon flow from grassland vegetation to soil microorganisms measured using 13C-labelling and signature fatty acids

Plant and Soil - Since the 1980s, numerous mutualistic Pseudomonas spp. strains have been used in studies on the biology of plant growth-promoting rhizobacteria (PGPR) and their interactions with...     

Zooplankton grazing effects on 14C-based phytoplankton production measurements: a theoretical study

A model constructed to describe carbon dynamics of phytoplanktongrowth during ¹⁴C-tracer incubations gives the range of errorin estimating specific growth rates and productivity rates causedby zooplankton grazing during the incubation. Error increaseswith increasing incubation times and higher specific growthrates. The range of these errors can be given as a functionof the specific growth rate calculated from measurements. Atthe low calculated specific growth rates of the oligotrophicPacific Ocean, 0.2 d^–1, errors are 16%. Similar argumentssuggest that bacterial uptake of excreted organics would notcause large errors at low oligotrophic ocean growth rates. Thereare, however, other possible ways that ¹⁴C-based productivityestimates could be wrong.     

Purification of a highly modified RNA-aptamer: Effect of complete denaturation during chromatography on product recovery and specific activity

To evaluate RNA-aptamers as potential drug candidates, efficient and scaleable purification protocols are needed. Because aptamers are highly structured and rigid molecules, denaturation during the purification process is a critical aspect to obtain a pure and active product. A two-step chromatographic procedure was developed to purify a synthetic anti-VEGF aptamer at the preparative scale. A reversed-phase chromatographic step was optimized with a highly hydrophobic ion pairing reagent, followed by ion-exchange chromatography in which heat and a chaotropic salt were used. Because of the presence of 2′-modified ribose, denaturation conditions had to be optimized in both chromatographic steps to achieve a fully active molecule.     

Fabrication of 14 different RNA nanoparticles for specific tumor targeting without accumulation in normal organs

Due to structural flexibility, RNase sensitivity, and serum instability, RNA nanoparticles with concrete shapes for in vivo application remain challenging to construct. Here we report the construction of 14 RNA nanoparticles with solid shapes for targeting cancers specifically. These RNA nanoparticles were resistant to RNase degradation, stable in serum for >36 h, and stable in vivo after systemic injection. By applying RNA nanotechnology and exemplifying with these 14 RNA nanoparticles, we have established the technology and developed “toolkits” utilizing a variety of principles to construct RNA architectures with diverse shapes and angles. The structure elements of phi29 motor pRNA were utilized for fabrication of dimers, twins, trimers, triplets, tetramers, quadruplets, pentamers, hexamers, heptamers, and other higher-order oligomers, as well as branched diverse architectures via hand-in-hand, foot-to-foot, and arm-on-arm interactions. These novel RNA nanostructures harbor resourceful functionalities for numerous applications in nanotechnology and medicine. It was found that all incorporated functional modules, such as siRNA, ribozymes, aptamers, and other functionalities, folded correctly and functioned independently within the nanoparticles. The incorporation of all functionalities was achieved prior, but not subsequent, to the assembly of the RNA nanoparticles, thus ensuring the production of homogeneous therapeutic nanoparticles. More importantly, upon systemic injection, these RNA nanoparticles targeted cancer exclusively in vivo without accumulation in normal organs and tissues. These findings open a new territory for cancer targeting and treatment. The versatility and diversity in structure and function derived from one biological RNA molecule implies immense potential concealed within the RNA nanotechnology field.     

Exponential growth rates of phytoplankton calculated from 14C uptake rates: a clarification

Phytoplankton growth rates can be estimated from measurementsof light-dependent [¹⁴C]bicarbonate uptake (photosynthesis)rates using the integrated (logarithmic) form of the exponentialgrowth law. A clanfication of this method is presented herein view of confusions in the recent literature.