Physicochemical assessment of Unio crassus habitat quality in a small upland stream and implications for conservation

It has been proposed that communities change from r to K strategies during primary succession. However, because strong-flying organisms are expected to arrive first in newly created habitats and they show trait characteristics associated more often with K strategies, we hypothesized that the r to K trajectories would be more closely followed by flightless and poorly-flying organisms. Moreover, we expected that macroinvertebrate communities would converge in their functional composition due to deterministic forces while diverging as taxonomic assemblages due to stochastic drift and biotic interactions. However, we also expected that dispersal abilities of the organisms would affect these tendencies. To address these issues, macroinvertebrates were sampled from isolated manmade ponds of different ages (1–22 years old) constructed at reclaimed opencast coal mines. In accordance with our expectations, only flightless and poorly-flying organisms exhibited a slight shift from r to K strategies, the community taxonomically diverged along the primary succession gradient, and stochastic drift showed greater effects on strong-flying organisms. In contrast, the community did not converge in its functional composition. The weak differences observed among the macroinvertebrates from ponds of different ages suggested that limiting environmental conditions prevented the organisms from evolving to a more structured community.     

An automated screening method for drugs and toxic compounds in human serum and urine using liquid chromatography–tandem mass spectrometry

A fully automated screening using liquid chromatography–mass spectrometric method applying data-dependent acquisition was developed to identify toxicologically relevant substances in serum and urine. A library including more than 405 spectra of about 365 compounds (main drugs and important metabolites) was established. An easy to use program was created to automate and accelerate library search. Drugs were identified based on their relative retention times, molecular ions and fragment ions. Limits of detection were tested with 100 of the 365 compounds the majority of these were lower than 100 μg/l (67%). The developed LC–MS–MS system seems to be a valuable alternative to other general unknown screening methods allowing fast and specific identification of drugs in serum and urine samples.     

Apoptosis initiation and angiogenesis inhibition: melanoma targets for nanosecond pulsed electric fields

Many effective anti-cancer strategies target apoptosis and angiogenesis mechanisms. Applications of non-ionizing, nanosecond pulsed electric fields (nsPEFs) induce apoptosis in vitro and eliminate cancer in vivo; however in vivo mechanisms require closer analysis. These studies investigate nsPEF-induced apoptosis and anti-angiogenesis examined by fluorescent microscopy, immunoblots, and morphology. Six hours after treatment with one hundred 300 ns pulses at 40 kV/cm, cells transiently expressed active caspases indicating that caspase-mediated mechanisms. Three hours after treatment transient peaks in Histone 2AX phosphorylation coincided with terminal deoxynucleotidyl transferase dUTP nick end labeling positive cells and pyknotic nuclei, suggesting caspase-independent mechanisms on nuclei/DNA. Large DNA fragments, but not 180 bp fragmentation ladders, were observed, suggesting incomplete apoptosis. Nevertheless, tumor weight and volume decreased and tumors disappeared. One week after treatment, vessel numbers, vascular endothelial growth factor (VEGF), platelet derived endothelial cell growth factor (PD-ECGF), CD31, CD35 and CD105 were decreased, indicating anti-angiogenesis. The nsPEFs activate multiple melanoma therapeutic targets, which is consistent with successes of nsPEF applications for tumor treatment in vivo as a new cancer therapeutic modality.     

Ultrahigh bioproductivity from algae

The potential for dramatic increases in bioproductivity in algal photobioreactors relative to current biomass approaches, e.g., for converting sunlight into biofuels, by an unorthodox integration of photonics and biotechnologies is described. The key to greater biomass yields—projected as high as 100 g dry weight m⁻² h⁻¹—is a pronounced heightening of algal flux tolerance, achieved by tailoring the photonic temporal, spectral and intensity characteristics with pulsed light-emitting diodes. Such tailored photonic input is applied in concert with thin-channel ultradense culture photobioreactors with flow patterns that produce rapid light/dark algae exposure cycles. The artificial-light scheme is globally feasible only with electricity generated from renewables. Recent advances in ultra-efficient concentrator photovoltaics, as well as high-performance light-emitting diodes, create a practical reality for converting sunlight into pulsed red light and delivering it to indoor photobioreactors, with characteristic pulse times and intensities optimally suited to the rate-limiting dark reactions of photosynthesis. Cellular engineering built upon recent progress in modifying algal chlorophyll antenna size, in combination with metabolic engineering, could further enhance bioproductivity. The proposed strategy requires no major advances for implementation and adopts existing technologies. Revision submitted to Applied Microbiology and Biotechnology on 25 June 2007.     

Intracellular Ca2+ oscillations, a potential pacemaking mechanism in early embryonic heart cells

Early (E9.5-E11.5) embryonic heart cells beat spontaneously, even though the adult pacemaking mechanisms are not yet fully established. Here we show that in isolated murine early embryonic cardiomyocytes periodic oscillations of cytosolic Ca(2+) occur and that these induce contractions. The Ca(2+) oscillations originate from the sarcoplasmic reticulum and are dependent on the IP(3) and the ryanodine receptor. The Ca(2+) oscillations activate the Na(+)-Ca(2+) exchanger, giving rise to subthreshold depolarizations of the membrane potential and/or action potentials. Although early embryonic heart cells are voltage-independent Ca(2+) oscillators, the generation of action potentials provides synchronization of the electrical and mechanical signals. Thus, Ca(2+) oscillations pace early embryonic heart cells and the ensuing activation of the Na(+)-Ca(2+) exchanger evokes small membrane depolarizations or action potentials.     

Molecular architecture of strictosidine glucosidase: the gateway to the biosynthesis of the monoterpenoid indole alkaloid family

Strictosidine beta-D-glucosidase (SG) follows strictosidine synthase (STR1) in the production of the reactive intermediate required for the formation of the large family of monoterpenoid indole alkaloids in plants. This family is composed of approximately 2000 structurally diverse compounds. SG plays an important role in the plant cell by activating the glucoside strictosidine and allowing it to enter the multiple indole alkaloid pathways. Here, we report detailed three-dimensional information describing both native SG and the complex of its inactive mutant Glu207Gln with the substrate strictosidine, thus providing a structural characterization of substrate binding and identifying the amino acids that occupy the active site surface of the enzyme. Structural analysis and site-directed mutagenesis experiments demonstrate the essential role of Glu-207, Glu-416, His-161, and Trp-388 in catalysis. Comparison of the catalytic pocket of SG with that of other plant glucosidases demonstrates the structural importance of Trp-388. Compared with all other glucosidases of plant, bacterial, and archaeal origin, SG's residue Trp-388 is present in a unique structural conformation that is specific to the SG enzyme. In addition to STR1 and vinorine synthase, SG represents the third structural example of enzymes participating in the biosynthetic pathway of the Rauvolfia alkaloid ajmaline. The data presented here will contribute to deciphering the structure and reaction mechanism of other higher plant glucosidases.     

Trophic relationships between the larvae of two freshwater mussels and their fish hosts

Freshwater mussels of the order Unionoida have life cycles that include larval attachment to and later metamorphosis on suitable host fishes. Information on the trophic relationship between unionoid larvae and their host fishes is scarce. We investigated the trophic interaction between fish hosts and encysted larvae of two species of freshwater mussels, Margaritifera margaritifera and Unio crassus, using stable isotope analyses of larvae and juvenile mussels as well as of host fish gill and muscle tissues before and after infestation. Due to different life histories and durations of host‐encystment, mass and size increase in M. margaritifera during the host‐dependent phase were greater than those of U. crassus. δ¹³C and δ¹⁵N signatures of juvenile mussels approached isotopic signatures of fish tissues, indicating a parasitic relationship between mussels and their hosts. Shifts were more pronounced for M. margaritifera, which had a five‐fold longer host‐dependent phase than U. crassus. The results of this study suggest that stable isotope analyses are a valuable tool for characterizing trophic relationships and life history strategies in host–parasite systems. In the case of unionoid mussels, stable isotopic shifts of the larvae are indicative of the nutritional versus phoretic importance of the host.     

Site-specific recombinases: from tag-and-target- to tag-and-exchange-based genomic modifications

Site-specific recombinases (SSRs) enable novel tag-and-target as well as tag-and-exchange strategies for tailoring mammalian genomes. If used in combination with homologous recombination, which per se is inefficient but can serve to introduce SSR sites, the tagged locus lends itself to repeated modification at largely increased efficiency and specificity. The more conventional SSR-based genetic modifications enable straightforward integration of a transgene with efficiencies depending on both the target locus and the vector composition. Only the more recent tag-and-exchange strategies in conjunction with advanced selection principles enable the clean replacement of a genomically anchored cassette by a donor cassette with the related architecture. Meanwhile this recombinase-mediated cassette exchange (RMCE) concept could be verified for two classes of SSRs, belonging to either the Tyr or the Ser family. Certain members of these open different fields of application that will be discussed with reference to the molecular properties of the respective enzymes. A major aim of our review is to characterize the RMCE-relevant components and describe their optimal utilization in the fields of gene therapy and molecular genomics. Early contributions to the field of experimental animal models will be mentioned considering in vivo modifications enabled by microinjection into oocytes.