Products of the lipoxygenase pathway in human natural killer cell cytotoxicity

As earlier data suggested the importance of lipoxygenase activation for expression of human NK cell cytotoxicity, four different lipoxygenase inhibitors were tested for suppression of natural killer (NK) cell lysis. All inhibitors were found active at nontoxic concentrations with 50% inhibition at approximately 15 microM for nordihydroguaiaretic acid (NDGA). NK cell lysis could be reconstituted to NDGA-suppressed cells with leukotriene B4 (LTB4), the all-trans isomers 6-trans-LTB4 and 12-epi-6-trans-LTB4, and 20-COOH-LTB4. LTB4 reconstitution was best in the concentration range 1-100 pM and near control levels at both higher and lower concentrations. Herpesvirus Ateles-transformed killer T cells could also be inhibited by NDGA. These data indicate that lipoxygenase activity is required for human NK cell lysis and that several different LTB4-related products can restore NK activity in inhibited cells; they also suggest that the lipoxygenase pathway is present in the killer cell population.     

The lepidopteran mitochondrial control region: structure and evolution

For several species of lepidoptera, most of the approximately 350-bp mitochondrial control-region sequences were determined. Six of these species are in one genus, Jalmenus; are closely related; and are believed to have undergone recent rapid speciation. Recent speciation was supported by the observation of low interspecific sequence divergence. Thus, no useful phylogeny could be constructed for the genus. Despite a surprising conservation of control-region length, there was little conservation of primary sequences either among the three lepidopteran genera or between lepidoptera and Drosophila. Analysis of secondary structure indicated only one possible feature in common--inferred stem loops with higher-than-random folding energies-- although the positions of the structures in different species were unrelated to regions of primary sequence similarity. We suggest that the conserved, short length of control regions is related to the observed lack of heteroplasmy in lepidopteran mitochondrial genomes. In addition, determination of flanking sequences for one Jalmenus species indicated (i) only weak support for the available model of insect 12S rRNA structure and (ii) that tRNA translocation is a frequent event in the evolution of insect mitochondrial genomes.      

Comparative genomics of duplicate γ-glutamyl transferase genes in teleosts: medaka (Oryzias latipes), stickleback (Gasterosteus aculeatus), green spotted pufferfish (Tetraodon nigroviridis), fugu (Takifugu rubripes), and zebrafish (Danio rerio)

The availability of multiple teleost (bony fish) genomes is providing unprecedented opportunities to understand the diversity and function of gene duplication events using comparative genomics. Here we examine multiple paralogous genes of γ-glutamyl transferase (GGT) in several distantly related teleost species including medaka, stickleback, green spotted pufferfish, fugu, and zebrafish. Through mining genome databases, we have identified multiple GGT orthologs. Duplicate (paralogous) GGT sequences for GGT1 (GGT1 a and b), GGTL1 (GGTL1 a and b), and GGTL3 (GGTL3 a and b) were identified for each species. Phylogenetic analysis suggests that GGTs are ancient proteins conserved across most metazoan phyla and those paralogous GGTs in teleosts likely arose from the serial 3R genome duplication events. A third GGTL1 gene (GGTL1c) was found in green spotted pufferfish; however, this gene is not present in medaka, stickleback, or fugu. Similarly, one or both paralogs of GGTL3 appear to have been lost in green spotted pufferfish, fugu, and zebrafish. Syntenic relationships were highly maintained between duplicated teleost chromosomes, among teleosts and across ray-finned (Actinopterygii) and lobe-finned (Sarcopterygii) species. To assess subfunction partitioning, six medaka GGT genes were cloned and assessed for developmental and tissue-specific expression. On the basis of these data, we propose a modification of the "duplication-degeneration-complementation" model of subfunction partitioning where quantitative differences rather than absolute differences in gene expression are observed between gene paralogs. Our results demonstrate that multiple GGT genes have been retained within teleost genomes. Questions remain, however, regarding the functional roles of multiple GGTs in these species.     

Composition and chirality of amino acids in aerosol/dust from laboratory and residential enclosures

Initial results from the analyses of geological and anthropological samples for amino acids were difficult to accept because of the high enantiomeric purities of the analytes (i.e., predominantly L-amino acids). Consequently, sources of contamination had to be considered. All sources were eliminated except for direct atmospheric contamination. Essentially invisible, microscopic, aerosol/dust was found to rapidly contaminate the surface of samples and sample containers even after brief exposure times in clean laboratories. Contamination increased with exposure time. The aerosol/dust amino acids were contained predominantly in a proteinaceous material. Aerosol/dust from different locations can contain different percentages of proteinoid/amino acid material. However, the relative concentrations of the amino acids were similar for both laboratory and residential samples. The enantiomeric purity of the L-amino acids studied in aerosol/dust appears to be 99% or greater for the samples examined. Thus, even slight contamination of any sample with microscopic dust or aerosol particles can skew the results of trace amino acid analyses and amino acid e.e. determinations.     

Molecular Cloning,Functional Characterization,and Evolutionary Analysis of Vitamin D Receptors Isolated from Basal Vertebrates

The vertebrate genome is a result of two rapid and successive rounds of whole genome duplication, referred to as 1R and 2R. Furthermore, teleost fish have undergone a third whole genome duplication (3R) specific to their lineage, resulting in the retention of multiple gene paralogs. The more recent 3R event in teleosts provides a unique opportunity to gain insight into how genes evolve through specific evolutionary processes. In this study we compare molecular activities of vitamin D receptors (VDR) from basal species that diverged at key points in vertebrate evolution in order to infer derived and ancestral VDR functions of teleost paralogs. Species include the sea lamprey (Petromyzon marinus), a 1R jawless fish; the little skate (Leucoraja erinacea), a cartilaginous fish that diverged after the 2R event; and the Senegal bichir (Polypterus senegalus), a primitive 2R ray-finned fish. Saturation binding assays and gel mobility shift assays demonstrate high affinity ligand binding and classic DNA binding characteristics of VDR has been conserved across vertebrate evolution. Concentration response curves in transient transfection assays reveal EC₅₀ values in the low nanomolar range, however maximum transactivational efficacy varies significantly between receptor orthologs. Protein-protein interactions were investigated using co-transfection, mammalian 2-hybrid assays, and mutations of coregulator activation domains. We then combined these results with our previous study of VDR paralogs from 3R teleosts into a bioinformatics analysis. Our results suggest that 1, 25D₃ acts as a partial agonist in basal species. Furthermore, our bioinformatics analysis suggests that functional differences between VDR orthologs and paralogs are influenced by differential protein interactions with essential coregulator proteins. We speculate that we may be observing a change in the pharmacodynamics relationship between VDR and 1, 25D₃ throughout vertebrate evolution that may have been driven by changes in protein-protein interactions between VDR and essential coregulators.     

Alpine flora dynamics – a critical review of responses to climate change in the Swedish Scandes since the early 1950s

Reports about changes of alpine plant species richness over the past 60 years in the Swedish Scandes are reviewed, synthesized and updated with data from recent reinventories. Methodologically, this endeavour is based on resurveys of the floristic composition on the uppermost 20 m of four high‐mountain summits. The key finding is that the species pool has increased by 60–170% since the 1950s and later. Some of the invading species are new to the alpine tundra, with more silvine and thermophilic properties than the extant alpine flora. Not a single species of the original flora has disappeared from any of the summits. This circumstance is discussed in perspective of widespread expectations of pending temperature‐driven extinction of alpine species in an alleged future warmer climate. These progressive changes coincided with distinct warming (summer and winter) since the late 1980s. During a short cooler period (1974–1994), the species numbers decreased and the upper elevational limits of some ground cover species descended. Thus, discernible responses, concurrent with both warming and cooling intervals, sustain a strong causal link between climate variability and alpine plant species richness. Methodologically, plot‐less revisitation studies of the present kind are beset with substantial uncertainties, which may overstate floristic changes over time. However, it is argued here that carefully executed and critically interpreted, no other method can equally effectively sense the earliest phases of plant invasions into alpine vegetation.     

Immigration of Picea abies into North-Central Sweden. New evidence of regional expansion and tree-limit evolution

Analyses of subfossil tree remains in peats and raw humus soils account for the immigration and spread of Picea abies (Norway spruce) into Sweden and the evolution of the alpine tree-limit ecotone. Picea abies is recorded for the first time about 11 000 BP, on an early emerging nunatak in the southern Swedish Scandes. Prior to c. 8 000 BP, Picea was strictly bound to high elevations in the west. Farther to the east in North-Central Sweden, Picea emerged in the subfossil record mainly after c. 6 000 BP. Later on, growth of local founder populations and landscape-scale expansion may have been forced chiefly by a successively less seasonal climate (the Milankovitch model of orbital forcing), promoting increased net soil moisture and possibly a deeper and more persistent snow cover. Already at the Weichselian/Holocene transition, an elevational tree-limit ecotone was established and arborescent Picea grew at least 400 m higher than the modern tree-limit. Until about 8 000 BP, the species-limit descended, whereafter it stabilized up to the present day. Lack of significant species-limit retraction after c. 8 000 BP may seem paradoxical in perspective of independently inferred climate cooling in response to reduced insolation and land uplift. This could be a consequence of substantial phenotypic plasticity and increased snow accumulation, mitigating the long-term cooling. Thus, the elevational species-limit (krummholz) of Picea abies is out of equilibrium with the modern thermal climate. The striking incongruence between the results exposed here and earlier palynological interpretations of the same biogeographical process will have implications for the use of pollen data for range-limit reconstructions within historical biogeography, and urges for re-evaluation of certain aspects of the Fennoscandian forest history.     

The fatty acid profile of vegetative Azotobacter vinelandii ATCC 12837: growth phase-dependence

Fatty acids of Azotobacter vinelandii ATCC 12837 were determined at various times during aerobic vegetative growth at 30°C to provide baseline data for studying the effects of chemical agents on the organism’s survival and fatty acid biosynthesis. Palmitate (16:0) was the highest at 36.7±4.3 mol% (mean±SD) after the first 5 h in fresh culture, decreasing slightly to 33.4±2.6 mol% at 49 h. The other fatty acids were therefore each normalized as a ratio of 16:0. At 5 h, as a ratio of 16:0, myristate (14:0) was 0.14±0.06, palmitoleate (16:1cΔ^9–10) 0.13±0.06, oleate (18:1cΔ^9–10) 0.21±0.12, cis-vaccenate (18:1cΔ^11–12) 0.30±0.17 and stearate (18:0) 0.68±0.02. As the growth phase advanced to 49 h, 14:0 and 16:1cΔ^9–10 increased, 18:1cΔ^9–10 decreased and cis-vaccenate reciprocally increased, whereas 18:0 decreased. These suggest that the saturated fatty acid biosynthesis pathway yielded 16:0 and 18:0 in the 5-h lag period. By desaturation, 18:0 formed the unsaturated fatty acid (UFA) 18:1cΔ^9–10. As the culture aged, the anaerobic UFA biosynthesis pathway formed 16:1cΔ^9–10, which was elongated to 18:1cΔ^11–12. These fatty acid alterations represent a homeoviscous adaptation, modulating the microbe’s membrane lipid viscosity for optimal cellular function.     

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