Novel ENU-Induced Point Mutation in Scavenger Receptor Class B,Member 1, Results in Liver Specific Loss of SCARB1 Protein

Cardiovascular disease (CVD) is the largest cause of premature death in human populations throughout the world. Circulating plasma lipid levels, specifically high levels of LDL or low levels of HDL, are predictive of susceptibility to CVD. The scavenger receptor class B member 1 (SCARB1) is the primary receptor for the selective uptake of HDL cholesterol by liver and steroidogenic tissues. Hepatic SCARB1 influences plasma HDL-cholesterol levels and is vital for reverse cholesterol transport. Here we describe the mapping of a novel N-ethyl-N-nitrosourea (ENU) induced point mutation in the Scarb1 gene identified in a C57BL/6J background. The mutation is located in a highly conserved amino acid in the extracellular loop and leads to the conversion of an isoleucine to an asparagine (I179N). Homozygous mutant mice express normal Scarb1 mRNA levels and are fertile. SCARB1 protein levels are markedly reduced in liver (∼90%), but not in steroidogenic tissues. This leads to ∼70% increased plasma HDL levels due to reduced HDL cholesteryl ester selective uptake. Pdzk1 knockout mice have liver-specific reduction of SCARB1 protein as does this mutant; however, in vitro analysis of the mutation indicates that the regulation of SCARB1 protein in this mutant is independent of PDZK1. This new Scarb1 model may help further our understanding of post-translational and tissue-specific regulation of SCARB1 that may aid the important clinical goal of raising functional HDL.     

Diversity of bacteria associated with grassland soil nematodes of different feeding groups

The bacterial diversity associated with soil nematodes and its relationship with their feeding habits are as yet poorly understood. In the present study the diversity and abundance of bacteria from nematodes and their surrounding soil were analysed and compared. The nematodes were collected from a grassland soil and sorted into bacterial, fungal, plant, predatory and omnivore feeding groups and assigned to taxonomic groups. Total DNA was extracted from the nematodes and partial bacterial 16S rRNA genes were PCR amplified, cloned and sequenced. The abundance and composition of bacterial taxa differed between and within feeding groups. The lowest bacterial diversity was found in the predatory nematodes Prionchulus sp., whereas the highest bacterial diversity was associated with the bacterial-feeding nematode Acrobeles sp. The soil had a more diverse bacterial community than the communities found in the nematode groups. The 16S rRNA gene sequences of bacteria associated with nematodes did not overlap with those detected in soil as determined using the cloning screening approach. However, bacterial sequences identified from nematodes could be detected in the soil with targeted PCR. Our data suggest that the nematodes do not feed on the most abundant bacteria present in soil. Furthermore, several nematodes contained suspected bacterial symbionts and parasites.     

A novel approach to identifying beta-secretase inhibitors: bis-statine peptide mimetics discovered using structure and spot synthesis

Beta-secretase 1 (BACE1) is an aspartic protease believed to play a critical role in Alzheimer's disease. Inhibitors of this enzyme have been designed by incorporating the non-cleavable hydroxyethylene and statine isosteres into peptides corresponding to BACE1 substrate sequences. We sought to develop new methods to quickly characterize and optimize inhibitors based on the statine core. Minimal sequence requirements for binding were first established using both crystallography and peptide spot synthesis. These shortened peptide inhibitors were then optimized by using spot synthesis to perform iterative cycles of substitution and deletion. The present study resulted in the identification of novel "bis-statine" inhibitors shown by crystallography to have a unique binding mode. Our results demonstrate the application of peptide spot synthesis as an effective method for enhancing peptidomimetic drug discovery.     

Effects of patch size and density on flower visitation and seed set of wild plants: a pan-European approach

1.  Habitat fragmentation can affect pollinator and plant population structure in terms of species composition, abundance, area covered and density of flowering plants. This, in turn, may affect pollinator visitation frequency, pollen deposition, seed set and plant fitness.
2.  A reduction in the quantity of flower visits can be coupled with a reduction in the quality of pollination service and hence the plants' overall reproductive success and long-term survival. Understanding the relationship between plant population size and/or isolation and pollination limitation is of fundamental importance for plant conservation.
3.  We examined flower visitation and seed set of 10 different plant species from five European countries to investigate the general effects of plant populations size and density, both within (patch level) and between populations (population level), on seed set and pollination limitation.
4.  We found evidence that the effects of area and density of flowering plant assemblages were generally more pronounced at the patch level than at the population level. We also found that patch and population level together influenced flower visitation and seed set, and the latter increased with increasing patch area and density, but this effect was only apparent in small populations.
5.   Synthesis. By using an extensive pan-European data set on flower visitation and seed set we have identified a general pattern in the interplay between the attractiveness of flowering plant patches for pollinators and density dependence of flower visitation, and also a strong plant species-specific response to habitat fragmentation effects. This can guide efforts to conserve plant–pollinator interactions, ecosystem functioning and plant fitness in fragmented habitats.     

Use of trace elements in feathers of sand martin Riparia riparia for identifying moulting areas

We investigated whether trace elements in tail feathers of an insectivorous and long-distance migratory bird species could be used to identify moulting areas and hence migratory pathways. We analysed tail feathers from birds of different age and sex collected from a range of different breeding sites across Europe. The site of moult had a large effect on elemental composition of feathers of birds, both at the European and African moulting sites. Analysis of feathers of nestlings with known origin suggested that the elemental composition of feathers depended largely upon the micro-geographical location of the colony. The distance between moulting areas could not explain the level of differences in trace elements. Analysis of feathers grown by the same individuals on the African wintering grounds and in the following breeding season in Europe showed a large difference in composition indicating that moulting site affects elemental composition. Tail feathers moulted in winter in Africa by adults breeding in different European regions differed markedly in elemental composition, indicating that they used different moulting areas. Analysis of tail feathers of the same adult individuals in two consecutive years showed that sand martins in their first and second wintering season grew feathers with largely similar elemental composition, although the amounts of several elements in tail feathers of the older birds was lower. There was no difference between the sexes in the elemental composition of their feathers grown in Africa. Investigation of the trace element composition of feathers could be a useful method for studying similarity among groups of individuals in their use of moulting areas.     

Cell division in <Emphasis Type="Italic">Escherichia coli</Emphasis>cultures monitored at single cell resolution

Background  

A fundamental characteristic of cells is the ability to divide. To date, most parameters of bacterial cultures, including cell division, have been measured as cell population averages, assuming that all bacteria divide at a uniform rate.     

Over-expression of xylulokinase in a xylose-metabolising recombinant strain of Zymomonas mobilis

The broad host range vector pBBR1MCS-2 has been evaluated as an expression vector for Zymomonas mobilis. The transformation efficiency of this vector was 2 x 10(3) CFU per mug of DNA in a recombinant strain of Z. mobilis ZM4/AcR containing the plasmid pZB5. Stable replication for this expression vector was demonstrated for 50 generations. This vector was used to study xylose metabolism in acetate resistant Z. mobilis ZM4/AcR (pZB5) by over-expression of xylulokinase (XK), as previous studies had suggested that XK could be the rate-limiting enzyme for such strains. Based on the above vector, a recombinant plasmid pJX1 harboring xylB (expressing XK) under control of a native Z. mobilis promotor Ppdc was constructed. When this plasmid was introduced into ZM4/AcR (pZB5) a 3-fold higher XK expression was found compared to the control strain. However, fermentation studies with ZM4/AcR (pZB5, pJX1) on xylose medium did not result in any increase in rate of growth or xylose metabolism, suggesting that XK expression was not rate-limiting for ZM4/AcR (pZB5) and related strains.     

Effects of Kainic Acid on Brain Calcium Fluxes Studied In Vivo and In Vitro

The effect of in vivo administration of kainic acid into the rabbit hippocampus was studied with brain dialysis and subsequent determination of the Ca2+ concentration in the dialysate. When included in the perfusing medium, kainic acid as well as veratridine induced a decrease in extracellular Ca2+. The effect of kainic acid (but not of veratridine) was insensitive to tetrodotoxin. In vitro studies revealed no effect of kainic acid on 45Ca2+ uptake by isolated astrocytes, but showed an enhancement of synaptosomal 45Ca2+ accumulation. This was, however, only 25% of the stimulatory effect of high K+ depolarization. Glutamate activated synaptosomal Ca2+ uptake, whereas dihydrokainate had no effect. The uptake evoked by kainate and glutamate was independent of the K+ level in the medium which indicates the involvement of other than voltage-sensitive Ca2+ channels. The results confirm previous finding that kainic acid promotes the uptake of Ca2+ in brain cells. Kainate affects Ca2+ fluxes pre- and postsynaptically. Presynaptic Ca2+ influx may be mediated by chemically gated mechanisms.     

Effects of copper,zinc, and cadmium ions on the production of phosphate from phytic acid by the phytase system in spruce forest soil

Summary The hydrolysis of phosphate from phytic acid by the acid soil phytase system was reduced in the presence of metal ions. Copper was most effective in this respect — zinc and cadmium were less inhibitory. Binding to metals did not completely inhibit the hydrolysis of phytic acid. At higher metal concentrations, where binding to other soil constituents, like humic acids, interfered less, the inhibition of the phytase activity was stronger than that of acid phosphatase.