A constant-volume ultrafiltration technique for the calculation of equilibrium binding data

A constant-volume ultrafiltration technique is described, and details of its assessment presented. The retention characteristics of two membranes were evaluated using molecules of known molecular weight.The technique is rapid, precise, economical of material and yields equilibrium data. In these respects, it compares favourably with conventional techniques such as equilibrium dialysis.     

Patterns of population genetic structure and diversity across bumble bee communities in the Pacific Northwest

Patterns of genetic structure and diversity are largely mediated by a species’ ecological niche and sensitivity to climate variation. Some species with narrow ecological niches have been found to exhibit increased population differentiation, limited gene flow across populations, and reduced population genetic diversity. In this study, we examine patterns of population genetic structure and diversity of four bumble bee species that are broadly sympatric, but do not necessarily inhabit the same ecological niche in the Pacific Northwest of the United States. Testing for the effect of isolation by geographic distance (IBD) with linearized F _st and D _est found that Bombus sylvicola and B. mixtus exhibited significant IBD across populations. In contrast, both B. melanopygus and B. flavifrons, two species that are distributed across a broad elevation gradient, exhibited no IBD, a result further corroborated by Bayesian a priori population assignment tests. Furthermore, we discovered that B. sylvicola populations distributed on the Olympic Peninsula have significantly less average allelic diversity than populations distributed in the Cascade Mountains. Our results suggest that populations distributed in the Olympic Mountains represent a distinct genetic cluster relative to the Cascade Mountains, with B. sylvicola and B. mixtus likely experiencing the greatest degree of population genetic differentiation relative to B. flavifrons and B. melanopygus. While bumble bees are known to co-exist across a diversity of habitats, our results demonstrate that underlying population genetic structure and diversity may not necessarily be similar across species, and are largely governed by their respective niches.     

Effects of predation risk on the nocturnal activity budgets of thin-billed prions Pachyptila belcheri on New Island, Falkland Islands

Predation is a major ecological and evolutionary driver of natural populations, greatly influencing fitness and behaviour of prey species. Small, long-lived petrels are vulnerable to predation at the breeding colonies and are expected to evolve behavioural strategies to minimize predation risks. Using an automatic nest monitoring system and nightly aerial counts, we examined the effect of vegetation cover and moonlight on colony attendance patterns and levels of burrow activity of breeding thin-billed prions, Pachyptila belcheri, on New Island, Falkland Islands. We further investigated how these parameters were related to predation by Falkland skuas. We monitored up to 32 nests in two habitats, one with Tussock grass and one with low vegetation cover. Individuals in both areas were more active at the nest before hatching, and those breeding in the low cover habitat were more active and arrived at the colony earlier, which might reflect an effect of reaction time over predation risk. Nocturnal activity peaks shifted in time as the season progressed, indicating behavioural adjustments to sunrise hours. Moon phase did not affect attendance and activity levels of breeders in either habitat or overall aerial activity, but influenced arrival time at the colony during chick-rearing, individuals arriving later in periods of full moon. Skua capture rates were positively correlated with aerial and nest activity but not with overall breeder attendance and were unaffected by moon phase. Thin-billed prions activity budgets are influenced by environmental parameters that affect their likelihood of being predated.     

The chemokine receptor, CCR5

The chemokine receptor, CCR5, is a G protein coupled receptor responsible for some of the effects of the chemokines CCL3, CCL4 and CCL5. It is also one of the co-receptors for the entry of human immunodeficiency virus-1 (HIV-1) into cells. Regulation of CCR5 number on cells is, therefore, important for determining the infection rate by HIV-1.     

CLH-3, a ClC-2 anion channel ortholog activated during meiotic maturation in C. elegans oocytes

BACKGROUND: ClC anion channels are ubiquitous and have been identified in organisms as diverse as bacteria and humans. Despite their widespread expression and likely physiological importance, the function and regulation of most ClCs are obscure. The nematode Caenorhabditis elegans offers significant experimental advantages for defining ClC biology. These advantages include a fully sequenced genome, cellular and molecular manipulability, and genetic tractability. RESULTS: We show by patch clamp electrophysiology that C. elegans oocytes express a hyperpolarization- and swelling-activated Cl(-) current with biophysical characteristics strongly resembling those of mammalian ClC-2. Double-stranded RNA-mediated gene interference (RNAi) and single-oocyte RT-PCR demonstrated that the channel is encoded by clh-3, one of six C. elegans ClC genes. CLH-3 is inactive in immature oocytes but can be triggered by cell swelling. However, CLH-3 plays no apparent role in oocyte volume homeostasis. The physiological signal for channel activation is the induction of oocyte meiotic maturation. During meiotic maturation, the contractile activity of gonadal sheath cells, which surround oocytes and are coupled to them via gap junctions, increases dramatically. These ovulatory sheath cell contractions are initiated prematurely in animals in which CLH-3 expression is disrupted by RNAi. CONCLUSIONS: The inwardly rectifying Cl(-) current in C. elegans oocytes is due to the activity of a ClC channel encoded by clh-3. Functional and structural similarities suggest that CLH-3 and mammalian ClC-2 are orthologs. CLH-3 is activated during oocyte meiotic maturation and functions in part to modulate ovulatory contractions of gap junction-coupled gonadal sheath cells.     

Public Support for Conserving Bird Species Runs Counter to Climate Change Impacts on Their Distributions

There is increasing evidence that global climate change will alter the spatiotemporal occurrences and abundances of many species at continental scales. This will have implications for efficient conservation of biodiversity. We investigate if the general public in Denmark are willing to pay for the preservation of birds potentially immigrating and establishing breeding populations due to climate change to the same extent that they are for native species populations currently breeding in Denmark, but potentially emigrating due to climate change. We find that Danish citizens are willing to pay much more for the conservation of birds currently native to Denmark, than for bird species moving into the country – even when they are informed about the potential range shifts associated with climate change. The only exception is when immigrating species populations are under pressure at European level. Furthermore, people believing climate change to be man-made and people more knowledgeable about birds tended to have higher WTP for conservation of native species, relative to other people, whereas their preferences for conserving immigrant species generally resembled those of other people. Conservation investments rely heavily on public funding and hence on public support. Our results suggest that cross-country coordination of conservation efforts under climate change will be challenging in terms of achieving an appropriate balance between cost-effectiveness in adaptation and the concerns of a general public who seem mostly worried about protecting currently-native species.     

Constraints on the transport and glycosylation of recombinant IFN-gamma in Chinese hamster ovary and insect cells.

In this study we compare intracellular transport and processing of a recombinant glycoprotein in mammalian and insect cells. Detailed analysis of the N-glycosylation of recombinant human IFN-gamma by matrix-assisted laser-desorption mass spectrometry showed that the protein secreted by Chinese hamster ovary and baculovirus-infected insect Sf9 cells was associated with complex sialylated or truncated tri-mannosyl core glycans, respectively. However, the intracellular proteins were predominantly associated with high-mannose type oligosaccharides (Man-6 to Man-9) in both cases, indicating that endoplasmic reticulum to cis-Golgi transport is a predominant rate-limiting step in both expression systems. In CHO cells, although there was a minor intracellular subpopulation of sialylated IFN-gamma glycoforms identical to the secreted product (therefore associated with late-Golgi compartments or secretory vesicles), no other intermediates were evident. Therefore, anterograde transport processes in the Golgi stack do not limit secretion. In Sf9 insect cells, there was no direct evidence of post-ER glycan-processing events other than core fucosylation and de-mannosylation, both of which were glycosylation site-specific. To investigate the influence of nucleotide-sugar availability on cell-specific glycosylation, the cellular content of nucleotide-sugar substrates in both mammalian and insect cells was quantitatively determined by anion-exchange HPLC. In both host cell types, UDP-hexose and UDP-N-acetylhexosamine were in greater abundance relative to other substrates. However, unlike CHO cells, sialyltransferase activity and CMP-NeuAc substrate were not present in uninfected or baculovirus-infected Sf9 cells. Similar data were obtained for other insect cell hosts, Sf21 and Ea4. We conclude that although the limitations on intracellular transport and secretion of recombinant proteins in mammalian and insect cells are similar, N-glycan processing in Sf insect cells is limited, and that genetic modification of N-glycan processing in these insect cell lines will be constrained by substrate availability to terminal galactosylation.