A quantitative assay for aggrecanase activity.

Aggrecanase activities of ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) proteinases were measured with a recombinant aggrecan fragment and two monoclonal antibodies. Recombinant human aggrecan interglobular domain was first incubated in the presence of ADAMTS enzymes. The aggrecan peptide with the N-terminal sequence ARGSVIL released upon hydrolysis was then quantified in an enzyme-linked immunosorbent assay (ELISA) with an anti-neoepitope antibody specific for the N-terminal ARGSVIL sequence and a second anti-aggrecan peptide antibody. For higher sensitivity of the assay, P1-P5 residues of the aggrecanase site within the aggrecan substrate were changed by in vitro mutagenesis. Specific activities of recombinant truncated ADAMTS1 and ADAMTS4 estimated with authentic aggrecan interglobular domain amounted to 2.4 +/- 0.4 and 21.7 +/- 9.5 nmoles hydrolyzed substrate/min.mg, respectively. The values were 10.3 +/- 5.1 and 151.5 +/- 93.5 nmoles/min.mg for hydrolysis of the modified substrate. The aggrecanase activity assay can be used for (1) kinetic characterization of aggrecanase activities of human and animal ADAMTS, (2) screening of inhibitors for aggrecan hydrolyzing ADAMTS, and (3) estimation of aggrecanase activities in biological samples.     

The effects of annual fertilization and complete competition control on fascicle morphology of different aged loblolly pine stands

The goal of this study was to determine how increased nutrient availability affects foliage morphology of loblolly pine (Pinus taeda L.) without introducing the confounding influence of light availability. Morphology of fascicles from the terminal leader (radius, length, specific needle area, density, and needles per fascicle) and terminal leader traits (length, foliated length, total leaf area, and total fascicles) were measured for different aged (ranged from 5-year-old to 12-year-old) loblolly pine stands growing on the Piedmont and lower Coastal Plain of Georgia, USA. A factorial combination of annual fertilization and complete interspecific competition control was applied as stand level treatments. Competition control and stand age generally did not affect fascicle morphology. In contrast, annual fertilization significantly increased fascicle length (5%), needles per fascicle (4%), and total leaf area of the terminal (18%), while decreasing specific needle area (4%). Fertilization also increased terminal leader growth (5%) and total number of fascicles (6%). Therefore, loblolly pine foliage morphology does have plasticity to respond to increased nutrient availability. However, the magnitude of these morphological changes is small compared to changes in total canopy leaf area.     

Repressing the neuron within

A myriad of coordinated signals control cellular differentiation. Reprogramming the cell's proteome drives global changes in cell morphology and function that define cell phenotype. A switch in alternative splicing of many pre-mRNAs encoding neuronal-specific proteins accompanies neuronal differentiation. Three groups recently showed that the global splicing repressor, polypyrimidine track-binding protein (PTB), regulates this switch.1-3 Although a subset of neuronal genes are turned on in both non-neuronal and neuronal cells, restricted expression of PTB in non-neuronal cells diverts their mRNAs to nonsense-mediated decay and prevents protein expression. When the PTB brake is released, the cell splices like a neuron.     

Effects of diets enriched in omega-3 and omega-6 polyunsaturated fatty acids on offspring sex-ratio and maternal behavior in mice

There have been many trials describing the effects of polyunsaturated fatty acids (PUFA) on fecundity, neonatal development, and maternal behavior in humans, but few controlled studies in rodents. We examined the effects of a maternal diet high in omega 3 (N-3) or omega 6 (N-6) PUFA on NIH Swiss mice. Female mice were ad libitum fed one of three complete and balanced diets (N-3, enriched in menhaden oil; N-6, enriched in corn oil; C, control diet, Purina 5015) from age 4 wk until the end of the study. Mice were bred at approximately 19 wk and 27 wk of age, providing a total of 838 pups from 129 litters in two experiments. After weaning their pups from parity 1, behavior of dams was assessed on elevated-plus and open-field mazes. Although the fraction of male pups from the N-3 and C groups was not different from 0.5, dams on the N-6 diet birthed more daughters than sons (213 vs. 133; P < 0.001). Although maternal stress has been reported to favor birth of daughters, the behavior of N-6 dams was not different from controls. By contrast, the N-3 dams displayed greater anxiety, spending less time in the open arms and more time in the closed arms of the elevated maze and traveling less distance and exhibiting less exploratory behavior in the open field (P < 0.05). N-3 dams tended to produce smaller litters than C dams, and N-3-suckled pups gained less weight (P < 0.05). In conclusion, the N-3 diet had negative effects on murine fecundity and maternal behavior, whereas the N-6 diet favored birth of daughters.     

Small-scale demographic variability of the biocolor damselfish, <Emphasis Type="Italic">Stegastes partitus</Emphasis>, in the Florida Keys USA

The demographic responses of reef fish to their environment can be complex and in many cases, quite strong. Growth, mortality, longevity, and even reproductive effort have been demonstrated to vary for the same species of reef fish over scales of 100s to 1,000s of kilometers due to physiological and ecological interactions. Though few studies have explicitly documented it, this sort of habitat-mediated demography can also exist at very local scales. Here we present the results of a 2-year study of the bicolor damselfish, Stegastes partitus, in the Florida Keys, USA. We measured density and distribution, calculated key demographic rates (growth, survival, and fecundity), and characterized the environment (resident fish assemblage, substrate type and complexity, and food availability) of populations living in two adjacent but different habitats, the continuous fore reef and patchy back reef. Fish on the fore reef had an elevated growth rate and asymptotic size, increased mortality, and higher fecundity than fish on the back reef. We identified four potential causative mechanisms for these differences: food availability; competition; intraspecific density-dependent effects; and predation risk. Our data did not support an effect of either food availability or intraspecific density-dependence, but rather suggested that demographic responses are affected by both competition and predation risk.     

Production of a fusion protein of sweet potato sporamin from recombinant E. coli XL1 Blue by fed-batch fermentations

Glucose-stat and pH-stat control strategies were employed in order to culture a recombinant E. coli XL1 Blue to produce a fusion protein of sweet potato sporamin (SPA) and glutathione S-transferase (GST) from the recombinant E. coli XL1 Blue. Cell densities up to 25 g l^–1 and 28.9 mg fusion protein (GST-SPA) g^–1 cell dry weight (CDW) was achieved from a fed-batch fermentation controlled by glucose-stat strategy. A pH-stat control fermentation using glycerol as a carbon source gave E. coli up to 27 g l^–1 and 31.5 mg GST-SPA g^–1 CDW. Additionally, a pH-stat control strategy using glucose as a carbon source gave E. coli up to 15 g l^–1 and about 22.7 mg g^–1 CDW of GST-SPA.     

Testing domains of danger in the selfish herd: sparrowhawks target widely spaced redshanks in flocks

The main tenet of Hamilton's 'selfish herd theory' for the evolution of group living is that individual risk of being killed upon attack by a predator is greater when relatively far from conspecifics. Here we examine the role of spacing using video analysis of encounters between redshanks, Tringa totanus, in flocks on saltmarsh, and sparrowhawks, Accipiter nisus, surprise hunting from adjacent woodland. Targeted redshanks were 35% (approx. 5 body lengths) more widely spaced than their nearest non-targeted neighbours, controlling for proximity to the hawk. Although targeted redshanks were also twice as slow to escape, the effect dropped out of a model containing spacing, which alone accounted for twice as much variation as escape delay. Redshanks were more tightly spaced on the riskiest side of the flock, suggesting they attempted to compensate for the greater risk, while birds on the edges of flocks were more widely spaced than those in the centre. Our analysis controls for most of the confounding effects associated with the edge-centre comparisons that are normally used in similar studies and provides strong support for spacing-dependent differential predation risk in the wild. In general, we suggest that positive selection for tight spacing when prey are stationary is largely due to domains of danger, but that this also leads to positive selection when prey are mobile because of predator confusion.     

Functional studies of an HIV-1 encoded glutathione peroxidase

In an alternate reading frame overlapping the viral envelope gene, HIV-1 has been shown to encoded a truncated glutathione peroxidase (GPx) module. Essential active site residues of the catalytic core regions of mammalian GPx sequences are conserved in the putative viral GPx (vGPx, encoded by the env-fs gene). Cells transfected with an HIV-1 env-fs construct show up to a 100% increase in GPx enzyme activity, and are protected against the loss of mitochondrial transmembrane potential and subsequent cell death induced by exogenous oxidants or mitochondrial reactive oxygen species. An intact vGPx gene was observed to be more common in HIV-1-infected long-term non-progressors, as compared to HIV-1 isolates from patients developing AIDS. An antioxidant/antiapoptotic protective role of the vGPx is also consistent with the observation that -1 frameshifting induced by the HIV-1 env-fs sequence AAAAAGA (which contains a potential "hungry" arginine codon, AGA) increases during arginine deficiency, which has been associated with increased oxidative stress. Under arginine-limited conditions, nitric oxide synthase generates superoxide, which rapidly combines with NO to form peroxynitrite, which can cause activated T-cells to undergo apoptosis. Thus, biosynthesis of the HIV-1 GPx as an adaptive response to low arginine conditions might delay oxidant-induced apoptotic cell death, providing an enhanced opportunity for viral replication.     

The Pleistocene archaeology and environments of the Wasiriya Beds, Rusinga Island, Kenya

Western Kenya is well known for abundant early Miocene hominoid fossils. However, the Wasiriya Beds of Rusinga Island, Kenya, preserve a Pleistocene sedimentary archive with radiocarbon age estimates of >33–45 ka that contains Middle Stone Age artifacts and abundant, well-preserved fossil fauna: a co-occurrence rare in eastern Africa, particularly in the region bounding Lake Victoria. Artifacts and fossils are associated with distal volcanic ash deposits that occur at multiple localities in the Wasiriya Beds, correlated on the basis of geochemical composition as determined by electron probe microanalysis. Sediment lithology and the fossil ungulates suggest a local fluvial system and associated riparian wooded habitat within a predominantly arid grassland setting that differs substantially from the modern environment, where local climate is strongly affected by moisture availability from Lake Victoria. In particular, the presence of oryx (Oryx gazella) and Grevy’s zebra (Equus grevyi) suggest a pre-Last Glacial Maximum expansion of arid grasslands, an environmental reconstruction further supported by the presence of several extinct specialized grazers (Pelorovis antiquus, Megalotragus sp., and a small alcelaphine) that are unknown from Holocene deposits in eastern Africa. The combination of artifacts, a rich fossil fauna, and volcaniclastic sediments makes the Wasiriya Beds a key site for examining the Lake Victoria basin, a biogeographically important area for understanding the diversification and dispersal of Homo sapiens from Africa, whose pre-Last Glacial Maximum history remains poorly understood.