Dual roles of Incenp crucial to the assembly of the acentrosomal metaphase spindle in female meiosis

Spindle formation in female meiosis differs from mitosis in many animals, as it takes place independently of centrosomes, and the molecular requirements of this pathway remain to be understood. Here, we report two crucial roles of Incenp, an essential subunit of the chromosomal passenger complex (the Aurora B complex), in centrosome-independent spindle formation in Drosophila female meiosis. First, the initial assembly of spindle microtubules is drastically delayed in an incenp mutant. This clearly demonstrates, for the first time, a crucial role for Incenp in chromosome-driven spindle microtubule assembly in living oocytes. Additionally, Incenp is necessary to stabilise the equatorial region of the metaphase I spindle, in contrast to mitosis, where the equivalent function becomes prominent after anaphase onset. Our analysis suggests that Subito, a kinesin-6 protein, cooperates with Incenp for this latter function, but not in microtubule assembly. We propose that the two functions of Incenp are part of the mechanisms that compensate for the lack of centrosomes during meiotic spindle formation.     

The biogeography of avian extinctions on oceanic islands

Aim To test the influences of island area, island isolation, and human‐introduced mammalian predators on avian extinctions that have occurred on oceanic islands worldwide. Location The oceanic islands of the world. Methods We augmented and re‐examined an existing data set for 218 oceanic islands by means of causal modelling using path analysis (a form of structural equation modelling) and a null model. Results The number of extinctions was not a simple function of the number of bird species on the various islands. Whereas introduced mammalian predators had an influence on the number of extinctions, island area (via indirect influences) and isolation (via direct influences) were equally or more important. Main conclusions The multiple influences of physical and biotic factors on past extinctions can be revealed through modelling the causal influences of physical attributes of islands on biological characteristics, and the causal influences of both physical and biological characteristics on extinctions.     

Weight regain after sustained weight reduction is accompanied by suppressed oxidation of dietary fat and adipocyte hyperplasia

A dual-tracer approach (dietary 14C-palmitate and intraperitoneal 3H-H2O) was used to assess the trafficking of dietary fat and net retention of carbon in triglyceride depots during the first 24 h of weight regain. Obesity-prone male Wistar rats were allowed to mature under obesogenic conditions for 16 wk. One group was switched to ad libitum feeding of a low-fat diet for 10 wk (Obese group). The remaining rats were switched to an energy-restricted, low-fat diet for 10 wk that reduced body weight by 14% and were then assessed in energy balance (Reduced group), with free access to the low-fat diet (Relapse-Day1 group), or with a provision that induced a minor imbalance (+10 kcal) equivalent to that observed in obese rats (Gap-Matched group). Fat oxidation remained at a high, steady rate throughout the day in Obese rats, but was suppressed in Reduced, Gap-Matched, and Relapse-Day1 rats though 9, 18, and 24 h, respectively. The same caloric excess in Obese and Gap-Matched rats led to less fat oxidation over the day and greater trafficking of dietary fat to visceral depots in the latter. In addition to trafficking nutrients to storage, Relapse-Day1 rats had more small, presumably new, adipocytes at the end of 24 h. Dietary fat oxidation at 24 h was related to the phosphorylation of skeletal muscle acetyl-CoA carboxylase and fatty acid availability. These observations provide evidence of adaptations in the oxidation and trafficking of dietary fat that extend beyond the energy imbalance, which facilitate rapid, efficient regain during the relapse to obesity.     

Adenosine A2A receptor activation reduces infarct size in the isolated, perfused mouse heart by inhibiting resident cardiac mast cell degranulation

Mast cells are found in the heart and contribute to reperfusion injury following myocardial ischemia. Since the activation of A2A adenosine receptors (A2AARs) inhibits reperfusion injury, we hypothesized that ATL146e (a selective A2AAR agonist) might protect hearts in part by reducing cardiac mast cell degranulation. Hearts were isolated from five groups of congenic mice: A2AAR+/+ mice, A2AAR(-/-) mice, mast cell-deficient (Kit(W-sh/W-sh)) mice, and chimeric mice prepared by transplanting bone marrow from A2AAR(-/-) or A2AAR+/+ mice to radiation-ablated A2AAR+/+ mice. Six weeks after bone marrow transplantation, cardiac mast cells were repopulated with >90% donor cells. In isolated, perfused hearts subjected to ischemia-reperfusion injury, ATL146e or CGS-21680 (100 nmol/l) decreased infarct size (IS; percent area at risk) from 38 +/- 2% to 24 +/- 2% and 22 +/- 2% in ATL146e- and CGS-21680-treated hearts, respectively (P < 0.05) and significantly reduced mast cell degranulation, measured as tryptase release into reperfusion buffer. These changes were absent in A2AAR(-/-) hearts and in hearts from chimeric mice with A2AAR(-/-) bone marrow. Vehicle-treated Kit(W-sh/W-sh) mice had lower IS (11 +/- 3%) than WT mice, and ATL146e had no significant protective effect (16 +/- 3%). These data suggest that in ex vivo, buffer-perfused hearts, mast cell degranulation contributes to ischemia-reperfusion injury. In addition, our data suggest that A2AAR activation is cardioprotective in the isolated heart, at least in part by attenuating resident mast cell degranulation.     

Assessment of ecological risks in weed biocontrol: Input from retrospective ecological analyses

Prediction of the outcomes of natural enemy introductions remains the most fundamental challenge in biological control. Quantitative retrospective analyses of ongoing biocontrol projects provide a systematic strategy to evaluate and further develop ecological risk assessment. In this review, we highlight a crucial assumption underlying a continued reliance on the host specificity paradigm as a quantitative prediction of ecological risk, summarize the status of our retrospective analyses of nontarget effects of two weevils used against exotic thistles in North America, and discuss our prospective assessment of risk to a federally listed, threatened species (Cirsium pitcheri) based on those studies. Our analyses quantify the fact that host range and preference from host specificity tests are not sufficient to predict ecological impact if the introduced natural enemy is not strictly monophagous. The implicit assumption when such use is made of the host specificity data in risk assessment is that population impacts are proportional to relative preference and performance, the key components of host specificity. However, in concert with shifting awareness in the field, our studies demonstrate that the environment influences and can alter host use and population growth, leading to higher than expected direct impacts on the less preferred native host species at several spatial scales. Further, we have found that straightforward, easily anticipated indirect effects, on intraguild foragers as well as on the less preferred native host plant species, can be both widespread and significant. We conclude that intensive retrospective ecological studies provide some guidance for the quantitative prospective studies needed to assess candidate biological control agent dynamics and impacts and, so, contribute to improved rigor in the evaluation of total ecological risk to native species.     

Development of multiplexed protein profiling and detection using near infrared detection of reverse-phase protein microarrays

Protein microarrays have been recently employed for signal pathway profiling and high-throughput protein expression analysis. Reversephase arrays, where the array consists of immobilized analytes and lysates has especially shown promise in low abundance analyte detection and signal pathway profiling using phospho-specific antibodies. A limitation to current reverse phase array methodology is the inability to multiplex proteomic-based endpoints as each array can only report one analyte endpoint. In this study, we report on the use of a dual dye based approach that can effectively double the number of endpoints observed per array allowing, for example, both phosphospecific and total protein levels to be measured and analyzed at once. The method utilizes antibody bound dyes that emit in the infrared spectral region as a means of sensitive and specific detection.     

Association Between Body Fat Response to Exercise Training and Multilocus ADR Genotypes

Objectives : To examine the contribution of adrenergic receptor (ADR) gene polymorphisms and their gene‐gene interactions to the variability of exercise training‐induced body fat response. Research Methods and Procedures : This was an intervention study that used a volunteer sample of 70 healthy, sedentary men (n = 29) and postmenopausal women (n = 41) 50 to 75 years of age, with a BMI ≤37 kg/m², from the Washington, DC, metropolitan area. Participants completed 6 weeks of dietary stabilization (American Heart Association diet) before 24 weeks of supervised aerobic exercise training. Diet was maintained throughout the intervention. Change in percent total body fat, percent trunk fat, and fat mass by DXA in ADR genotype groups (Glu¹²/Glu⁹ α2b‐ADR, Trp64Arg β3‐ADR, and Gln27Glu β2‐ADR) at baseline and after 24 weeks of aerobic exercise training was measured. Results : In multivariate analysis (covariates: age, gender, and baseline value of phenotype), best fit models for percent total body and trunk fat response to exercise training retained main effects of all three ADR gene loci and the effects of each gene‐gene interaction (p = 0.009 and 0.003, respectively). Similarly, there was a trend for the fat mass response model (p = 0.03). The combined genetic factors explained 17.5% of the overall model variability for percent total body fat, 22% for percent trunk fat, and 10% for fat mass. Discussion : The body fat response to exercise training in older adults is associated with the combined effects of the Glu¹²/Glu⁹ α2b‐, Trp64Arg β3‐, and Gln27Glu β2‐ADR gene variants and their gene‐gene interactions.     

Foliar physiology of yellow-poplar ( Liriodendron tulipifera L.) exposed to O3 and elevated CO2 over five seasons.

The chronic effects of ozone (O₃) alone or combined with elevated carbon dioxide (CO₂) on the foliar physiology of unfertilized field-grown yellow-poplar ( Liriodendron tulipifera L.) seedlings were studied from 1992 to 1996. Within open-top chambers, juvenile trees were exposed to the following: charcoal-filtered air (CF); 1× ambient ozone (1XO₃); 1.5× ambient ozone (1.5XO₃); 1.5× ambient ozone plus 700 ppm carbon dioxide (1.5XO₃+CO₂); or chamberless open-air (OA). Seasonal 24-h mean ambient O₃ concentrations ranged from 32 to 46 ppm over the five seasons. Averaged over 5 years, midseason net photosynthesis at saturating light ( A _sat) was reduced by 14% ( P =0.029) and stomatal conductance ( g _s) was reduced, albeit non-significant, by 13% ( P =0.096) in upper canopy foliage exposed to 1.5XO₃-air relative to CF controls. There were no significant differences over the 5 years in A _sat and g _s between trees grown in 1XO₃- and 1.5XO₃-air. Our results support the hypothesis that the magnitude of O₃ effects on A _sat and g _s decreases as saplings age. When averaged over the five seasons of exposure, total chlorophyll concentration ( chl) was not significantly affected by exposure to elevated O₃; however, in 1.5XO₃+CO₂-air, foliar chl was reduced by 33% relative to all others ( P <0.001). In 1.5XO₃+CO₂-air, A _sat was 1.4–1.9 times higher ( P <0.001) and g _s was 0.7 times lower ( P =0.022) than all others. O₃ uptake in juvenile trees exposed to elevated O₃ plus elevated CO₂ (1.5XO₃+CO₂-air) was most comparable to trees exposed to ambient air (1XO₃) throughout the study. These findings suggest that elevated CO₂ may minimize the negative effects of O₃ by reducing O₃ uptake through decreased stomatal conductance.     

Near-infrared fluorescence detection permits accurate imaging of loading controls for Western blot analysis

Housekeeping proteins are typically chosen as internal loading controls for Western blot analysis because of their high, relatively constant expression. It was previously reported that antibodies against beta-actin did not reliably identify differences in sample loading, and extended antibody incubations caused a failure to discriminate differences in target protein levels. Here, beta-actin and GAPDH were evaluated as loading controls using near-infrared fluorescence. A load-dependent response in signal intensity was observed over a 250-fold range of sample concentrations, with R(2) values as high as 0.9939. Longer antibody incubations continued to detect differences in protein level and load-dependent responses became more linear.     

Identifying and characterizing the biological targets of metallotherapeutics: Two approaches using Au(I)-protein interactions as model systems

A significant challenge in the field of medicinal inorganic chemistry is the identification of biological targets of metal-based drugs and the characterization of the metal–biomolecule adducts. A classic example is Au(I), which has long been used to treat rheumatoid arthritis despite a poor understanding of its biological targets due to the lability, reactivity, and “spectroscopic silence” that are characteristic of Au(I). Here, we report two qualitative methods for characterizing Au(I)–protein adducts: a thiol-reactive probe that facilitates the identification of biological cysteine–Au(I) adducts and a photoreactive Au(I) complex that produces a covalent bond between the Au(I) complex and the biomolecule.