A novel approach to characterize branching network: Application to termite tunnel patterns

We defined a novel “branch length similarity” (BLS) entropy, S, on a simple network consisting of a single node and branches. This simple network is referred to as “unit branching network” (UBN) because UBNs are components of larger networks. As an application of BLS entropy, we considered the characterization of termite tunnel patterns because termite tunnel patterns can be broken down into a collection of simple units consisting of a single node and branches. These simple units correspond to UBNs. To this end, in additional to the entropy, we introduced the standard deviation (σ) of the difference in S between UBNs connected by a single tunnel branch. Forty simulated tunnel patterns were created for each of two termite species, Reticulitermes flavipes (Kollar) and Coptotermes formosanus Shiraki. These patterns were projected into <S>–σ phase space in order to assess their topological properties. This approach showed that for R. flavipes, their coordinates were relatively more clustered than those of C. formosanus. This result reflected that these two species were differently constrained by emergent property resulting from simple worker's tunneling behavior. We believe that the approach proposed in this study can be a useful tool to explore termite tunnel systems, but not limited to termite system.     

A novel meta‐analytical approach to improve systematic review of rates and patterns of microevolution

A classic topic in ecology and evolution, phenotypic microevolution of quantitative traits has received renewed attention in the face of rapid global environmental change. However, for plants, synthesis has been hampered by the limited use of standard metrics, which makes it difficult to systematize empirical information. Here we demonstrate the advantages of incorporating meta‐analysis tools to the review of microevolutionary rates. We perform a systematic survey of the plant literature on microevolution of quantitative traits over known periods of time, based on the scopus database. We quantify the amount of change by standard mean difference and develop a set of effect sizes to analyze such data. We show that applying meta‐analysis tools to a systematic literature review allows the extraction of a much larger volume of information than directly calculating microevolutionary rates. We also propose derived meta‐analysis effect sizes (h, LG and LR) which are appropriate for the study of evolutionary patterns, the first being similar to haldanes, the second and third allowing the application of a preexisting analytical framework for the inference of evolutionary mechanisms. This novel methodological development is applicable to the study of microevolution in any taxa. To pilot test it, we built an open‐access database of 1,711 microevolutionary rates of 152 angiosperm species from 128 studies documenting population changes in quantitative traits following an environmental novelty with a known elapsed time (<260 years). The performance of the metrics proposed (h, LG and LR) is similar to that of preexisting ones, and at the same time they bring the advantages of lower estimation bias and higher number of usable observations typical of meta‐analysis.     

A novel solid phase approach to Aia‐containing peptides

A strategy was developed to directly assemble 4‐amino‐1,2,4,5‐tetrahydro‐indolo[2,3‐c]‐azepin‐3‐ones on solid‐phase‐supported peptide sequences. Fmoc‐ and Boc‐based strategies were investigated. The Fmoc‐strategy approach strongly depends on the peptide sequence being synthesized while the Boc‐based synthesis leads to excellent results for all the selected peptide analogs. The method was applied to prepare Aia‐analogs of several bioactive peptides containing one or more Trp‐residues which were shown to be important for biological recognition. Copyright © 2008 European Peptide Society and John Wiley & Sons, Ltd.     

A proteomic approach to characterize protein shedding

Shedding (i.e. proteolysis of ectodomains of membrane proteins) plays an important pathophysiological role. In order to study the feasibility of identifying shed proteins, we analyzed serum-free media of human mammary epithelial cells by mass spectrometry following induction of shedding by the phorbol ester, 4 beta-phorbol 12-myristate 13-acetate (PMA). Different means of sample preparation, including biotinylation of cell surface proteins, isolation of glycosylated proteins, and preparation of crude protein fractions, were carried out to develop the optimal method of sample processing. The collected proteins were digested with trypsin and analyzed by reversed-phase capillary liquid chromatography interfaced to an ion-trap mass spectrometer. The resulting peptide spectra were interpreted using the program SEQUEST. Analyzing the sample containing the crude protein mixture without chemical modification or separation resulted in the greatest number of identifications, including putatively shed proteins. Overall, 45 membrane-associated proteins were identified including 22 that contain at least one transmembrane domain and 23 that indirectly associate with the extracellular surface of the plasma membrane. Of the 22 transmembrane proteins, 18 were identified by extracellular peptides providing strong evidence they originate from regulated proteolysis or shedding processes. We combined results from the different experiments and used a peptide count method to estimate changes in protein abundance. Using this approach, we identified two proteins, syndecan-4 and hepatoma-derived growth factor, whose abundances increased in media of cells treated with PMA. We also detected proteins whose abundances decreased after PMA treatment such as 78 kDa glucose-regulated protein and lactate dehydrogenase A. Further analysis using immunoblotting validated the abundance changes for syndecan-4 and 78 kDa glucose-regulated protein as a result of PMA treatment. These results demonstrate that tandem mass spectrometry can be used to identify shed proteins and to estimate changes in protein abundance.     

A model‐based approach to characterize individual inbreeding at both global and local genomic scales

Inbreeding results from the mating of related individuals and may be associated with reduced fitness because it brings together deleterious variants in one individual. In general, inbreeding is estimated with respect to an arbitrary base population consisting of ancestors that are assumed unrelated. We herein propose a model‐based approach to estimate and characterize individual inbreeding at both global and local genomic scales by assuming the individual genome is a mosaic of homozygous‐by‐descent (HBD) and non‐HBD segments. The HBD segments may originate from ancestors tracing back to different periods in the past defining distinct age‐related classes. The lengths of the HBD segments are exponentially distributed with class‐specific parameters reflecting that inbreeding of older origin generates on average shorter stretches of observed homozygous markers. The model is implemented in a hidden Markov model framework that uses marker allele frequencies, genetic distances, genotyping error rates and the sequences of observed genotypes. Note that genotyping errors, low‐fold sequencing or genotype‐by‐sequencing data are easily accommodated under this framework. Based on simulations under the inference model, we show that the genomewide inbreeding coefficients and the parameters of the model are accurately estimated. In addition, when several inbreeding classes are simulated, the model captures them if their ages are sufficiently different. Complementary analyses, either on data sets simulated under more realistic models or on human, dog and sheep real data, illustrate the range of applications of the approach and how it can reveal recent demographic histories among populations (e.g., very recent bottlenecks or founder effects). The method also allows to clearly identify individuals resulting from extreme consanguineous matings.     

A novel fluorescence imaging approach to monitor salt stress‐induced modulation of ouabain‐sensitive ATPase activity in sunflower seedling roots

Seedlings exposed to salt stress are expected to show modulation of intracellular accumulation of sodium ions through a variety of mechanisms. Using a new methodology, this work demonstrates ouabain (OU)‐sensitive ATPase activity in the roots of sunflower seedlings subjected to salt stress (120 mM NaCl). 9‐Anthroylouabain (a derivative of ouabain known to inhibit Na⁺,K⁺‐ATPase activity in animal systems, EC 3.6.3.9) has been used as a probe to analyze OU‐sensitive ATPase activity in sunflower (Helianthus annuus) seedling roots by spectrofluorometric estimation and localization of its spatial distribution using confocal laser scanning microscopy. Salt stress for 48 h leads to a significant induction of OU‐sensitive ATPase activity in the meristematic region of the seedling roots. Calcium ions (10 mM) significantly inhibit enzyme activity and a parallel accumulation of sodium ions in the cytosol of the columella cells, epidermis and in the cells of the meristematic region of the roots is evident. As a rapid response to NaCl stress, the activity of OU‐sensitive ATPase gets localized in the nuclear membrane of root protoplasts and it gets inhibited after treatment with calcium ions. Nuclear membrane localization of the OU‐sensitive ATPase activity highlights a possible mechanism to efflux sodium ions from the nucleus. Thus, a correlation between OU‐sensitive ATPase activity, its modulation by calcium ions and accumulation of sodium ions in various regions of the seedling roots, has been demonstrated using a novel approach in a plant system.     

A novel approach to rapidly prevent age‐related cognitive decline

The loss of cognitive function is a pervasive and often debilitating feature of the aging process for which there are no effective therapeutics. We hypothesized that a novel metal chaperone (PBT2; Prana Biotechnology, Parkville, Victoria, Australia) would enhance cognition in aged rodents. We show here that PBT2 rapidly improves the performance of aged C57Bl/6 mice in the Morris water maze, concomitant with increases in dendritic spine density, hippocampal neuron number and markers of neurogenesis. There were also increased levels of specific glutamate receptors (alpha‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid and N‐methyl‐d ‐aspartate), the glutamate transporter (VGLUT1) and glutamate itself. Markers of synaptic plasticity [calmodulin‐dependent protein kinase II (CaMKII) and phosphorylated CaMKII, CREB, synaptophysin] were also increased following PBT2 treatment. We also demonstrate that PBT2 treatment results in a subregion‐specific increase in hippocampal zinc, which is increasingly recognized as a potent neuromodulator. These data demonstrate that metal chaperones are a novel approach to the treatment of age‐related cognitive decline.     

A simple protocol to characterize bacterial cell‐envelope lipoproteins in a native‐like environment

Physiological conditions in living cells are strictly regulated to allow, optimize, and coordinate biological processes. The bacterial cell envelope is the compartment where the communication with the external environment takes place. This involves membrane proteins, key players in many biological processes that ensure bacterial survival. The biochemical characterization of membrane proteins, either integral, lipidated or peripheral is challenging due to their mixed protein‐lipid nature, making it difficult to purify and obtain considerable amounts of samples. In contrast to integral membrane proteins, lipidated proteins are usually purified as truncated soluble versions, neglecting the impact of the membrane environment. Here we report a simple and robust protocol to characterize bacterial lipidated proteins in spheroplasts from Escherichia coli using a β‐lactamase as a model. The Metallo‐β‐lactamase NDM‐1 is an enzyme anchored to the inner leaflet of the outer membrane of Gram‐negative bacteria. Kinetic parameters and stability of the lipidated NDM‐1 and the soluble unbound version (NDM‐1 C26A) were measured in spheroplasts and periplasm, respectively. These studies revealed that membrane anchoring increases the K_M of the enzyme, consequently decreasing the catalytic efficiency, while not affecting its kinetic stability. This approach can be used to characterize lipidated proteins avoiding the purification step while mimicking its native environment. This approach also helps in filling the gap between in vitro and in vivo studies.     

Analysis of bacteriolytic activity patterns, a novel approach to the taxonomy of enterococci.

The bacteriolytic activities of different group D streptococcal species on various media and substrates were studied. Our results showed that all of the enterococcal species which we tested had bacteriolytic activity on at least one of the media used, while the group D nonenterococcal species had no such activity. In addition, using culture media containing different additives and different pH values, we defined seven major groups of bacteriolytic activity (lyogroups), each of which overlapped with one species (four lyogroups), two species (two lyogroups), or four species (one lyogroup). The detection of enterococcal lyogroups proved to be as reliable for species identification as the conventional methods presently in use.     

A simulation‐based approach to understand how metacommunity characteristics influence emergent biodiversity patterns

To understand controls over biodiversity, it is necessary to take a multi‐scale approach to understand how local and regional factors affect the community assembly processes that drive emergent patterns. This need is reflected in the growing use of the metacommunity concept to interpret multi‐scale measures of biodiversity, including metrics derived from diversity partitioning (e.g. α, β and γ diversity) and variation partitioning (e.g. spatial and environmental components of compositional turnover) techniques. However, studies have shown limited success using these metrics to characterize underlying community assembly dynamics. Here we demonstrate how a metacommunity simulation package (MCSim) can be used to evaluate when and how biodiversity metrics can be used to make inferences about metacommunity characteristics. We examined a wide range of parameter settings representing ecologically relevant scenarios. We used artificial neural networks (ANNs) to assess the sensitivity of diversity and variation partitioning metrics (calculated from simulation outcomes) to metacommunity parameter settings. In the scenarios examined in this study, the niche‐neutral gradient strongly influenced most biodiversity metrics, metacommunity size exhibited a marginal influence over some metrics, and dispersal dynamics only affected a subset of variation partitioning outcomes. Variation partitioning response curves along the niche‐neutral gradient were not monotonic; however, simulation outcomes suggest other biodiversity metrics (e.g. dissimilarity saturation) can be used in combination with variation partitioning metrics to make inferences about metacommunity properties. With the growing availability of archived ecological data, we expect future work will apply simulation‐based techniques to better understand links between biodiversity and the metacommunity characteristics that are presumed to control the underlying community assembly processes.     

Urban gulls adapt foraging schedule to human‐activity patterns

Numerous animals are able to adapt to temporal patterns in natural food availability, but whether species living in relatively novel environments such as cities can adapt to anthropogenic activity cycles is less well understood. We aimed to assess the extent to which urban gulls have adapted their foraging schedule to anthropogenic food source fluctuations related to human activity by combining field observations at three distinct urban feeding grounds (park, school and waste centre) with global positioning system (GPS) tracking data of gulls visiting similar types of feeding grounds throughout the same city. We found that the birds' foraging patterns closely matched the timing of school breaks and the opening and closing times of the waste centre, but gull activity in the park appeared to correspond to the availability of natural food sources. Overall, this suggests that gulls may have the behavioural flexibility to adapt their foraging behaviour to human time schedules when beneficial and that this trait could potentially enable them to thrive in cities.     

Changing the school environment to increase physical activity in children

We examined the hypothesis that elementary school-age children will be more physically active while attending school in a novel, activity-permissive school environment compared to their traditional school environment. Twenty-four children were monitored with a single-triaxial accelerometer worn on the thigh. The students attended school in three different environments: traditional school with chairs and desks, an activity-permissive environment, and finally their traditional school with desks which encouraged standing. Data from the school children were compared with another group of age-matched children (n = 16) whose physical activity was monitored during summer vacation. When children attended school in their traditional environment, they moved an average (mean +/- s.d.) of 71 +/- 0.4 m/s(2). When the children attended school in the activity-permissive environment, they moved an average of 115 +/- 3 m/s(2). The children moved 71 +/- 0.7 m/s(2) while attending the traditional school with standing desks. Children moved significantly more while attending school in the activity-permissive environment compared to the amount that they moved in either of the traditional school environments (P < 0.0001 for both). Comparing children's activity while they were on summer vacation (113 +/- 8 m/s(2)) to school-bound children in their traditional environment showed significantly more activity for the children on summer vacation (P < 0.0001). The school children in the activity-permissive environment were as active as children on summer vacation. Children will move more in an activity-permissive environment. Strategies to increase the activity of school children may involve re-designing the school itself.     

Racial differences in subcutaneous fat patterns in children aged 7–15 years

In a stratified random sample of 278 children aged 7–15 years of an entire biracial community, skinfold measurements were taken on six standard body sites. We found that white children had generally thicker skinfolds than blacks for the same body weight, with a consistent exception: the subscapular skinfold was relatively thicker in blacks. It is suggested that this racial difference in distribution of fat may manifest a genetic adaptive trait developed under circumstances demanding both a caloric reserve and facilitation of convective heat loss in tropical climates.     

A novel model to characterize structure and function of BRCA1

BRCA1 plays a central role in DNA repair. Although N‐terminal RING and C‐terminal BRCT domains are studied well, the functions of the central region of BRCA1 are poorly characterized. Here, we report a structural and functional analysis of BRCA1 alleles and functional human BRCA1 in chicken B‐lymphocyte cell line DT40. The combination of “homologous recombineering” and “RT‐cassette” enables modifications of chicken BRCA1 gene in Escherichia coli. Mutant BRCA1 knock‐in DT40 cell lines were generated using BRCA1 mutation constructs by homologous recombination with a targeting efficiency of up to 100%. Our study demonstrated that deletion of motifs 2–9 BRCA1^{Δ/Δ181‐1415} (Caenorhabditis elegans BRCA1 mimic) or deletion of motif 1 BRCA1^{Δ/Δ126‐136} decreased cell viability following cisplatin treatment. Furthermore, deletion of motifs 5 and 6 BRCA1^{Δ/Δ525‐881} within DNA‐binding region, even the conserved 7‐amino acid deletion BRCA1^{Δ/Δ872‐878} within motif 6, caused a decreased cell viability upon cisplatin treatment. Surprisingly, human BRCA1 is functional in DT40 cells as indicated by DNA damage‐induced Rad 51 foci formation in human BRCA1 knock‐in DT40 cells. These results demonstrate that those conserved motifs within the central region are essential for DNA repair functions of BRCA1. These findings provide a valuable tool for the development of new therapeutic modalities of breast cancer linked to BRCA1.     

Sensation‐seeking genes and physical activity in youth

Many studies examining genetic influences on physical activity (PA) have evaluated the impact of single nucleotide polymorphisms (SNPs) related to the development of lifestyle‐related chronic diseases, under the hypothesis that they would be associated with PA. However, PA is a multidetermined behavior and associated with a multitude of health consequences. Thus, examining a broader range of candidate genes associated with a broader range of PA correlates may provide new insights into the genetic underpinnings of PA. In this study, we focus on one such correlate – sensation‐seeking behavior. Participants (N = 1130 Mexican origin youth) provided a saliva sample and data on PA and sensation‐seeking tendencies in 2008–2009. Participants were genotyped for 630 functional and tagging variants in the dopamine, serotonin and cannabinoid pathways. Overall 30% of participants (males – 37.6% and females – 22.0%) reported ≥60 min of PA on 5 of 7 days. After adjusting for gender, age and population stratification, and applying the Bayesian False Discovery Probability approach for assessing noteworthiness, four gene variants were significantly associated with PA. In a multivariable model, being male, having higher sensation‐seeking tendencies and at least one copy of the minor allele for SNPs in angiotensin I‐converting enzyme gene [ACE; rs8066276 odds ratio (OR) = 1.44; P = 0.012] and tryptophan hydroxylase 2 gene (TPH2; rs11615016 OR = 1.73; P = 0.021) were associated with increased likelihood of meeting PA recommendations. Participants with at least one copy of the minor allele for SNPs in synaptosomal‐associated protein 25 gene (SNAP25; rs363035 OR = 0.53; P = 0.005) and cannabinoid receptor 1 gene (CNR1; rs6454672 OR = 0.62; P = 0.022) have decreased likelihood of meeting PA recommendations. Our findings extend current knowledge of the complex relationship between PA and possible genetic underpinnings.