Alkaline Sucrose Gradient Sedimentation of Chromosomal Deoxyribonucleic Acid from Escherichia coli PolA+ and PolA− Strains During Thymine Starvation

Single-strand breaks were not detected in the deoxyribonucleic acid of Escherichia coli after thymine starvation for up to 180 min, even in a sensitive PolA(-) strain.     

Stoichiometry in an ecological context: testing for links between <Emphasis Type="Italic">Daphnia</Emphasis> P-content,growth rate and habitat preference

We used laboratory experiments with ten Daphnia taxa to test for links between Daphnia P-content, growth rate and habitat preference. The taxa represent a wide range of body sizes and most show distinct preferences for one of three habitats: shallow lakes, deep, stratified lakes or fishless ponds. Previous studies show that taxa from shallow lakes and fishless ponds experience high predation risk and rich food resources, whereas taxa from deep lakes experience low predation risk, strong food limitation and potentially P-deficient resources. Thus, we predicted higher P-content and higher maximal growth rates in taxa from ponds and shallow lakes and lower P-content, lower maximal growth but reduced sensitivity to P-limitation in taxa preferring stratified lakes. In each of 25 experiments, a clonal Daphnia cohort was cultured for 4 days on a P-sufficient (molar C:P ratio 70) or a P-deficient (C:P 1,000) diet of a green alga at a high concentration (1 mg C l^–1). The P-content of adult Daphnia fed the P-sufficient diet ranged from 1.52 to 1.22% mass. Small-bodied taxa from shallow lakes had higher P-content than larger-bodied taxa from deep lakes or fishless ponds. However, we found a nonsignificant negative correlation between P-content and growth on the P-sufficient diet, rather than the positive relationship predicted by the growth rate hypothesis. The P-deficient diet resulted in declines in both growth rate and P-content compared with the P-sufficient controls and the extent of the declines differed between taxa. Taxa from ponds showed a marginally greater decline in growth with the P-deficient diet compared with taxa from shallow or deep lakes. However, contrary to stoichiometric theory, no relationship was found between a species P-content and growth depression on the P-deficient diet. Although we found evidence for habitat adaptations, our results show that factors other than Daphnia P-content are important in determining differences between Daphnia species in both maximal growth rate and sensitivity to P-limited growth.     

Sexual Orientation Disparities in Weight Status in Adolescence: Findings From a Prospective Study

A growing number of studies among adult women have documented disparities in overweight adversely affecting lesbian and bisexual women, but few studies have examined sexual orientation–related patterns in weight status among men or adolescents. We examined sexual orientation group trends in BMI (kg/m²), BMI Z‐scores, and overweight using 56,990 observations from 13,785 adolescent females and males in the Growing Up Today Study (GUTS), a large prospective cohort of US youth. Participants provided self‐reported information from six waves of questionnaire data collection from 1998 to 2005. Gender‐stratified linear regression models were used to estimate BMI and BMI Z‐scores and modified Poisson regression models to estimate risk ratios for overweight, controlling for age and race/ethnicity, with heterosexuals as the referent group. Among females, we observed fairly consistently elevated BMI in all sexual orientation minority groups relative to heterosexual peers. In contrast, among males we documented a sexual‐orientation‐by‐age interaction indicating steeper increases in BMI with age from early‐to‐late adolescence in heterosexuals relative to sexual orientation minorities. Additional prospective research is needed to understand the determinants of observed sexual orientation disparities and to inform appropriate preventive and treatment interventions. The long‐term health consequences of overweight are well‐documented and over time are likely to exact a high toll on populations with elevated rates.     

Climate–ecosystem modelling made easy: The Land Sites Platform

Dynamic Global Vegetation Models (DGVMs) provide a state-of-the-art process-based approach to study the complex interplay between vegetation and its physical environment. For example, they help to predict how terrestrial plants interact with climate, soils, disturbance and competition for resources. We argue that there is untapped potential for the use of DGVMs in ecological and ecophysiological research. One fundamental barrier to realize this potential is that many researchers with relevant expertize (ecology, plant physiology, soil science, etc.) lack access to the technical resources or awareness of the research potential of DGVMs. Here we present the Land Sites Platform (LSP): new software that facilitates single-site simulations with the Functionally Assembled Terrestrial Ecosystem Simulator, an advanced DGVM coupled with the Community Land Model. The LSP includes a Graphical User Interface and an Application Programming Interface, which improve the user experience and lower the technical thresholds for installing these model architectures and setting up model experiments. The software is distributed via version-controlled containers; researchers and students can run simulations directly on their personal computers or servers, with relatively low hardware requirements, and on different operating systems. Version 1.0 of the LSP supports site-level simulations. We provide input data for 20 established geo-ecological observation sites in Norway and workflows to add generic sites from public global datasets. The LSP makes standard model experiments with default data easily achievable (e.g., for educational or introductory purposes) while retaining flexibility for more advanced scientific uses. We further provide tools to visualize the model input and output, including simple examples to relate predictions to local observations. The LSP improves access to land surface and DGVM modelling as a building block of community cyberinfrastructure that may inspire new avenues for mechanistic ecosystem research across disciplines.     

Metaphase arrest by cyclin E-Cdk2 requires the spindle-checkpoint kinase Mps1

Cytostatic factor (CSF) arrests vertebrate eggs in metaphase of meiosis II through several pathways that inhibit activation of the anaphase-promoting complex/cyclosome (APC/C). In Xenopus, the Mos-MEK1-MAPK-p90(Rsk) cascade utilizes spindle-assembly-checkpoint components to effect metaphase arrest. Another pathway involves cyclin E-Cdk2, and sustained cyclin E-Cdk2 activity in egg extracts causes metaphase arrest in the absence of Mos; this latter finding suggests that an independent pathway contributes to CSF arrest. Here, we demonstrate that metaphase arrest with cyclin E-Cdk2, but not with Mos, requires the spindle-checkpoint kinase monopolar spindles 1 (Mps1), a cyclin E-Cdk2 target that is also implicated in centrosome duplication. xMps1 is synthesized and activated during oocyte maturation and inactivated upon CSF release. In egg extracts, CSF release by calcium was inhibited by constitutively active cyclin E-Cdk2 and delayed by wild-type xMps1. Ablation of cyclin E by antisense oligonucleotides blocked accumulation of xMps1, suggesting that cyclin E-Cdk2 controls Mps1 levels. During meiosis II, activated cyclin E-Cdk2 significantly inhibited the APC/C even in the absence of the Mos-MAPK pathway, but this inhibition was not sufficient to suppress S phase between meiosis I and II. These results uniquely place xMps1 downstream of cyclin E-Cdk2 in mediating a pathway of APC/C inhibition and metaphase arrest.     

Correlated Inter-Domain Motions in Adenylate Kinase

Correlated inter-domain motions in proteins can mediate fundamental biochemical processes such as signal transduction and allostery. Here we characterize at structural level the inter-domain coupling in a multidomain enzyme, Adenylate Kinase (AK), using computational methods that exploit the shape information encoded in residual dipolar couplings (RDCs) measured under steric alignment by nuclear magnetic resonance (NMR). We find experimental evidence for a multi-state equilibrium distribution along the opening/closing pathway of Adenylate Kinase, previously proposed from computational work, in which inter-domain interactions disfavour states where only the AMP binding domain is closed. In summary, we provide a robust experimental technique for study of allosteric regulation in AK and other enzymes.     

Long-Term Weight-Loss in Gastric Bypass Patients Carrying Melanocortin 4 Receptor Variants

Background

The melanocortin 4 receptor (MC4R) critically regulates feeding and satiety. Rare variants in MC4R are predominantly found in obese individuals. Though some rare variants in MC4R discovered in patients have defects in localization, ligand binding and signaling to cAMP, many have no recognized defects.

Subjects/Methods

In our cohort of 1433 obese subjects that underwent Roux-en-Y Gastric Bypass (RYGB) surgery, we found fifteen variants of MC4R. We matched rare variant carriers to patients with the MC4R reference alleles for gender, age, starting BMI and T2D to determine the variant effect on weight-loss post-RYGB. In vitro, we determined expression of mutant receptors by ELISA and western blot, and cAMP production by microscopy.

Results

While carrying a rare MC4R allele is associated with obesity, carriers of rare variants exhibited comparable weight-loss after RYGB to non-carriers. However, subjects carrying three of these variants, V95I, I137T or L250Q, lost less weight after surgery. In vitro, the R305Q mutation caused a defect in cell surface expression while only the I137T and C326R mutations showed impaired cAMP signaling. Despite these apparent differences, there was no correlation between in vitro signaling and pre- or post-surgery clinical phenotype.

Conclusions

These data suggest that subtle differences in receptor signaling conferred by rare MC4R variants combined with additional factors predispose carriers to obesity. In the absence of complete MC4R deficiency, these differences can be overcome by the powerful weight-reducing effects of bariatric surgery. In a complex disorder such as obesity, genetic variants that cause subtle defects that have cumulative effects can be overcome after appropriate clinical intervention.