The population genomics of multiple tsetse fly (Glossina fuscipes fuscipes) admixture zones in Uganda

Understanding the mechanisms that enforce, maintain or reverse the process of speciation is an important challenge in evolutionary biology. This study investigates the patterns of divergence and discusses the processes that form and maintain divergent lineages of the tsetse fly Glossina fuscipes fuscipes in Uganda. We sampled 251 flies from 18 sites spanning known genetic lineages and the four admixture zones between them. We apply population genomics, hybrid zone and approximate Bayesian computation to the analysis of three types of genetic markers: 55,267 double‐digest restriction site‐associated DNA (ddRAD) SNPs to assess genome‐wide admixture, 16 microsatellites to provide continuity with published data and accurate biogeographic modelling, and a 491‐bp fragment of mitochondrial cytochrome oxidase I and II to infer maternal inheritance patterns. Admixture zones correspond with regions impacted by the reorganization of Uganda's river networks that occurred during the formation of the West African Rift system over the last several hundred thousand years. Because tsetse fly population distributions are defined by rivers, admixture zones likely represent both old and new regions of secondary contact. Our results indicate that older hybrid zones contain mostly parental types, while younger zones contain variable hybrid types resulting from multiple generations of interbreeding. These findings suggest that reproductive barriers are nearly complete in the older admixture zones, while nearly absent in the younger admixture zones. Findings are consistent with predictions of hybrid zone theory: Populations in zones of secondary contact transition rapidly from early to late stages of speciation or collapse all together.     

Structural and functional fingerprint of the mitochondrial ATP-binding cassette transporter Mdl1 from Saccharomyces cerevisiae

The ATP-binding cassette half-transporter Mdl1 from Saccharomyces cerevisiae has been proposed to be involved in the quality control of misassembled respiratory chain complexes by exporting degradation products generated by the m-AAA proteases from the matrix. Direct functional or structural data of the transport complex are, however, not known so far. After screening expression in various hosts, Mdl1 was overexpressed 100-fold to 1% of total mitochondrial membrane protein in S. cerevisiae. Based on detergent screens, Mdl1 was solubilized and purified to homogeneity. Mdl1 showed a high binding affinity for MgATP (Kd = 0.26 microm) and an ATPase activity with a Km of 0.86 mm (Hill coefficient of 0.98) and a turnover rate of 2.6 ATP/s. Mutagenesis of the conserved glutamate downstream of the Walker B motif (E599Q) or the conserved histidine of the H-loop (H631A) abolished ATP hydrolysis, whereas ATP binding was not affected. Mdl1 reconstituted into liposomes showed an ATPase activity similar to the solubilized complex. By single particle electron microscopy, a first three-dimensional structure of the mitochondrial ATP-binding cassette transporter was derived at 2.3-nm resolution, revealing a homodimeric complex in an open conformation.     

Species‐specific ontogenetic diet shifts attenuate trophic cascades and lengthen food chains in exploited ecosystems

Ontogenetic diet shifts are pervasive in food websbut rules governing their emergence and the implications for trophic cascades are only partly understood. Recent theoretical advances in multispecies size spectrum models (MSSMs) predict that the emergence of ontogenetic diet shifts are driven primarily by size‐selective predation and changes in the relative abundances of suitably sized prey. Howeverthese assumptions have not yet been tested with data. Herewe developed alternative MSSMs based on different assumptions about the nature of species and size‐based preferences and tested them using an extensive dietary database for the Eastern Bering Sea (EBS). MSSMs with both size and species‐specific prey preferences correctly predicted approximately three‐fold more of the diet links than those that assumed fixed species preferences. Importantlythese model assumptions also had a profound effect on the strength of fishing‐induced trophic cascades and the emergent trophic structure of the community with and without fishing. The diet‐informed models exhibited lower predation mortality ratesparticularly for small individuals (less than 1 g) whichin turnreduced the intensity and reach of fishing‐induced trophic cascades up the size spectrum. If the level and size dependency of piscivory observed in EBS predators is typical of other systemsthe potential for fishing‐induced trophic cascades may be over‐stated in MSSMs as they are currently formulated and parameterized. Representation of species‐specific ontogenetic shifts in diet can strongly influence system responses to perturbationsand the extensions we propose should accelerate adoption of MSSMs as frameworks for exploring size‐based food web theory and developing modeling tools to support strategic management decisions.     

Retinoic acid–induced survival effects in SH-SY5Y neuroblastoma cells

Neuroblastoma is a malignant childhood cancer arising from the embryonic sympathoadrenal lineage of the neural crest. Retinoic acid (RA) is included in the multimodal therapy of patients with high-risk neuroblastoma to eliminate minimal residual disease. However, the formation of RA-resistant cells substantially lowers 5-year overall survival rates. To examine mechanisms that lead to treatment failure, we chose human SH-SY5Y cells, which are known to tolerate incubation with RA by activating the survival kinases Akt and extracellular signal-regulated kinase 1/2. Characterization of downstream pathways showed that both kinases increased the phosphorylation of the ubiquitin ligase mouse double minute homolog 2 (Mdm2) and thereby enhanced p53 degradation. When p53 signaling was sustained by blocking complex formation with Mdm2 or enhancing c-Jun N-terminal kinase (JNK) activation, cell viability was significantly reduced. In addition, Akt-mediated phosphorylation of the cell-cycle regulator p21 stimulated complex formation with caspase-3, which also contributed to cell protection. Thus, treatment with RA augmented survival signaling and attenuated basal apoptotic pathways in SH-SY5Y cells, which increased cell viability.     

Effect of selected recovery conditions on performance of repeated bouts of intermittent cycling separated by 24 hours

The purpose of this study was to examine the effects of active recovery (AR), massage (MR), and cold water immersion (CR) on performance of repeated bouts of high-intensity cycling separated by 24 hours. For each recovery condition, subjects were asked to take part in 2 intermittent cycling sessions; 18 minutes of varying work intervals performed in succession at a resistance of 80 g/kg body weight separated by 24 hours. One of four 15-minute recovery conditions immediately followed the first session and included: (a) AR, cycling at 30% Vo(2)max; (b) CR, immersion of legs in a 15 degrees C water bath; (c) MR, massage of the legs; and (d) control, seated rest. Only the control condition showed a significant decline in the total work completed between the first and second exercise sessions (108.1 +/- 5.4 kJ vs. 106.0 +/- 5.0 kJ, p < 0.05). Thus, AR, MR, and CR appeared to facilitate the recovery process between 2 high-intensity, intermittent exercise sessions separated by 24 hours.     

CD28 costimulation mediates down-regulation of p27kip1 and cell cycle progression by activation of the PI3K/PKB signaling pathway in primary human T cells

CD28 provides a costimulatory signal that cooperates with the TCR/CD3 complex to induce T cell activation, cytokine production, and clonal expansion. We have recently shown that CD28 directly regulates progression of T lymphocytes through the cell cycle. Although a number of signaling pathways have been linked to the TCR/CD3 and to CD28, it is not known how these two receptors cooperate to induce cell cycle progression. Here, using cell-permeable pharmacologic inhibitors of phosphatidylinositol 3-hydroxykinase (PI3K) and mitogen-activated protein kinase kinase (MEK1/2), we show that cell cycle progression of primary T lymphocytes requires simultaneous activation of PI3K- and MEK1/2-dependent pathways. Decreased abundance of cyclin-dependent kinase inhibitor p27(kip1), which requires simultaneous TCR/CD3 and CD28 ligation, was dependent upon both MEK and PI3K activity. Ligation of TCR/CD3, but not CD28 alone, resulted in activation of MEK targets extracellular signal-related kinase 1/2, whereas ligation of CD28 alone was sufficient for activation of PI3K target protein kinase B (PKB; c-Akt). CD28 ligation alone was also sufficient to mediate inactivating phosphorylation of PKB target glycogen synthase kinase-3 (GSK-3). Moreover, direct inactivation of GSK-3 by LiCl in the presence of anti-CD3, but not in the presence of anti-CD28, resulted in down-regulation of p27(kip1), hyperphosphorylation of retinoblastoma tumor suppressor gene product, and cellular proliferation. Thus, inactivation of the PI3K-PKB target GSK-3 could substitute for CD28 but not for CD3 signals. These results show that the PI3K-PKB pathway links CD28 to cell cycle progression and suggest that p27(kip1) integrates mitogenic MEK- and PI3K-dependent signals from TCR and CD28 in primary T lymphocytes.     

The involvement of the left motor cortex in learning of a novel action word lexicon

Current theoretical positions assume that action-related word meanings are established by functional connections between perisylvian language areas and the motor cortex (MC) according to Hebb's associative learning principle. To test this assumption, we probed the functional relevance of the left MC for learning of a novel action word vocabulary by disturbing neural plasticity in the MC with transcranial direct current stimulation (tDCS). In combination with tDCS, subjects learned a novel vocabulary of 76 concrete, body-related actions by means of an associative learning paradigm. Compared with a control condition with "sham" stimulation, cathodal tDCS reduced success rates in vocabulary acquisition, as shown by tests of novel action word translation into the native language. The analysis of learning behavior revealed a specific effect of cathodal tDCS on the ability to associatively couple actions with novel words. In contrast, we did not find these effects in control experiments, when tDCS was applied to the prefrontal cortex or when subjects learned object-related words. The present study lends direct evidence to the proposition that the left MC is causally involved in the acquisition of novel action-related words.     

Post-Stroke Inhibition of Induced NADPH Oxidase Type 4 Prevents Oxidative Stress and Neurodegeneration

Ischemic stroke is the second leading cause of death worldwide. Only one moderately effective therapy exists, albeit with contraindications that exclude 90% of the patients. This medical need contrasts with a high failure rate of more than 1,000 pre-clinical drug candidates for stroke therapies. Thus, there is a need for translatable mechanisms of neuroprotection and more rigid thresholds of relevance in pre-clinical stroke models. One such candidate mechanism is oxidative stress. However, antioxidant approaches have failed in clinical trials, and the significant sources of oxidative stress in stroke are unknown. We here identify NADPH oxidase type 4 (NOX4) as a major source of oxidative stress and an effective therapeutic target in acute stroke. Upon ischemia, NOX4 was induced in human and mouse brain. Mice deficient in NOX4 (Nox4 ^−/−) of either sex, but not those deficient for NOX1 or NOX2, were largely protected from oxidative stress, blood-brain-barrier leakage, and neuronal apoptosis, after both transient and permanent cerebral ischemia. This effect was independent of age, as elderly mice were equally protected. Restoration of oxidative stress reversed the stroke-protective phenotype in Nox4 ^−/− mice. Application of the only validated low-molecular-weight pharmacological NADPH oxidase inhibitor, VAS2870, several hours after ischemia was as protective as deleting NOX4. The extent of neuroprotection was exceptional, resulting in significantly improved long-term neurological functions and reduced mortality. NOX4 therefore represents a major source of oxidative stress and novel class of drug target for stroke therapy.     

An experimental and theoretical study of the morphine binding capacity and kinetics of an engineered opioid receptor

Electrochemical real-time monitoring of ligand binding to an engineered opioid receptor specific for morphine is reported. In the particular systems studied, 90% of the binding was found to be completed after only 85-120 s. Thus, the binding kinetics has proven to be more rapid than previously believed. The observed association rate constant for the morphine binding reaction was calculated to be 215 M(-1)s(-1). A theoretical analysis of the experimental binding data suggested that the binding sites of the engineered opioid receptor could best be described by a model having two populations of binding sites: K(D)=40 microM (13 micromol/g) and K(D)=205 microM (29 micromol/g). Furthermore, a theoretical model was developed in order to explain the observed binding of the engineered opioid receptor. This model suggested that the binding sites on the polymer surface are up to 5.1A deep and they allow 100% of the ligand (morphine) to anchor itself into the site. The predicted theoretical maximum binding capacity for the reported receptor is calculated to be approximately 2 mmol/g polymer (based on an increase of cavity density).