Parasitoid pressure and the radiation of a gallforming group (Cecidomyiidae: Asphondylia spp.) on creosote bush (Larrea tridentata)

Summary We tested the Enemy Impact Hypothesis, which predicts that communities of one tropic level are organized by the tropic level above. In the case of gallforming insect communities, the hypothesis predicts that gall morphology will diverge, minimizing the number of parasitoids shared among species. We used the monophyletic group of gallforming cecidomyiids (Asphondylia spp.) on creosote bush (Larrea tridentata) to test this hypothesis, predicting that species with thicker gall walls should exclude species of parasitoids with shorter ovipositors and have lower levels of parasitism. Of 17 parasitoid species reared from Asphondylia galls on creosote bush, 9 accounted for over 98% of parasitism. Seven of these 9 species had ovipositors long enough to penetrate 10 of 13 gall morphs measured. There was no significant relationship between gall wall thickness and number of associated parasitoid species (r ²=0.01, P>0.05, n=13). There was no relationship between gall wall thickness and types of parasitoid species colonizing galls: parasitoids with the shortest ovipositors colonized all types of gall morphs and were dominant members of the parasitoid assemblages in galls with the thickest walls. Ultimately, there were no significant differences in percent parasitism among Asphondylia species, regardless of gall wall thickness. We found no difference in numbers of associated parasitoids or percent parasitism in galls with different textures (e.g. hairy versus smooth), different locations on the plant or different phenologies. Our results suggest that enemy impact has not influenced the diversity of this gall community. Gall wall thickness, phenology, location on the plant and surface structure do not appear to influence the distribution of parasitoid species. Other explanations are offered to account for diversity in gall morphology among these species.     

STN1 OB Fold Mutation Alters DNA Binding and Affects Selective Aspects of CST Function

Mammalian CST (CTC1-STN1-TEN1) participates in multiple aspects of telomere replication and genome-wide recovery from replication stress. CST resembles Replication Protein A (RPA) in that it binds ssDNA and STN1 and TEN1 are structurally similar to RPA2 and RPA3. Conservation between CTC1 and RPA1 is less apparent. Currently the mechanism underlying CST action is largely unknown. Here we address CST mechanism by using a DNA-binding mutant, (STN1 OB-fold mutant, STN1-OBM) to examine the relationship between DNA binding and CST function. In vivo, STN1-OBM affects resolution of endogenous replication stress and telomere duplex replication but telomeric C-strand fill-in and new origin firing after exogenous replication stress are unaffected. These selective effects indicate mechanistic differences in CST action during resolution of different replication problems. In vitro binding studies show that STN1 directly engages both short and long ssDNA oligonucleotides, however STN1-OBM preferentially destabilizes binding to short substrates. The finding that STN1-OBM affects binding to only certain substrates starts to explain the in vivo separation of function observed in STN1-OBM expressing cells. CST is expected to engage DNA substrates of varied length and structure as it acts to resolve different replication problems. Since STN1-OBM will alter CST binding to only some of these substrates, the mutant should affect resolution of only a subset of replication problems, as was observed in the STN1-OBM cells. The in vitro studies also provide insight into CST binding mechanism. Like RPA, CST likely contacts DNA via multiple OB folds. However, the importance of STN1 for binding short substrates indicates differences in the architecture of CST and RPA DNA-protein complexes. Based on our results, we propose a dynamic DNA binding model that provides a general mechanism for CST action at diverse forms of replication stress.     

Linking leaf veins to growth and mortality rates: an example from a subtropical tree community

A fundamental goal in ecology is to link variation in species function to performance, but functional trait–performance investigations have had mixed success. This indicates that less commonly measured functional traits may more clearly elucidate trait–performance relationships. Despite the potential importance of leaf vein traits, which are expected to be related to resource delivery rates and photosynthetic capacity, there are few studies, which examine associations between these traits and demographic performance in communities. Here, we examined the associations between species traits including leaf venation traits and demographic rates (Relative Growth Rate, RGR and mortality) as well as the spatial distributions of traits along soil environment for 54 co‐occurring species in a subtropical forest. Size‐related changes in demographic rates were estimated using a hierarchical Bayesian approach. Next, Kendall's rank correlations were quantified between traits and estimated demographic rates at a given size and between traits and species‐average soil environment. Species with denser venation, smaller areoles, less succulent, or thinner leaves showed higher RGR for a wide range of size classes. Species with leaves of denser veins, larger area, cheaper construction costs or thinner, or low‐density wood were associated with high mortality rates only in small size classes. Lastly, contrary to our expectations, acquisitive traits were not related to resource‐rich edaphic conditions. This study shows that leaf vein traits are weakly, but significantly related to tree demographic performance together with other species traits. Because leaf traits associated with an acquisitive strategy such as denser venation, less succulence, and thinner leaves showed higher growth rate, but similar leaf traits were not associated with mortality, different pathways may shape species growth and survival. This study suggests that we are still not measuring some of key traits related to resource‐use strategies, which dictate the demography and distributions of species.     

Role of the JNK pathway in NMDA-mediated excitotoxicity of cortical neurons

Excitotoxic insults induce c-Jun N-terminal kinase (JNK) activation, which leads to neuronal death and contributes to many neurological conditions such as cerebral ischemia and neurodegenerative disorders. The action of JNK can be inhibited by the D-retro-inverso form of JNK inhibitor peptide (D-JNKI1), which totally prevents death induced by N-methyl-D-aspartate (NMDA) in vitro and strongly protects against different in vivo paradigms of excitotoxicity. To obtain optimal neuroprotection, it is imperative to elucidate the prosurvival action of D-JNKI1 and the death pathways that it inhibits. In cortical neuronal cultures, we first investigate the pathways by which NMDA induces JNK activation and show a rapid and selective phosphorylation of mitogen-activated protein kinase kinase 7 (MKK7), whereas the only other known JNK activator, mitogen-activated protein kinase kinase 4 (MKK4), was unaffected. We then analyze the action of D-JNKI1 on four JNK targets containing a JNK-binding domain: MAPK-activating death domain-containing protein/differentially expressed in normal and neoplastic cells (MADD/DENN), MKK7, MKK4 and JNK-interacting protein-1 (IB1/JIP-1).     

Locating an antagonist in the 5-HT3 receptor binding site using modeling and radioligand binding

We have used a homology model of the extracellular domain of the 5-HT(3) receptor to dock granisetron, a 5-HT(3) receptor antagonist, into the binding site using AUTODOCK. This yielded 13 alternative energetically favorable models. The models fell into 3 groups. In model type A the aromatic rings of granisetron were between Trp-90 and Phe-226 and its azabicyclic ring was between Trp-183 and Tyr-234, in model type B this orientation was reversed, and in model type C the aromatic rings were between Asp-229 and Ser-200 and the azabicyclic ring was between Phe-226 and Asn-128. Residues located no more than 5 A from the docked granisetron were identified for each model; of 26 residues identified, 8 were found to be common to all models, with 18 others being represented in only a subset of the models. To identify which of the docking models best represents the ligand-receptor complex, we substituted each of these 26 residues with alanine and a residue with similar chemical properties. The mutant receptors were expressed in human embryonic kidney (HEK)293 cells and the affinity of granisetron determined using radioligand binding. Mutation of 2 residues (Trp-183 and Glu-129) ablated binding, whereas mutation of 14 other residues caused changes in the [(3)H]granisetron binding affinity in one or both mutant receptors. The data showed that residues both in and close to the binding pocket can affect antagonist binding and overall were found to best support model B.     

Partial alanine scan of mast cell degranulating peptide (MCD): importance of the histidine- and arginine residues.

The influence of the two histidine and two arginine residues of mast cell degranulating peptide (MCD) in activity and binding was studied by replacing these amino acids in the MCD sequence with L-alanine. Their histamine releasing activity was determined on rat peritoneal mast cells. Their binding affinity to the FcepsilonRIalpha binding subunit of the human mast cell receptor protein, was carried out using fluorescence polarization. The histamine assay showed that replacement of His13 by Ala o ccurred without loss of activity compared with the activity of MCD. Alanine substitutions for Arg7 and His8 resulted in an approximately 40 fold increase, and for Arg16 in a 14-fold increase in histamine-releasing activity of MCD. The binding affinities of the analogs were tested by competitive displacement of bound fluorescent MCD peptide from the FcepsilonRIalpha binding protein of the mast cell receptor by the Ala analogs using fluorescence polarization. The analogs Ala8 (for His) and Ala16 (for Arg) showed the same binding affinities as MCD, whereas analog Ala7 (for Arg) and analog Ala13 (for His) showed slightly better binding affinity than the parent compound. This study showed that the introduction of alanine residues in these positions resulted in MCD agonists of diverse potency. These findings will be useful in further MCD structure-activity studies.     

Effect of Seasonal Variation on Adult Clinical Laboratory Parameters in Rwanda,Zambia, and Uganda: Implications for HIV Biomedical Prevention Trials

Objectives

To investigate the effect of seasonal variation on adult clinical laboratory parameters in Rwanda, Zambia, and Uganda and determine its implications for HIV prevention and other clinical trials.

Methods

Volunteers in a cross-sectional study to establish laboratory reference intervals were asked to return for a seasonal visit after the local season had changed from dry to rainy or vice versa. Volunteers had to be clinically healthy, not pregnant and negative for HIV, Hepatitis B and C, and syphilis infection at both visits. At each visit, blood was taken for measurement of hemoglobin, haematocrit, mean corpuscular volume, red blood cells, platelets, total white blood cells (WBC), neutrophils, lymphocytes, monocytes, eosinophils, basophils, CD4/CD8 T cells, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, direct bilirubin, total bilirubin, total immunoglobulin gamma, total protein, creatinine, total amylase, creatine phosphokinase and lactate dehydrogenase (LDH). Consensus dry season reference intervals were applied to rainy season values (and vice versa) and the proportion of ‘out-of-range’ values determined. Percentage differences between dry and rainy season parameter mean values were estimated.

Results

In this cohort of 903 volunteers, less than 10.0% of consensus parameter (except LDH) values in one season were “out-of-range” in the other. Twenty-two (22) percent of rainy season LDH values fell outside of the consensus dry season interval with the higher values observed in the rainy season. Variability between consensus seasonal means ranged from 0.0% (total WBC, neutrophils, monocytes, basophils, and direct bilirubin) to 40.0% (eosinophils). Within sites, the largest seasonal variations were observed for monocytes (Masaka, 11.5%), LDH (Lusaka, 21.7%), and basophils (Kigali, 22.2%).

Conclusions

Seasonality had minimal impact on adult clinical laboratory parameter values in Rwanda, Zambia, and Uganda. Seasonal variation may not be an important factor in the evaluation of adult clinical laboratory parameters in HIV prevention and other clinical trials in these countries.     

Accomplishments in genome-scale in silico modeling for industrial and medical biotechnology

Driven by advancements in high-throughput biological technologies and the growing number of sequenced genomes, the construction of in silico models at the genome scale has provided powerful tools to investigate a vast array of biological systems and applications. Here, we review comprehensively the uses of such models in industrial and medical biotechnology, including biofuel generation, food production, and drug development. While the use of in silico models is still in its early stages for delivering to industry, significant initial successes have been achieved. For the cases presented here, genome-scale models predict engineering strategies to enhance properties of interest in an organism or to inhibit harmful mechanisms of pathogens. Going forward, genome-scale in silico models promise to extend their application and analysis scope to become a transformative tool in biotechnology.