Separating Instability from Aggregation Propensity in γS-Crystallin Variants

Molecular dynamics (MD) simulations, circular dichroism (CD), and dynamic light scattering (DLS) measurements were used to investigate the aggregation propensity of the eye-lens protein γS-crystallin. The wild-type protein was investigated along with the cataract-related G18V variant and the symmetry-related G106V variant. The MD simulations suggest that local sequence differences result in dramatic differences in dynamics and hydration between these two apparently similar point mutations. This finding is supported by the experimental measurements, which show that although both variants appear to be mostly folded at room temperature, both display increased aggregation propensity. Although the disease-related G18V variant is not the most strongly destabilized, it aggregates more readily than either the wild-type or the G106V variant. These results indicate that γS-crystallin provides an excellent model system for investigating the role of dynamics and hydration in aggregation by locally unfolded proteins.     

Positive feedback favors invasion by a submersed freshwater plant

The submersed macrophyte Utricularia inflata has invaded lakes in northern New York State, thereby threatening native isoetids such as Eriocaulon aquaticum. Isoetids often dominate and modify softwater lakes due to their capacity to oxidize sediment and thus influence solute mobilization. Greenhouse experiments tested the hypotheses that U. inflata invasion could result in higher porewater iron (Fe) concentrations and greater ammonium (NH₄ ⁺) and Fe release from the sediment into the water column, and that this mobilization would stimulate further U. inflata growth. In the first experiment, three levels of U. inflata impact on E. aquaticum were imposed using sediment cores overlain by lake water: E. aquaticum alone, E. aquaticum with a cover of U. inflata, and bare sediment—the latter to simulate local extirpation of the isoetid by the invasive. After 16 weeks, sediment porewater NH₄ ⁺ and total dissolved Fe concentrations were significantly higher (P < 0.05) for the U. inflata and bare sediment treatments. Water column concentrations of these solutes were five-fold higher (P < 0.05) for the bare sediment treatment than E. aquaticum alone, indicating that isoetid extirpation by U. inflata can compromise water quality. A second experiment demonstrated that U. inflata grew faster over bare sediment than over sediment with E. aquaticum (P < 0.05), likely due to greater solute mobilization in the absence of E. aquaticum. Where U. inflata causes a decline of native isoetids in Adirondack Mountain lakes, changes to lake sediment and water chemistry can create a positive feedback loop further escalating the impact of this invasive species.     

A network of conserved interactions regulates the allosteric signal in a glutamine amidotransferase

We have combined equilibrium and steered molecular dynamics (SMD) simulations with principal component and correlation analyses to probe the mechanism of allosteric regulation in imidazole glycerol phosphate (IGP) synthase. An evolutionary analysis of IGP synthase revealed a conserved network of interactions leading from the effector binding site to the glutaminase active site, forming conserved communication pathways between the remote active sites. SMD simulations of the undocking of the ribonucleotide effector N1-[(5'-phosphoribulosyl)-formino]-5'-aminoimidazole carboxamide ribonucleotide (PRFAR) resulted in a large scale hinge-opening motion at the interface. Principal component analysis and a correlation analysis of the equilibration protein motion indicate that the dynamics involved in the allosteric transition are mediated by coupled motion between sites that are more than 25 A apart. Furthermore, conserved residues at the substrate-binding site, within the barrel, and at the interface were found to exhibit highly correlated motion during the allosteric transition. The coupled motion between PRFAR unbinding and the directed opening of the interface is interpreted in combination with kinetic assays for the wild-type and mutant systems to develop a model of allosteric regulation in IGP synthase that is monitored and investigated with atomic resolution.     

Social defeat: impact on fear extinction and amygdala-prefrontal cortical theta synchrony in 5-HTT deficient mice

Emotions, such as fear and anxiety, can be modulated by both environmental and genetic factors. One genetic factor is for example the genetically encoded variation of the serotonin transporter (5-HTT) expression. In this context, the 5-HTT plays a key role in the regulation of central 5-HT neurotransmission, which is critically involved in the physiological regulation of emotions including fear and anxiety. However, a systematic study which examines the combined influence of environmental and genetic factors on fear-related behavior and the underlying neurophysiological basis is missing. Therefore, in this study we used the 5-HTT-deficient mouse model for studying emotional dysregulation to evaluate consequences of genotype specific disruption of 5-HTT function and repeated social defeat for fear-related behaviors and corresponding neurophysiological activities in the lateral amygdala (LA) and infralimbic region of the medial prefrontal cortex (mPFC) in male 5-HTT wild-type (+/+), homo- (-/-) and heterozygous (+/-) mice. Naive males and experienced losers (generated in a resident-intruder paradigm) of all three genotypes, unilaterally equipped with recording electrodes in LA and mPFC, underwent a Pavlovian fear conditioning. Fear memory and extinction of conditioned fear was examined while recording neuronal activity simultaneously with fear-related behavior. Compared to naive 5-HTT+/+ and +/- mice, 5-HTT-/- mice showed impaired recall of extinction. In addition, 5-HTT-/- and +/- experienced losers showed delayed extinction learning and impaired recall of extinction. Impaired behavioral responses were accompanied by increased theta synchronization between the LA and mPFC during extinction learning in 5-HTT-/- and +/- losers. Furthermore, impaired extinction recall was accompanied with increased theta synchronization in 5-HTT-/- naive and in 5-HTT-/- and +/- loser mice. In conclusion, extinction learning and memory of conditioned fear can be modulated by both the 5-HTT gene activity and social experiences in adulthood, accompanied by corresponding alterations of the theta activity in the amygdala-prefrontal cortex network.     

Cyclooxygenase-2 deficiency leads to intestinal barrier dysfunction and increased mortality during polymicrobial sepsis

Sepsis remains the leading cause of death in critically ill patients, despite modern advances in critical care. Intestinal barrier dysfunction may lead to secondary bacterial translocation and the development of the multiple organ dysfunction syndrome during sepsis. Cyclooxygenase (COX)-2 is highly upregulated in the intestine during sepsis, and we hypothesized that it may be critical in the maintenance of intestinal epithelial barrier function during peritonitis-induced polymicrobial sepsis. COX-2(-/-) and COX-2(+/+) BALB/c mice underwent cecal ligation and puncture (CLP) or sham surgery. Mice chimeric for COX-2 were derived by bone marrow transplantation and underwent CLP. C2BBe1 cells, an intestinal epithelial cell line, were treated with the COX-2 inhibitor NS-398, PGD(2), or vehicle and stimulated with cytokines. COX-2(-/-) mice developed exaggerated bacteremia and increased mortality compared with COX-2(+/+) mice following CLP. Mice chimeric for COX-2 exhibited the recipient phenotype, suggesting that epithelial COX-2 expression in the ileum attenuates bacteremia following CLP. Absence of COX-2 significantly increased epithelial permeability of the ileum and reduced expression of the tight junction proteins zonula occludens-1, occludin, and claudin-1 in the ileum following CLP. Furthermore, PGD(2) attenuated cytokine-induced hyperpermeability and zonula occludens-1 downregulation in NS-398-treated C2BBe1 cells. Our findings reveal that absence of COX-2 is associated with enhanced intestinal epithelial permeability and leads to exaggerated bacterial translocation and increased mortality during peritonitis-induced sepsis. Taken together, our results suggest that epithelial expression of COX-2 in the ileum is a critical modulator of tight junction protein expression and intestinal barrier function during sepsis.     

Historical demography and colonization pathways of the widespread intertidal seaweed Hormosira banksii (Phaeophyceae) in southeastern Australia

The palaeoceanography of southern Australia has been characterized by fluctuating sea levels during glacial periods, changing temperature regimes and modified boundary currents. Previous studies on genetic structuring of species in southeastern Australia have focused mainly on the differentiation of eastern and western populations while the potential role of Bass Strait as a region of overlap for three biogeographic provinces (Peronia, Maugea, and Flindersia) has been largely ignored. This study aimed to explore the likely roles of historic and contemporary factors in determining divergence patterns in the habitat‐forming intertidal seaweed Hormosira banksii in southeastern Australia with a special focus on postglacial dispersal into Bass Strait. We examined the genetic diversity of 475 Hormosira specimens collected from 19 sites around southern Australia using DNA sequence analysis of cytochrome oxidase 1. Three major haplotype groups were identified (western, centre and eastern) corresponding with the three existing biogeographical provinces in this region. Historic break points appeared to be retained and reinforced by modern day dispersal barriers. Phylogeographic grouping of Hormosira reflected a combination of historic and contemporary oceanography. As western and eastern group haplotypes were largely absent within Bass Strait, re‐colonization after the last glacial maximum appeared to have originated from refuges within or near present day Bass Strait. Patterns of genetic structure for Hormosira are consistent with other marine species in this region and highlight the importance of biogeographical barriers in contributing to modern genetic structure.     

Consumer–plant interaction strength: importance of body size,density and metabolic biomass

Explaining variability in the strength and sign of trophic interactions between primary consumers and plants is a long‐standing research challenge. Consumer density and body size vary widely in space and time and are predicted to have interactive effects on consumer–plant interactions. In a southern US salt marsh, we used replicate field enclosures to orthogonally manipulate the body size (mass) and density of a dominant consumer (a snail). We investigated impacts (leaf damage and biomass) on monocultures of cordgrass, the foundation species, over three months. Increasing consumer density and body size increased leaf damage additively and, as predicted, multiplicatively reduced plant biomass. Notably, size and density determined the sign of consumer impact on plants: low to medium densities of small consumers enhanced, while high densities of large consumers strongly suppressed, plant biomass. Finally, total consumer metabolic biomass (mass^0.75) within an enclosure parsimoniously explained plant biomass response, supporting theoretical predictions and suggesting that multiplicative effects of density and body size resulted from their effects on total metabolic biomass. The consequences of changes in consumer density and body size resulting from anthropogenic perturbations may therefore be predicted based on metabolic biomass. Synthesis Consumer size, density and biomass can all strongly affect consumer–plant interactions. Though density and body size have been extensively studied as drivers of variation in interaction strength, the role of biomass as the ultimate driver has been less appreciated. We manipulated body size and density of a single consumer species and, based on metabolic theory, integrated these into a single variable: total metabolic biomass. Our results suggest that changes in interaction strength attributed to size or density may in fact be due to changes in metabolic biomass. This metric could thus serve as a useful tool in further understanding species interactions.     

The mousetrap: what we can learn when the mouse model does not mimic the human disease

In recent years, mouse models for human metabolic diseases have become commonplace because the information gained from in vivo study of biochemical pathways is invaluable, and many metabolic diseases are relatively easy to recreate in mice through gene knockout technology in embryonic stem cells. In certain cases, however, the knockout mice may reproduce only some of the human disease phenotype, may be more severely affected than human cases, or may have no clinical phenotype at all. Under these circumstances, the disease pathology can become more complex, causing the researcher to evaluate basic differences in mouse and human biology as well as questions of genetic background, alternate pathways, and possible gene interactions. This review is a brief analysis of gene knockout models for Lesch-Nyhan syndrome, Lowe syndrome, X-linked adrenoleukodystrophy, Fabry disease, galactosemia, glycogen storage disease type II, metachromatic leukodystrophy, and Tay-Sachs disease, which produce a biochemical model of disease but often do not reproduce clinical symptoms. These mice may be useful for studying the biochemical and physiological pathways in which certain metabolites function toward embryonic and fetal development, as well as specific functions in various organs, and they may provide an inexpensive and useful model system for development of new therapeutic techniques.     

Phenological change in a spring ephemeral: implications for pollination and plant reproduction

Climate change has had numerous ecological effects, including species range shifts and altered phenology. Altering flowering phenology often affects plant reproduction, but the mechanisms behind these changes are not well‐understood. To investigate why altering flowering phenology affects plant reproduction, we manipulated flowering phenology of the spring herb Claytonia lanceolata (Portulacaceae) using two methods: in 2011–2013 by altering snow pack (snow‐removal vs. control treatments), and in 2013 by inducing flowering in a greenhouse before placing plants in experimental outdoor arrays (early, control, and late treatments). We measured flowering phenology, pollinator visitation, plant reproduction (fruit and seed set), and pollen limitation. Flowering occurred approx. 10 days earlier in snow‐removal than control plots during all years of snow manipulation. Pollinator visitation patterns and strength of pollen limitation varied with snow treatments, and among years. Plants in the snow removal treatment were more likely to experience frost damage, and frost‐damaged plants suffered low reproduction despite lack of pollen limitation. Plants in the snow removal treatment that escaped frost damage had higher pollinator visitation rates and reproduction than controls. The results of the array experiment supported the results of the snow manipulations. Plants in the early and late treatments suffered very low reproduction due either to severe frost damage (early treatment) or low pollinator visitation (late treatment) relative to control plants. Thus, plants face tradeoffs with advanced flowering time. While early‐flowering plants can reap the benefits of enhanced pollination services, they do so at the cost of increased susceptibility to frost damage that can overwhelm any benefit of flowering early. In contrast, delayed flowering results in dramatic reductions in plant reproduction through reduced pollination. Our results suggest that climate change may constrain the success of early‐flowering plants not through plant‐pollinator mismatch but through the direct impacts of extreme environmental conditions.     

Landscape simplification increases vineyard pest outbreaks and insecticide use

Diversifying agricultural landscapes may mitigate biodiversity declines and improve pest management. Yet landscapes are rarely managed to suppress pests, in part because researchers seldom measure key variables related to pest outbreaks and insecticides that drive management decisions. We used a 13‐year government database to analyse landscape effects on European grapevine moth (Lobesia botrana) outbreaks and insecticides across c. 400 Spanish vineyards. At harvest, we found pest outbreaks increased four‐fold in simplified, vineyard‐dominated landscapes compared to complex landscapes in which vineyards are surrounded by semi‐natural habitats. Similarly, insecticide applications doubled in vineyard‐dominated landscapes but declined in vineyards surrounded by shrubland. Importantly, pest population stochasticity would have masked these large effects if numbers of study sites and years were reduced to typical levels in landscape pest‐control studies. Our results suggest increasing landscape complexity may mitigate pest populations and insecticide applications. Habitat conservation represents an economically and environmentally sound approach for achieving sustainable grape production.