bluuebunny/crossref_metadata_2025 huggingface dataset:
Dynamic sexual dichromatism in an explosively breeding Neotropical toad
Authors: Doucet Stéphanie M. , Mennill Daniel J.
Published: October 2009

Sexual selection often promotes the evolution of elaborate colour signals in males, but the importance of sexually selected colour signals remains poorly studied in amphibians. We used reflectance spectrometry to document pronounced sexual dichromatism and dramatic colour change inBufo luetkenii, a toad that breeds in large aggregations at the onset of the rainy season in Costa Rica. Our observations suggest that males fade rapidly from a vibrant lemon yellow to a dull brown once they have paired with a female. We demonstrate this by showing that males are much brighter than females and that unpaired males are more colourful than males in amplexus. We also show that coloration fades rapidly when males are briefly held captive. This is, to our knowledge, the first study to document such dynamic change in male coloration and sexual dichromatism in anurans.
Sharks shape the geometry of a selfish seal herd: experimental evidence from seal decoys
Authors: De Vos Alta , O'Riain M. Justin
Published: October 2009

Many animals respond to predation risk by forming groups. Evolutionary explanations for group formation in previously ungrouped, but loosely associated prey have typically evoked the selfish herd hypothesis. However, despite over 600 studies across a diverse array of taxa, the critical assumptions of this hypothesis have remained collectively untested, owing to several confounding problems in real predator–prey systems. To solve this, we manipulated the domains of danger of Cape fur seal (Arctocephalus pusillus pusillus) decoys to provide evidence that a selfish reduction in a seals' domain of danger results in a proportional reduction in its predation risk from ambush shark attacks. This behaviour confers a survival advantage to individual seals within a group and explains the evolution of selfish herds in a prey species. These findings empirically elevate Hamilton's selfish herd hypothesis to more than a ‘theoretical curiosity’.
Adaptation as organism design
Authors: Gardner Andy
Published: October 2009

The problem of adaptation is to explain the apparent design of organisms. Darwin solved this problem with the theory of natural selection. However, population geneticists, whose responsibility it is to formalize evolutionary theory, have long neglected the link between natural selection and organismal design. Here, I review the major historical developments in theory of organismal adaptation, clarifying what adaptation is and what it is not, and I point out future avenues for research.
Climate change increases the likelihood of catastrophic avian mortality events during extreme heat waves
Authors: McKechnie Andrew E. , Wolf Blair O.
Published: October 2009

Severe heat waves have occasionally led to catastrophic avian mortality in hot desert environments. Climate change models predict increases in the intensity, frequency and duration of heat waves. A model of avian evaporative water requirements and survival times during the hottest part of day reveals that the predicted increases in maximum air temperatures will result in large fractional increases in water requirements (in small birds, equivalent to 150–200 % of current values), which will severely reduce survival times during extremely hot weather. By the 2080s, desert birds will experience reduced survival times much more frequently during mid-summer, increasing the frequency of catastrophic mortality events.
Resolving the impact of waiting time distributions on the persistence of measles
Authors: Conlan Andrew J. K. , Rohani Pejman , Lloyd Alun L. , Keeling Matthew , Grenfell Bryan T.
Published: October 2009

Measles epidemics in human populations exhibit what is perhaps the best empirically characterized, and certainly the most studied, stochastic persistence threshold in population biology. A critical community size (CCS) of around 250 000–500 000 separates populations where measles is predominantly persistent from smaller communities where there are frequent extinctions of measles between major epidemics. The fundamental mechanisms contributing to this pattern of persistence, which are long-lasting immunity to re-infection, recruitment of susceptibles, seasonality in transmission, age dependence of transmission and the spatial coupling between communities, have all been quantified and, to a greater or lesser level of success, captured by theoretical models. Despite these successes there has not been a consensus over whether simple models can successfully predict the value of the CCS, or indeed which mechanisms determine the persistence of measles over a broader range of population sizes. Specifically, the level of the CCS has been thought to be particularly sensitive to the detailed stochastic dynamics generated by the waiting time distribution (WTD) in the infectious and latent periods. We show that the relative patterns of persistence between models with different WTDs are highly sensitive to the criterion of comparison—in particular, the statistical measure of persistence that is employed. To this end, we introduce two new statistical measures of persitence—fade-outs post epidemic and fade-outs post invasion. Contrary to previous reports, we demonstrate that, no matter the choice of persistence measure, appropriately parametrized models of measles demonstrate similar predictions for the level of the CCS.
An impaired alveolar-capillary barrier
in vitro
: effect of proinflammatory cytokines and consequences on nanocarrier interaction
Authors: Hermanns Maria Iris , Kasper Jennifer , Dubruel Peter , Pohl Christine , Uboldi Chiara , Vermeersch Vincent , Fuchs Sabine , Unger Ronald E. , Kirkpatrick C. James
Published: October 2009

The alveolar region of the lung is an important target for drug and gene delivery approaches. Treatment with drugs is often necessary under pathophysiological conditions, in which there is acute inflammation of the target organ. Therefore,
in vitro
models of the alveolar-capillary barrier, which mimic inflammatory conditions in the alveolar region, would be useful to analyse and predict effects of novel drugs on healthy or inflamed tissues. The epithelial cell line H441 was cultivated with primary isolated human pulmonary microvascular endothelial cells (HPMECs) or the endothelial cell line ISO-HAS-1 on opposite sides of a permeable filter support under physiological and inflammatory conditions. Both epithelial and endothelial cell types grew as polarized monolayers in bilayer coculture and were analysed in the presence and absence of the proinflammatory stimuli tumour necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ). In addition, the nanocarrier polyethyleneimine (PEI) was chosen as a model compound to study cell uptake (Oregon Green (OG)-labelled PEI) and gene transfer (PEI–pDNA complex). Upon treatment with TNF-α and IFN-γ, both cocultures exhibited comparable effects on the trans-bilayer electrical resistance, the transport of sodium fluorescein and the increase in secondary cytokine release. Basolateral (endothelial side) exposure to TNF-α or simultaneous exposure to TNF-α and IFN-γ generated an alveolar-capillary barrier with inflammation-like characteristics, impaired barrier function and a local disruption of the continuous apical labelling of the tight junction plaque protein zonula occludens-1 (ZO-1). Although transfection rates of 8 per cent were obtained for H441 cells in non-polarized monocultures, apical–basolateral-differentiated (polarized) H441 in coculture could not be transfected. After basolateral cytokine exposure, uptake of fluorescently labelled PEI in polarized H441 was predominantly detected in those areas with a local disruption of ZO-1 expression. Accordingly, transfected cells were only sparsely found in coculture after basolateral costimulation with TNF-α and IFN-γ. We designed a coculture model that mimics both the structural architecture of the alveolar-capillary barrier and inflammatory mechanisms with consequences on barrier characteristics, cytokine production and nanoparticle interaction. Our model will be suitable to systematically study adsorption, uptake and trafficking of newly synthesized nanosized carriers under different physiological conditions.
Constructing tensegrity structures from one-bar elementary cells
Authors: Li Yue , Feng Xi-Qiao , Cao Yan-Ping , Gao Huajian
Published: October 2009

This study aimed to develop a method to construct tensegrity structures from elementary cells, defined as structures consisting of only one bar connected with a few strings. Comparison of various elementary cells leads to the further selection of the so-called ‘Z-shaped’ cell, which contains one bar and three interconnected strings, as the elementary module to assemble the Z-based spatial tensegrity structures. The graph theory is utilized to analyse the topology of strings required to construct this type of tensegrity structures. It is shown that ‘a string net can be used to construct a Z-based tensegrity structure if and only if its topology is a simple and bridgeless cubic graph’. Once the topology of strings has been determined, one can easily design the associated tensegrity structure by adding a deterministic number of bars. Two schemes are suggested for this design strategy. One is to enumerate all possible topologies of Z-based tensegrity for a specified number of bars or cells, and the other is to determine the tensegrity structure from a vertex-truncated polyhedron. The method developed in this paper allows us to construct various types of novel tensegrity structures.
Balance between maternal and paternal alleles sets the timing of resource accumulation in the maize endosperm
Authors: Li Na , Dickinson Hugh G.
Published: October 2009

Key aspects of seed development in flowering plants are held to be under epigenetic control and to have evolved as a result of conflict between the interests of the male and female gametes (kinship theory). Attempts to identify the genes involved have focused on imprinted sequences, although imprinting is only one mechanism by which male or female parental alleles may be exclusively expressed immediately post-fertilization. We have studied the expression of a subset of endosperm gene classes immediately following interploidy crosses in maize and show that departure from the normal 2 : 1 ratio between female and male genomes exerts a dramatic effect on the timing of expression of some, but not all, genes investigated. Paternal genomic excess prolongs the expression of early genes and delays accumulation of reserves, while maternal genomic excess foreshortens the expression period of early genes and dramatically brings forward endosperm maturation. Our data point to a striking interdependence between the phases of endosperm development, and are consonant with previous work from maize showing progression from cell proliferation to endoreduplication is regulated by the balance between maternal and paternal genomes, and from Arabidopsis suggesting that this ‘phasing’ is regulated by maternally expressed imprinted genes. Our findings are discussed in context of the kinship theory.
Drosophila
male sex peptide inhibits siesta sleep and promotes locomotor activity in the post-mated female
Authors: Isaac R. Elwyn , Li Chenxi , Leedale Amy E. , Shirras Alan D.
Published: October 2009

Quiescence, or a sleep-like state, is a common and important feature of the daily lives of animals from both invertebrate and vertebrate taxa, suggesting that sleep appeared early in animal evolution. Recently,
Drosophila melanogaster
has been shown to be a relevant and powerful model for the genetic analysis of sleep behaviour. The sleep architecture of
D. melanogaster
is sexually dimorphic, with females sleeping much less than males during day-time, presumably because reproductive success requires greater foraging activity by the female as well as the search for egg-laying sites. However, this loss of sleep and increase in locomotor activity will heighten the risk for the female from environmental and predator hazards. In this study, we show that virgin females can minimize this risk by behaving like males, with an extended afternoon ‘siesta’. Copulation results in the female losing 70 per cent of day-time sleep and becoming more active. This behaviour lasts for at least 8 days after copulation and is abolished if the mating males lack sex peptide (SP), normally present in the seminal fluid. Our results suggest that SP is the molecular switch that promotes wakefulness in the post-mated female, a change of behaviour compatible with increased foraging and egg-laying activity. The stress resulting from SP-dependent sleep deprivation might be an important contribution to the toxic side-effects of male accessory gland products that are known to reduce lifespan in post-mated females.
Nymphalid butterflies diversify following near demise at the Cretaceous/Tertiary boundary
Authors: Wahlberg Niklas , Leneveu Julien , Kodandaramaiah Ullasa , Peña Carlos , Nylin Sören , Freitas André V. L. , Brower Andrew V. Z.
Published: October 2009

The butterfly family Nymphalidae contains some of the most important non-drosophilid insect model systems for evolutionary and ecological studies, yet the evolutionary history of the group has remained shrouded in mystery. We have inferred a robust phylogenetic hypothesis based on sequences of 10 genes and 235 morphological characters for exemplars of 400 of the 540 valid nymphalid genera representing all major lineages of the family. By dating the branching events, we infer that Nymphalidae originated in the Cretaceous at 90 Ma, but that the ancestors of 10–12 lineages survived the end-Cretaceous catastrophe in the Neotropical and Oriental regions. Patterns of diversification suggest extinction of lineages at the Cretaceous/Tertiary boundary (65 Ma) and subsequent elevated speciation rates in the Tertiary.
Sauropod dinosaurs evolved moderately sized genomes unrelated to body size
Authors: Organ Chris L. , Brusatte Stephen L. , Stein Koen
Published: October 2009

Sauropodomorph dinosaurs include the largest land animals to have ever lived, some reaching up to 10 times the mass of an African elephant. Despite their status defining the upper range for body size in land animals, it remains unknown whether sauropodomorphs evolved larger-sized genomes than non-avian theropods, their sister taxon, or whether a relationship exists between genome size and body size in dinosaurs, two questions critical for understanding broad patterns of genome evolution in dinosaurs. Here we report inferences of genome size for 10 sauropodomorph taxa. The estimates are derived from a Bayesian phylogenetic generalized least squares approach that generates posterior distributions of regression models relating genome size to osteocyte lacunae volume in extant tetrapods. We estimate that the average genome size of sauropodomorphs was 2.02 pg (range of species means: 1.77–2.21 pg), a value in the upper range of extant birds (mean = 1.42 pg, range: 0.97–2.16 pg) and near the average for extant non-avian reptiles (mean = 2.24 pg, range: 1.05–5.44 pg). The results suggest that the variation in size and architecture of genomes in extinct dinosaurs was lower than the variation found in mammals. A substantial difference in genome size separates the two major clades within dinosaurs, Ornithischia (large genomes) and Saurischia (moderate to small genomes). We find no relationship between body size and estimated genome size in extinct dinosaurs, which suggests that neutral forces did not dominate the evolution of genome size in this group.
The Celtic fringe of Britain: insights from small mammal phylogeography
Authors: Searle Jeremy B. , Kotlík Petr , Rambau Ramugondo V. , Marková Silvia , Herman Jeremy S. , McDevitt Allan D.
Published: October 2009

Recent genetic studies have challenged the traditional view that the ancestors of British Celtic people spread from central Europe during the Iron Age and have suggested a much earlier origin for them as part of the human recolonization of Britain at the end of the last glaciation. Here we propose that small mammals provide an analogue to help resolve this controversy. Previous studies have shown that common shrews (
Sorex araneus
) with particular chromosomal characteristics and water voles (
Arvicola terrestris
) of a specific mitochondrial (mt) DNA lineage have peripheral western/northern distributions with striking similarities to that of Celtic people. We show that mtDNA lineages of three other small mammal species (bank vole
Myodes glareolus
, field vole
Microtus agrestis
and pygmy shrew
Sorex minutus
) also form a ‘Celtic fringe’. We argue that these small mammals most reasonably colonized Britain in a two-phase process following the last glacial maximum (LGM), with climatically driven partial replacement of the first colonists by the second colonists, leaving a peripheral geographical distribution for the first colonists. We suggest that these natural Celtic fringes provide insight into the same phenomenon in humans and support its origin in processes following the end of the LGM.
Androgen receptor gene sequence and basal cortisol concentrations predict men's hormonal responses to potential mates
Authors: Roney James R. , Simmons Zachary L. , Lukaszewski Aaron W.
Published: October 2009

Exposure to potential mates triggers rapid elevations of testosterone and glucocorticoid concentrations in males of many non-human species, and preliminary studies support similar effects in human males. The human studies have all reported large individual differences in these responses, however, and the present study tested whether specific biological variables may help explain these differences. Replicating past research, the present study found that men's salivary testosterone and cortisol concentrations increased after a brief conversation with a young woman, but did not change (or slightly decreased) after a conversation with a young man. In addition, smaller numbers of CAG repeats in men's androgen receptor gene, and lower baseline cortisol concentrations, each predicted larger testosterone responses to the interactions with women. The CAG repeat finding demonstrates that a specific genetic polymorphism predicts physiological responses to social interactions that may in turn have important downstream consequences on men's mating behaviour. The effects of cortisol are consistent with past demonstrations of glucocorticoid inhibition of testosterone production and show that such inhibition also affects testosterone responses to social stimuli. In sum, the present study both confirms men's hormonal reactions to potential mates and identifies novel biological variables that predict individual differences in these responses.
Both population size and patch quality affect local extinctions and colonizations
Authors: Franzén Markus , Nilsson Sven G.
Published: October 2009

Currently, the habitat of many species is fragmented, resulting in small local populations with individuals occasionally dispersing between the remaining habitat patches. In a solitary bee metapopulation, extinction probability was related to both local bee population sizes and pollen resources measured as host plant population size. Patch size, on the other hand, had no additional predictive power. The turnover rate of local bee populations in 63 habitat patches over 4 years was high, with 72 extinction events and 31 colonization events, but the pollen plant population was stable with no extinctions or colonizations. Both pollen resources and bee populations had strong and independent effects on extinction probability, but connectivity was not of importance. Colonizations occurred more frequently within larger host plant populations. For metapopulation survival of the bee, large pollen plant populations are essential, independent of current bee population size.
The annealing helicase HARP is recruited to DNA repair sites via an interaction with RPA
Authors: Yusufzai Timur , Kong Xiangduo , Yokomori Kyoko , Kadonaga James T.
Published: October 2009

HepA-related protein (HARP) (also known as SMARCAL1) is an ATP-driven annealing helicase that catalyzes the formation of dsDNA from complementary Replication protein A (RPA)-bound ssDNA. Here we find that HARP contains a conserved N-terminal motif that is necessary and sufficient for binding to RPA. This RPA-binding motif is not required for annealing helicase activity, but is essential for the recruitment of HARP to sites of laser-induced DNA damage. These findings suggest that the interaction of HARP with RPA increases the concentration of annealing helicase activity in the vicinity of ssDNA regions to facilitate processes such as DNA repair.
The SIOD disorder protein SMARCAL1 is an RPA-interacting protein involved in replication fork restart
Authors: Ciccia Alberto , Bredemeyer Andrea L. , Sowa Mathew E. , Terret Marie-Emilie , Jallepalli Prasad V. , Harper J. Wade , Elledge Stephen J.
Published: October 2009

The integrity of genomic DNA is continuously challenged by the presence of DNA base lesions or DNA strand breaks. Here we report the identification of a new DNA damage response protein, SMARCAL1 (SWI/SNF-related, matrix associated, actin-dependent regulator of chromatin, subfamily a-like 1), which is a member of the SNF2 family and is mutated in Schimke immunoosseous dysplasia (SIOD). We demonstrate that SMARCAL1 directly interacts with Replication protein A (RPA) and is recruited to sites of DNA damage in an RPA-dependent manner. SMARCAL1-depleted cells display sensitivity to DNA-damaging agents that induce replication fork collapse, and exhibit slower fork recovery and delayed entry into mitosis following S-phase arrest. Furthermore, SIOD patient fibroblasts reconstituted with SMARCAL1 exhibit faster cell cycle progression after S-phase arrest. Thus, the symptoms of SIOD may be caused, at least in part, by defects in the cellular response to DNA replication stress.
The annealing helicase HARP protects stalled replication forks
Authors: Yuan Jingsong , Ghosal Gargi , Chen Junjie
Published: October 2009

Mutations in HepA-related protein (HARP) are the only identified causes of Schimke immunoosseous dysplasia (SIOD). HARP has a unique annealing helicase activity in vitro, but the in vivo functional significance remains unknown. Here, we demonstrated that HARP is recruited to stalled replication forks via its direct interaction with Replication protein A (RPA). Cells with HARP depletion displayed increased spontaneous DNA damage and G2/M arrest, suggesting that HARP normally acts to stabilize stalled replication forks. Our data place the annealing helicase activity of HARP at replication forks and propose that SIOD syndrome may be caused by the destabilization of replication forks during cell proliferation.
A heritable switch in carbon source utilization driven by an unusual yeast prion
Authors: Brown Jessica C.S. , Lindquist Susan
Published: October 2009

Several well-characterized fungal proteins act as prions, proteins capable of multiple conformations, each with different activities, at least one of which is self-propagating. Through such self-propagating changes in function, yeast prions act as protein-based elements of phenotypic inheritance. We report a prion that makes cells resistant to the glucose-associated repression of alternative carbon sources, [GAR+] (for “resistant to glucose-associated repression,” with capital letters indicating dominance and brackets indicating its non-Mendelian character). [GAR+] appears spontaneously at a high rate and is transmissible by non-Mendelian, cytoplasmic inheritance. Several lines of evidence suggest that the prion state involves a complex between a small fraction of the cellular complement of Pma1, the major plasma membrane proton pump, and Std1, a much lower-abundance protein that participates in glucose signaling. The Pma1 proteins from closely related Saccharomyces species are also associated with the appearance of [GAR+]. This allowed us to confirm the relationship between Pma1, Std1, and [GAR+] by establishing that these proteins can create a transmission barrier for prion propagation and induction in Saccharomyces cerevisiae. The fact that yeast cells employ a prion-based mechanism for heritably switching between distinct carbon source utilization strategies, and employ the plasma membrane proton pump to do so, expands the biological framework in which self-propagating protein-based elements of inheritance operate.
Long-range spreading of dosage compensation in Drosophila captures transcribed autosomal genes inserted on X
Authors: Gorchakov Andrey A. , Alekseyenko Artyom A. , Kharchenko Peter , Park Peter J. , Kuroda Mitzi I.
Published: October 2009

Dosage compensation in Drosophila melanogaster males is achieved via targeting of male-specific lethal (MSL) complex to X-linked genes. This is proposed to involve sequence-specific recognition of the X at ∼150–300 chromatin entry sites, and subsequent spreading to active genes. Here we ask whether the spreading step requires transcription and is sequence-independent. We find that MSL complex binds, acetylates, and up-regulates autosomal genes inserted on X, but only if transcriptionally active. We conclude that a long-sought specific DNA sequence within X-linked genes is not obligatory for MSL binding. Instead, linkage and transcription play the pivotal roles in MSL targeting irrespective of gene origin and DNA sequence.
Autophagy for the avoidance of neurodegeneration
Authors: Madeo Frank , Eisenberg Tobias , Kroemer Guido
Published: October 2009

Cellular defense mechanisms, including the unfolded protein response (UPR) and autophagy, attempt to resolve toxic protein aggregates, which are common denominators of neurodegenerative diseases. In this issue of Genes & Development, Hetz and colleagues (pp. 2294–2306) surprisingly show that inhibition of the UPR by knockout of XBP-1 causes a massive increase in autophagy, enhances clearance of superoxide dismutase 1 (SOD1) aggregates, and delays the development of amyotrophic lateral sclerosis. These findings suggest the existence of a homeostatic—if not hormetic—balance between distinct cellular defense mechanisms.
Behavioral deficits and subregion-specific suppression of LTP in mice expressing a population of mutant NMDA receptors throughout the hippocampus
Authors: Chen Philip E. , Errington Michael L. , Kneussel Matthias , Chen Guiquan , Annala Alexander J. , Rudhard York H. , Rast Georg F. , Specht Christian G. , Tigaret Cezar M. , Nassar Mohammed A. , Morris Richard G.M. , Bliss Timothy V.P. , Schoepfer Ralf
Published: October 2009

The NMDA receptor (NMDAR) subunit GluN1 is an obligatory component of NMDARs without a known functional homolog and is expressed in almost every neuronal cell type. The NMDAR system is a coincidence detector with critical roles in spatial learning and synaptic plasticity. Its coincidence detection property is crucial for the induction of hippocampal long-term potentiation (LTP). We have generated a mutant mouse model expressing a hypomorph of the Grin1N598R allele, which leads to a minority (about 10%) of coincidence detection-impaired NMDARs. Surprisingly, these animals revealed specific functional changes in the dentate gyrus (DG) of the hippocampal formation. Early LTP was expressed normally in area CA1 in vivo, but was completely suppressed at perforant path-granule cell synapses in the DG. In addition, there was a pronounced reduction in the amplitude of the evoked population spike in the DG. These specific changes were accompanied by behavioral impairments in spatial recognition, spatial learning, reversal learning, and retention. Our data show that minor changes in GluN1-dependent NMDAR physiology can cause dramatic consequences in synaptic signaling in a subregion-specific fashion despite the nonredundant nature of the GluN1 gene and its global expression.
Long-term memory shapes the primary olfactory center of an insect brain
Authors: Hourcade Benoît , Perisse Emmanuel , Devaud Jean-Marc , Sandoz Jean-Christophe
Published: October 2009

The storage of stable memories is generally considered to rely on changes in the functional properties and/or the synaptic connectivity of neural networks. However, these changes are not easily tractable given the complexity of the learning procedures and brain circuits studied. Such a search can be narrowed down by studying memories of specific stimuli in a given sensory modality and by working on networks with a modular and relatively simple organization. We have therefore focused on associative memories of individual odors and the possible related changes in the honeybee primary olfactory center, the antennal lobe (AL). As this brain structure is organized in well-identified morpho-functional units, the glomeruli, we looked for evidence of structural and functional plasticity in these units in relation with the bees' ability to store long-term memories (LTMs) of specific odors. Restrained bees were trained to form an odor-specific LTM in an appetitive Pavlovian conditioning protocol. The stability and specificity of this memory was tested behaviorally 3 d after conditioning. At that time, we performed both a structural and a functional analysis on a subset of 17 identified glomeruli by measuring glomerular volume under confocal microscopy, and odor-evoked activity, using in vivo calcium imaging. We show that long-term olfactory memory for a given odor is associated with volume increases in a subset of glomeruli. Independent of these structural changes, odor-evoked activity was not modified. Lastly, we show that structural glomerular plasticity can be predicted based on a putative model of interglomerular connections.
Novelty-induced arousal enhances memory for cued classical fear conditioning: Interactions between peripheral adrenergic and brainstem glutamatergic systems
Authors: King Stanley O. , Williams Cedric L.
Published: October 2009

Exposure to novel contexts produce heightened states of arousal and biochemical changes in the brain to consolidate memory. However, processes permitting simple exposure to unfamiliar contexts to elevate sympathetic output and to improve memory are poorly understood. This shortcoming was addressed by examining how novelty-induced changes in peripheral and/or central arousal modulates memory for Pavlovian fear conditioning. Male rats were either exposed to the conditioning chamber for 5-min or given no exposure 24 h before conditioning with five tone–shock (0.35 mA) pairings. Retention was assessed 48 h later in a different context. Non-pre-exposed animals exhibited significantly greater freezing during conditioned stimulus (CS) presentations than did pre-exposed animals (P< 0.05). The improvement in retention produced by novelty was attenuated by pretraining a blockade of peripheral β-adrenergic receptors with sotalol (6 mg/kg, i.p.). Study 2 revealed that novelty-induced increases in peripheral autonomic output are conveyed to the brain by visceral afferents that synapse upon brainstem neurons in the nucleus tractus solitarius (NTS). Blocking AMPA receptor activity in the NTS with CNQX (1.0 μg) significantly reduced freezing to the CS in non-pre-exposed animals (P< 0.01). Study 3 showed that elevating epinephrine levels in habituated animals influences learning through mechanisms similar to those produced by novelty-induced arousal. Pre-exposed animals given epinephrine (0.1 mg/kg) froze significantly more than saline controls (P< 0.01), and this effect was attenuated by intra-NTS infusion of CNQX. The findings demonstrate that novelty-induced arousal or increasing sympathetic activity with epinephrine in pre-exposed animals enhances memory through adrenergic mechanisms initiated in the periphery and transmitted centrally via the vagus/NTS complex.
Maize (Zea mays): A Model Organism for Basic and Applied Research in Plant Biology
Authors: Strable Josh , Scanlon Michael J.
Published: October 2009

INTRODUCTION

Zea mays ssp. mays is one of the world’s most important crop plants, boasting a multibillion dollar annual revenue. In addition to its agronomic importance, maize has been a keystone model organism for basic research for nearly a century. Within the cereals, which include other plant model species such as rice (Oryza sativa), sorghum (Sorghum bicolor), wheat (Triticum spp.), and barley (Hordeum vulgare), maize is the most thoroughly researched genetic system. Several attributes of the maize plant, including a vast collection of mutant stocks, large heterochromatic chromosomes, extensive nucleotide diversity, and genic colinearity within related grasses, have positioned this species as a centerpiece for genetic, cytogenetic, and genomic research. As a model organism, maize is the subject of such far-ranging biological investigations as plant domestication, genome evolution, developmental physiology, epigenetics, pest resistance, heterosis, quantitative inheritance, and comparative genomics. These and other studies will be advanced by the completed sequencing and annotation of the maize gene space, which will be realized during 2009. Here we present an overview of the use of maize as a model system and provide links to several protocols that enable its genetic and genomic analysis.
High-Resolution Dual-Trap Optical Tweezers with Differential Detection: Instrument Design
Authors: Bustamante Carlos , Chemla Yann R. , Moffitt Jeffrey R.
Published: October 2009

INTRODUCTION

Optical traps or “optical tweezers” have become an indispensable tool in understanding fundamental biological processes. In this article, we present the design of a high-resolution optical tweezers, which is designed to minimize sensitivity to environmental noise. When the tether is attached to a stage, resolution is typically limited by drift and fluctuations in the relative position of the stage and trap-forming objective, cued by environmental noise. In a dual-trap design, however, the biological system is decoupled entirely from the stage by utilizing two optical traps to hold both ends of the tether. By holding each end of the molecule in an identical fashion, such dual-trap optical tweezers are less sensitive to drift and fluctuations of mechanical components. In our instrument, we form two traps from two orthogonally polarized beams generated by a single laser. As a result, many optical components and much of the optical path traveled are common to both trapping beams, thus minimizing differential movements of one optical trap relative to the other. Additionally, this economical use of optics creates an instrument that is simple and compact, reducing the number of elements susceptible to environmental fluctuations.
Phenotypic Analysis of Arabidopsis Mutants: Oomycete Pathogens
Authors: Clarke Joseph D.
Published: October 2009

INTRODUCTION

Various fungal pathogens are used in Arabidopsis pathogen studies, including Fusarium oxysporum, Alternaria brassicicola, Botrytis cinerea, and others. The oomycete pathogen Peronospora parasitica has been used by several groups and is described in this protocol. Working with Peronospora is complicated by the fact that it is an obligate biotroph, and consequently cultures must be maintained on living plants. There is no central repository for Peronospora stocks, but most investigators who work with them are willing to provide samples of infected tissue. These can be used to initiate new stock cultures, or they can be maintained as live cultures on seedlings. One of the most important factors in maintaining Peronospora is the humidity of the growth chamber, which must be kept at a minimum of 80%. Various Peronospora isolates are available. These vary with respect to which Arabidopsis ecotypes they can infect, because some combinations trigger gene-for-gene resistance. Thus, it is important that the appropriate ecotype is inoculated with the appropriate strain of pathogen. The extent of infections can be rated or quantitatively measured as the number of spores produced per plant, and frozen tissue stocks can be prepared from heavily infected tissue.
Fixation and Paraffin-Embedding of Mouse Tissues for GFP Visualization
Authors: Vankelecom Hugo
Published: October 2009

INTRODUCTION

Green fluorescent protein (GFP) and its variants are used in many applications, including protein localization, high-throughput screening, and gene discovery. Transgenes that use GFP-based proteins as a reporter provide a convenient way to follow protein expression over the course of development and disease progression in the mouse. The basic principle of visualizing fluorescent proteins is the same as for the fluorophores used in fluorescent microscopy. This protocol describes a method for fixation and paraffin-embedding of fresh mouse embryos or tissue for use in GFP microscopy.
Analysis of In Vivo Protein Complexes by Coimmunoprecipitation from Caenorhabditis elegans
Authors: Jedamzik Britta , Eckmann Christian R.
Published: October 2009

INTRODUCTION

Protein coimmunoprecipitation (co-IP) is a method used to analyze in vivo complex formation of various proteins. Although such an analysis supports the coexistence of proteins in a complex, a direct protein-protein interaction cannot be concluded unless further in vitro data are available. This protocol describes how to perform co-IPs from C. elegans whole-worm extracts using protein-specific antibodies. First, we describe how to culture a large number of worms while maintaining their overall appearance and wild-type fertility rates, which are important factors when analyzing the germline tissue. Next, we present a gentle and effective method to generate worm extracts with high protein concentrations that maintain protein complexes of high quality. Finally, we describe how to purify the protein of choice along with its associated complex members. The precipitated protein complex can be analyzed by either immunoblot analysis or mass spectrometry to identify the copurified protein components. When working with RNA-binding proteins, it is of interest to assess whether RNA molecules, rather than a direct interaction between the proteins, might mediate complex formation. For this purpose, an optional RNase digestion step to degrade the RNA in the extract is described.
Analysis of RNA-Protein Complexes by RNA Coimmunoprecipitation and RT-PCR Analysis from Caenorhabditis elegans
Authors: Jedamzik Britta , Eckmann Christian R.
Published: October 2009

INTRODUCTION

RNA coimmunoprecipitation (co-IP) experiments are an extension of protein co-IP experiments in which in vivo RNA-protein complexes are investigated. This protocol describes how to perform RNA co-IPs from C. elegans whole-worm extracts. In principle, a protein-specific antibody is used to purify the protein of choice and its associated complex members from worm extract. This may also include RNA molecules associated with other protein components. To identify a specific mRNA molecule, all RNA molecules are first separated from the protein components after immunopurification. The mRNAs are then converted into cDNA by reverse transcription. Candidate mRNAs are detected by sensitive gene-specific amplification via polymerase chain reaction (PCR) in a semiquantitative manner. Since RNA molecules are very prone to degradation, it is crucial to avoid any kind of contamination with RNase activity in this experiment.
In Situ Hybridization in the Plant Kalanchoë daigremontiana
Authors: Garcês Helena , Sinha Neelima
Published: October 2009

INTRODUCTION

Here we describe in detail the detection of gene expression in plant tissues of Kalanchoë daigremontiana by in situ hybridization analyses. Included are methods for making RNA transcript probes, probe-tissue hybridization, and detection of antisense RNA probes. The in situ hybridization technique is used to determine which cells or group of cells in particular tissue(s) express a gene of interest.
Strain Maintenance of Nasonia vitripennis (Parasitoid Wasp)
Authors: Werren John H. , Loehlin David W.
Published: October 2009

INTRODUCTION

Nasonia is a complex of four closely related species of wasps with several features that make it an excellent system for a variety of genetic studies. These include a short generation time, ease of rearing, interfertile species, visible and molecular markers, and a sequenced genome. Furthermore, its parasitoid lifestyle allows investigations of questions relating to parasitoid/host dynamics, host preference, and specialist versus generalist biology. It also can serve as a behavior model for studies of courtship, male aggression and territoriality, female dispersal, and sex ratio control. This protocol describes standard rearing of Nasonia vitripennis strains on Sarcophaga bullata hosts.
Egg Collection for Nasonia (Parasitoid Wasp)
Authors: Werren John H. , Loehlin David W.
Published: October 2009

INTRODUCTION

Nasonia is a complex of four closely related species of wasps with several features that make it an excellent system for a variety of genetic studies. These include a short generation time, ease of rearing, interfertile species, visible and molecular markers, and a sequenced genome. Furthermore, its parasitoid lifestyle allows investigations of questions relating to parasitoid/host dynamics, host preference, and specialist versus generalist biology. It also can serve as a behavior model for studies of courtship, male aggression and territoriality, female dispersal, and sex ratio control. This protocol describes methods to count and/or collect Nasonia eggs. Fly hosts are placed in a foam plug such that only one oviposition site is available to the female wasp.
Virgin Collection and Haplodiploid Crossing Methods in Nasonia (Parasitoid Wasp): Figure 1.
Authors: Werren John H. , Loehlin David W.
Published: October 2009

INTRODUCTION

This protocol covers most procedures used by the Nasonia (wasp) geneticist, including virgin collection. Nasonia are easily sexed as pupae, so collecting virgins can easily be performed within a window of several days. The female-specific ovipositor is visible at even the earliest pupal stages as a longitudinal ridge on the ventral posterior abdomen. To prolong generations, yellow pupae can be stored at 4°C-8°C for up to 2 mo and adults for 2-3 wk. In the example presented here, haplodiploid crossing methods are used to make a new Nasonia vitripennis mutant homozygous, using an aunt/nephew mating approach. The protocol can be extended to inbreeding strains. Inbreeding depression generally is not a problem in Nasonia unless the mutation itself is female-sterile. Mother/son mating and sib mating are also reasonable approaches to inbreeding.
Curing Wolbachia Infections in Nasonia (Parasitoid Wasp)
Authors: Werren John H. , Loehlin David W.
Published: October 2009

INTRODUCTION

Nasonia is a complex of four closely related species of wasps with several features that make it an excellent system for a variety of genetic studies. These include a short generation time, ease of rearing, interfertile species, visible and molecular markers, and a sequenced genome. However, the Nasonia species normally are completely or partially reproductively isolated owing to Wolbachia, widespread intracellular bacteria found in arthropods and nematodes. Wolbachia cause cytoplasmic incompatibility (CI) in Nasonia. Although the current genome-sequenced strains of Nasonia (AsymCX, IV7[u], RV2X[u]) are all Wolbachia-free, crosses between infected and uninfected wasps, or between infected wasps of different species, will fail because of Wolbachia. The sperm modification that induces CI appears to be established before the adult stage, so it is generally not possible to cure an infection and break the CI barrier in the same generation. However, a Wolbachia infection can be removed from a strain over a number of generations by feeding antibiotics and selecting on females showing partial CI. This protocol describes the methods to cure a Nasonia strain of its Wolbachia symbionts.
Field Collection of Nasonia (Parasitoid Wasp) Using Baits
Authors: Werren John H. , Loehlin David W.
Published: October 2009

INTRODUCTION

This protocol describes a standard method for collecting Nasonia wasps using “flyliver”: liver remains fed upon by fly maggots (e.g., Sarcophaga) and collected after the mature larvae have dispersed. The flyliver, which contains a volatile substance that is very attractive to the wasps, is placed in a large (~15-cm2) mesh bag hung in appropriate collection spots (e.g., near birds’ nests or under culverts). Within the large mesh bag is a smaller mesh bag placed abutting the flyliver and containing four to six Sarcophaga pupae. These retain the wasps, which will enter the bag and begin stinging the hosts. The mesh bags can be made with standard nylon window screening, or any other material with mesh width large enough to permit entry of the wasps.
High-Resolution Dual-Trap Optical Tweezers with Differential Detection: An Introduction: Figure 1.
Authors: Bustamante Carlos , Chemla Yann R. , Moffitt Jeffrey R.
Published: October 2009

INTRODUCTION

Optical traps or “optical tweezers” have become an indispensable tool in understanding fundamental biological processes. The ability to manipulate and probe individual molecules or molecular complexes has led to a new, more refined understanding of the mechanical properties of the fundamental building blocks of the cell, and of the mechanism by which molecular machines function. The field has seen a steady stream of technological advances that have greatly refined the technique. One major effort has been in developing methods to resolve motions at the angstrom level--the fundamental length scale for many biological processes. This drive has only recently come to fruition with the advent of high-resolution optical trapping techniques that can now detect movements on the scale of a single base pair of DNA, 3.4 Å. Here we briefly review the basic concepts and components of optical traps and the single-molecule experiments in which they are used.
Mechanical Stimuli Modulate Lateral Root Organogenesis
Authors: Richter Gregory L. , Monshausen Gabriele B. , Krol Alexandra , Gilroy Simon
Published: October 2009

Unlike mammals, whose development is limited to a short temporal window, plants produce organs de novo throughout their lifetime in order to adapt their architecture to the prevailing environmental conditions. The production of lateral roots represents one example of such postembryonic organogenesis. An endogenous developmental program likely imposes an ordered arrangement on the position of new lateral roots. However, environmental stimuli such as nutrient levels also affect the patterning of lateral root production. In addition, we have found that mechanical forces can act as one of the triggers that entrain lateral root production to the environment of the Arabidopsis (Arabidopsis thaliana) plant. We observed that physical bending of the root recruited new lateral root formation to the convex side of the resultant bend. Transient bending of 20 s was sufficient to elicit this developmental program. Such bending triggered a Ca2+ transient within the pericycle, and blocking this change in Ca2+ also blocked recruitment of new lateral root production to the curved region of the root. The initial establishment of the mechanically induced lateral root primordium was independent of an auxin supply from the shoot and was not disrupted by mutants in a suite of auxin transporters and receptor/response elements. These results suggest that Ca2+ may be acting to translate the mechanical forces inherent in growth to a developmental response in roots.