Cloning, expression and characterization of an ethanol tolerant GH3 β-glucosidase from Myceliophthora thermophila
The β-glucosidase gene bgl3a from Myceliophthora thermophila, member of the fungal glycosyl hydrolase (GH) family 3, was cloned and expressed in Pichia pastoris. The mature β-glucosidase gene, which results after the excision of one intron and the secreting signal peptide, was placed under the control of the strong alcohol oxidase promoter (AOX1) in the plasmid pPICZαC. The recombinant enzyme (90 kDa) was purified and characterized in order to evaluate its biotechnological potential. Recombinant P. pastoris efficiently secreted β-glucosidase into the medium and produced high level of enzymatic activity (41 U/ml) after 192 h of growth, under methanol induction. MtBgl3a was able to hydrolyze low molecular weight substrates and polysaccharides containing β-glucosidic residues. The Km was found to be 0.39 mM on p-β-NPG and 2.64 mM on cellobiose. Optimal pH and temperature for the p-β-NPG hydrolysis were 5.0 and 70 °C. The β-glucosidase exhibits a half life of 143 min at 60 °C. Kinetic parameters of inhibition were determined for D-glucose, D-xylose and D-gluconic acid, indicating tolerance of the enzyme for these sugars and oxidized products. The recombinant enzyme was stimulated by short chain alcohols and has been shown to efficiently synthesize methyl-D-glucoside in the presence of methanol due to its transglycosylation activity. The stability of MtBgl3a in ethanol was prominent, and it retained most of its original activity after we exposed it to 50% ethanol for 6 h. The high catalytic performance, good thermal stability and tolerance to elevated concentrations of ethanol, D-xylose and D-glucose qualify this enzyme for use in the hydrolysis of lignocellulosic biomass for biofuel production, as part of an efficient complete multi-enzyme cocktail.
Karnaouri et al. (2013) Cloning, expression and characterization of an ethanol tolerant GH3 β-glucosidase from Myceliophthora thermophila. PeerJ 1:e46 https://doi.org/10.7717/peerj.46
The co-evolution of multiply-informed dispersal: information transfer across landscapes from neighbors and immigrants
Dispersal plays a key role in natural systems by shaping spatial population and evolutionary dynamics. Dispersal has been largely treated as a population process with little attention to individual decisions and the influence of information use on the fitness benefits of dispersal despite clear empirical evidence that dispersal behavior varies among individuals. While information on local density is common, more controversial is the notion that indirect information use can easily evolve. We used an individual-based model to ask under what conditions indirect information use in dispersal will evolve. We modeled indirect information provided by immigrant arrival into a population which should be linked to overall metapopulation density. We also modeled direct information use of density which directly impacts fitness. We show that immigrant-dependent dispersal evolves and does so even when density dependent information is available. Use of two sources of information also provides benefits at the metapopulation level by reducing extinction risk and prolonging the persistence of populations. Our results suggest that use of indirect information in dispersal can evolve under conservative conditions and thus could be widespread.
Chaine et al. (2013) The co-evolution of multiply-informed dispersal: information transfer across landscapes from neighbors and immigrants. PeerJ 1:e44 https://doi.org/10.7717/peerj.44
Pattern classification of brain activation during emotional processing in subclinical depression: psychosis proneness as potential confounding factor
We used Support Vector Machine (SVM) to perform multivariate pattern classification based on brain activation during emotional processing in healthy participants with subclinical depressive symptoms. Six-hundred undergraduate students completed the Beck Depression Inventory II (BDI-II). Two groups were subsequently formed: (i) subclinical (mild) mood disturbance (n = 17) and (ii) no mood disturbance (n = 17). Participants also completed a self-report questionnaire on subclinical psychotic symptoms, the Community Assessment of Psychic Experiences Questionnaire (CAPE) positive subscale. The functional magnetic resonance imaging (fMRI) paradigm entailed passive viewing of negative emotional and neutral scenes. The pattern of brain activity during emotional processing allowed correct group classification with an overall accuracy of 77% (p = 0.002), within a network of regions including the amygdala, insula, anterior cingulate cortex and medial prefrontal cortex. However, further analysis suggested that the classification accuracy could also be explained by subclinical psychotic symptom scores (correlation with SVM weights r = 0.459, p = 0.006). Psychosis proneness may thus be a confounding factor for neuroimaging studies in subclinical depression.
Modinos et al. (2013) Pattern classification of brain activation during emotional processing in subclinical depression: psychosis proneness as potential confounding factor. PeerJ 1:e42 https://doi.org/10.7717/peerj.42
Does haemosporidian infection affect hematological and biochemical profiles of the endangered Black-fronted piping-guan (Aburria jacutinga)?
Infectious diseases can cause deleterious effects on bird species, leading to population decline and extinction. Haemosporidia can be recognized by their negative effects on host fitness, including reproductive success and immune responses. In captivity, outbreaks of haemosporidian infection have been observed in birds in zoos and aviaries. The endemic Brazilian Atlantic rainforest species Aburria jacutinga is one of the most endangered species in the Cracidae family, and wild populations of this species are currently found mainly in conservation areas in only two Brazilian states. In this study, we aimed to evaluate the effects of avian haemosporidia on hematological and biochemical parameters in two captive populations of A. jacutinga. Forty-two animals were assessed, and the haemosporidian prevalence was similar for males and females. The occurrence of haemosporidian infection in captive A. jacutinga observed in this study was similar to results found in other captive and wild birds in Brazil. We found three different lineages of haemosporidia. Two lineages were identified as Plasmodium sp., one of which was previously detected in Europe and Asia, and the other is a new lineage closely related to P. gallinaceum. A new third lineage was identified as Haemoproteus sp. We found no significant differences in hematological and biochemical values between infected and non-infected birds, and the haemosporidian lineage did not seem to have an impact on the clinical and physiological parameters of A. jacutinga. This is the first report on an evaluation of natural haemosporidian infections diagnosed by microscopic and molecular methods in A. jacutinga by hematology, blood biochemistry, and serum protein values. Determining physiological parameters, occurrence and an estimation of the impact of haemosporidia in endangered avian species may contribute to the management of species rehabilitation and conservation.
Motta et al. (2013) Does haemosporidian infection affect hematological and biochemical profiles of the endangered Black-fronted piping-guan (Aburria jacutinga)?. PeerJ 1:e45 https://doi.org/10.7717/peerj.45
Development of guidelines for tertiary education institutions to assist them in supporting students with a mental illness: a Delphi consensus study with Australian professionals and consumers
Background. The age at which most young people are in tertiary education is also the age of peak onset for mental illness. Because mental health problems can have adverse effects on students’ academic performance and welfare, institutions require guidance how they can best provide support. However, the scientific evidence for how best to do this is relatively limited. Therefore a Delphi expert consensus study was carried out with professional and consumer experts. Methods. A systematic review of websites, books and journal articles was conducted to develop a 172 item survey containing strategies that institutions might use to support students with a mental illness. Two panels of Australian experts (74 professionals and 35 consumers) were recruited and independently rated the items over three rounds, with strategies reaching consensus on importance written into the guidelines. Results. The overall response rate across three rounds was 83% (80% consumers, 85% professionals). 155 strategies were endorsed as essential or important by at least 80% of panel members. The endorsed strategies provided information on policy, measures to promote support services, service provision, accessibility of support services, relationships between services, other types of support and issues associated with reasonable adjustments. They also provided guidance on the procedures the institutions should have for making staff aware of issues associated with mental illness, mental illness training, support for staff and communicating with a student with a mental illness. They also covered student rights and responsibilities, the procedures the institutions should have for making students aware of issues associated with mental illness, dealing with mental health crises, funding and research and evaluation. Conclusions. The guidelines provide guidance for tertiary institutions to assist them in supporting students with a mental illness. It is hoped that they may be used to inform policy and practice in tertiary institutions.
Reavley et al. (2013) Development of guidelines for tertiary education institutions to assist them in supporting students with a mental illness: a Delphi consensus study with Australian professionals and consumers. PeerJ 1:e43 https://doi.org/10.7717/peerj.43
Associations between the human intestinal microbiota, Lactobacillus rhamnosus GG and serum lipids indicated by integrated analysis of high-throughput profiling data
Accumulating evidence indicates that the intestinal microbiota regulates our physiology and metabolism. Bacteria marketed as probiotics confer health benefits that may arise from their ability to affect the microbiota. Here high-throughput screening of the intestinal microbiota was carried out and integrated with serum lipidomic profiling data to study the impact of probiotic intervention on the intestinal ecosystem, and to explore the associations between the intestinal bacteria and serum lipids. We performed a comprehensive intestinal microbiota analysis using a phylogenetic microarray before and after Lactobacillus rhamnosus GG intervention. While a specific increase in the L. rhamnosus-related bacteria was observed during the intervention, no other changes in the composition or stability of the microbiota were detected. After the intervention, lactobacilli returned to their initial levels. As previously reported, also the serum lipid profiles remained unaltered during the intervention. Based on a high-resolution microbiota analysis, intake of L. rhamnosus GG did not modify the composition of the intestinal ecosystem in healthy adults, indicating that probiotics confer their health effects by other mechanisms. The most prevailing association between the gut microbiota and lipid profiles was a strong positive correlation between uncultured phylotypes of Ruminococcus gnavus-group and polyunsaturated serum triglycerides of dietary origin. Moreover, a positive correlation was detected between serum cholesterol and Collinsella (Coriobacteriaceae). These associations identified with the spectrometric lipidome profiling were corroborated by enzymatically determined cholesterol and triglyceride levels. Actinomycetaceae correlated negatively with triglycerides of highly unsaturated fatty acids while a set of Proteobacteria showed negative correlation with ether phosphatidylcholines. Our results suggest that several members of the Firmicutes, Actinobacteria and Proteobacteria may be involved in the metabolism of dietary and endogenous lipids, and provide a scientific rationale for further human studies to explore the role of intestinal microbes in host lipid metabolism.
Lahti et al. (2013) Associations between the human intestinal microbiota, Lactobacillus rhamnosus GG and serum lipids indicated by integrated analysis of high-throughput profiling data. PeerJ 1:e32 https://doi.org/10.7717/peerj.32
Foundation species loss affects vegetation structure more than ecosystem function in a northeastern USA forest
Loss of foundation tree species rapidly alters ecological processes in forested ecosystems. Tsuga canadensis, an hypothesized foundation species of eastern North American forests, is declining throughout much of its range due to infestation by the nonnative insect Adelges tsugae and by removal through pre-emptive salvage logging. In replicate 0.81-ha plots, T. canadensis was cut and removed, or killed in place by girdling to simulate adelgid damage. Control plots included undisturbed hemlock and mid-successional hardwood stands that represent expected forest composition in 50–100 years. Vegetation richness, understory vegetation cover, soil carbon flux, and nitrogen cycling were measured for two years prior to, and five years following, application of experimental treatments. Litterfall and coarse woody debris (CWD), including snags, stumps, and fallen logs and branches, have been measured since treatments were applied. Overstory basal area was reduced 60%–70% in girdled and logged plots. Mean cover and richness did not change in hardwood or hemlock control plots but increased rapidly in girdled and logged plots. Following logging, litterfall immediately decreased then slowly increased, whereas in girdled plots, there was a short pulse of hemlock litterfall as trees died. CWD volume remained relatively constant throughout but was 3–4× higher in logged plots. Logging and girdling resulted in small, short-term changes in ecosystem dynamics due to rapid regrowth of vegetation but in general, interannual variability exceeded differences among treatments. Soil carbon flux in girdled plots showed the strongest response: 35% lower than controls after three years and slowly increasing thereafter. Ammonium availability increased immediately after logging and two years after girdling, due to increased light and soil temperatures and nutrient pulses from leaf-fall and reduced uptake following tree death. The results from this study illuminate ecological processes underlying patterns observed consistently in region-wide studies of adelgid-infested hemlock stands. Mechanisms of T. canadensis loss determine rates, magnitudes, and trajectories of ecological changes in hemlock forests. Logging causes abrupt, large changes in vegetation structure whereas girdling (and by inference, A. tsugae) causes sustained, smaller changes. Ecosystem processes depend more on vegetation cover per se than on species composition. We conclude that the loss of this late-successional foundation species will have long-lasting impacts on forest structure but subtle impacts on ecosystem function.
Orwig et al. (2013) Foundation species loss affects vegetation structure more than ecosystem function in a northeastern USA forest. PeerJ 1:e41 https://doi.org/10.7717/peerj.41
A novel control of human keratin expression: cannabinoid receptor 1-mediated signaling down-regulates the expression of keratins K6 and K16 in human keratinocytes in vitro and in situ
Cannabinoid receptors (CB) are expressed throughout human skin epithelium. CB1 activation inhibits human hair growth and decreases proliferation of epidermal keratinocytes. Since psoriasis is a chronic hyperproliferative, inflammatory skin disease, it is conceivable that the therapeutic modulation of CB signaling, which can inhibit both proliferation and inflammation, could win a place in future psoriasis management. Given that psoriasis is characterized by up-regulation of keratins K6 and K16, we have investigated whether CB1 stimulation modulates their expression in human epidermis. Treatment of organ-cultured human skin with the CB1-specific agonist, arachidonoyl-chloro-ethanolamide (ACEA), decreased K6 and K16 staining intensity in situ. At the gene and protein levels, ACEA also decreased K6 expression of cultured HaCaT keratinocytes, which show some similarities to psoriatic keratinocytes. These effects were partly antagonized by the CB1-specific antagonist, AM251. While CB1-mediated signaling also significantly inhibited human epidermal keratinocyte proliferation in situ, as shown by K6/Ki-67-double immunofluorescence, the inhibitory effect of ACEA on K6 expression in situ was independent of its anti-proliferative effect. Given recent appreciation of the role of K6 as a functionally important protein that regulates epithelial wound healing in mice, it is conceivable that the novel CB1-mediated regulation of keratin 6/16 revealed here also is relevant to wound healing. Taken together, our results suggest that cannabinoids and their receptors constitute a novel, clinically relevant control element of human K6 and K16 expression.
Ramot et al. (2013) A novel control of human keratin expression: cannabinoid receptor 1-mediated signaling down-regulates the expression of keratins K6 and K16 in human keratinocytes in vitro and in situ. PeerJ 1:e40 https://doi.org/10.7717/peerj.40
Personality change after Internet-delivered cognitive behavior therapy for depression
Background. The Temperament and Character Inventory (TCI) by Cloninger is a widely used instrument to measure personality dimensions. Two dimensions of the TCI, Harm avoidance (HA) and Self-Directedness (SD), are known to be influenced by depressed mood. This study investigated changes in HA and SD after 10 weeks of Internet-delivered cognitive behavior therapy (ICBT) in a sample of clinically depressed subjects (N = 108). Differences in personality changes among treatment responders and non-responders were also investigated. Exploratory investigations on changes for other TCI dimensions, were also conducted. Methods. Depressed subjects were randomized either to ICBT or to a moderated online discussion group, which served as an active control group. The interventions lasted for 10 weeks. TCI was measured at baseline and after treatment. Depressive symptoms were assessed using the Beck Depression Inventory-II. Results. There were significant changes on HA and SD after ICBT. However, when comparing post-treatment HA and SD to the control, no differences were found. Among responders, larger changes compared to non-responders were found in HA and in SD, as well as in Cooperativeness. Conclusions. The study showed that HA and SD changed after ICBT. The changes in personality seem related to improvement in depression rather than a direct effect of ICBT.
Johansson et al. (2013) Personality change after Internet-delivered cognitive behavior therapy for depression. PeerJ 1:e39 https://doi.org/10.7717/peerj.39
Phenotypic characterization of patients with deletions in the 3’-flanking SHOX region
Context. Leri–Weill dyschondrosteosis is a clinically variable skeletal dysplasia, caused by SHOX deletion or mutations, or a deletion of enhancer sequences in the 3’-flanking region. Recently, a 47.5 kb recurrent PAR1 deletion downstream of SHOX was reported, but its frequency and clinical importance are still unknown. Objective. This study aims to compare the clinical features of different sizes of deletions in the 3’-flanking SHOX region in order to determine the relevance of the regulatory sequences in this region. Design. We collected DNA from 28 families with deletions in the 3’-PAR1 region. Clinical data were available from 23 index patients and 21 relatives. Results. In 9 families (20 individuals) a large deletion ( ∼ 200–900 kb) was found and in 19 families (35 individuals) a small deletion was demonstrated, equal to the recently described 47.5 kb PAR1 deletion. Median height SDS, sitting height/height ratio SDS and the presence of Madelung deformity in patients with the 47.5 kb deletion were not significantly different from patients with larger deletions. The index patients had a median height SDS which was slightly lower than in their affected family members (p = 0.08). No significant differences were observed between male and female patients. Conclusions. The phenotype of patients with deletions in the 3’-PAR1 region is remarkably variable. Height, sitting height/height ratio and the presence of Madelung deformity were not significantly different between patients with the 47.5 kb recurrent PAR1 deletion and those with larger deletions, suggesting that this enhancer plays an important role in SHOX expression.
Kant et al. (2013) Phenotypic characterization of patients with deletions in the 3’-flanking SHOX region. PeerJ 1:e35 https://doi.org/10.7717/peerj.35
Validation of the Emotiv EPOC® EEG gaming system for measuring research quality auditory ERPs
Background. Auditory event-related potentials (ERPs) have proved useful in investigating the role of auditory processing in cognitive disorders such as developmental dyslexia, specific language impairment (SLI), attention deficit hyperactivity disorder (ADHD), schizophrenia, and autism. However, laboratory recordings of auditory ERPs can be lengthy, uncomfortable, or threatening for some participants – particularly children. Recently, a commercial gaming electroencephalography (EEG) system has been developed that is portable, inexpensive, and easy to set up. In this study we tested if auditory ERPs measured using a gaming EEG system (Emotiv EPOC®, www.emotiv.com) were equivalent to those measured by a widely-used, laboratory-based, research EEG system (Neuroscan). Methods. We simultaneously recorded EEGs with the research and gaming EEG systems, whilst presenting 21 adults with 566 standard (1000 Hz) and 100 deviant (1200 Hz) tones under passive (non-attended) and active (attended) conditions. The onset of each tone was marked in the EEGs using a parallel port pulse (Neuroscan) or a stimulus-generated electrical pulse injected into the O1 and O2 channels (Emotiv EPOC®). These markers were used to calculate research and gaming EEG system late auditory ERPs (P1, N1, P2, N2, and P3 peaks) and the mismatch negativity (MMN) in active and passive listening conditions for each participant. Results. Analyses were restricted to frontal sites as these are most commonly reported in auditory ERP research. Intra-class correlations (ICCs) indicated that the morphology of the research and gaming EEG system late auditory ERP waveforms were similar across all participants, but that the research and gaming EEG system MMN waveforms were only similar for participants with non-noisy MMN waveforms (N = 11 out of 21). Peak amplitude and latency measures revealed no significant differences between the size or the timing of the auditory P1, N1, P2, N2, P3, and MMN peaks. Conclusions. Our findings suggest that the gaming EEG system may prove a valid alternative to laboratory ERP systems for recording reliable late auditory ERPs (P1, N1, P2, N2, and the P3) over the frontal cortices. In the future, the gaming EEG system may also prove useful for measuring less reliable ERPs, such as the MMN, if the reliability of such ERPs can be boosted to the same level as late auditory ERPs.
Badcock et al. (2013) Validation of the Emotiv EPOC® EEG gaming system for measuring research quality auditory ERPs. PeerJ 1:e38 https://doi.org/10.7717/peerj.38
Development and evolution of the unique cetacean dentition
The evolutionary success of mammals is rooted in their high metabolic rate. A high metabolic rate is sustainable thanks to efficient food processing and that in turn is facilitated by precise occlusion of the teeth and the acquisition of rhythmic mastication. These major evolutionary innovations characterize most members of the Class Mammalia. Cetaceans are one of the few groups of mammals in which precise occlusion has been secondarily lost. Most toothed whales have an increased number of simple crowned teeth that are similar along the tooth row. Evolution toward these specializations began immediately after the time cetaceans transitioned from terrestrial-to-marine environments. The fossil record documents the critical aspects of occlusal evolution of cetaceans, and allows us to pinpoint the evolutionary timing of the macroevolutionary events leading to their unusual dental morphology among mammals. The developmental controls of tooth differentiation and tooth number have been studied in a few mammalian clades, but nothing is known about how these controls differ between cetaceans and mammals that retain functional occlusion. Here we show that pigs, a cetacean relative with regionalized tooth morphology and complex tooth crowns, retain the typical mammalian gene expression patterns that control early tooth differentiation, expressing Bmp4 in the rostral (mesial, anterior) domain of the jaw, and Fgf8 caudally (distal, posterior). By contrast, dolphins have lost these regional differences in dental morphology and the Bmp4 domain is extended into the caudal region of the developing jaw. We hypothesize that the functional constraints underlying mammalian occlusion have been released in cetaceans, facilitating changes in the genetic control of early dental development. Such major developmental changes drive morphological evolution and are correlated with major shifts in diet and food processing during cetacean evolution.
Armfield et al. (2013) Development and evolution of the unique cetacean dentition. PeerJ 1:e24 https://doi.org/10.7717/peerj.24
A metagenomic insight into freshwater methane-utilizing communities and evidence for cooperation between the Methylococcaceae and the Methylophilaceae
We investigated microbial communities active in methane oxidation in lake sediment at different oxygen tensions and their response to the addition of nitrate, via stable isotope probing combined with deep metagenomic sequencing. Communities from a total of four manipulated microcosms were analyzed, supplied with 13C-methane in, respectively, ambient air, ambient air with the addition of nitrate, nitrogen atmosphere and nitrogen atmosphere with the addition of nitrate, and these were compared to the community from an unamended sediment sample. We found that the major group involved in methane oxidation in both aerobic and microaerobic conditions were members of the family Methylococcaceae, dominated by species of the genus Methylobacter, and these were stimulated by nitrate in aerobic but not microaerobic conditions. In aerobic conditions, we also noted a pronounced response to both methane and nitrate by members of the family Methylophilaceae that are non-methane-oxidizing methylotrophs, and predominantly by the members of the genus Methylotenera. The relevant abundances of the Methylococcaceae and the Methylophilaceae and their coordinated response to methane and nitrate suggest that these species may be engaged in cooperative behavior, the nature of which remains unknown.
Beck et al. (2013) A metagenomic insight into freshwater methane-utilizing communities and evidence for cooperation between the Methylococcaceae and the Methylophilaceae. PeerJ 1:e23 https://doi.org/10.7717/peerj.23
Iodine nutrition and toxicity in Atlantic cod (Gadus morhua) larvae
Copepods as feed promote better growth and development in marine fish larvae than rotifers. However, unlike rotifers, copepods contain several minerals such as iodine (I), at potentially toxic levels. Iodine is an essential trace element and both under and over supply of I can inhibit the production of the I containing thyroid hormones. It is unknown whether marine fish larvae require copepod levels of I or if mechanisms are present that prevent I toxicity. In this study, larval Atlantic cod (Gadus morhua) were fed rotifers enriched to intermediate (26 mg I kg-1 dry weight; MI group) or copepod (129 mg I kg-1 DW; HI group) I levels and compared to cod larvae fed control rotifers (0.6 mg I kg-1 DW). Larval I concentrations were increased by 3 (MI) and 7 (HI) fold compared to controls during the rotifer feeding period. No differences in growth were observed, but the HI diet increased thyroid follicle colloid to epithelium ratios, and affected the essential element concentrations of larvae compared to the other groups. The thyroid follicle morphology in the HI larvae is typical of colloid goitre, a condition resulting from excessive I intake, even though whole body I levels were below those found previously in copepod fed cod larvae. This is the first observation of dietary induced I toxicity in fish, and suggests I toxicity may be determined to a greater extent by bioavailability and nutrient interactions than by total body I concentrations in fish larvae. Rotifers with 0.6 mg I kg-1 DW appeared sufficient to prevent gross signs of I deficiency in cod larvae reared with continuous water exchange, while modelling of cod larvae versus rotifer I levels suggests that optimum I levels in rotifers for cod larvae is 3.5 mg I kg-1 DW.
Penglase et al. (2013) Iodine nutrition and toxicity in Atlantic cod (Gadus morhua) larvae. PeerJ 1:e20 https://doi.org/10.7717/peerj.20
Evaluation of animal control measures on pet demographics in Santa Clara County, California, 1993–2006
The measurable benefits of animal control programs are unknown and the aim of this study was to determine the impact of these programs on pet population changes. A prospective cross-sectional study of 1000 households was implemented in 2005 to evaluate characteristics of the owned and unowned population of dogs and cats in Santa Clara County, California. The same population was previously studied 12 years earlier. During this time period, the county instituted in 1994 and then subsequently disestablished a municipal spay/neuter voucher program for cats. Dog intakes declined from 1992–2005, as they similarly did for an adjacent county (San Mateo). However, cat intakes declined significantly more in Santa Clara County than San Mateo, with an average annual decline of approximately 700 cats for the 12 year period. Time series analysis showed a greater than expected decline in the number of cats surrendered to shelters in Santa Clara County during the years the voucher program was in effect (1994–2005). The net savings to the county by reducing the number of cat shelter intakes was estimated at approximately $1.5 million. The measurable benefits of animal control programs are unknown and the aim of this study was to determine the impact of these programs on pet population changes.
Kass et al. (2013) Evaluation of animal control measures on pet demographics in Santa Clara County, California, 1993–2006. PeerJ 1:e18 https://doi.org/10.7717/peerj.18
Loss of CclA, required for histone 3 lysine 4 methylation, decreases growth but increases secondary metabolite production in Aspergillus fumigatus
Secondary metabolite (SM) production in filamentous fungi is mechanistically associated with chromatin remodeling of specific SM clusters. One locus recently shown to be involved in SM suppression in Aspergillus nidulans was CclA, a member of the histone 3 lysine 4 methylating COMPASS complex. Here we examine loss of CclA and a putative H3K4 demethylase, HdmA, in the human pathogen Aspergillus fumigatus. Although deletion of hdmA showed no phenotype under the conditions tested, the cclA deletant was deficient in tri- and di-methylation of H3K4 and yielded a slowly growing strain that was rich in the production of several SMs, including gliotoxin. Similar to deletion of other chromatin modifying enzymes, ΔcclA was sensitive to 6-azauracil indicating a defect in transcriptional elongation. Despite the poor growth, the ΔcclA mutant had wild-type pathogenicity in a murine model and the Toll-deficient Drosophila model of invasive aspergillosis. These data indicate that tri- and di-methylation of H3K4 is involved in the regulation of several secondary metabolites in A. fumigatus, however does not contribute to pathogenicity under the conditions tested.
Palmer et al. (2013) Loss of CclA, required for histone 3 lysine 4 methylation, decreases growth but increases secondary metabolite production in Aspergillus fumigatus. PeerJ 1:e4 https://doi.org/10.7717/peerj.4
Pain assessment in children undergoing venipuncture: the Wong–Baker faces scale versus skin conductance fluctuations
The aim of this study was to evaluate the efficacy of the subjective Wong–Baker faces pain rating scale (WBFS) and of the objective skin conductance fluctuation (SCF) test in assessing pain in children undergoing venipuncture. One-hundred and fifty children (aged 5–16 years) entered the study. All underwent venipuncture at the antecubital fossa to collect blood specimens for routine testing in the same environmental conditions. After venipuncture, the children indicated their pain intensity using the WBFS, whereas the number of SCFs was recorded before, during and after venipuncture. So, pain level was measured in each child with WBFS and SCF. We found that the level of WBFS-assessed pain was lower in all children, particularly those above 8 years of age, than SCF-assessed pain (p < 0.0001). Moreover, the number of SCFs was significantly higher during venipuncture than before or after venipuncture (p < 0.0001). At multivariate regression analysis, age and previous experience of venipuncture influenced the WBFS (β = −1.81, p < 0.001, and β = −0.86, p < 0.001, respectively) but not SCFs. In conclusion, although both procedures can be useful for research and clinical practice, our findings show that WBFS was affected by age and previous venipuncture, whereas SCF produced uniform data. If verified in other studies, our results should be taken into account when using these tools to evaluate pain in children.
Savino et al. (2013) Pain assessment in children undergoing venipuncture: the Wong–Baker faces scale versus skin conductance fluctuations. PeerJ 1:e37 https://doi.org/10.7717/peerj.37
Why sauropods had long necks; and why giraffes have short necks
The necks of the sauropod dinosaurs reached 15 m in length: six times longer than that of the world record giraffe and five times longer than those of all other terrestrial animals. Several anatomical features enabled this extreme elongation, including: absolutely large body size and quadrupedal stance providing a stable platform for a long neck; a small, light head that did not orally process food; cervical vertebrae that were both numerous and individually elongate; an efficient air-sac-based respiratory system; and distinctive cervical architecture. Relevant features of sauropod cervical vertebrae include: pneumatic chambers that enabled the bone to be positioned in a mechanically efficient way within the envelope; and muscular attachments of varying importance to the neural spines, epipophyses and cervical ribs. Other long-necked tetrapods lacked important features of sauropods, preventing the evolution of longer necks: for example, giraffes have relatively small torsos and large, heavy heads, share the usual mammalian constraint of only seven cervical vertebrae, and lack an air-sac system and pneumatic bones. Among non-sauropods, their saurischian relatives the theropod dinosaurs seem to have been best placed to evolve long necks, and indeed their necks probably surpassed those of giraffes. But 150 million years of evolution did not suffice for them to exceed a relatively modest 2.5 m.
Taylor et al. (2013) Why sauropods had long necks; and why giraffes have short necks. PeerJ 1:e36 https://doi.org/10.7717/peerj.36
Coronatine inhibits stomatal closure and delays hypersensitive response cell death induced by nonhost bacterial pathogens
Pseudomonas syringae is the most widespread bacterial pathogen in plants. Several strains of P. syringae produce a phytotoxin, coronatine (COR), which acts as a jasmonic acid mimic and inhibits plant defense responses and contributes to disease symptom development. In this study, we found that COR inhibits early defense responses during nonhost disease resistance. Stomatal closure induced by a nonhost pathogen, P. syringae pv. tabaci, was disrupted by COR in tomato epidermal peels. In addition, nonhost HR cell death triggered by P. syringae pv. tabaci on tomato was remarkably delayed when COR was supplemented along with P. syringae pv. tabaci inoculation. Using isochorismate synthase (ICS)-silenced tomato plants and transcript profiles of genes in SA- and JA-related defense pathways, we show that COR suppresses SA-mediated defense during nonhost resistance.
Lee et al. (2013) Coronatine inhibits stomatal closure and delays hypersensitive response cell death induced by nonhost bacterial pathogens. PeerJ 1:e34 https://doi.org/10.7717/peerj.34
A perfusion study of the handling of urea and urea analogues by the gills of the dogfish shark (Squalus acanthias)
The branchial mechanism of urea retention in elasmobranchs was investigated using an in vitro isolated-perfused head preparation, as well as in vivo samples, in the spiny dogfish shark. Both in vivo and in control saline perfusions containing 350 mmol L−1 urea, calculated intracellular urea concentrations in gill epithelial cells were close to extracellular concentrations. Urea efflux to the external water fell only non-significantly, and calculated gill intracellular urea concentration did not change when perfusate urea concentration was reduced from 350 to 175 mmol L−1 with osmotic compensation by 175 mmol L−1 mannitol. However, when the urea analogues thiourea or acetamide were present in the perfusate at concentrations equimolar (175 mmol L−1) to those of urea (175 mmol L−1), urea efflux rates were increased 4-fold and 6.5-fold respectively, and calculated gill intracellular urea concentrations were depressed by about 55%. Analogue efflux rates were similar to urea efflux rates. Previous studies have argued that either the basolateral or apical membranes provided the limiting permeability barrier, and/or that a back-transporter on the basolateral membranes of gill cells is responsible for urea retention. The present results provide new evidence that the apical membrane is the limiting factor in maintaining gill urea impermeability, and raise the prospect that a urea back-transporter, which can be competitively inhibited by thiourea and acetamide, operates at the apical membrane.
Wood et al. (2013) A perfusion study of the handling of urea and urea analogues by the gills of the dogfish shark (Squalus acanthias). PeerJ 1:e33 https://doi.org/10.7717/peerj.33
Analysis of innate and acquired resistance to anti-CD20 antibodies in malignant and nonmalignant B cells
The anti-CD20 monoclonal antibody, rituximab, provides a significant therapeutic benefit for patients with B-cell disorders. However, response to therapy varies and relapses are common, so an understanding of both inherited and acquired rituximab resistance is needed. In order to identify mechanisms of inherited resistance, sensitive versus resistant individuals were selected from a survey of 92 immortalized lymphoblastoid B-cell lines from normal individuals. Levels of CD20 protein and surface expression were lower in the resistant group. In contrast, CD20 mRNA levels were not correlated with susceptibility, suggesting regulation at a post-transcriptional level. To examine acquired resistance, resistant sublines were selected from both lymphoblastoid as well as lymphoma cell lines. Confirming previous findings, there was significant down-regulation of CD20 protein expression in all the resistant sublines. CD20 mRNA splice variants are reported to be associated with development of resistance. Three splice variants were observed in our cell lines, each lacking the binding epitope for rituximab, but none were associated with rituximab resistance. The second generation anti-CD20 mAb, ofatumumab, was more active compared with rituximab in vitro in the survey of all B-cell lines, mirroring results that have been reported previously with malignant B-cells. These studies show that normal B-lymphoblastoid cell lines can be used to model both innate and acquired mechanisms of resistance. They validate the important role of CD20 expression and enable future genetic studies to identify additional mediators of anti-CD20 mAb resistance.
Small et al. (2013) Analysis of innate and acquired resistance to anti-CD20 antibodies in malignant and nonmalignant B cells. PeerJ 1:e31 https://doi.org/10.7717/peerj.31
A practical implementation of de-Pake-ing via weighted Fourier transformation
We provide an NMRPipe macro to meet an increasing need in membrane biophysics for facile de-Pake-ing of axially symmetric deuterium, and to an extent phosphorous, static lineshapes. The macro implements the development of McCabe & Wassall (1997), and is run as a simple replacement for the usual Fourier transform step in an NMRPipe processing procedure.
Sani et al. (2013) A practical implementation of de-Pake-ing via weighted Fourier transformation. PeerJ 1:e30 https://doi.org/10.7717/peerj.30
Anti-apoptotic signaling as a cytoprotective mechanism in mammalian hibernation
In the context of normal cell turnover, apoptosis is a natural phenomenon involved in making essential life and death decisions. Apoptotic pathways balance signals which promote cell death (pro-apoptotic pathways) or counteract these signals (anti-apoptotic pathways). We proposed that changes in anti-apoptotic proteins would occur during mammalian hibernation to aid cell preservation during prolonged torpor under cellular conditions that are highly injurious to most mammals (e.g. low body temperatures, ischemia). Immunoblotting was used to analyze the expression of proteins associated with pro-survival in six tissues of thirteen-lined ground squirrels, Ictidomys tridecemlineatus. The brain showed a concerted response to torpor with significant increases in the levels of all anti-apoptotic targets analyzed (Bcl-2, Bcl-xL, BI-1, Mcl-1, cIAP1/2, xIAP) as well as enhanced phosphorylation of Bcl-2 at S70 and T56. Heart responded similarly with most anti-apoptotic proteins elevated significantly during torpor except for Bcl-xL and xIAP that decreased and Mcl-1 that was unaltered. In liver, BI-1 increased whereas cIAP1/2 decreased. In kidney, there was an increase in BI-1, cIAP and xIAP but decreases in Bcl-xL and p-Bcl-2(T56) content. In brown adipose tissue, protein levels of BI-1, cIAP1/2, and xIAP decreased significantly during torpor (compared with euthermia) whereas Bcl-2, Bcl-xL, Mcl-1 were unaltered; however, Bcl-2 showed enhanced phosphorylation at Thr56 but not at Ser70. In skeletal muscle, only xIAP levels changed significantly during torpor (an increase). The data show that anti-apoptotic pathways have organ-specific responses in hibernators with a prominent potential role in heart and brain where coordinated enhancement of anti-apoptotic proteins occurred in response to torpor.
Rouble et al. (2013) Anti-apoptotic signaling as a cytoprotective mechanism in mammalian hibernation. PeerJ 1:e29 https://doi.org/10.7717/peerj.29
Reduced expression of glycolate oxidase leads to enhanced disease resistance in rice
Glycolate oxidase (GLO) is a key enzyme in photorespiration, catalyzing the oxidation of glycolate to glyoxylate. Arabidopsis GLO is required for nonhost defense responses to Pseudomonas syringae and for tobacco Pto/AvrPto-mediated defense responses. We previously described identification of rice GLO1 that interacts with a glutaredoxin protein, which in turn interacts with TGA transcription factors. TGA transcription factors are well known to participate in NPR1/NH1-mediated defense signaling, which is crucial to systemic acquired resistance in plants. Here we demonstrate that reduction of rice GLO1 expression leads to enhanced resistance to Xanthomonas oryzae pv oryzae (Xoo). Constitutive silencing of GLO1 leads to programmed cell death, resulting in a lesion-mimic phenotype and lethality or reduced plant growth and development, consistent with previous reports. Inducible silencing of GLO1, employing a dexamethasone-GVG (Gal4 DNA binding domain-VP16 activation domain-glucocorticoid receptor fusion) inducible system, alleviates these detrimental effects. Silencing of GLO1 results in enhanced resistance to Xoo, increased expression of defense regulators NH1, NH3, and WRKY45, and activation of PR1 expression.
Chern et al. (2013) Reduced expression of glycolate oxidase leads to enhanced disease resistance in rice. PeerJ 1:e28 https://doi.org/10.7717/peerj.28
Novel enzyme-polymer conjugates for biotechnological applications
In the present research, a rapid, simple and efficient chemoselective method for the site-directed incorporation of tailor-made polymers into protein to create biocatalysts with excellent properties for pharmaceutical industrial purpose has been performed. First we focused on the protein engineering of the Geobacillus thermocatenulatus lipase 2 (BTL2) to replace the two cysteines (Cys65, Cys296) in the wild type enzyme (BTL-WT) by two serines. Then, by similar mode, a unique cysteine was introduced in the lid area of the protein. For the site-directed polymer incorporation, a set of different tailor-made thiol-ionic-polymers were synthesized and the protein cysteine was previously activated with 2,2-dithiodipyridine (2-PDS) to allow the disulfide exchange. The protected BTL variants were specifically modified with the different polymers in excellent yields, creating a small library of new biocatalysts. Different and important changes in the catalytic properties, possible caused by structural changes in the lid region, were observed. The different modified biocatalysts were tested in the synthesis of intermediates of antiviral and antitumor drugs, like nucleoside analogues and derivatives of phenylglutaric acid. In the hydrolysis of per-acetylated thymidine, the best biocatalyst was the BTL*-193-DextCOOH , where the activity was increased in 3-fold and the regioselectivity was improved, reaching a yield of 92% of 3’-O-acetyl-thymidine. In the case of the asymmetric hydrolysis of dimethyl phenylglutarate, the best result was found with BTL*-193-DextNH2-6000, where the enzyme activity was increased more than 5-fold and the enantiomeric excess was >99%.
Romero et al. (2013) Novel enzyme-polymer conjugates for biotechnological applications. PeerJ 1:e27 https://doi.org/10.7717/peerj.27
Organ homologies in orchid flowers re-interpreted using the Musk Orchid as a model
Background and Aims. The presence of novel structures in orchid flowers, including auricles, rostellum and bursicles on the gynostemium and a lobed labellum, has prompted long-standing homology disputes, fuelled by conflicting evidence from a wide range of sources. Re-assessment of this debate using an improved model is timely, following recent phylogenetic insights and on the cusp of a revolution in developmental genetics. Methods. We use new data from floral development and anatomy in the small-flowered terrestrial orchid Herminium monorchis as a model to explore organ homologies in orchid flowers within the context of a review of recent literature on developmental genetics. Key Results. The apex of the median carpel of Herminium is trilobed, and the bursicles develop from its lateral lobes, relatively late in flower ontogeny. The bursicles enclose the viscidia, which adhere to the tapetal remnants to form a caudicle linking the viscidium with the pollinium. The auricles are initiated earlier than the bursicles, but they also remain unvascularized. The deeply trilobed labellum possesses three vascular traces, in contrast with the lateral petals, each of which contains a single vascular trace. The two lateral labellum traces diverge from the traces supplying the two adjacent lateral sepals. Data from flower ontogeny and anatomy conflict with respect to organ homologies. Conclusions. Much progress has recently been made in understanding the exceptional differentiation shown by orchids among perianth segments, focusing on multiple copies of the DEF/AP3 subclass of B-class MADS-box genes. In contrast, untangling homologies of profound congenital union of multiple floral organs forming the orchid gynostemium is hampered by their profound congenital union, which we ascribe to overlap in gene expression between organs. Thus, the functional morphology of the orchid flower could ultimately reflect extreme synorganization and associated genetic integration. Analogizing the deeply lobed orchid labellum with a compound leaf, we speculate that KNOX genes could be implicated not only in their demonstrated role in spur development but also in the development of both the characteristic lobed morphology of the orchid labellum and the lobing of the median carpel that differentiates the bursicles and rostellum.
Rudall et al. (2013) Organ homologies in orchid flowers re-interpreted using the Musk Orchid as a model. PeerJ 1:e26 https://doi.org/10.7717/peerj.26
Repeated hands-and-knees positioning during labour: a randomized pilot study
Background. Caesarean birth rates in North America continue to rise, in the absence of benefit for mothers and babies. One reason may be that hospitalized labouring women spend most of their labours in recumbent or semi-recumbent positions. Although hands-and-knees position has theoretical advantages, efforts to encourage its adoption in practice are severely hampered by the lack of compelling evidence that it is beneficial. Before a definitive, large scale trial, with spontaneous vaginal birth as the primary outcome, could be justified in terms of time, effort, and expense, several feasibility and acceptability questions had to be addressed. We aimed to enrol 60 women in a pilot study to assess feasibility and acceptability of the trial protocol, and to obtain estimates of treatment effects on method of birth and persistent back pain. Methods. We conducted a pilot study at two North American hospitals. In ten months of recruitment, 30 nulliparous women in labour at term were randomly allocated to either usual care (use of any position during labour except hands-and-knees) or to try hands-and-knees for 15 min every hour during labour. Data were collected about compliance, acceptability, persistent back pain, intrapartum interventions, and women’s views of their experiences. Results. Although mean length of time from randomization to delivery was over 12 hours, only 9 of the 16 women allocated to repeated hands-and-knees used it more than twice. Two of the 14 in the usual care group used hands-and-knees once. Twenty-seven women had regional analgesia (15 in the hands-and-knees group and 12 in the usual care group). Eleven in the hands-and-knees group and 14 in the usual care group had spontaneous vaginal births. One woman (in the hands-and-knees group) had a vacuum extraction. Four women in the hands-and-knees group and none in the usual care group gave birth by caesarean section. Hourly back pain ratings were highly variable in both groups, covering the full range of possible scores. Given the low compliance with the hands-and-knees position, it was not possible to explore relationships between use of the position and persistent back pain scores. When asked to rate their overall satisfaction with their birth experiences, the hands-and-knees group’s ratings tended to be lower than those in the usual care group, although 11 in the hands-and-knees group and 8 in the usual care group stated they would probably or definitely try the position in a subsequent labour. Conclusion. We concluded that we could not justify the time and expense associated with a definitive trial. However such a trial could be feasible with modifications to eligibility criteria and careful selection of suitable settings.
Hodnett et al. (2013) Repeated hands-and-knees positioning during labour: a randomized pilot study. PeerJ 1:e25 https://doi.org/10.7717/peerj.25
Poorer verbal working memory for a second language selectively impacts academic achievement in university medical students
Working memory (WM) is often poorer for a second language (L2). In low noise conditions, people listening to a language other than their first language (L1) may have similar auditory perception skills for that L2 as native listeners, but do worse in high noise conditions, and this has been attributed to the poorer WM for L2. Given that WM is critical for academic success in children and young adults, these speech in noise effects have implications for academic performance where the language of instruction is L2 for a student. We used a well-established Speech-in-Noise task as a verbal WM (vWM) test, and developed a model correlating vWM and measures of English proficiency and/or usage to scholastic outcomes in a multi-faceted assessment medical education program. Significant differences in Speech-Noise Ratio (SNR50 ) values were observed between medical undergraduates who had learned English before or after five years of age, with the latter group doing worse in the ability to extract whole connected speech in the presence of background multi-talker babble (Student-t tests, p < 0.001). Significant negative correlations were observed between the SNR50 and seven of the nine variables of English usage, learning styles, stress, and musical abilities in a questionnaire administered to the students previously. The remaining two variables, Perceived Stress Scale (PSS) and the Age of Acquisition of English (AoAoE) were significantly positively correlated with the SNR50 , showing that those with a poorer capacity to discriminate simple English sentences from noise had learnt English later in life and had higher levels of stress – all characteristics of the international students. Local students exhibited significantly lower SNR50 scores and were significantly younger when they first learnt English. No significant correlation was detected between the SNR50 and the students’ Visual/Verbal Learning Style (r = −0.023). Standard multiple regression was carried out to assess the relationship between language proficiency and verbal working memory (SNR50 ) using 5 variables of L2 proficiency, with the results showing that the variance in SNR50 was significantly predicted by this model (r2 = 0.335). Hierarchical multiple regression was then used to test the ability of three independent variable measures (SNR50 , age of acquisition of English and English proficiency) to predict academic performance as the dependent variable in a factor analysis model which predicted significant performance differences in an assessment requiring communications skills (p = 0.008), but not on a companion assessment requiring knowledge of procedural skills, or other assessments requiring factual knowledge. Thus, impaired vWM for an L2 appears to affect specific communications-based assessments in university medical students.
Mann et al. (2013) Poorer verbal working memory for a second language selectively impacts academic achievement in university medical students. PeerJ 1:e22 https://doi.org/10.7717/peerj.22
Glucose-6-phosphate dehydrogenase regulation in the hepatopancreas of the anoxia-tolerant marine mollusc, Littorina littorea
Glucose-6-phosphate dehydrogenase (G6PDH) gates flux through the pentose phosphate pathway and is key to cellular antioxidant defense due to its role in producing NADPH. Good antioxidant defenses are crucial for anoxia-tolerant organisms that experience wide variations in oxygen availability. The marine mollusc, Littorina littorea, is an intertidal snail that experiences daily bouts of anoxia/hypoxia with the tide cycle and shows multiple metabolic and enzymatic adaptations that support anaerobiosis. This study investigated the kinetic, physical and regulatory properties of G6PDH from hepatopancreas of L. littorea to determine if the enzyme is differentially regulated in response to anoxia, thereby providing altered pentose phosphate pathway functionality under oxygen stress conditions. Several kinetic properties of G6PDH differed significantly between aerobic and 24 h anoxic conditions; compared with the aerobic state, anoxic G6PDH (assayed at pH 8) showed a 38% decrease in Km G6P and enhanced inhibition by urea, whereas in pH 6 assays Km NADP and maximal activity changed significantly between the two states. The mechanism underlying anoxia-responsive changes in enzyme properties proved to be a change in the phosphorylation state of G6PDH. This was documented with immunoblotting using an anti-phosphoserine antibody, in vitro incubations that stimulated endogenous protein kinases versus protein phosphatases and significantly changed Km G6P, and phosphorylation of the enzyme with 32P-ATP. All these data indicated that the aerobic and anoxic forms of G6PDH were the high and low phosphate forms, respectively, and that phosphorylation state was modulated in response to selected endogenous protein kinases (PKA or PKG) and protein phosphatases (PP1 or PP2C). Anoxia-induced changes in the phosphorylation state of G6PDH may facilitate sustained or increased production of NADPH to enhance antioxidant defense during long term anaerobiosis and/or during the transition back to aerobic conditions when the reintroduction of oxygen causes a rapid increase in oxidative stress.
Lama et al. (2013) Glucose-6-phosphate dehydrogenase regulation in the hepatopancreas of the anoxia-tolerant marine mollusc, Littorina littorea. PeerJ 1:e21 https://doi.org/10.7717/peerj.21
Perceptual elements in Penn & Teller’s “Cups and Balls” magic trick
Magic illusions provide the perceptual and cognitive scientist with a toolbox of experimental manipulations and testable hypotheses about the building blocks of conscious experience. Here we studied several sleight-of-hand manipulations in the performance of the classic “Cups and Balls” magic trick (where balls appear and disappear inside upside-down opaque cups). We examined a version inspired by the entertainment duo Penn & Teller, conducted with three opaque and subsequently with three transparent cups. Magician Teller used his right hand to load (i.e. introduce surreptitiously) a small ball inside each of two upside-down cups, one at a time, while using his left hand to remove a different ball from the upside-down bottom of the cup. The sleight at the third cup involved one of six manipulations: (a) standard maneuver, (b) standard maneuver without a third ball, (c) ball placed on the table, (d) ball lifted, (e) ball dropped to the floor, and (f) ball stuck to the cup. Seven subjects watched the videos of the performances while reporting, via button press, whenever balls were removed from the cups/table (button “1”) or placed inside the cups/on the table (button “2”). Subjects’ perception was more accurate with transparent than with opaque cups. Perceptual performance was worse for the conditions where the ball was placed on the table, or stuck to the cup, than for the standard maneuver. The condition in which the ball was lifted displaced the subjects’ gaze position the most, whereas the condition in which there was no ball caused the smallest gaze displacement. Training improved the subjects’ perceptual performance. Occlusion of the magician’s face did not affect the subjects’ perception, suggesting that gaze misdirection does not play a strong role in the Cups and Balls illusion. Our results have implications for how to optimize the performance of this classic magic trick, and for the types of hand and object motion that maximize magic misdirection.
Rieiro et al. (2013) Perceptual elements in Penn & Teller’s “Cups and Balls” magic trick. PeerJ 1:e19 https://doi.org/10.7717/peerj.19
Timing of molt of barn swallows is delayed in a rare Clock genotype
Photoperiodic responses are major factors entraining circannual life-cycles, functioning to adaptively synchronize annual routines to seasonal fluctuations in ecological conditions. Photoperiodism in physiology and behaviour is enforced by genes, including the vertebrate Clock orthologues, which are associated, for example, with phenology of migration in fish and breeding in birds. However, the role of Clock in photoperiodic plumage molt processes is unknown. We analyzed variation in molt schedules in relation to Clock genotype, using the long-distance migratory barn swallow (Hirundo rustica) as a model and by identifying males and females using molecular sexing techniques. Consistently with previous studies, we found one very common (Q7) and two rare (Q6, Q8) variants of a functionally significant Clock polyglutamine repeat. Molt schedules of primary wing feathers of swallows during their wintering period in Nigeria differed among Clock genotypes: rare (1.1%) Q7/Q8 heterozygotes had significantly delayed molt compared to the other genotypes. Molt schedules did not differ between males and females, and no differential association between molt and Clock in relation to sex emerged. The same rare Clock genotype that exhibited delayed breeding in Europe was here found to delay molt in Africa. Though based on a limited number of Q7/Q8 individuals from an otherwise very large sample, these novel results suggest that Clock is involved in the photoperiodic control of both molt and breeding, potentially also via reciprocal carry-over effects. If confirmed in species with higher Clock polymorphism, present results may have far-reaching consequences for the study of photoperiodic control of molt and expression of annual routines.
Saino et al. (2013) Timing of molt of barn swallows is delayed in a rare Clock genotype. PeerJ 1:e17 https://doi.org/10.7717/peerj.17
Na+/Ca2+ selectivity in the bacterial voltage-gated sodium channel NavAb
The recent publication of a number of high resolution bacterial voltage-gated sodium channel structures has opened the door for the mechanisms employed by these channels to distinguish between ions to be elucidated. The way these channels select between Na+ and K+ has been investigated in computational studies, but the selectivity for Na+ over Ca2+ has not yet been studied in this way. Here we use molecular dynamics simulations to calculate the energetics of Na+ and Ca2+ transport through the channel. Single ion profiles show that Ca2+ experiences a large barrier midway through the selectivity filter that is not seen by Na+. This barrier is caused by the need for Ca2+ to partly dehydrate to pass through this region and the lack of compensating interactions with the protein. Multi-ion profiles show that ions can pass each other in the channel, which is why the presence of Ca2+ does not block Na+ conduction despite binding more strongly in the pore.
Corry (2013) Na+/Ca2+ selectivity in the bacterial voltage-gated sodium channel NavAb. PeerJ 1:e16 https://doi.org/10.7717/peerj.16
Fluorescent protein tagging confirms the presence of ribosomal proteins at Drosophila polytene chromosomes
Most ribosomal proteins (RPs) are stoichiometrically incorporated into ribosomal subunits and play essential roles in ribosome biogenesis and function. However, a number of RPs appear to have non-ribosomal functions, which involve direct association with pre-mRNA and transcription factors at transcription sites. The consensus is that the RPs found at these sites are off ribosomal subunits, but observation that different RPs are usually found together suggests that ribosomal or ribosomal-like subunits might be present. Notably, it has previously been reported that antibodies against 20 different RPs stain the same Pol II transcription sites in Drosophila polytene chromosomes. Some concerns, however, were raised about the specificity of the antibodies. To investigate further whether RPs are present at transcription sites in Drosophila, we have generated several transgenic flies expressing RPs (RpS2, RpS5a, RpS9, RpS11, RpS13, RpS18, RpL8, RpL11, RpL32, and RpL36) tagged with either green or red fluorescent protein. Imaging of salivary gland cells showed that these proteins are, as expected, abundant in the cytoplasm as well as in the nucleolus. However, these RPs are also apparent in the nucleus in the region occupied by the chromosomes. Indeed, polytene chromosome immunostaining of a representative subset of tagged RPs confirms the association with transcribed loci. Furthermore, characterization of a strain expressing RpL41 functionally tagged at its native genomic locus with YFP, also showed apparent nuclear accumulation and chromosomal association, suggesting that such a nuclear localization pattern might be a shared feature of RPs and is biologically important. We anticipate that the transgenes described here should provide a useful research tool to visualize ribosomal subunits in Drosophila tissues and to study the non-ribosomal functions of RPs.
Rugjee et al. (2013) Fluorescent protein tagging confirms the presence of ribosomal proteins at Drosophila polytene chromosomes. PeerJ 1:e15 https://doi.org/10.7717/peerj.15
Simultaneous recordings of ocular microtremor and microsaccades with a piezoelectric sensor and a video-oculography system
Our eyes are in continuous motion. Even when we attempt to fix our gaze, we produce so called “fixational eye movements”, which include microsaccades, drift, and ocular microtremor (OMT). Microsaccades, the largest and fastest type of fixational eye movement, shift the retinal image from several dozen to several hundred photoreceptors and have equivalent physical characteristics to saccades, only on a smaller scale (Martinez-Conde, Otero-Millan & Macknik, 2013). OMT occurs simultaneously with drift and is the smallest of the fixational eye movements (∼1 photoreceptor width, >0.5 arcmin), with dominant frequencies ranging from 70 Hz to 103 Hz (Martinez-Conde, Macknik & Hubel, 2004). Due to OMT’s small amplitude and high frequency, the most accurate and stringent way to record it is the piezoelectric transduction method. Thus, OMT studies are far rarer than those focusing on microsaccades or drift. Here we conducted simultaneous recordings of OMT and microsaccades with a piezoelectric device and a commercial infrared video tracking system. We set out to determine whether OMT could help to restore perceptually faded targets during attempted fixation, and we also wondered whether the piezoelectric sensor could affect the characteristics of microsaccades. Our results showed that microsaccades, but not OMT, counteracted perceptual fading. We moreover found that the piezoelectric sensor affected microsaccades in a complex way, and that the oculomotor system adjusted to the stress brought on by the sensor by adjusting the magnitudes of microsaccades.
McCamy et al. (2013) Simultaneous recordings of ocular microtremor and microsaccades with a piezoelectric sensor and a video-oculography system. PeerJ 1:e14 https://doi.org/10.7717/peerj.14
Dissecting the mechanisms of squirrel monkey (Saimiri boliviensis) social learning
Although the social learning abilities of monkeys have been well documented, this research has only focused on a few species. Furthermore, of those that also incorporated dissections of social learning mechanisms, the majority studied either capuchins (Cebus apella) or marmosets (Callithrix jacchus). To gain a broader understanding of how monkeys gain new skills, we tested squirrel monkeys (Saimiri boliviensis) which have never been studied in tests of social learning mechanisms. To determine whether S. boliviensis can socially learn, we ran “open diffusion” tests with monkeys housed in two social groups (N = 23). Over the course of 10 20-min sessions, the monkeys in each group observed a trained group member retrieving a mealworm from a bidirectional task (the “Slide-box”). Two thirds (67%) of these monkeys both learned how to operate the Slide-box and they also moved the door significantly more times in the direction modeled by the trained demonstrator than the alternative direction. To tease apart the underlying social learning mechanisms we ran a series of three control conditions with 35 squirrel monkeys that had no previous experience with the Slide-box. The first replicated the experimental open diffusion sessions but without the inclusion of a trained model, the second was a no-information control with dyads of monkeys, and the third was a ‘ghost’ display shown to individual monkeys. The first two controls tested for the importance of social support (mere presence effect) and the ghost display showed the affordances of the task to the monkeys. The monkeys showed a certain level of success in the group control (54% of subjects solved the task on one or more occasions) and paired controls (28% were successful) but none were successful in the ghost control. We propose that the squirrel monkeys’ learning, observed in the experimental open diffusion tests, can be best described by a combination of social learning mechanisms in concert; in this case, those mechanisms are most likely object movement reenactment and social facilitation. We discuss the interplay of these mechanisms and how they related to learning shown by other primate species.
Hopper et al. (2013) Dissecting the mechanisms of squirrel monkey (Saimiri boliviensis) social learning. PeerJ 1:e13 https://doi.org/10.7717/peerj.13
Novel control of lactate dehydrogenase from the freeze tolerant wood frog: role of posttranslational modifications
Lactate dehydrogenase (LDH), the terminal enzyme of anaerobic glycolysis, plays a crucial role both in sustaining glycolytic ATP production under oxygen-limiting conditions and in facilitating the catabolism of accumulated lactate when stress conditions are relieved. In this study, the effects on LDH of in vivo freezing and dehydration stresses (both of which impose hypoxia/anoxia stress on tissues) were examined in skeletal muscle of the freeze-tolerant wood frog, Rana sylvatica. LDH from muscle of control, frozen and dehydrated wood frogs was purified to homogeneity in a two-step process. The kinetic properties and stability of purified LDH were analyzed, revealing no significant differences in Vmax, Km and I50 values between control and frozen LDH. However, control and dehydrated LDH differed significantly in Km values for pyruvate, lactate, and NAD, I50 urea, and in temperature, glucose, and urea effects on these parameters. The possibility that posttranslational modification of LDH was responsible for the stable differences in enzyme behavior between control and dehydrated states was assessed using ProQ diamond staining to detect phosphorylation and immunoblotting to detect acetylation, methylation, ubiquitination, SUMOylation and nitrosylation of the enzyme. LDH from muscle of dehydrated wood frogs showed significantly lower levels of acetylation, providing one of the first demonstrations of a potential role for protein acetylation in the stress-responsive control of a metabolic enzyme.
Abboud et al. (2013) Novel control of lactate dehydrogenase from the freeze tolerant wood frog: role of posttranslational modifications. PeerJ 1:e12 https://doi.org/10.7717/peerj.12
Assessing insect responses to climate change: What are we testing for? Where should we be heading?
To understand how researchers are tackling globally important issues, it is crucial to identify whether current research is comprehensive enough to make substantive predictions about general responses. We examined how research on climate change affecting insects is being assessed, what factors are being tested and the localities of studies, from 1703 papers published between 1985 and August 2012. Most published research (64%) is generated from Europe and North America and being dedicated to core data analysis, with 29% of the studies analysed dedicated to Lepidoptera and 22% Diptera: which are well above their contribution to the currently identified insect species richness (estimated at 13% and 17% respectively). Research publications on Coleoptera fall well short of their proportional contribution (19% of publications but 39% of insect species identified), and to a lesser extent so do Hemiptera, and Hymenoptera. Species specific responses to changes in temperature by assessing distribution/range shifts or changes in abundance were the most commonly used methods of assessing the impact of climate change on insects. Research on insects and climate change to date is dominated by manuscripts assessing butterflies in Europe, insects of economic and/or environmental concern in forestry, agriculture, and model organisms. The research on understanding how insects will respond to a rapidly changing climate is still in its infancy, but the current trends of publications give a good basis for how we are attempting to assess insect responses. In particular, there is a crucial need for broader studies of ecological, behavioural, physiological and life history responses to be addressed across a greater range of geographic locations, particularly Asia, Africa and Australasia, and in areas of high human population growth and habitat modification. It is still too early in our understanding of taxa responses to climate change to know if charismatic taxa, such as butterflies, or disease vectors, including Diptera, can be used as keystone taxa to generalise other insect responses to climate change. This is critical as the basic biology of most species is still poorly known, and dominant, well studied taxa may show variable responses to climate change across their distribution due to regional biotic and abiotic influences. Indeed identifying if insect responses to climate change can be generalised using phylogeny, functional traits, or functional groups, or will populations and species exhibit idiosyncratic responses, should be a key priority for future research.
Andrew et al. (2013) Assessing insect responses to climate change: What are we testing for? Where should we be heading?. PeerJ 1:e11 https://doi.org/10.7717/peerj.11
Influence of the experimental design of gene expression studies on the inference of gene regulatory networks: environmental factors
The inference of gene regulatory networks gained within recent years a considerable interest in the biology and biomedical community. The purpose of this paper is to investigate the influence that environmental conditions can exhibit on the inference performance of network inference algorithms. Specifically, we study five network inference methods, Aracne, BC3NET, CLR, C3NET and MRNET, and compare the results for three different conditions: (I) observational gene expression data: normal environmental condition, (II) interventional gene expression data: growth in rich media, (III) interventional gene expression data: normal environmental condition interrupted by a positive spike-in stimulation. Overall, we find that different statistical inference methods lead to comparable, but condition-specific results. Further, our results suggest that non-steady-state data enhance the inferability of regulatory networks.
Emmert-Streib (2013) Influence of the experimental design of gene expression studies on the inference of gene regulatory networks: environmental factors. PeerJ 1:e10 https://doi.org/10.7717/peerj.10
The effects of fixation target size and luminance on microsaccades and square-wave jerks
A large amount of classic and contemporary vision studies require subjects to fixate a target. Target fixation serves as a normalizing factor across studies, promoting the field’s ability to compare and contrast experiments. Yet, fixation target parameters, including luminance, contrast, size, shape and color, vary across studies, potentially affecting the interpretation of results. Previous research on the effects of fixation target size and luminance on the control of fixation position rendered conflicting results, and no study has examined the effects of fixation target characteristics on square-wave jerks, the most common type of saccadic intrusion. Here we set out to determine the effects of fixation target size and luminance on the characteristics of microsaccades and square-wave jerks, over a large range of stimulus parameters. Human subjects fixated a circular target with varying luminance and size while we recorded their eye movements with an infrared video tracker (EyeLink 1000, SR Research). We detected microsaccades and SWJs automatically with objective algorithms developed previously. Microsaccade rates decreased linearly and microsaccade magnitudes increased linearly with target size. The percent of microsaccades forming part of SWJs decreased, and the time from the end of the initial SWJ saccade to the beginning of the second SWJ saccade (SWJ inter-saccadic interval; ISI) increased with target size. The microsaccadic preference for horizontal direction also decreased moderately with target size . Target luminance did not affect significantly microsaccades or SWJs, however. In the absence of a fixation target, microsaccades became scarcer and larger, while SWJ prevalence decreased and SWJ ISIs increased. Thus, the choice of fixation target can affect experimental outcomes, especially in human factors and in visual and oculomotor studies. These results have implications for previous and future research conducted under fixation conditions, and should encourage forthcoming studies to report the size of fixation targets to aid the interpretation and replication of their results.
McCamy et al. (2013) The effects of fixation target size and luminance on microsaccades and square-wave jerks. PeerJ 1:e9 https://doi.org/10.7717/peerj.9
Mutations changing tropomodulin affinity for tropomyosin alter neurite formation and extension
Assembly of the actin cytoskeleton is an important part of formation of neurites in developing neurons. Tropomodulin, a tropomyosin-dependent capping protein for the pointed end of the actin filament, is one of the key players in this process. Tropomodulin binds tropomyosin in two binding sites. Tmod1 and Tmod2, tropomodulin isoforms found in neurons, were overexpressed in PC12 cells, a model system for neuronal differentiation. Tmod1 did not affect neuronal differentiation; while cells expressing Tmod2 showed a significant reduction in the number and the length of neurites. Both tropomodulins bind short α-, γ- and δ-tropomyosin isoforms. Mutations in one of the tropomyosin-binding sites of Tmod1, which increased its affinity to short γ- and δ-tropomyosin isoforms, caused a decrease in binding short α-tropomyosin isoforms along with a 2-fold decrease in the length of neurites. Our data demonstrate that Tmod1 is involved in neuronal differentiation for proper neurite formation and outgrowth, and that Tmod2 inhibits these processes. The mutations in the tropomyosin-binding site of Tmod1 impair neurite outgrowth, suggesting that the integrity of this binding site is critical for the proper function of Tmod1 during neuronal differentiation.
Moroz et al. (2013) Mutations changing tropomodulin affinity for tropomyosin alter neurite formation and extension. PeerJ 1:e7 https://doi.org/10.7717/peerj.7
Dealing with the unexpected: consumer responses to direct-access BRCA mutation testing
Background. Inherited BRCA gene mutations convey a high risk for breast and ovarian cancer, but current guidelines limit BRCA mutation testing to women with early-onset cancer and relatives of mutation-positive cases. Benefits and risks of providing this information directly to consumers are unknown. Methods. To assess and quantify emotional and behavioral reactions of consumers to their 23andMe Personal Genome Service® report of three BRCA mutations that are common in Ashkenazi Jews, we invited all 136 BRCA1 and BRCA2 mutation-positive individuals in the 23andMe customer database who had chosen to view their BRCA reports to participate in this IRB-approved study. We also invited 160 mutation-negative customers who were matched for age, sex and ancestry. Semi-structured phone interviews were completed for 32 mutation carriers, 16 women and 16 men, and 31 non-carriers. Questions addressed personal and family history of cancer, decision and timing of viewing the BRCA report, recollection of the result, emotional responses, perception of personal cancer risk, information sharing, and actions taken or planned. Results. Eleven women and 14 men had received the unexpected result that they are carriers of a BRCA1 185delAG or 5382insC, or BRCA2 6174delT mutation. None of them reported extreme anxiety and four experienced moderate anxiety that was transitory. Remarkably, five women and six men described their response as neutral. Most carrier women sought medical advice and four underwent risk-reducing procedures after confirmatory mutation testing. Male carriers realized that their test results implied genetic risk for female relatives, and several of them felt considerably burdened by this fact. Sharing mutation information with family members led to screening of at least 30 relatives and identification of 13 additional carriers. Non-carriers did not report inappropriate actions, such as foregoing cancer screening. All but one of the 32 mutation-positive participants appreciated learning their BRCA mutation status. Conclusions. Direct access to BRCA mutation tests, considered a model for high-risk actionable genetic tests of proven clinical utility, provided clear benefits to participants. The unexpected information demonstrated a cascade effect as relatives of newly identified carriers also sought testing and more mutation carriers were identified. Given the absence of evidence for serious emotional distress or inappropriate actions in this subset of mutation-positive customers who agreed to be interviewed for this study, broader screening of Ashkenazi Jewish women for these three BRCA mutations should be considered.
Francke et al. (2013) Dealing with the unexpected: consumer responses to direct-access BRCA mutation testing. PeerJ 1:e8 https://doi.org/10.7717/peerj.8
Soil carbon determination by thermogravimetrics
Determination of soil constituents and structure has a vital role in agriculture generally. Methods for the determination of soil carbon have in particular gained greater currency in recent times because of the potential that soils offer in providing offsets for greenhouse gas (CO2-equivalent) emissions. Ideally, soil carbon which can also be quite diverse in its makeup and origin, should be measureable by readily accessible, affordable and reliable means. Loss-on-ignition is still a widely used method being suitably simple and available but may have limitations for soil C monitoring. How can these limitations be better defined and understood where such a method is required to detect relatively small changes during soil-C building? Thermogravimetric (TGA) instrumentation to measure carbonaceous components has become more interesting because of its potential to separate carbon and other components using very precise and variable heating programs. TGA related studies were undertaken to assist our understanding in the quantification of soil carbon when using methods such as loss-on-ignition. Combining instrumentation so that mass changes can be monitored by mass spectrometer ion currents has elucidated otherwise hidden features of thermal methods enabling the interpretation and evaluation of mass-loss patterns. Soil thermogravimetric work has indicated that loss-on-ignition methods are best constrained to temperatures from 200 to 430 °C for reliable determination for soil organic carbon especially where clay content is higher. In the absence of C-specific detection where mass only changes are relied upon, exceeding this temperature incurs increasing contributions from inorganic sources adding to mass losses with diminishing contributions related to organic matter. The smaller amounts of probably more recalcitrant organic matter released at the higher temperatures may represent mineral associated material and/or simply more refractory forms.
Pallasser et al. (2013) Soil carbon determination by thermogravimetrics. PeerJ 1:e6 https://doi.org/10.7717/peerj.6
Bacterial curli protein promotes the conversion of PAP248-286 into the amyloid SEVI: cross-seeding of dissimilar amyloid sequences
Fragments of prostatic acid phosphatase (PAP248-286) in human semen dramatically increase HIV infection efficiency by increasing virus adhesion to target cells. PAP248-286 only enhances HIV infection in the form of amyloid aggregates termed SEVI (Semen Enhancer of Viral Infection), however monomeric PAP248-286 aggregates very slowly in isolation. It has therefore been suggested that SEVI fiber formation in vivo may be promoted by exogenous factors. We show here that a bacterially-produced extracellular amyloid (curli or Csg) acts as a catalytic agent for SEVI formation from PAP248-286 at low concentrations in vitro, producing fibers that retain the ability to enhance HIV (Human Immunodeficiency Virus) infection. Kinetic analysis of the cross-seeding effect shows an unusual pattern. Cross-seeding PAP248-286 with curli only moderately affects the nucleation rate while significantly enhancing the growth of fibers from existing nuclei. This pattern is in contrast to most previous observations of cross-seeding, which show cross-seeding partially bypasses the nucleation step but has little effect on fiber elongation. Seeding other amyloidogenic proteins (IAPP (islet amyloid polypeptide) and Aβ1−40) with curli showed varied results. Curli cross-seeding decreased the lag-time of IAPP amyloid formation but strongly inhibited IAPP elongation. Curli cross-seeding exerted a complicated concentration dependent effect on Aβ1−40 fibrillogenesis kinetics. Combined, these results suggest that the interaction of amyloidogenic proteins with preformed fibers of a different type can take a variety of forms and is not limited to epitaxial nucleation between proteins of similar sequence. The ability of curli fibers to interact with proteins of dissimilar sequences suggests cross-seeding may be a more general phenomenon than previously supposed.
Hartman et al. (2013) Bacterial curli protein promotes the conversion of PAP248-286 into the amyloid SEVI: cross-seeding of dissimilar amyloid sequences. PeerJ 1:e5 https://doi.org/10.7717/peerj.5
The roles of STP and LTP in synaptic encoding
Long-term potentiation (LTP), a cellular model of learning and memory, is generally regarded as a unitary phenomenon that alters the strength of synaptic transmission by increasing the postsynaptic response to the release of a quantum of neurotransmitter. LTP, at CA3-CA1 synapses in the hippocampus, contains a stimulation-labile phase of short-term potentiation (STP, or transient LTP, t-LTP) that decays into stable LTP. By studying the responses of populations of neurons to brief bursts of high-frequency afferent stimulation before and after the induction of LTP, we found that synaptic responses during bursts are potentiated equally during LTP but not during STP. We show that STP modulates the frequency response of synaptic transmission whereas LTP preserves the fidelity. Thus, STP and LTP have different functional consequences for the transfer of synaptic information.
Volianskis et al. (2013) The roles of STP and LTP in synaptic encoding. PeerJ 1:e3 https://doi.org/10.7717/peerj.3
Malleable ribonucleoprotein machine: protein intrinsic disorder in the Saccharomyces cerevisiae spliceosome
Recent studies revealed that a significant fraction of any given proteome is presented by proteins that do not have unique 3D structures as a whole or in significant parts. These intrinsically disordered proteins possess dramatic structural and functional variability, being especially enriched in signaling and regulatory functions since their lack of fixed structure defines their ability to be involved in interaction with several proteins and allows them to be re-used in multiple pathways. Among recognized disorder-based protein functions are interactions with nucleic acids and multi-target binding; i.e., the functions ascribed to many spliceosomal proteins. Therefore, the spliceosome, a multimegadalton ribonucleoprotein machine catalyzing the excision of introns from eukaryotic pre-mRNAs, represents an attractive target for the focused analysis of the abundance and functionality of intrinsic disorder in its proteinaceous components. In yeast cells, spliceosome consists of five small nuclear RNAs (U1, U2, U4, U5, and U6) and a range of associated proteins. Some of these proteins constitute cores of the corresponding snRNA-protein complexes known as small nuclear ribonucleoproteins (snRNPs). Other spliceosomal proteins have various auxiliary functions. To gain better understanding of the functional roles of intrinsic disorder, we have studied the prevalence of intrinsically disordered proteins in the yeast spliceosome using a wide array of bioinformatics methods. Our study revealed that similar to the proteins associated with human spliceosomes (Korneta & Bujnicki, 2012), proteins found in the yeast spliceosome are enriched in intrinsic disorder.
Coelho Ribeiro et al. (2013) Malleable ribonucleoprotein machine: protein intrinsic disorder in the Saccharomyces cerevisiae spliceosome. PeerJ 1:e2 https://doi.org/10.7717/peerj.2
Loops are irregular structures which connect two secondary structure elements in proteins. They often play important roles in function, including enzyme reactions and ligand binding. Despite their importance, their structure remains difficult to predict. Most protein loop structure prediction methods sample local loop segments and score them. In particular protein loop classifications and database search methods depend heavily on local properties of loops. Here we examine the distance between a loop’s end points (span). We find that the distribution of loop span appears to be independent of the number of residues in the loop, in other words the separation between the anchors of a loop does not increase with an increase in the number of loop residues. Loop span is also unaffected by the secondary structures at the end points, unless the two anchors are part of an anti-parallel beta sheet. As loop span appears to be independent of global properties of the protein we suggest that its distribution can be described by a random fluctuation model based on the Maxwell–Boltzmann distribution. It is believed that the primary difficulty in protein loop structure prediction comes from the number of residues in the loop. Following the idea that loop span is an independent local property, we investigate its effect on protein loop structure prediction and show how normalised span (loop stretch) is related to the structural complexity of loops. Highly contracted loops are more difficult to predict than stretched loops.
Choi et al. (2013) How long is a piece of loop?. PeerJ 1:e1 https://doi.org/10.7717/peerj.1