bluuebunny/crossref_metadata_2025_split huggingface dataset:
Redesigned upstream processing enables a 24-hour workflow from E. coli cells to cell-free protein synthesis
Authors: Levine Max Z. , So Byungcheol , Mullin Alissa C. , Watts Katharine R. , Oza Javin P. 
Published: August 2019

AbstractCell-free protein synthesis (CFPS) platforms have undergone numerous workflow improvements to enable diverse applications in research, biomanufacturing, point-of-care detection, therapeutics, and education using affordable laboratory equipment and reagents. The Escherichia coli cell extract-based platform, being one of the most affordable and versatile CFPS platforms, has been broadly adopted. In spite of the promise of simplicity, the cell-free platform remains technically nuanced, posing challenges to reproducible implementation and broad adoption. Additionally, while the CFPS reaction itself can be implemented on-demand, the upstream processing of cells to generate crude cell lysate remains time-intensive, representing one of the largest sources of cost associated with the biotechnology. To circumvent the lengthy and tedious upstream workflow, we have redesigned the processes by developing a long-lasting autoinduction media formulation for cell-free that obviates human intervention between inoculation and harvest. Cell-free autoinduction (CFAI) media supports these advantages through the production of highly robust cell extracts from high cell density cultures nearing stationary phase of growth. Growth of cells to high density and autoinduction of T7 RNAP expression can be achieved by incubation overnight, eliminating the need for user intervention for the entirety of the process. The total mass of cells obtained is substantially increased, which directly results in a 400% increase in total extract volume obtained compared to past workflows. Based on these advances, we outline a new upstream processing workflow that allows researchers to go from cells on a streak plate to completing CFPS reactions within 24 hours while maintaining robust reaction yields of sfGFP (>1 mg/ml). We hope this advance will improve the time and cost-efficiency for existing CFPS researchers, increase the simplicity and reproducibility, and reduce the barrier-to-entry for new researchers interested in implementing CFPS.
Are tropical reptiles really declining? A six-year survey of snakes in Drake Bay, Costa Rica, and the role of temperature, rain and light
Authors: Barquero-González Jose Pablo , Stice Tracie L. , Gómez Gianfranco , Monge-Nájera Julián 
Published: August 2019

AbstractIntroductionstudies in the last two decades have found declining snake populations in both temperate and tropical sites, including informal reports from Drake Bay, Costa Rica.Objectiveto investigate if reports of decreasing snake populations in Drake Bay had a real basis, and if environmental factors, particularly temperature, rain and light, have played a role in that decrease.Methodswe worked at Drake Bay from 2012 through 2017 and made over 4000 h of transect counts. Using head flashlights we surveyed a transect covered by lowland tropical rainforest at an altitude of 12–38 m above sea level, near the Agujas River, mostly at 1930–2200 hours. We counted all the snakes that we could see along the transect.Resultssnake counts increase from August to September and then decline rapidly. The May snakes/rainfall peaks coincide, but the second snake peak occurs one month before the rain peak; we counted more snakes in dry nights, with the exception of Imantodes cenchoa which was equally common despite rain conditions. We saw less Leptodeira septentrionalis on bright nights, but all other species were unaffected. Along the six years, the number of species with each diet type remained relatively constant, but the number of individuals declined sharply for those that feed on amphibians and reptiles. We report Rhadinella godmani, a highland species, at 12–38 m of altitude.Conclusionnight field counts of snakes in Drake Bay, Costa Rica, show a strong decline from 2012 through 2017.
Dynamics of the energy seascape can explain intra-specific variations in sea-crossing behaviour of soaring birds
Authors: Nourani E. , Vansteelant W. M. G. , Byholm P. , Safi K. 
Published: August 2019

AbstractThermal soaring birds extract energy from the atmosphere to achieve energetically low-cost movement. When encountering regions that are energetically costly to fly over, such as open seas, they attempt to adjust the spatio-temporal pattern of their passage to maximize energy extraction from the atmosphere over these ecological barriers. We apply the concept of energy landscapes to investigate the spatio-temporal dynamics of energy availability over the open sea for soaring flight. We specifically investigated how the “energy seascape” may shape age-specific sea-crossing behaviour of European honey buzzards Pernis apivorus over the Mediterranean Sea in autumn. We found uplift potential over the sea to be the main determinant of sea-crossing length, rather than wind conditions. Considering this variable as a proxy for available energy over the sea, we constructed the energy seascape for the autumn migration season using forty years of temperature data. Our results indicate that early-migrating adult buzzards are likely to encounter adverse energy subsidence over the Mediterranean, whereas late-migrating juveniles face less adverse flight conditions, and even conditions conducive to soaring flight. Our study provides evidence that the dynamics of the energy landscape can explain intra-specific variation in migratory behaviour also at sea.
NADPH consumption by L-cystine reduction creates a metabolic vulnerability upon glucose deprivation
Authors: Joly James H. , Delfarah Alireza , Phung Philip S. , Parrish Sydney , Graham Nicholas A. 
Published: August 2019

AbstractThe consequences of metabolic reprogramming in cancer can include an increased dependence on metabolic substrates such as glucose for survival. As such, the vulnerability of cancer cells to glucose deprivation creates an attractive opportunity for therapeutic intervention. Because it is not possible to starve tumors of glucosein vivo, we sought to identify the mechanisms regulating cancer cell death upon glucose deprivation and then design combinations of inhibitors to mimic glucose deprivation-induced cell death. Using metabolomic profiling, we found that cells undergoing glucose deprivation-induced cell death exhibited dramatic accumulation of intracellular L-cysteine and its oxidized dimer, L-cystine, and depletion of the antioxidant glutathione. Building on this observation, we show that glucose deprivation-induced cell death is driven not by lack of glucose but rather by L-cystine import. Following glucose deprivation, the import of L-cystine and subsequent reduction to L-cysteine depleted both NADPH and glutathione, thereby allowing toxic accumulation of reactive oxygen species. Consistent with this model, we found that the glutamate/cystine antiporter, xCT, was required for sensitivity to glucose deprivation. We searched for glycolytic enzymes whose expression is essential for survival of cancer cells with high xCT expression and identified the glucose transporter GLUT1. We therefore tested a drug combination co-targeting GLUT1 and glutathione synthesis and found that these drugs induced synthetic lethal cell death in high xCT-expressing cell lines susceptible to glucose deprivation. These results indicate that co-targeting GLUT1 and glutathione synthesis is a potential therapeutic approach in tumors dependent on glucose for survival.
Neutrophilic granule protein is a novel murine LPS antagonist
Authors: Hong Jaewoo , Qu Peng , Wuest Todd R. , Lin P. Charles 
Published: August 2019

AbstractNGP was previously reported as a granular protein of neutrophils in mouse but the function has not been known clearly. We found the presence of the possible signal peptide in NGP and we hypothesized this protein is being secreted to the blood stream. Since this protein has sequence similarity with an antimicrobial protein cathelicidin, we observed the aspect of inflammation of NGP. Interestingly, NGP interacts with the complex of LPS and LBP and blocked the inflammatory function of NGP. This inhibitory effect of NGP was through the inhibition of LPS-LBP complex to interact with TLR4 molecule. Furthermore, the inhibitory function of NGP could be repeated on the inflammatory effect of LPS in both in vitro and in vivo. With these findings, we report NGP is a novel secretory protein to mask LPS and inhibits its function.
Measurement of skeletal muscle fiber contractility with high-speed traction microscopy
Authors: Rausch M , Böhringer D , Steinmann M , Schubert DW , Schrüfer S , Mark C , Fabry B 
Published: August 2019

AbstractWe describe a technique for simultaneous quantification of the contractile forces and cytosolic calcium dynamics of muscle fibers embedded in three-dimensional biopolymer gels. We derive a scaling law for linear elastic matrices such as basement membrane extract hydrogels (Matrigel) that allows us to measure contractile force from the shape of the relaxed and contracted muscle cell and the Young’s modulus of the matrix, without further knowledge of the matrix deformations surrounding the cell and without performing computationally intensive inverse force reconstruction algorithms. We apply our method to isolated mouse flexor digitorum brevis (FDB) fibers that are embedded in 10 mg/ml Matrigel. Upon electrical stimulation, individual FDB fibers show twitch forces of 0.37 µN ± 0.15 µN and tetanic forces (100 Hz stimulation frequency) of 2.38 µN ± 0.71 µN, corresponding to a tension of 0.44 kPa ± 0.25 kPa and 2.53 kPa ± 1.17 kPa, respectively. Contractile forces of FDB fibers increase in response to caffeine and the troponin-calcium-stabilizer Tirasemtiv, similar to responses measured in whole muscle. From simultaneous high-speed measurements of cell length changes and cytosolic calcium concentration using confocal line scanning at a frequency of 2048 Hz, we show that twitch and tetanic force responses to electric pulses follow the low-pass filtered calcium signal. In summary, we present a technically simple high speed and high throughput method for measuring contractile forces and cytosolic calcium dynamics of single muscle fibers. We expect that our method will help to reduce preparation time, costs, and the number of sacrificed animals needed for experiments such as drug testing.Statement of significanceWe describe a high speed, high throughput method for the simultaneous measurement of contractile force and cytoplasmic calcium dynamics following electrical pulse stimulation of muscle fibers embedded in a 3-dimensional biopolymer matrix. In contrast to the classical approach of attaching muscle fibers to a force-transducer, our method allows for a highly efficient, parallel analysis of large numbers of fibers under different treatment conditions.
In situ activation and heterologous production of a cryptic lantibiotic from a plant-ant derived Saccharopolyspora species
Authors: Vikeli Eleni , Widdick David A. , Batey Sibyl F. , Heine Daniel , Holmes Neil A. , Bibb Mervyn J. , Martins Dino J. , Pierce Naomi E. , Hutchings Matthew I. , Wilkinson Barrie 
Published: August 2019

AbstractMost clinical antibiotics are derived from actinomycete natural products (NPs) discovered at least 60 years ago. Repeated rediscovery of known compounds led the pharmaceutical industry to largely discard microbial NPs as a source of new chemical diversity but advances in genome sequencing revealed that these organisms have the potential to make many more NPs than previously thought. Approaches to unlock NP biosynthesis by genetic manipulation of the strain, by the application of chemical genetics, or by microbial co-cultivation have resulted in the identification of new antibacterial compounds. Concomitantly, intensive exploration of coevolved ecological niches, such as insect-microbe defensive symbioses, has revealed these to be a rich source of chemical novelty. Here we report the novel lanthipeptide antibiotic kyamicin generated through the activation of a cryptic biosynthetic gene cluster identified by genome mining Saccharopolyspora species found in the obligate domatia-dwelling ant Tetraponera penzigi of the ant plant Vachellia drepanolobium. Heterologous production and purification of kyamicin allowed its structural characterisation and bioactivity determination. Our activation strategy was also successful for the expression of lantibiotics from other genera, paving the way for a synthetic heterologous expression platform for the discovery of lanthipeptides that are not detected under laboratory conditions or that are new to nature.ImportanceThe discovery of novel antibiotics to tackle the growing threat of antimicrobial resistance is impeded by difficulties in accessing the full biosynthetic potential of microorganisms. The development of new tools to unlock the biosynthesis of cryptic bacterial natural products will greatly increase the repertoire of natural product scaffolds. Here we report an activation strategy that can be rapidly applied to activate the biosynthesis of cryptic lanthipeptide biosynthetic gene clusters. This allowed the discovery of a new lanthipeptide antibiotic directly from the native host and via heterologous expression.
Diversity of responses of soil saprobic fungi to recurring heat events
Authors: Szymczak Aleksandra , Ryo Masahiro , Rillig Matthias C. 
Published: August 2019

AbstractAs a consequence of ongoing climate change, the frequency of extreme heat events is expected to increase. Recurring heat pulses may disrupt functions supported by soil microorganisms, thus affecting the entire ecosystem. However, most perturbation experiments only test effects of single heat events, and therefore it remains largely unknown how soil microorganisms react to repeated pulse events. Here we present data from a lab experiment exposing 32 filamentous fungi, originally isolated from the same soil, to sequential heat perturbations. Soil saprobic fungi isolates were exposed to one or two heat pulses: mild (35°C/2h), strong (45°C/1h), or both in sequence (35°C/2h+45°C/1h), and we assessed growth rate. Out of the 32 isolates 13 isolates showed an antagonistic response, 3 isolates a synergistic response and 16 isolates responded in an additive manner. These differences in species responses to the thermal environment may contribute to species coexistence, and such dissimilarities in thermal perturbation responses may be a key aspect influencing ecosystem services that soil saprobic fungi support.
Direct observation of independently moving replisomes in Escherichia coli
Authors: Japaridze Aleksandre , Gogou Christos , Kerssemakers Jacob W. J. , Nguyen Huyen My , Dekker Cees 
Published: August 2019

AbstractThe replication and transfer of genomic material from a cell to its progeny are vital processes in all living systems. Here we visualize the process of chromosome replication in E. coli cells with an increased width. Monitoring the replication of single chromosomes yields clear examples of replication bubbles that reveal that the two replisomes move independently from the origin to the terminus of replication along each of the two arms of the circular chromosome, providing direct support for the so-called train-track model, and against a factory model for replisomes. The origin of replication duplicates near midcell, initially splitting to random directions and subsequently towards the poles. The probability of successful segregation of chromosomes significantly decreases with increasing cell width, indicating that chromosome confinement by the cell boundary is an important driver of DNA segregation. Our findings resolve long standing questions in bacterial chromosome organization.
Genetic Associations in Four Decades of Multi-Environment Trials Reveal Agronomic Trait Evolution in Common Bean
Authors: MacQueen Alice H. , White Jeffrey W. , Lee Rian , Osorno Juan M. , Schmutz Jeremy , Miklas Phillip N. , Myers Jim , McClean Phillip E. , Juenger Thomas E. 
Published: August 2019

AbstractMulti-environment trials (METs) are widely used to assess the performance of promising crop germplasm. Though seldom designed to elucidate genetic mechanisms, MET datasets are often much larger than could be duplicated for genetic research and, given proper interpretation, may offer valuable insights into the genetics of adaptation across time and space. The Cooperative Dry Bean Nursery (CDBN) is a MET for common bean (Phaseolus vulgaris) grown for over 70 years in the United States and Canada, consisting of 20 to 50 entries each year at 10 to 20 locations. The CBDN provides a rich source of phenotypic data across entries, years, and locations that is amenable to genetic analysis. To study stable genetic effects segregating in this MET, we conducted genome-wide association (GWAS) using best linear unbiased predictions (BLUPs) derived across years and locations for 21 CDBN phenotypes and genotypic data (1.2M SNPs) for 327 CDBN genotypes. The value of this approach was confirmed by the discovery of three candidate genes and genomic regions previously identified in balanced GWAS. Multivariate adaptive shrinkage (mash) analysis, which increased our power to detect significant correlated effects, found significant effects for all phenotypes. The first use of mash on an agricultural dataset discovered two genomic regions with pleiotropic effects on multiple phenotypes, likely selected on in pursuit of a crop ideotype. Overall, our results demonstrate that by applying multiple statistical genomic approaches on data mined from MET phenotypic data sets, significant genetic effects that define genomic regions associated with crop improvement can be discovered.
Potential Utility of Routine Programmatic Data in Monitoring National and State-Level HIV Epidemic in Nigeria: Data Triangulation Analysis
Authors: Henry Debem , Yakubu Aminu , Ahmed Mukhtar , Jerry Gwamna , Ibrahim Dalhatu 
Published: August 2019

AbstractNigeria relies on data from periodic resource-intensive surveys such as antenatal HIV seroprevalence sentinel surveys (ANC-HSS) and population-based National AIDS and Reproductive Health Surveys (NARHS) for its HIV control efforts. Nigeria has not explored the use of readily available routine programmatic data (RPD) to easily inform and monitor epidemic control efforts at local settings in near real time. This study aimed to determine the utility of RPDs (Prevention of Mother-To-Child Transmission [PMTCT] and HIV Testing and Counseling [HTC]) as a proxy for monitoring HIV epidemic in Nigeria. Using World Health Organization 12 step triangulation procedures, we compared state-level seropositivity data from PMTCT and HTC programs to HIV prevalence data from NARHS and ANC-HSS reports in relevant pairs from 2010 to 2014 in Nigeria. The study population was pregnant women and general population. We abstracted relevant data from PEPFAR Nigeria data source and published national survey reports. We compared visual (scatterplots and maps) patterns and trends, and performed Pearson correlation and univariate linear regression models of the estimates for best matched/contiguous years for which data were available. Correlation between PMTCT2014 and ANC-HSS2014 was positive and significant (R=0.7,p<0.001). ANC-HSS2014 and HTC2014 were slightly correlated (R=0.4,p<0.05). Significant correlation was observed between ANC-HSS2010 and PMTCT2013 (R=0.8,p<0.001) and between ANC-HSS2010 and HTC2013 (R=0.6, p<0.001). All RPD sources and ANC-HSS indicated a decreasing trend in national HIV prevalence in Nigeria. PMTCT2014 data showed strong capability of predicting HIV prevalence in ANC-HSS2014 in regression model (B=2.09,p<0.0001). Use of routine PMTCT data in monitoring HIV prevalence among women of reproductive age could be more valid and reliable in local settings than the use of HTC data. Use of RPD to monitor national and sub-national-level HIV epidemic in between national surveys in Nigeria could maximize program resources, and promote a more responsive and efficient actions toward epidemic control.
Adaptation and competition in deteriorating environments
Authors: Limberger Romana , Fussmann Gregor F. 
Published: August 2019

AbstractEvolution might rescue populations from extinction in changing environments. Using experimental evolution with microalgae, we investigated if competition influences adaptation to an abiotic stressor, and vice versa, if adaptation to abiotic change influences competition. In a first set of experiments, we propagated monocultures of five species with and without increasing salt stress for ~180 generations. When assayed in monoculture, two of the five species showed signatures of adaptation, that is, lines with a history of salt stress had higher population growth rates at high salt than lines without prior exposure to salt. When assayed in mixtures of species, however, only one of these two species had increased population size at high salt, indicating that competition can alter how adaptation to abiotic change influences population dynamics. In a second experiment, we cultivated two species in monocultures and in pairs, with and without increasing salt. While we found no effect of competition on adaptation to salt, our experiment revealed that evolutionary responses to salt can influence competition. Specifically, one of the two species had reduced competitive ability in the no-salt environment after long-term exposure to salt stress. Collectively, our results highlight the complex interplay of adaptation to abiotic change and competitive interactions.
Casein Kinase 1 dynamics underlie the PER2 circadian phosphoswitch
Authors: Philpott Jonathan M. , Narasimamurthy Rajesh , Ricci Clarisse G. , Freeberg Alfred M. , Hunt Sabrina R. , Yee Lauren E. , Pelofsky Rebecca S. , Tripathi Sarvind , Virshup David M. , Partch Carrie L. 
Published: August 2019

SummaryPost-translational control of PERIOD stability by Casein Kinase 1δ and ε (CK1) plays a key regulatory role in metazoan circadian rhythms. Despite the deep evolutionary conservation of CK1 in eukaryotes, little is known about its regulation and the factors that influence substrate selectivity on functionally antagonistic sites in PERIOD that directly control circadian period. Here we describe a molecular switch involving a highly conserved anion binding site in CK1. This switch controls conformation of the activation loop to define substrate selectivity on mammalian PER2, thereby directly regulating its stability. Integrated experimental and computational studies shed light on the allosteric linkage between two anion binding sites that dynamically regulate kinase activity. We show that period-altering kinase mutations from humans toDrosophiladifferentially modulate this activation loop switch to elicit predictable changes in PER2 stability, providing a foundation to understand and further manipulate CK1 regulation of circadian rhythms.