bluuebunny/crossref_metadata_2025 huggingface dataset:
Benchmarking bacterial genome-wide association study methods using simulated genomes and phenotypes
Authors: Saber Morteza M. , Shapiro B. Jesse
Published: February 2020

Genome-wide association studies (GWASs) have the potential to reveal the genetics of microbial phenotypes such as antibiotic resistance and virulence. Capitalizing on the growing wealth of bacterial sequence data, microbial GWAS methods aim to identify causal genetic variants while ignoring spurious associations. Bacteria reproduce clonally, leading to strong population structure and genome-wide linkage, making it challenging to separate true ‘hits’ (i.e. mutations that cause a phenotype) from non-causal linked mutations. GWAS methods attempt to correct for population structure in different ways, but their performance has not yet been systematically and comprehensively evaluated under a range of evolutionary scenarios. Here, we developed a bacterial GWAS simulator (BacGWASim) to generate bacterial genomes with varying rates of mutation, recombination and other evolutionary parameters, along with a subset of causal mutations underlying a phenotype of interest. We assessed the performance (recall and precision) of three widely used single-locus GWAS approaches (cluster-based, dimensionality-reduction and linear mixed models, implemented inplink, pyseer andgemma) and one relatively new multi-locus model implemented in pyseer, across a range of simulated sample sizes, recombination rates and causal mutation effect sizes. As expected, all methods performed better with larger sample sizes and effect sizes. The performance of clustering and dimensionality reduction approaches to correct for population structure were considerably variable according to the choice of parameters. Notably, the multi-locus elastic net (lasso) approach was consistently amongst the highest-performing methods, and had the highest power in detecting causal variants with both low and high effect sizes. Most methods reached the level of good performance (recall >0.75) for identifying causal mutations of strong effect size [log odds ratio (OR) ≥2] with a sample size of 2000 genomes. However, only elastic nets reached the level of reasonable performance (recall=0.35) for detecting markers with weaker effects (log OR ~1) in smaller samples. Elastic nets also showed superior precision and recall in controlling for genome-wide linkage, relative to single-locus models. However, all methods performed relatively poorly on highly clonal (low-recombining) genomes, suggesting room for improvement in method development. These findings show the potential for multi-locus models to improve bacterial GWAS performance. BacGWASim code and simulated data are publicly available to enable further comparisons and benchmarking of new methods.
Coinfinder: detecting significant associations and dissociations in pangenomes
Authors: Whelan Fiona Jane , Rusilowicz Martin , McInerney James Oscar
Published: February 2020

The accessory genes of prokaryote and eukaryote pangenomes accumulate by horizontal gene transfer, differential gene loss, and the effects of selection and drift. We have developed Coinfinder, a software program that assesses whether sets of homologous genes (gene families) in pangenomes associate or dissociate with each other (i.e. are ‘coincident’) more often than would be expected by chance. Coinfinder employs a user-supplied phylogenetic tree in order to assess the lineage-dependence (i.e. the phylogenetic distribution) of each accessory gene, allowing Coinfinder to focus on coincident gene pairs whose joint presence is not simply because they happened to appear in the same clade, but rather that they tend to appear together more often than expected across the phylogeny. Coinfinder is implemented in C++, Python3 and R and is freely available under the GNU license from https://github.com/fwhelan/coinfinder.
Rapid Detection of Novel Coronavirus (COVID-19) by Reverse Transcription-Loop-Mediated Isothermal Amplification
Authors: Lamb Laura E. , Bartolone Sarah N. , Ward Elijah , Chancellor Michael B.
Published: February 2020

Novel Corona virus (COVID-19 or 2019-nCoV) is an emerging global health concern that requires a rapid diagnostic test. Quantitative reverse transcription PCR (qRT-PCR) is currently the standard for COVID-19 detection; however, Reverse Transcription Loop-Mediated Isothermal Amplification (RT-LAMP) may allow for faster and cheaper field based testing at point-of-risk. The objective of this study was to develop a rapid screening diagnostic test that could be completed in under 30 minutes. Simulated patient samples were generated by spiking serum, urine, saliva, oropharyngeal swabs, and nasopharyngeal swabs with a portion of the COVID-19 nucleic sequence. The samples were tested using RT-LAMP as well as by conventional qRT-PCR. Specificity of the RT-LAMP was evaluated by also testing against other related coronaviruses. RT-LAMP specifically detected COVID-19 in simulated patient samples. This test was performed in under 30 minutes. This approach could be used for monitoring of exposed individuals or potentially aid with screening efforts in the field and potential ports of entry.
Genome-scale metabolic rewiring to achieve predictable titers rates and yield of a non-native product at scale
Authors: Banerjee Deepanwita , Eng Thomas , Lau Andrew K. , Wang Brenda , Sasaki Yusuke , Herbert Robin A. , Chen Yan , Liu Yuzhong , Prahl Jan-Philip , Singan Vasanth R. , Tanjore Deepti , Petzold Christopher J. , Keasling Jay D. , Mukhopadhyay Aindrila
Published: February 2020

Achieving high titer rates and yields (TRY) remains a bottleneck in the production of heterologous products through microbial systems, requiring elaborate engineering and many iterations. Reliable scaling of engineered strains is also rarely addressed in the first designs of the engineered strains. Both high TRY and scale are challenging metrics to achieve due to the inherent trade-off between cellular use of carbon towards growthvs.target metabolite production. We hypothesized that being able to strongly couple product formation with growth may lead to improvements across both metrics. In this study, we use elementary mode analysis to predict metabolic reactions that could be targeted to couple the production of indigoidine, a sustainable pigment, with the growth of the chosen host,Pseudomonas putidaKT2440. We then filtered the set of 16 predicted reactions using -omics data. We implemented a total of 14 gene knockdowns using a CRISPRi method optimized forP. putidaand show that the resulting engineeredP. putidastrain could achieve high TRY. The engineered pairing of product formation with carbon use also shifted production from stationary to exponential phase and the high TRY phenotype was maintained across scale. In one design cycle, we constructed an engineeredP. putidastrain that demonstrates close to 50% maximum theoretical yield (0.33 g indigoidine/g glucose consumed), reaching 25.6 g/L indigoidine and a rate of 0.22g/l/h in exponential phase. These desirable phenotypes were maintained from batch to fed-batch cultivation mode, and from 100ml shake flasks to 250 mL ambr® and 2 L bioreactors.
Hyper-connectivity of the striatum related to restricted and repetitive behaviors’ severity in children with ASD
Authors: Dupong Irene , Di Martino Adriana
Published: February 2020

Restricted and repetitive behaviors (RRB) are a core feature of autism spectrum disorder (ASD). But little is known about the underlying neurobiology of the disorder, preventing from having specific therapeutic targets. Based on the literature, we explored the correlates between a clinical score of RRB, using the Repetitive Behaviour Scale – Revised, and the intrinsic connectivity of seven striatal regions in a sample of 157 children with ASD. The sample was acquired from the ABIDE consortium. We found a significant correlation between the severity of our clinical scale and several cortico-striatal networks. Specifically, stronger connections were found between striatal seeds and two cortical areas, an occipital area and a frontal area in the left hemisphere. Intrinsic functional connectivity of the striatum could serve as a potential biomarker for improved detection of RRB severity.
Loss of graded enemy recognition in a Whitehead population allopatric with brood parasitic Long-tailed Cuckoos
Authors: Lawson Shelby L. , Leuschner Nora , Gill Brian J. , Enos Janice K. , Hauber Mark E.
Published: February 2020

Many avian hosts of brood parasitic birds discriminate between different types of threats and may respond with categorically different, specifically anti-predatory or anti-parasitic behaviors. Alternatively, hosts may adjust their responses to threat level in a graded manner, responding more aggressively to brood parasites during the laying and incubation stages of nesting, when nests are most susceptible to parasitism, and more aggressively to nest predators during the nestling and fledgling stages when predation would be more costly than parasitism. In New Zealand, endemic host Whiteheads act inconspicuously around their nests in the presence of sympatric Long-tailed Cuckoos, their obligate brood parasite, perhaps to avoid disclosing nest location. We tested behavioral responses of a Whitehead population on Tiritiri Matangi Island that has been breeding allopatrically from cuckoos for 17 years. We presented models of the allopatric parasite, a sympatric predator (Morepork Owl), and a sympatric non-threatening heterospecific (Song Thrush) during the egg and chick stages, and to groups of cooperatively breeding Whiteheads. We compared responses across nest stage and stimulus type. We found that, unlike sympatric Whiteheads elsewhere in New Zealand, Whiteheads on Tiritiri Matangi produced alarm calls in response to the cuckoo model. Furthermore, the rate of alarm calling was similar towards the cuckoo and the owl and across egg and chick stage and higher than to the control stimulus. These results are consistent with allopatric Whiteheads having lost their specific anti-parasitic defense tactics in response to brood parasitic cuckoos.
Simple post-translational circadian clock models from selective sequestration
Authors: Byrne Mark
Published: February 2020

It is possible that there are post-translational circadian oscillators that continue functioning in the absence of negative feedback transcriptional repression in many cell types from diverse organisms. Apart from the KaiABC system from cyanobacteria, the minimal molecular components and interactions required to potentially create “test-tube” circadian oscillations in different cell types are currently unknown. Inspired by the KaiABC system, I provide proof-of-principle mathematical models that a protein with 2 (or more) modification sites which selectively sequesters an effector/cofactor molecule can function as a circadian time-keeper. The 2-site mechanism can be implemented using two relatively simple coupled non-linear ODEs in terms of site occupancy; the models do not require overly special fine-tuning of parameters for generating stable limit cycle oscillations.
In silico integration of thousands of epigenetic datasets into 707 cell type regulatory annotations improves the trans-ethnic portability of polygenic risk scores
Authors: Amariuta Tiffany , Ishigaki Kazuyoshi , Sugishita Hiroki , Ohta Tazro , Matsuda Koichi , Murakami Yoshinori , Price Alkes L. , Kawakami Eiryo , Terao Chikashi , Raychaudhuri Soumya
Published: February 2020

Poor trans-ethnic portability of polygenic risk score (PRS) models is a critical issue that may be partially due to limited knowledge of causal variants shared among populations. Hence, leveraging noncoding regulatory annotations that capture genetic variation across populations has the potential to enhance the trans-ethnic portability of PRS. To this end, we constructed a unique resource of 707 cell-type-specific IMPACT regulatory annotations by aggregating 5,345 public epigenetic datasets to predict binding patterns of 142 cell-type-regulating transcription factors across 245 cell types. With this resource, we partitioned the common SNP heritability of diverse polygenic traits and diseases from 111 GWAS summary statistics of European (EUR, average N=180K) and East Asian (EAS, average N=157K) origin. For 95 traits, we were able to identify a single IMPACT annotation most strongly enriched for trait heritability. Across traits, these annotations captured an average of 43.3% of heritability (se = 13.8%) with the top 5% of SNPs. Strikingly, we observed highly concordant polygenic trait regulation between populations: the same regulatory annotations captured statistically indistinguishable SNP heritability (fitted slope = 0.98, se = 0.04). Since IMPACT annotations capture both large and consistent proportions of heritability across populations, prioritizing variants in IMPACT regulatory elements may improve the trans-ethnic portability of PRS. Indeed, we observed that EUR PRS models more accurately predicted 21 tested phenotypes of EAS individuals when variants were prioritized by key IMPACT tracks (49.9% mean relative increase in R2). Notably, the improvement afforded by IMPACT was greater in the trans-ethnic EUR-to-EAS PRS application than in the EAS-to-EAS application (47.3% vs 20.9%, P < 1.7e-4). Overall, our study identifies a crucial role for functional annotations such as IMPACT to improve the trans-ethnic portability of genetic data, and this has important implications for future risk prediction models that work across populations.
High-throughput cellular-resolution synaptic connectivity mappingin vivowith concurrent two-photon optogenetics and volumetric Ca2+imaging
Authors: McRaven Christopher , Tanese Dimitrii , Zhang Lixia , Yang Chao-Tsung , Ahrens Misha B. , Emiliani Valentina , Koyama Minoru
Published: February 2020

The ability to measure synaptic connectivity and properties is essential for understanding neuronal circuits. However, existing methods that allow such measurements at cellular resolution are laborious and technically demanding. Here, we describe a system that allows such measurements in a high-throughput way by combining two-photon optogenetics and volumetric Ca2+imaging with whole-cell recording. We reveal a circuit motif for generating fast undulatory locomotion in zebrafish.
Inhibitory parvalbumin basket cell activity is selectively reduced during hippocampal sharp wave ripples in a mouse model of familial Alzheimer’s disease
Authors: Caccavano Adam , Bozzelli P. Lorenzo , Forcelli Patrick A. , Pak Daniel T.S. , Wu Jian-Young , Conant Katherine , Vicini Stefano
Published: February 2020

Memory disruption in mild cognitive impairment (MCI) and Alzheimer’s disease (AD) is poorly understood, particularly at early stages preceding neurodegeneration. In mouse models of AD, there are disruptions to sharp wave ripples (SWRs), hippocampal population events with a critical role in memory consolidation. However, the micro-circuitry underlying these disruptions is under-explored. We tested if a selective reduction in parvalbumin-expressing (PV) inhibitory interneuron activity underlies hyperactivity and SWR disruption. We employed the 5xFAD model of familial AD crossed with mouse lines labeling excitatory pyramidal cells (PCs) and inhibitory PV cells. We observed a 33% increase in frequency, 58% increase in amplitude, and 8% decrease in duration of SWRs inex vivoslices from male and female 3-month 5xFAD mice versus littermate controls. 5xFAD mice of the same age were impaired in a hippocampal-dependent memory task. Concurrent with SWR recordings, we performed calcium imaging, cell-attached, and whole-cell recordings of PC and PV cells within the CA1 region. PCs in 5xFAD mice participated in enlarged ensembles, with superficial PCs having a higher probability of spiking during SWRs. Both deep and superficial PCs displayed an increased synaptic E/I ratio, suggesting a disinhibitory mechanism. In contrast, we observed a 46% spike rate reduction during SWRs in PV basket cells (PVBCs), while PV bistratified and axo-axonic cells were unimpaired. Excitatory synaptic drive to PVBCs was selectively reduced by 50%, resulting in decreased E/I ratio. Considering prior studies of intrinsic PV cell dysfunction in AD, these findings suggest alterations to the PC-PVBC micro-circuit also contribute to impairment.

Significance Statement

We demonstrate that a specific sub-type of inhibitory neuron, parvalbumin-expressing basket cells, have selectively reduced activity in a model of Alzheimer’s disease during activity critical for the consolidation of memory. These results identify a potential cellular target for therapeutic intervention to restore aberrant network activity in early amyloid pathology. While parvalbumin cells have previously been identified as a potential therapeutic target, this study for the first time recognizes that other parvalbumin neuronal sub-types, including bistratified and axo-axonic cells, are spared. These experiments are the first to record synaptic and spiking activity during sharp wave ripple events in early amyloid pathology and reveal that a selective decrease in excitatory synaptic drive to parvalbumin basket cells likely underlies reduced function.
An autoimmune disease risk variant has atransmaster regulatory effect mediated by IRF1 under immune stimulation
Authors: Brandt Margot , Kim-Hellmuth Sarah , Ziosi Marcello , Gokden Alper , Wolman Aaron , Lam Nora , Recinos Yocelyn , Hornung Veit , Schumacher Johannes , Lappalainen Tuuli
Published: February 2020

Functional mechanisms remain unknown for most genetic loci associated to complex human traits and diseases. In this study, we first mappedtrans-eQTLs in a data set of primary monocytes stimulated with LPS, and discovered that a risk variant for autoimmune disease, rs17622517 in an intron ofC5ORF56, affects the expression of the transcription factorIRF120 kb away. The cis-regulatory effect onIRF1is active under early immune stimulus, with a large number oftrans-eQTL effects across the genome under late LPS response. Using CRISPRi silencing, we showed that the SNP locus indeed functions as anIRF1enhancer with widespread transcriptional effects. Genome editing by CRISPR further indicated that rs17622517 is indeed a causal variant in this locus, and recapitulated the LPS-specifictrans-eQTL signal. Our results suggest that this common genetic variant affects inter-individual response to immune stimuli via regulation ofIRF1. For this autoimmune GWAS locus, our work provides evidence of the causal variant, demonstrates a condition-specific enhancer effect, identifiesIRF1as the likely causal gene incis, and indicates that overactivation of the downstream immune-related pathway may be the cellular mechanism increasing disease risk. This work not only provides rare experimental validation of a master-regulatorytrans-eQTL, but also demonstrates the power of eQTL mapping to build mechanistic hypotheses amenable for experimental follow-up using the CRISPR toolkit.
Divergence, gene flow and the origin of leapfrog geographic distributions: The history of color pattern variation inPhyllobatespoison-dart frogs
Authors: Márquez Roberto , Linderoth Tyler P. , Mejía-Vargas Daniel , Nielsen Rasmus , Amézquita Adolfo , Kronforst Marcus R.
Published: February 2020

The geographic distribution of phenotypic variation among closely related populations is a valuable source of information about the evolutionary processes that generate and maintain biodiversity. Leapfrog distributions, in which phenotypically similar populations are disjunctly distributed and separated by one or more phenotypically distinct populations, represent geographic replicates for the existence of a phenotype, and are therefore especially informative. These geographic patterns have mostly been studied from phylogenetic perspectives to understand how common ancestry and divergent evolution drive their formation. Other processes, such as gene flow between populations, have not received as much attention. Here we investigate the roles of divergence and gene flow between populations in the origin and maintenance of a leapfrog distribution inPhyllobatespoison frogs. We found evidence for high levels of gene flow between neighboring populations but not over long distances, indicating that gene flow between populations exhibiting the central phenotype may have a homogenizing effect that maintains their similarity, and that introgression between “leapfroging” taxa has not played a prominent role as a driver of phenotypic diversity inPhyllobates. Although phylogenetic analyses suggest that the leapfrog distribution was formed through independent evolution of the peripheral (i.e. leapfrogging) populations, the elevated levels of gene flow between geographically close populations poise alternative scenarios, such as the history of phenotypic change becoming decoupled from genome-averaged patterns of divergence, which we cannot rule out. These results highlight the importance of incorporating gene flow between populations into the study of geographic variation in phenotypes, both as a driver of phenotypic diversity and as a confounding factor of phylogeographic inferences.
Ovarian Cancer Risk Variants are Enriched in Histotype-Specific Enhancers that Disrupt Transcription Factor Binding Sites
Authors: Jones Michelle R. , Peng Pei-Chen , Coetzee Simon G. , Tyrer Jonathan , Reyes Alberto L. , de la Fuente Rosario I. Corona , Davis Brian , Chen Stephanie , Dezem Felipe , Seo Ji-Heui ,   Ovarian Cancer Association Consortium, Berman Benjamin P. , Freedman Matthew L. , Plummer Jasmine T. , Lawrenson Kate , Pharoah Paul , Hazelett Dennis J. , Gayther Simon A.
Published: February 2020

Quantifying the functional effects of complex disease risk variants can provide insights into mechanisms underlying disease biology. Genome wide association studies (GWAS) have identified 39 regions associated with risk of epithelial ovarian cancer (EOC). The vast majority of these variants lie in the non-coding genome, suggesting they mediate their function through the regulation of gene expression by their interaction with tissue specific regulatory elements (REs). In this study, by intersecting germline genetic risk data with regulatory landscapes of active chromatin in ovarian cancers and their precursor cell types, we first estimated the heritability explained by known common low penetrance risk alleles. The narrow sense heritabilityof both EOC overall and high grade serous ovarian cancer (HGSOCs) was estimated to be 5-6%. Partitioned SNP-heritability across broad functional categories indicated a significant contribution of regulatory elements to EOC heritability. We collated epigenomic profiling data for 77 cell and tissue types from public resources (Roadmap Epigenomics and ENCODE), and H3K27Ac ChIP-Seq data generated in 26 ovarian cancer-relevant cell types. We identified significant enrichment of risk SNPs in active REs marked by H3K27Ac in HGSOCs. To further investigate how risk SNPs in active REs influence predisposition to ovarian cancer, we used motifbreakR to predict the disruption of transcription factor binding sites. We identified 469 candidate causal risk variants in H3K27Ac peaks that break TF motifs (enrichment P-Value < 1×10−5compared to control variants). The most frequently broken motif was REST (P-Value = 0.0028), which has been reported as both a tumor suppressor and an oncogene. These systematic functional annotations with epigenomic data highlight the specificity of the regulatory landscape and demonstrate functional annotation of germline risk variants is most informative when performed in highly relevant cell types.
DNA methylation epigenetically regulates gene expression inBurkholderia cenocepacia
Authors: Vandenbussche Ian , Sass Andrea , Pinto-Carbó Marta , Mannweiler Olga , Eberl Leo , Coenye Tom
Published: February 2020

Respiratory tract infections by the opportunistic pathogenBurkholderia cenocepaciaoften lead to severe lung damage in cystic fibrosis (CF) patients. New insights in how to tackle these infections might emerge from the field of epigenetics, as DNA methylation has shown to be an important regulator of gene expression. The present study focused on two DNA methyltransferases (MTases) inB. cenocepaciastrains J2315 and K56-2, and their role in regulating gene expression.In silicopredicted DNA MTase genes BCAL3494 and BCAM0992 were deleted in both strains, and the phenotypes of the resulting deletion mutants were studied: deletion mutant ΔBCAL3494 showed changes in biofilm structure and cell aggregation, ΔBCAM0992 was less motile.B. cenocepaciawild type cultures treated with sinefungin, a known DNA MTase inhibitor, exhibited the same phenotype as DNA MTase deletion mutants. Single-Molecule Real-Time sequencing was used to characterize the methylome ofB. cenocepacia, including methylation at the origin of replication, and motifs CACAG and GTWWAC were identified as targets of BCAL3494 and BCAM0992, respectively. All genes with methylated motifs in their putative promoter region were identified and qPCR experiments showed an upregulation of several genes, including biofilm and motility related genes, in MTase deletion mutants with unmethylated motifs, explaining the observed phenotypes in these mutants. In summary, our data confirm that DNA methylation plays an important role in regulating the expression ofB. cenocepaciagenes involved in biofilm formation and motility.

Importance

CF patients diagnosed withB. cenocepaciainfections often experience rapid deterioration of lung function, known ascepacia syndrome. B. cenocepaciahas a large multi-replicon genome and a lot remains to be learned about regulation of gene expression in this organism. From studies in other (model) organisms, it is known that epigenetic changes through DNA methylation play an important role in this regulation. The identification ofB. cenocepaciagenes of which the expression is regulated by DNA methylation and identification of the regulatory systems involved in this methylation are likely to lead to new insights in how to tackleB. cenocepaciainfections in CF patients.
Red-light (670 nm) therapy reduces mechanical sensitivity and neuronal cell death, and alters glial responses following spinal cord injury in rats
Authors: Hu Di , Moalem-Taylor Gila , Potas Jason R
Published: February 2020

Individuals with spinal cord injury (SCI) often develop debilitating neuropathic pain, which may be driven by neuronal damage and neuroinflammation. We have previously demonstrated that treatment using 670 nm (red) light irradiation alters microglia/macrophage responses and alleviates mechanical hypersensitivity at 7-days post-injury. Here, we investigated the effect of red-light on the development of mechanical hypersensitivity, neuronal markers, and glial response in the subacute stage (days 1-7) following SCI. Wistar rats were subjected to a mild T10 hemi-contusion SCI or sham surgery followed by daily red-light treatment (30 min/day; 670 nm LED; 35mW/cm2) or sham treatment. Mechanical sensitivity of the rat dorsum was assessed from 1-day post-injury and repeated every second day. Spinal cords were collected at 1, 3, 5 and 7-days post-injury for analysis of myelination, neurofilament protein NF200 expression, neuronal cell death, reactive astrocytes (GFAP+ cells), interleukin1β (IL1β) expression, and inducible nitric oxide synthase (iNOS) production in IBA1+microglia/macrophages. Red-light treatment significantly reduced the cumulative mechanical sensitivity and the hypersensitivity incidence following SCI. This effect was accompanied by significantly reduced neuronal cell death, reduced astrocyte activation and reduced iNOS expression in IBA1+cells at the level of the injury. However, myelin and NF200 immunoreactivity and IL1β expression in GFAP+and IBA1+cells were not altered by red-light treatment. Thus, red-light therapy may represent a useful non-pharmacological approach for treating pain during the subacute period after SCI by decreasing neuronal loss and modulating the inflammatory glial response.
Creating outbred and inbred populations of haplodiploid mites to measure adaptive responses in the lab
Authors: Godinho Diogo P. , Cruz Miguel A. , de la Masselière Maud Charlery , Teodoro-Paulo Jéssica , Eira Cátia , Fragata Inês , Rodrigues Leonor R. , Zélé Flore , Magalhães Sara
Published: February 2020

Laboratory studies are often criticized for not being representative of processes occurring in natural populations. This can be partially mitigated by using lab populations that capture large amounts of variation. Additionally, many studies addressing adaptation of organisms to their environment are done with laboratory populations, using quantitative genetics or experimental evolution methodologies. Such studies rely on populations that are either highly outbred or inbred. However, the methodology underlying the generation of such biological resources are usually not explicitly documented.

Given their small size, short generation time, amenability to laboratory experimentation and knowledge of their ecological interactions, haplodiploid spider mites are becoming a widely used model organism. Here, we describe the creation of outbred populations of two species of spider mites,Tetranychus urticaeandT. evansi, obtained by performing controlled crosses between individuals from field-collected populations. Subsequently, from the outbred population ofT. evansi, we derived inbred lines, by performing several generations of sib-mating. These can be used to measure broad-sense heritability as well as correlations among traits. Finally, we outline an experimental evolution protocol that can be widely used in other systems. Sharing these biological resources with other laboratories and combining them with the available powerful genetic tools forT. urticae(and other species) will allow consistent and comparable studies that greatly contribute to our understanding of ecological and evolutionary processes.
SimpylCellCounter: An Automated Solution for Quantifying Cells in Brain Tissue
Authors: Bal Aneesh , Maureira Fidel , Arguello Amy A.
Published: February 2020

Rationale & Objective

Manual quantification of activated cells can provide valuable information about stimuli-induced changes within brain regions; however, this analysis remains time intensive. Therefore, we created SimpylCellCounter (SCC), an automated method to quantify cells that express Cfos protein, an index of neuronal activity, in brain tissue and benchmarked it against two widely-used methods: OpenColonyFormingUnit (OCFU) and ImageJ Edge Detection Macro (IMJM).

Methods

In Experiment 1, manually-obtained counts were compared to those detected via OCFU, IMJM and SCC. The absolute error in counts (manualversusautomated method) was calculated, and error types were categorized as false positives or negatives. In Experiment 2, performance analytics of OCFU, IMJM and SCC were compared. In Experiment 3, SCC performed analysis on images it was not trained on, to assess its general utility.

Results & Conclusions

We found SCC to be highly accurate and efficient in quantifying both cells with circular morphologies and those expressing Cfos. Additionally, SCC utilizes a new approach for counting overlapping cells with a pretrained convolutional neural network classifier. The current study demonstrates that SCC is a novel, automated tool to quantify cells in brain tissue, complementing current, open-sourced quantification methods designed to detect cellsin vitro.
The costs of competition: high social status males experience accelerated epigenetic aging in wild baboons
Authors: Anderson Jordan A. , Johnston Rachel A. , Lea Amanda J. , Campos Fernando A. , Voyles Tawni N. , Akinyi Mercy Y. , Alberts Susan C. , Archie Elizabeth A. , Tung Jenny
Published: February 2020

Aging, for virtually all life, is inescapable. However, within populations, biological aging rates vary. Understanding sources of variation in this process is central to understanding the biodemography of natural populations. We constructed a DNA methylation-based age predictor for an intensively studied wild baboon population in Kenya. Consistent with findings in humans, the resulting “epigenetic clock” closely tracks chronological age, but individuals are predicted to be somewhat older or younger than their known ages. Surprisingly, these deviations are not explained by the strongest predictors of lifespan in this population, early adversity and social integration. Instead, they are best predicted by male dominance rank: high-ranking males are predicted to be older than their true ages, and epigenetic age tracks changes in rank over time. Our results argue that achieving high rank for male baboons—the best predictor of reproductive success—imposes costs consistent with a “live fast, die young” life history strategy.
Specialized DNA structures act as genomic beacons for integration by evolutionarily diverse retroviruses
Authors: Kohio Hinissan P. , Ajoge Hannah O. , Coleman Macon D. , Ndashimye Emmanuel , Gibson Richard M. , Arts Eric J. , Barr Stephen D.
Published: February 2020

Retroviral integration site targeting is not random and plays a critical role in expression and long-term survival of the integrated provirus. To better understand the genomic environment surrounding retroviral integration sites, we performed an extensive comparative analysis of new and previously published integration site data from evolutionarily diverse retroviruses from seven genera, including different HIV-1 subtypes. We showed that evolutionarily divergent retroviruses exhibited distinct integration site profiles with strong preferences for non-canonical B-form DNA (non-B DNA). Whereas all lentiviruses and most retroviruses integrate within or near genes and non-B DNA, MMTV and ERV integration sites were highly enriched in heterochromatin and transcription-silencing non-B DNA features (e.g. G4, triplex and Z-DNA). Compared toin vitro-derived HIV-1 integration sites,in vivo-derived sites are significantly more enriched in transcriptionally silent regions of the genome and transcription-silencing non-B DNA features. Integration sites from individuals infected with HIV-1 subtype A, C or D viruses exhibited different preferences for non-B DNA and were more enriched in transcriptionally active regions of the genome compared to subtype B virus. In addition, we identified several integration site hotspots shared between different HIV-1 subtypes with specific non-B DNA sequence motifs present at these hotspots. Together, these data highlight important similarities and differences in retroviral integration site targeting and provides new insight into how retroviruses integrate into genomes for long-term survival.

Graphical Abstract

Schematic comparing integration site profiles from evolutionarily diverse retroviruses. Upper left, heatmaps showing the fold-enrichment (blue) and fold-depletion (red) of integration sites near non-B DNA features (lower left). Lower right, circa plot showing integration site hotspots shared between HIV-1 subtype A, B, C and D virus.
Lower Soil Carbon Loss Due to Persistent Microbial Adaptation to Climate Warming
Authors: Guo Xue , Gao Qun , Yuan Mengting , Wang Gangsheng , Zhou Xishu , Feng Jiajie , Shi Zhou , Hale Lauren , Wu Linwei , Zhou Aifen , Tian Renmao , Liu Feifei , Wu Bo , Chen Lijun , Jung Chang Gyo , Niu Shuli , Li Dejun , Xu Xia , Jiang Lifen , Escalas Arthur , Wu Liyou , He Zhili , Van Nostrand Joy D. , Ning Daliang , Liu Xueduan , Yang Yunfeng , Schuur Edward. A.G. , Konstantinidis Konstantinos T. , Cole James R. , Penton C. Ryan , Luo Yiqi , Tiedje James M. , Zhou Jizhong
Published: February 2020

Soil microbial respiration is an important source of uncertainty in projecting future climate and carbon (C) cycle feedbacks. Despite intensive studies for two decades, the magnitude, direction, and duration of such feedbacks are uncertain, and their underlying microbial mechanisms are still poorly understood. Here we examined the responses of soil respiration and microbial community structure to long-term experimental warming in a temperate grassland ecosystem. Our results indicated that the temperature sensitivity of soil microbial respiration (i.e.,Q10) persistently decreased by 12.0±3.7% across 7 years of warming. Integrated metagenomic and functional analyses showed that microbial community adaptation played critical roles in regulating respiratory acclimation. Incorporating microbial functional gene abundance data into a microbially-enabled ecosystem model significantly improved the modeling performance of soil microbial respiration by 5–19%, compared to the traditional non-microbial model. Model parametric uncertainty was also reduced by 55–71% when gene abundances were used. In addition, our modeling analyses suggested that decreased temperature sensitivity could lead to considerably less heterotrophic respiration (11.6±7.5%), and hence less soil C loss. If such microbially mediated dampening effects occur generally across different spatial and temporal scales, the potential positive feedback of soil microbial respiration in response to climate warming may be less than previously predicted.
Complete reconstruction of the unbinding pathway of an anticancer drug by conventional unbiased molecular dynamics simulation
Authors: Sohraby Farzin , Moghadam Mostafa Javaheri , Aliyar Masoud , Aryapour Hassan
Published: February 2020

Understanding the details of unbinding mechanism of small molecule drugs is an inseparable part of rational drug design. Reconstruction of the unbinding pathway of small molecule drugs, todays, can be achieved through molecular dynamics simulations. Nonetheless, simulating a process in which a drug unbinds from its receptor demands lots of time, mostly up to several milliseconds. This amount of time is neither reasonable nor affordable; therefore, many researchers utilize various biases that there are still many doubts about their trustworthiness. In this work we have utilized short-run simulations, replicas, to make such time-consuming process cost effective. By replicating those snapshots of the trajectories which, after careful analyses, were selected as potential candidates we increased our system’s efficiency considerably. As a matter of fact, we have implemented a sort of human bias, inspecting trajectories visually, to achieve multiple unbinding events. We would like to call this stratagem, replicating of potent snapshots, “rational sampling” as it is, in fact, benefiting from human logic. In our case, an anticancer drug, the dasatinib, completely unbounded from its target protein, c-Src kinase, in only 392.6 ns, and this was gained without applying any internal biases and potentials which can increase error level. Thus, we achieved important structural details that can alter our viewpoint as well as assist drug designers.
Enzyme Kinetics and Inhibition Parameters of Human Leukocyte Glucosylceramidase
Authors: Karataş Mesut , Doğan Şenol , Spahiu Emrulla , Ašić Adna , Bešić Larisa , Turan Yusuf
Published: February 2020

Glucosylceramidase (GCase) is a lysosomal enzyme that catalyzes the cleavage of β-glucosidic linkage of glucocerebroside (GC) into glucose and ceramide; thereby, plays an essential function in the degradation of complex lipids and the turnover of cellular membranes.

The growing list of 460 mutations in the gene coding for it—glucosylceramidase beta acid 1 (GBA1)—is reported to abolish its catalytic activity and decrease its enzyme stability, associating it with severe health conditions such as Gaucher disease (GD), Parkinson Disease and Dementia with Lewy bodies.

Although the three-dimensional structure of wild type glucosylceramidase is elucidated, little is known about its features in human cells. Moreover, alternative sources of GCase that prove to be effective in the treatment of diseases with enzyme treatment therapies, impose the need for simple and cost-effective procedures to study the enzyme behaviour. This work, for the first time, shows a well established, yet simple, cost- and time-efficient protocol for the study of GCase enzyme in human leukocytes by the artificial substrate PNPG. Characterization of the enzyme in human leukocytes for activation parameters (optimal pH, Km, and Vmax) and enzyme inhibition, was done. The results indicate that the optimum pH of GCase enzyme with PNPG is 5.1. The Kmand Vmaxvalues were 12.6mM and 333 U/mg, respectively. Gluconolactone successfully inhibits GCase in a competitive manner, with a Kivalue of 0.023 mM and IC50of 0.047 mM. Glucose inhibition was uncompetitive with a Kiof 1.94 mM and IC50of 55.3 mM. This is the first report for the inhibitory effect of glucose, δ-gluconolactone on leukocyte GCase activity.
Brainrender: a python-based software for visualizing anatomically registered data
Authors: Claudi F. , Tyson A. L. , Petrucco L. , Margrie T.W. , Portugues R. , Branco T.
Published: February 2020

The recent development of high-resolution three-dimensional (3D) digital brain atlases and high-throughput brain wide imaging techniques has fueled the generation of large datasets that can be registered to a common reference frame. This registration facilitates integrating data from different sources and resolutions to assemble rich multidimensional datasets. Generating insights from these new types of datasets depends critically on the ability to easily visualize and explore the data in an interactive manner. This is, however, a challenging task. Currently available software is dedicated to single atlases, model species or data types, and generating 3D renderings that merge anatomically registered data from diverse sources requires extensive development and programming skills. To address this challenge, we have developedbrainrender: a generic, open-source Python package for simultaneous and interactive visualization of multidimensional datasets registered to brain atlases.Brainrenderhas been designed to facilitate the creation of complex custom renderings and can be used programmatically or through a graphical user interface. It can easily render different data types in the same visualization, including user-generated data, and enables seamless use of different brain atlases using the same code base. In addition,brainrendergenerates high-quality visualizations that can be used interactively and exported as high-resolution figures and animated videos. By facilitating the visualization of anatomically registered data,brainrendershould accelerate the analysis, interpretation, and dissemination of brain-wide multidimensional data.
A zebrafish functional genomics model to investigate the role of human A20 variantsin vivo
Authors: Cultrone Daniele , Zammit W. Nathan , Self Eleanor , Postert Benno , Han Jeremy ZR , Bailey Jacqueline , Warren Joanna , Croucher David R , Kikuchi Kazu , Bogdanovic Ozren , Chtanova Tatyana , Hesselson Daniel , Grey Shane T.
Published: February 2020

Germline loss-of-function variation inTNFAIP3, encoding A20, has been implicated in a wide variety of autoinflammatory and autoimmune conditions, with acquired somatic missense mutations linked to cancer progression. Furthermore, human sequence data reveals that the A20 locus contains ~400 non-synonymous coding variants which are largely uncharacterised. The growing number of A20 coding variants with unknown function, but potential clinical impact, poses a challenge to traditional mouse-based approaches. Here we report the development of a novel functional genomics approach that utilises the new A20-deficient zebrafish (Danio rerio) model to investigate the impact ofTNFAIP3genetic variantsin vivo. Similar to A20-deficient mice, A20-deficient zebrafish are hyper-responsive to inflammatory triggers and exhibit spontaneous early lethality. While ectopic addition of human A20 rescued A20-null zebrafish from lethality, missense mutations at two conserved A20 residues, S381A and C243Y reversed this protective effect. Ser381 represents a phosphorylation site important for enhancing A20 activity that is abrogated by its mutation to alanine, or by a C243Y mutation that associates with human autoimmune disease. These data reveal an evolutionarily conserved role for A20, but also demonstrate how a zebrafish functional genomics pipeline can be utilized to investigate thein vivosignificance of medically relevant TNFAIP3 gene variants. This approach could be utilised to investigate genetic variation for other conserved genes.
Ecological memory preserves phage resistance mechanisms in bacteria
Authors: Skanata Antun , Kussell Edo
Published: February 2020

Defense mechanisms against pathogens are prevalent in nature, and their maintenance is critical for long-term survival of a species. Such mechanisms, which include CRISPR-mediated immunity in bacteria and the R genes in plants, carry substantial costs to organisms and can be rapidly lost when pathogens are eliminated. How a species preserves its molecular defenses despite their costs, in the face of variable pathogen levels, and across an ecology of localized patches remains a major unsolved problem in epidemiology and evolutionary biology. Using techniques of game theory and non-linear dynamical systems, we show that by maintaining a non-zero failure rate of immunity, hosts sustain sufficient levels of pathogen across an ecology to select against loss of the defense. Thisresistance switching strategyis evolutionarily stable, and provides a powerful evolutionary mechanism that maintains host-pathogen interactions and enables co-evolutionary dynamics in a wide range of systems.
Mapping proteolytic neo-N termini at the surface of living cells
Authors: Weeks Amy M. , Byrnes James R. , Lui Irene , Wells James A.
Published: February 2020

N terminomics is a powerful strategy for profiling proteolytic neo-N termini, but its application to cell surface proteolysis has been limited by the low relative abundance of plasma membrane proteins. Here we apply plasma membrane-targeted subtiligase variants to efficiently and specifically capture cell surface N termini in live cells. Using this approach, we sequenced 807 cell surface N termini and quantified changes in their abundance in response to stimuli that induce proteolytic remodeling of the cell surface proteome. This technology will facilitate greater understanding of extracellular protease biology and reveal neo-N termini biomarkers and targets in disease.