bluuebunny/crossref_metadata_2025 huggingface dataset:
Inference of a genome-wide protein-coding gene set of the inshore hagfishEptatretus burgeri
Authors: Yamaguchi Kazuaki , Hara Yuichiro , Tatsumi Kaori , Nishimura Osamu , Smith Jeramiah J. , Kadota Mitsutaka , Kuraku Shigehiro
Published: July 2020

The group of hagfishes (Myxiniformes) arose from agnathan (jawless vertebrate) lineages and is one of the only two extant cyclostome taxa, together with lampreys (Petromyzontiformes). Even though whole genome sequencing has been achieved for diverse vertebrate taxa, genome-wide sequence information has been highly limited for cyclostomes. Here we sequenced the genome of the inshore hagfishEptatretus burgeriusing DNA extracted from the testis, with a short-read sequencing platform, aiming at reconstructing a high-coverage coding gene catalogue. The obtained genome assembly, scaffolded with mate-pair reads and paired RNA-seq reads, exhibited an N50 scaffold length of 293 Kbp, which allowed the genome-wide prediction of coding genes. This computation resulted in the gene models whose completeness was estimated at the complete coverage of more than 83 % and the partial coverage of more than 93 % by referring to evolutionarily conserved single-copy orthologs. The high contiguity of the assembly and completeness of resulting gene models promises a high utility in various comparative analyses including phylogenomics and phylome exploration.
Loss of resistance to punishment of cocaine use after prior experience
Authors: Durand Audrey , Girardeau Paul , Freese Luana , Ahmed Serge H.
Published: July 2020

One behavioral feature of drug addiction is continued drug use despite awareness that this causes negative consequences. Attempts to model this feature in animals typically involve punishing drug self-administration with a brief electrical footshock and look for resistance to punishment. Though all individual animals eventually stop self-administering the drug with increasing intensity of punishment, some individuals do so at higher intensities than other individuals. The greater relative resistance to punishment of the former individuals is generally interpreted as evidence for a compulsion-like behavior. Here we show that resistance to footshock punishment is in fact not a stable individual behavioral feature. Specifically, when rats are retested for their resistance to increasing intensity of footshock punishment, they become much less resistant. As a result, they suppress their cocaine intake even when punished with an initially low and ineffective intensity. A series of original behavioral experiments reveals that this low resistance to footshock punishment is rapidly acquired after rats experience a punishment intensity that leads them to near-completely suppress their cocaine intake. Passive exposure to the same intensity does not induce this effect. Once acquired, low resistance to punishment persists during at least one month, but can nevertheless be extinguished by retesting rats on a daily basis. Interestingly, this acquired low resistance to footshock punishment does not generalize to a non-painful form of punishment (i.e., histamine) that is also seldom used in animal drug self-administration studies. We discuss some possible theoretical and methodological implications of these findings for future research on animal models of compulsion-like behavior.
Triplanar ensemble U-Net model for white matter hyperintensities segmentation on MR images
Authors: Sundaresan Vaanathi , Zamboni Giovanna , Rothwell Peter M. , Jenkinson Mark , Griffanti Ludovica
Published: July 2020

White matter hyperintensities (WMHs) have been associated with various cerebrovascular and neurodegenerative diseases. Reliable quantification of WMHs is essential for understanding their clinical impact in normal and pathological populations. Automated segmentation of WMHs is highly challenging due to heterogeneity in WMH characteristics between deep and periventricular white matter, presence of artefacts and differences in the pathology and demographics of populations. In this work, we propose an ensemble triplanar network that combines the predictions from three different planes of brain MR images to provide an accurate WMH segmentation. Also, the network uses anatomical information regarding WMH spatial distribution in loss functions for improving the efficiency of segmentation and to overcome the contrast variations between deep and periventricular WMHs. We evaluated our method on 5 datasets, of which 3 are part of a publicly available dataset (training data for MICCAI WMH Segmentation Challenge 2017 - MWSC 2017) consisting of subjects from three different cohorts. On evaluating our method separately in deep and periventricular regions, we observed robust and comparable performance in both regions. Our method performed better than most of the existing methods, including FSL BIANCA, and on par with the top ranking deep learning method of MWSC 2017.
Temperature Dependence of theKrokinobacterrhodopsin 2 Kinetics
Authors: Eberhardt P. , Slavov C. , Sörmann J. , Bamann C. , Braun M. , Wachtveitl J.
Published: July 2020

Here we applied target analysis to a temperature dependent flash photolysis dataset of the light-driven sodium ion pumpKrokinobacterrhodopsin 2 (KR2) at sodium pumping conditions. With an increase in temperature from 10 – 40 °C, the overall photocycle duration was accelerated by a factor of six, while single transitions like the L to M transition increased by a factor of 40. Using kinetic modeling with the Eyring constraint as well as spectral corrections on the datasets the spectral position as well as the equilibria of the different photointermediates could be resolved. The results provide further insight into KR2s photocycle and energetics.

STATEMENT OF SIGNIFICANCE

KR2 is the most prominent member of the new class of non-proton cation pumps, as it represents an interesting new optogenetic tool. Despite widespread biophysical investigations, the molecular mechanisms of light-induced sodium pumping in KR2 are still not sufficiently understood. Therefore, an expanded set of thermodynamic parameters is essential for a complete picture. Our study of the KR2 photocycle shows that different steps in the photocycle are affected differently by temperature changes. Rigorous data analysis provides strong evidence that the transient states observed in time-resolved experiments represent rather equilibria between the different photocycle intermediates than pure intermediates. Gaining access to the dynamics and energetics of KR2 helps to answer long standing open questions concerning the molecular mechanism of cation pumping.
Specific changes in sleep oscillations after blocking human metabotropic glutamate receptor 5 in the absence of altered memory function
Authors: Feld Gordon B. , Bergmann Til Ole , Alizadeh-Asfestani Marjan , Stuke Viola , Wriede Jan-Philipp , Soekadar Surjo , Born Jan
Published: July 2020

Background

Sleep consolidates declarative memory by repeated replay linked to the cardinal oscillations of NonREM sleep. However, there is so far little evidence of classical glutamatergic plasticity induced by this replay. Rather, we have previously reported that blocking NMDA or AMPA receptors does not affect sleep-dependent consolidation of declarative memory.

Aims

Investigate the role of metabotropic glutamate receptor 5 (mGluR5) on memory processing during sleep.

Methods

In two placebo-controlled within-subject cross-over experiments with 20 healthy humans each, we used fenobam to block mGluR5 during sleep. In Experiment I, participants learned word-pairs (declarative task) and a finger sequence (procedural task) in the evening, then received the drug and recall was tested in the next morning. To cover possible effects on synaptic renormalization processes during sleep, in Experiment II, participants learned new word-pairs in the morning after sleep.

Results/Outcomes

Surprisingly, fenobam neither reduced retention of memory across sleep nor new learning after sleep, although it severely altered sleep architecture and memoryrelevant EEG oscillations. In NonREM sleep, fenobam suppressed 12-15 Hz spindles but augmented 2-4 Hz delta waves, whereas in REM sleep it suppressed 4-8 Hz theta and 16-22 Hz beta waves. Notably, under Fenobam NonREM spindles became more consistently phase-coupled to the slow oscillation.

Conclusions/Interpretations

Our findings indicate that mGluR5-related plasticity is not essential for memory processing during sleep, even though mGlurR5 are strongly implicated in the regulation of the cardinal sleep oscillations.

Declaration of interest/Funding

The authors have nothing to disclose and funders had no influence on the research presented here.
A simple strategy to reduce the salivary gland and kidney uptake of PSMA targeting small molecule radiopharmaceuticals
Authors: Kalidindi Teja Muralidhar , Lee Sang-Gyu , Jou Katerina , Chakraborty Goutam , Skafida Myrto , Tagawa Scott T. , Bander Neil H. , Schoder Heiko , Bodei Lisa , Pandit-Taskar Neeta , Lewis Jason S. , Larson Steven M. , Osborne Joseph R. , Kishore Pillarsetty Naga Vara
Published: July 2020

The past five years have seen an increasing acceptance of peptide-based prostate-specific membrane antigen (PSMA)-targeted radionuclide therapy (TRT) agents for treatment of metastatic castration-resistant prostate cancer (mCRPC), with [177Lu]-DKFZ-PSMA-617 ([177Lu]-PSMA-617) emerging as the leading candidate. [177Lu]-PSMA-617 and other PSMA ligands have shown efficacy in reducing the tumor burden in mCRPC patients but irradiation to salivary gland and kidneys is a concern and dose limiting factor. Therefore, methods to reduce non-target organ toxicity are needed to safely treat patients and preserve their quality of life. Here, we report the effects of the addition of the cold PSMA ligand DKFZ-PSMA-11 (PSMA-11) on the uptake of [177Lu]-PSMA-617 in tumor, salivary glands and kidneys. Groups of athymic nude mice (n = 4) bearing PC3-PIP (PSMA+) tumor xenografts were administered with [177Lu]-PSMA-617 along with 0, 5, 100, 500, 1000 and 2000 pmoles of PSMA-11. Biodistribution studies 1 h post-administration revealed that [177Lu]-PSMA-617 uptake in PSMA-expressing PC3-PIP tumors was 21.71±6.13, 18.7±2.03, 26.44±2.94, 16.21±3.5, 13.52±3.68, and 12.03±1.96 %ID/g when 0, 5, 100, 500, 1000 and 2000 pmoles of PSMA-11 were added, respectively. Corresponding kidney uptake values were 123.14±52.52, 132.31±47.4, 84.29±78.25, 2.12±1.88, 1.16±0.36, 0.64±0.23 %ID/g, respectively. Corresponding salivary gland uptake values were 0.48±0.11, 0.45±0.15, 0.38±0.3, 0.08±0.03, 0.09±0.07, 0.05±0.02 % ID/g, respectively. Thus, uptake of PSMA TRT agents in salivary gland and kidney can be substantially reduced without impact on tumor uptake by adding cold PSMA-11. Our data provides proof-of-concept and we propose that similar strategy be pursued in future clinical trials to prevent xerostomia and renal toxicity arising from [177Lu]-PSMA-617.
WaveletSEG: Automatic wavelet-based 3D nuclei segmentation and analysis for multicellular embryo quantification
Authors: Wu Tzu-Ching , Wang Xu , Li Linlin , Bu Ye , Umulis David M.
Published: July 2020

Identification of individual cells in tissues, organs, and in various developing systems is a well-studied problem because it is an essential part of objectively analyzing quantitative images in numerous biological contexts. We developed a size-dependent wavelet-based segmentation method that provides robust segmentation without any preprocessing, filtering or fine-tuning steps, and is robust to the signal-to-noise ratio (SNR). The wavelet-based method achieves robust segmentation results with respect to True Positive rate, Precision, and segmentation accuracy compared with other commonly used methods. We applied the segmentation program to zebrafish embryonic development IN TOTO for nuclei segmentation, image registration, and nuclei shape analysis. These new approaches to segmentation provide a means to carry out quantitative patterning analysis with single-cell precision throughout three dimensional tissues and embryos and they have a high tolerance for non-uniform and noisy image data sets.
A Simple Optimization Workflow to Enable Precise and Accurate Imputation of Missing Values in Proteomic Datasets
Authors: Dabke Kruttika , Kreimer Simion , Jones Michelle R. , Parker Sarah J.
Published: July 2020

Missing values in proteomic data sets have real consequences on downstream data analysis and reproducibility. Although several imputation methods exist to handle missing values, there is no single imputation method that is best suited for a diverse range of data sets and no clear strategy exists for evaluating imputation methods for large-scale DIA-MS data sets, especially at different levels of protein quantification. To navigate through the different imputation strategies available in the literature, we have established a workflow to assess imputation methods on large-scale label-free DIA-MS data sets. We used two distinct DIA-MS data sets with real missing values to evaluate eight different imputation methods with multiple parameters at different levels of protein quantification; dilution series data set and an independent data set with actual experimental samples. We found that imputation methods based on local structures within the data, like local least squares (LLS) and random forest (RF), worked well in our dilution series data set whereas, imputation methods based on global structures within the data, like BPCA performed well in our independent data set. We also found that imputation at the most basic level of protein quantification – fragment level-improved accuracy and number of proteins quantified. Overall, this study indicates that the most suitable imputation method depends on the overall structure and correlations of proteins within the data set and can be identified with the workflow presented here.
Evaluating SARS-CoV-2 spike and nucleocapsid proteins as targets for IgG antibody detection in severe and mild COVID-19 cases using a Luminex bead-based assay
Authors: Mariën Joachim , Michiels Johan , Heyndrickx Leo , Kerkhof Karen , Foque Nikki , Widdowson Marc-Alain , Desombere Isabelle , Jansens Hilde , Van Esbroeck Marjan , Ariën Kevin K.
Published: July 2020

Large-scale serosurveillance of severe acute respiratory syndrome coronavirus type 2 (SARS- CoV-2) will only be possible if serological tests are sufficiently reliable, rapid and inexpensive. Current assays are either labour-intensive and require specialised facilities (e.g. virus neutralization assays), or expensive with suboptimal specificity (e.g. commercial ELISAs). Bead-based assays offer a cost-effective alternative and allow for multiplexing to test for antibodies of other pathogens. Here, we compare the performance of four antigens for the detection of SARS-CoV-2 specific IgG antibodies in a panel of sera that includes both severe (n=40) and mild (n=52) cases, using a neutralization and a Luminex bead-based assay. While we show that neutralising antibody levels are significantly lower in mild than in severe cases, we demonstrate that a combination of recombinant nucleocapsid protein (NP), receptor- binding domain (RBD) and the whole spike protein (S1S2) results in a highly sensitive (96%) and specific (99%) bead-based assay that can detect IgG antibodies in both groups. Although S1-specific IgG levels correlate most strongly with neutralizing antibody levels, they fall below the detection threshold in 10% of the cases in our Luminex assay. In conclusion, our data supports the use of RBD, NP and S1S2 for the development of SARS-CoV-2 serological bead- based assays. Finally, we argue that low antibody levels in mild/asymptomatic cases might complicate the epidemiological assessment of large-scale surveillance studies.
Assessing the quality of nontraditional N95 filtering face-piece respirators available during the COVID-19 pandemic
Authors: Plana Deborah , Tian Enze , Cramer Avilash K. , Yang Helen , Carmack Mary M. , Sinha Michael S. , Bourgeois Florence T. , Yu Sherry H. , Masse Peter , Boyer Jon , Kim Minjune , Mo Jinhan , LeBoeuf Nicole R. , Li Ju , Sorger Peter K.
Published: July 2020

Background

During the current COVID-19 pandemic, supply chains for Personal Protective Equipment (PPE) have been severely disrupted and many products, particularly surgical N95 filtering facepiece respirators (FFRs; “masks”) are in short supply. As a consequence, an Emergency Use Authorization (EUA) from the FDA has allowed importation of N95-type masks manufactured to international standards; these include KN95 masks from China and FFP2 masks from the European Union.

Methods

We conducted a survey of mask in the inventory of major academic medical centers in Boston, MA to determine provenance and manufacturer. We then assembled a simple apparatus for performing a necessary (but not sufficient) test of filtration performance and tested masks from the inventory; an accompanying website shows how to build and use the testing apparatus.

Results

Our survey showed that, seven months after the start of the COVID-19 pandemic, over 100 different makes and models of N95-type masks are in the inventory of local hospitals as opposed to 2-5 models under normal circumstances. A substantial number of unfamiliar masks are from unknown manufacturers. Many did not perform to accepted standards and are likely to be counterfeit. Due to the absence of publicly available information on mask suppliers in the FDA EUA and confusing or inconsistent labeling of KN95 masks, it is difficult to distinguish legitimate and counterfeit products.

Conclusions

Many of the FFR masks available for procurement during the COVID-19 pandemic do not provide levels of fit and filtration similar to those of N95 masks and are not acceptable for use in healthcare settings. Based on these results, and in consultation with occupational health officers, we make six recommendations for end users to assist in acquiring legitimate products. In particular, institutions should always assess masks from non-traditional supply chains by checking their markings and manufacturer information against data provided by NIOSH and the latest FDA EUA Appendix A. In the absence of verifiable information on the legitimacy of mask source, institutions should consider measuring mask fit and filtration directly. We also make suggestions for U.S and Chinese regulatory agencies with regard to labeling and public disclosure aimed at increase pandemic resilience.
Enabling out-of-clinic human immunity studies via single-cell profiling of capillary blood
Authors: Dobreva Tatyana , Brown David , Park Jong Hwee , Thomson Matt
Published: July 2020

An individual’s immune system is driven by both genetic and environmental factors that vary over time. To better understand the temporal and inter-individual variability of gene expression within distinct immune cell types, we developed a platform that leverages multiplexed single-cell sequencing and out-of-clinic capillary blood extraction to enable simplified, cost-effective profiling of the human immune system across people and time at single-cell resolution. Using the platform, we detect widespread differences in cell type-specific gene expression between subjects that are stable over multiple days.

Increasing evidence implicates the immune system in an overwhelming number of diseases, and distinct cell types play specific roles in their pathogenesis.1,2Studies of peripheral blood have uncovered a wealth of associations between gene expression, environmental factors, disease risk, and therapeutic efficacy.4For example, in rheumatoid arthritis, multiple mechanistic paths have been found that lead to disease, and gene expression of specific immune cell types can be used as a predictor of therapeutic non-response.12Furthermore, vaccines, drugs, and chemotherapy have been shown to yield different efficacy based on time of administration, and such findings have been linked to the time-dependence of gene expression in downstream pathways.21,22,23However, human immune studies of gene expression between individuals and across time remain limited to a few cell types or time points per subject, constraining our understanding of how networks of heterogeneous cells making up each individual’s immune system respond to adverse events and change over time.
Exploring Efficient Linear Mixed Models to Detect Quantitative Trait Locus-by-Environment Interactions
Authors: Yamamoto Eiji , Matsunaga Hiroshi
Published: July 2020

Genotype-by-environment interactions (G×E) are important for understanding genotype–phenotype relationships. To date, various statistical models have been proposed to account for G×E effects, especially in genomic selection (GS) studies. Generally, GS does not focus on the detection of each quantitative trait locus (QTL), while the genome-wide association study (GWAS) was designed for QTL detection. G×E modeling methods in GS can be included as covariates in GWAS using unified linear mixed models (LMMs). However, the efficacy of G×E modeling methods in GS studies has not been evaluated for GWAS. In this study, we performed a comprehensive comparison of LMMs that integrate the G×E modeling methods to detect both QTL and QTL-by-environment interaction (Q×E) effects. Model efficacy was evaluated using simulation experiments. For the fixed effect terms representing Q×E effects, simultaneous scoring of specific and non-specific environmental effects was recommended because of the higher recall and improved genomic inflation factor value. For random effects, it was necessary to account for both G×E and genotype-by-trial (G×T) effects to control genomic inflation factor value. Thus, the recommended LMM includes fixed QTL effect terms that simultaneously score specific and non-specific environmental effects and random effects accounting for both G×E and G×T. The LMM was applied to real tomato phenotype data obtained from two different cropping seasons. We detected not only QTLs with persistent effects across the cropping seasons but also QTLs with Q×E effects. The optimal LMM identified in this study successfully detected more QTLs with Q×E effects.
Spatially guided distractor suppression during visual search
Authors: Feldmann-Wüstefeld Tobias , Weinberger Marina , Awh Edward
Published: July 2020

Past work has demonstrated that active suppression of salient distractors is a critical part of visual selection. Evidence for goal-driven suppression includes below-baseline visual encoding at the position of salient distractors (Gaspelin and Luck, 2015) and neural signals such as the Pd that track the position and number of distractors in the visual field (Feldmann-Wustefeld and Vogel, 2019). One basic question regarding distractor suppression is whether it is inherently spatial or nonspatial in character. Indeed, past work has shown that distractors evoke both spatial (Theeuwes, 1992) and nonspatial forms of interference (Folk and Remington, 1998), motivating a direct examination of whether space is integral to goal-driven distractor suppression. Here, we provide clear evidence for a spatial gradient of suppression surrounding salient singleton distractors. Replicating past work, both reaction time and neural indices of target selection improved monotonically as the distance between target and distractor increased. Importantly, these target selection effects were paralleled by a monotonic decline in the amplitude of the Pd, an electrophysiological index of distractor suppression. Moreover, multivariate analyses revealed spatially selective activity in the theta band that tracked the position of the target and – critically – revealed suppressed activity at spatial channels centered on distractor positions. Thus, goal-driven selection of relevant over irrelevant information benefits from a spatial gradient of suppression surrounding salient distractors.
Neuroanatomical underpinning of diffusion kurtosis measurements in the cerebral cortex of healthy macaque brains
Authors: Zhu Tianjia , Peng Qinmu , Ouyang Austin , Huang Hao
Published: July 2020

Purpose

To investigate the neuroanatomical underpinning of healthy macaque brain cortical microstructure measured by diffusion kurtosis imaging (DKI) which characterizes non-Gaussian water diffusion.

Methods

High-resolution DKI was acquired from 6 postmortem macaque brains. Neurofilament density (ND) was quantified based on structure tensor from neurofilament histological images of a different macaque brain sample. After alignment of DKI-derived mean kurtosis (MK) maps to the histological images, MK and histology-based ND were measured at corresponding regions of interests characterized by distinguished cortical MK values in the prefrontal/precentral-postcentral and temporal cortices. Pearson correlation was performed to test significant correlation between these cortical MK and ND measurements.

Results

Heterogeneity of cortical MK across different cortical regions was revealed, with significantly and consistently higher MK measurements in the prefrontal/precentral-postcentral cortex compared to those in the temporal cortex across all 6 scanned macaque brains. Corresponding higher ND measurements in the prefrontal/precentral-postcentral cortex than in the temporal cortex were also found. The heterogeneity of cortical MK is associated with heterogeneity of histology-based ND measurements, with significant correlation between cortical MK and corresponding ND measurements (P <0.005).

Conclusion

These findings suggested that DKI-derived MK can potentially be an effective noninvasive biomarker quantifying underlying neuroanatomical complexity inside the cerebral cortical mantle for clinical and neuroscientific research.
AggreCount: An unbiased image analysis tool for identifying and quantifying cellular aggregates in a spatial manner
Authors: Klickstein Jacob Aaron , Mukkavalli Sirisha , Raman Malavika
Published: July 2020

Protein quality control is maintained by a number of integrated cellular pathways that monitor the folding and functionality of the cellular proteome. Defects in these pathways lead to the accumulation of misfolded or faulty proteins that may become insoluble and aggregate over time. Protein aggregates significantly contribute to the development of a number of human diseases such as Amyotrophic lateral sclerosis, Huntington’s and Alzheimer’s Disease.In vitro, imaging-based, cellular studies have defined key components that recognize and clear aggregates; however, no unifying method is available to quantify cellular aggregates. Here we describe an ImageJ macro called AggreCount to identify and measure protein aggregates in cells. AggreCount is designed to be intuitive, easy to use and customizable for different types of aggregates observed in cells. Minimal experience in coding is required to utilize the script. Based on a user defined image, AggreCount will report a number of metrics: (i) total number of cellular aggregates, (ii) percent cells with aggregates, (iii) aggregates per cell, (iv) area of aggregates and (v) localization of aggregates (cytosol, perinuclear or nuclear). A data table of aggregate information on a per cell basis as well as a summary table is provided for further data analysis. We demonstrate the versatility of AggreCount by analyzing a number of different cellular aggregates including aggresomes, stress granules and inclusion bodies caused by Huntingtin polyQ expansion.
Arabidopsis thalianainteraction withEnsifer melilotican support plant growth under N-deficiency
Authors: Armijo Grace , Kraiser Tatiana , Medina María P. , Gras Diana E. , Zúñiga Ana , González Bernardo , Gutiérrez Rodrigo A.
Published: July 2020

Nitrogen (N) is an essential macronutrient for plants. Some plant species obtain this nutrient by interacting with N-fixing bacteria. These beneficial interactions are well described in legumes but have also been observed in non-legume plant species that are unable to form root nodules. We studied the expanding role of beneficial plant-bacteria interactions for N-nutrition in the widely used model plantArabidopsis thaliana. We found that the bacteriaEnsifer melilotienhancedA. thalianagrowth under severe N-deficiency conditions, allowing plants to complete their life cycle. Our results showed that bacteria colonize the rhizosphere associated with the epidermis of the plant root. We also demonstrated thatA. thalianapossesses genes that are critical for this beneficial interaction and are required for plant-growth promotion byE. melilotiunder N-deficiency.

This work shows association betweenA. thalianaandE. melilotifor plant nutrition under severe N-deficiency, and suggests that plants have conserved-molecular mechanisms to interact with N-fixing bacteria to procure N and escape adverse conditions. Under these circumstances, the supply of N via N-fixation is critical for survival, allowing the plant to complete its life cycle. Our findings provide a new framework and an experimental model system that expand our understanding of plant-rhizobia interactions for plant N-nutrition.
A 3-D Constitutive Model for Finite Element Analyses of Agarose with a Range of Gel Concentrations
Authors: Wang Xiaogang , June Ronald K. , Pierce David M.
Published: July 2020

Hydrogels have seen widespread application across biomedical sciences and there is considerable interest in using hydrogels, including agarose, for creatingin vitrothree-dimensional environments to grow cells and study mechanobiology and mechanotransduction. Recent advances in the preparation of agarose gels enable successful encapsulation of viable cells at gel concentrations as high as 5%. Agarose with a range of gel concentrations can thus serve as an experimental model mimicking changes in the 3-D microenvironment of cells during disease progression and can facilitate experiments aimed at probing the corresponding mechanobiology, e.g. the evolving mechanobiology of chondrocytes during the progression of osteoarthritis. Importantly, whether stresses (forces) or strains (displacement) drive mechanobiology and mechanotransduction is currently unknown. We can use experiments to quantify mechanical properties of hydrogels, and imaging to estimate microstructure and even strains; however, only computational models can estimate intra-gel stresses in cell-seeded agarose constructs because the requiredin vitroexperiments are currently impossible. Finite element modeling is well-established for (computational) mechanical analyses, but accurate constitutive models for modeling the 3-D mechanical environments of cells within high-stiffness agarose are currently unavailable. In this study we aimed to establish a 3-D constitutive model of high-stiffness agarose with a range of gel concentrations. We applied a multi-step, physics-based optimization approach to separately fit subsets of model parameters and help achieve robust convergence. Our constitutive model, fitted to experimental data on progressive stress-relaxations, was able to predict reaction forces determined from independent experiments on cyclical loading. Our model has broad applications in finite element modeling aimed at interpreting mechanical experiments on agarose specimens seeded with cells, particularly in predicting distributions of intra-gel stresses. Our model and fitted parameters enable more accurate finite element simulations of high-stiffness agarose constructs, and thus better understanding of experiments aimed at mechanobiology, mechanotransduction, or other applications in tissue engineering.
Full genome characterization of Laem Singh virus (LSNV) in shrimpPenaeus monodon
Authors: Taengchaiyaphum Suparat , Srisala Jiraporn , Sanguanrut Piyachat , Ongvarrasopone Chalermporn , Flegel Timothy W. , Sritunyalucksana Kallaya
Published: July 2020

Laem Singh virus (LSNV) was discovered in 2006 and proposed as a necessary but insufficient cause of retarded growth in the giant tiger shrimpPenaeus monodon.Its closest relatives were plant viruses including an unassignedSobemovirusand viruses in the familyLuteoviridae. During succeeding years, attempts to obtain the full LSNV genome sequence by genome walking failed. However, recent publication of the full sequence of Wenzhou shrimp virus 9 (WZSV 9) at GenBank revealed that LSNV sequences in our database shared 99% sequence identity with it. Thus, we hypothesized that LSNV and WZSV 9 were different isolates of the same virus species. Here we confirm that hypothesis by cloning and sequencing of the full genome of LSNV fromP. monodonand by showing that it consists of two fragments each with 99% identity to the matching fragments of WZSV.
Differential activation of JAK-STAT signaling in blood cell progenitors reveals functional compartmentalization of theDrosophilalymph gland
Authors: Rodrigues Diana , Renaud Yoan , VijayRaghavan K. , Waltzer Lucas , Inamdar Maneesha S.
Published: July 2020

Blood cells arise from diverse pools of stem and progenitor cells. Understanding progenitor heterogeneity is a major challenge. TheDrosophilalarval lymph gland is a well-studied model to understand blood progenitor maintenance and recapitulates several aspects of vertebrate hematopoiesis. However in-depth analysis has focused on progenitors located in lymph gland anterior lobes (AP), ignoring the progenitors from the posterior lobes (PP). Usingin situexpression mapping and transcriptome analysis we reveal PP heterogeneity and identify molecular-genetic tools to study this abundant progenitor population. Functional analysis shows that PP resist differentiation upon immune challenge, in a JAK-STAT-dependent manner. Upon wasp parasitism, AP downregulate JAK-STAT signaling and form lamellocytes. In contrast, we show that PP activate STAT92E and remain undifferentiated.Stat92Eknockdown in PP or genetically reducing JAK-STAT signaling permits PP lamellocyte differentiation. We discuss how heterogeneity and compartmentalization allow functional segregation in response to systemic cues and could be widely applicable.

Highlights

We provide anin situand transcriptome map of larval blood progenitors Posterior lymph gland progenitors are refractory to immune challenge STAT activation after wasp parasitism maintains posterior progenitors
The SARS-CoV-2 Spike Variant D614G Favors an Open Conformational State
Authors: Mansbach Rachael A. , Chakraborty Srirupa , Nguyen Kien , Montefiori David C. , Korber Bette , Gnanakaran S.
Published: July 2020

The COVID-19 pandemic underwent a rapid transition with the emergence of a SARS-CoV-2 variant that carried the amino acid substitution D614G in the Spike protein that became globally prevalent. The G-form is both more infectiousin vitroand associated with increased viral loads in infected people. To gain insight into the mechanism underlying these distinctive characteristics, we employed multiple replicas of microsecond all-atom simulations to probe the molecular-level impact of this substitution on Spike’s closed and open states. The open state enables Spike interactions with its human cellular receptor, ACE2. Here we show that changes in the inter-protomer energetics due to the D614G substitution favor a higher population of infection-capable (open) states. The inter-protomer interactions between S1 and S2 subunits in the open state of the D-form are asymmetric. This asymmetry is resolved in the G-form due to the release of tensile hydrogen bonds resulting in an increased population of open conformations. Thus, the increased infectivity of the G-form is likely due to a higher rate of profitable binding encounters with the host receptor. It is also predicted to be more neutralization sensitive due to enhanced exposure of the receptor binding domain, a key target region for neutralizing antibodies.
Regulated distribution of mitochondria in touch receptor neurons ofC. elegansinfluences touch response
Authors: Awasthi Anjali , Modi Souvik , Hegde Sneha , Chatterjee Anusheela , Mondal Sudip , Romero Eva , Sure Guruprasad R. , Koushika Sandhya P.
Published: July 2020

Density of mitochondria and their localization at specific sub-cellular regions of the neurons is regulated by molecular motors, their adaptors and the cytoskeleton. However, the regulation of the mitochondrial density, the positioning of mitochondria along the neuronal process and the role of axonal mitochondria in neuronal function remain poorly understood. This study shows that the density of mitochondria inC. eleganstouch receptor neuron processes remains constant through development. Simulations show that mitochondrial positioning along parts of the neuronal process that are devoid of synapses is regulated. Additionally, we also demonstrate that axonal mitochondria are necessary for maintaining touch responsiveness.
Using genetically encoded heme sensors to probe the mechanisms of heme uptake and homeostasis inCandida albicans
Authors: Weissman Ziva , Pinsky Mariel , Donegan Rebecca K. , Reddi Amit R. , Kornitzer Daniel
Published: July 2020

Candida albicansis a major fungal pathogen that can utilize hemin and hemoglobin as iron sources in the iron-scarce host environment. WhileC. albicansis a heme prototroph, we show here that it can also efficiently utilize external heme as a cellular heme source. Using genetically encoded ratiometric fluorescent heme sensors, we show that heme extracted from hemoglobin and free hemin enter the cells with different kinetics. Heme supplied as hemoglobin is taken up via the CFEM (Common in Fungal Extracellular Membrane) hemophore cascade, and reaches the cytoplasm over several hours, whereas entry of free hemin via CFEM-dependent and independent pathways is much faster, less than an hour. To prevent an influx of extracellular heme from reaching toxic levels in the cytoplasm, the cells deploy Hmx1, a heme oxygenase. Hmx1 was previously suggested to be involved in utilization of hemoglobin and hemin as iron sources, but we find that it is primarily required to prevent heme toxicity. Taken together, the combination of novel heme sensors with genetic analysis revealed new details of the fungal mechanisms of heme import and homeostasis, necessary to balance the uses of heme as essential cofactor and potential iron source against its toxicity.
Fluffy feathers: how neoptile feathers contribute to camouflage in precocial chicks
Authors: Rohr Veronika A. , Volkmer Tamara , Metzler Dirk , Küpper Clemens
Published: July 2020

Camouflage is a widespread strategy to increase survival. The plumage of precocial chicks often contains elements of disruptive colouration and background matching to enhance concealment. Chick plumage also features fringed feathers as appendages that may contribute to camouflage. Here, we examine whether and how neoptile feathers conceal the outline of chicks. We first conducted a digital experiment to test two potential mechanisms for outline diffusion through appendages: 1) edge intensity reduction and 2) luminance transition. Local Edge Intensity Analysis (LEIA) showed that appendages decreased edge intensity and a mean luminance comparison revealed that the appendages created an intermediate transition zone to conceal the object’s outline. The outline was most diffused through an intermediate number of interspersed thin appendages. Increased appendage thickness resulted in fewer appendages improving camouflage, whereas increased transparency required more appendages for best concealment. For edge intensity, the outline diffusion was strongest for a vision system with low spatial acuity, which is characteristic of many mammalian predators. We then analysed photographs of young snowy plover (Charadrius nivosus) chicks to examine whether neoptile feathers increase outline concealment in a natural setting. Consistent with better camouflage, the outline of digitally cropped chicks with protruding feathers showed lower edge intensities than the outline of chicks cropped without those feathers. However, the observed mean luminance changes were not consistent with better concealment. Taken together, our results suggest that thin skin appendages such as neoptile feathers improve camouflage. As skin appendages are widespread, this mechanism may apply to a large variety of organisms.
Structure of voltage-modulated sodium-selective NALCN-FAM155A channel complex
Authors: Kang Yunlu , Wu Jing-Xiang , Chen Lei
Published: July 2020

Resting membrane potential determines the excitability of the cell and is essential for the cellular electrical activities. NALCN channel mediates sodium leak currents, which positively tune the resting membrane potential towards depolarization. NALCN channel is involved in many important neurological processes and is implicated in a spectrum of neurodevelopmental diseases. Despite its functional importance, the mechanisms of ion permeation and voltage-modulation for NALCN channel remain elusive. Here, we report the cryo-EM structure of rat NALCN and mouse FAM155A complex to 2.7 Å resolution. The structure reveals detailed interactions between NALCN and extracellular cysteine-rich domain of FAM155A. The non-canonical architecture of NALCN selectivity filter dictates its sodium selectivity and calcium block. The asymmetric arrangement of two functional voltage-sensors confers the modulation by membrane potential. Moreover, mutations found in human diseases were mapped to the domain-domain interfaces or the pore domain of NALCN, intuitively suggesting their pathological mechanisms.
Practical Design and Implementation of Animal Movements Tracking System for Neuroscience Trials
Authors: Sorkhabi Majid Memarian
Published: July 2020

Abstract

Background

The nervous system functions of an animal are predominantly reflected in the behaviour and the movement, therefore the movement-related data and measuring behavior quantitatively are crucial for behavioural analyses. The animal movement is traditionally recorded, and human observers follow the animal behaviours; if they recognize a certain behaviour pattern, they will note it manually, which may suffer from observer fatigue or drift.

Objective

Automating behavioural observations with computer-vision algorithms are becoming essential equipment to the brain function characterization in neuroscience trials. In this study, the proposed tracking module is eligible to measure the locomotor behaviour (such as speed, distance, turning) over longer time periods that the operator is unable to precisely evaluate. For this aim, a novel animal cage is designed and implemented to track the animal movement. The frames received from the camera are analyzed by the 2D bior 3.7 Wavelet transform and SURF feature points.

Results

Implemented video tracking device can report the location, duration, speed, frequency and latency of each behavior of an animal. Validation tests were conducted on the auditory stimulation trial and the magnetic stimulation treatment of hemi-Parkinsonian rats.

Conclusion/ Significance

The proposed toolkit can provide qualitative and quantitative data on animal behaviour in an automated fashion, and precisely summarize an animal’s movement at an arbitrary time and allows operators to analyse movement patterns without requiring to check full records for every experiment.
ClpAP proteolysis does not require rotation of the ClpA unfoldase relative to ClpP
Authors: Kim Sora , Zuromski Kristin L , Bell Tristan A , Sauer Robert T , Baker Tania A
Published: July 2020

AAA+ proteases, which perform regulated protein degradation in all kingdoms of life, consist of a hexameric AAA+ unfoldase/translocase in complex with a self-compartmentalized peptidase. Based on asymmetric features of cryo-EM structures and a sequential hand-over-hand model of substrate translocation, recent publications have proposed that the AAA+ unfoldases ClpA and ClpX must rotate with respect to their partner peptidase ClpP to allow function. Here, we test this model by covalently crosslinking ClpA to ClpP to prevent rotation. We find that crosslinked ClpAP omplexes unfold, translocate, and degrade protein substrates, albeit modestly slower han uncrosslinked enzyme controls. Rotation of ClpA with respect to ClpP therefore is ot required for ClpAP protease activity, although some flexibility in how the AAA+ ring ocks on ClpP may be necessary for optimal function.
Bioremediation Potential of Select Bacterial Species for the Neonicotinoid Insecticides, Thiamethoxam and Imidacloprid
Authors: Zamule Stephanie M. , Dupre Cassandra , Mendola Meghan , Widmer Julia , Shebert Jane , Roote Carol E. , Das Padmini
Published: July 2020

The neonicotinoid insecticides, including thiamethoxam (THM) and imidacloprid (IMI), have become increasingly favored in the past decade due to their specificity as insect neurotoxicants. However, neonicotinoids have been implicated as a potential contributing factor in Colony Collapse Disorder (CCD), the widespread disappearance of honey bees, which affects produce production on a global scale. The environmental persistence of neonicotinoids underscores the importance of developing a sustainable, ecologically-friendly remediation technique to remove residual insecticides from the environment. The present study characterizes the neonicotinoid bioremediation potential of six bacterial species:Pseudomonas fluorescens, Pseudomonas putida, Pseudomonas aeruginosa, Alcaligenes faecalis, Escherichia coli, andStreptococcus lactis. In Phase I, we evaluated the utilization of IMI or THM as the sole carbon or nitrogen source byP. fluorescens, P. putida, andP. aeruginosa. All three species were better able to utilize THM over IMI as their sole carbon or nitrogen source, and better growth was noted when THM was used as the sole nitrogen source compared to the sole carbon source. Thus, further studies proceeded with THM only. In Phase II, we assessed the kinetics of THM removal from aqueous media by the six species. Significant (p<0.0001) reductions in 70 mg/L THM concentration were observed forP. fluorescens(67%),P. putida(65%),P. aeruginosa(52%), andA. faecalis(39%) over the 24-day study period, and forE. coli(60%) andS. lactis(12%) over the 14-day study period. The amount of time required to remove 50% of the THM in the media (T50) was: 12 days (d) (E. coli), 18 d (P. fluorescens), 19 d (P. putida), and 23 d (P. aeruginosa). NeitherA. faecalisnorS. lactisachieved 50% removal during the study periods. The THM removal by all species followed a first-order kinetic reaction and half-lives were calculated accordingly. HPLC chromatograms ofP. fluorescens, P. putida, andE. colicultures revealed that as the area of the THM peak decreased over time, the area of an unidentified metabolite peak increased. In Phase II, we sought to characterize this metabolite and the overall metabolic efficiency of these three species. Maximal THM removal occurred at 30°C for all bacterial species assessed. Identification of the metabolite is currently underway, which will allow determination of whether the metabolite is less toxic than the parent compound, a prerequisite for this remediation technique to be viable. If the metabolite is found to be less hazardous than THM, further testing will follow to evaluate the use of this bioremediation technique in the field.