bluuebunny/crossref_metadata_2025 huggingface dataset:
Tyr is Responsible for the Cctq1a QTL and Links Developmental Environment to Central Corneal Thickness Determination
Authors: Meyer Kacie J. , Larson Demelza R. , Whitmore S. Scott , van der Heide Carly J. , Hedberg-Buenz Adam , Dutca Laura M. , Koli Swanand , Pomernackas Nicholas , Mercer Hannah E. , Mansaray Maurisa N. , Paradee William J. , Wang Kai , Nair K. Saidas , Scheetz Todd E. , Anderson Michael G.
Published: September 2021

Central corneal thickness is a quantitative trait with important associations to human health. In a phenotype-driven approach studying corneal thickness of congenic derivatives of C57BLKS/J and SJL/J mice, the critical region for a quantitative trait locus influencing corneal thickness, Cctq1a, was delimited to a 10-gene interval. Exome sequencing, RNAseq, and studying independent mutations eliminated multiple candidate genes and confirmed one. Though the causative gene, Tyr, has no obvious direct function in the transparent cornea, studies with multiple alleles on matched genetic backgrounds, both in isolation and genetic complementation crosses, confirmed allelism of Tyr-Cctq1a; albino mice lacking Tyr function had thin corneas. Albino mice also had increased axial length. Because albinism exposes eyes to increased light, the effect of dark-rearing was tested and found to rescue central corneal thickness. In sum, the results point to an epiphenomenon; developmental light exposure interacts with genotype as an important determinate of adult corneal thickness.
Peptide fusion improves prime editing efficiency
Authors: Velimirovic Minja , Zanetti Larissa C. , Shen Max W. , Fife James D. , Lin Lin , Cha Minsun , Akinci Ersin , Barnum Danielle , Yu Tian , Sherwood Richard I.
Published: September 2021

Prime editing enables search-and-replace genome editing but is limited by low editing efficiency. We present a high-throughput approach, PepSEq, to measure how fusion of 12,000 85-amino acid peptides derived from human DNA repair-related proteins influences prime editing efficiency. We show that peptide fusion can enhance prime editing, prime-enhancing peptides combine productively, and a top dual peptide-prime editor increases prime editing significantly in multiple cell lines across dozens of target sites.
Brain Injury Localization in Electromagnetic Imaging using Symmetric Crossing Lines Method
Authors: Zhu Guohun , Bialkowski Alina , Crozier Stuart , Guo Lei , Nguyen Phong , Stancombe Anthony , Abbosh Amin
Published: September 2021

To avoid death or disability, patients with brain injury should undertake a diagnosis at the earliest time and accept frequent monitoring after starting any medical intervention. This paper presents a novel approach to localize brain injury using the intersection of pairs of signals from symmetrical antennas based on the hypothesis that healthy brains are approximately symmetric that the bleeding targets will lead to significantly different amplitude and phase changes if one of pair of transmit signals cross targets. The scattered signals (S-parameters) are acquired using 100 realistic brain models and 150 experimental data measurements. Firstly, three pair of horizontal antennas are used to detect target crossing which line and in which hemisphere in low frequency bands and estimate the size using high frequency bands. Then, an intersection of two pairs of antennas are identified the position of the target. Finally, a heat map is used to visualise the stroke brain. The results indicate that crossing pairs of antenna signals from the hemisphere with a blood mass exhibit significantly different signal amplitude in the graph features compared to those without the target (p<0.003). The experiments show that our novel localization algorithm can achieve an accuracy of 0.85±0.08 Dice similarity coefficient based on 150 experimental measurements using an elliptical container, which is suitable for brain injury localization.
Nonparametric Interrogation of Transcriptional Regulation in Single-Cell RNA and Chromatin Accessibility Multiomic Data
Authors: Jiang Yuchao , Harigaya Yuriko , Zhang Zhaojun , Zhang Hongpan , Zang Chongzhi , Zhang Nancy R
Published: September 2021

Epigenetic control of gene expression is highly cell-type- and context-specific. Yet, despite its complexity, gene regulatory logic can be broken down into modular components consisting of a transcription factor (TF) activating or repressing the expression of a target gene through its binding to a cis-regulatory region. Recent advances in joint profiling of transcription and chromatin accessibility with single-cell resolution offer unprecedented opportunities to interrogate such regulatory logic. Here, we propose a nonparametric approach, TRIPOD, to detect and characterize three-way relationships between a TF, its target gene, and the accessibility of the TF’s binding site, using single-cell RNA and ATAC multiomic data. We apply TRIPOD to interrogate cell-type-specific regulatory logic in peripheral blood mononuclear cells and contrast our results to detections from enhancer databases, cis-eQTL studies, ChIP-seq experiments, and TF knockdown/knockout studies. We then apply TRIPOD to mouse embryonic brain data during neurogenesis and gliogenesis and identified known and novel putative regulatory relationships, validated by ChIP-seq and PLAC-seq. Finally, we demonstrate TRIPOD on SHARE-seq data of differentiating mouse hair follicle cells and identify lineage-specific regulation supported by histone marks for gene activation and super-enhancer annotations.
The necrotrophic effector ToxA fromParastagonospora nodoruminteracts with wheat NHL proteins to facilitateTsn1-mediated necrosis
Authors: Dagvadorj Bayantes , Outram Megan A. , Williams Simon J. , Solomon Peter S.
Published: September 2021

The plant pathogenParastagonospora nodorumsecretes necrotrophic effectors to promote disease. These effectors induce cell death on wheat cultivars carrying dominant susceptibility genes in an inverse gene-for-gene manner. However, the molecular mechanisms underpinning these interactions and resulting cell death remain unclear. Here, we used a yeast-two-hybrid library approach to identify wheat proteins that interact with the necrotrophic effector ToxA. Using this strategy, we identified an interaction between ToxA and a wheat transmembrane NDR/HIN1-like protein (TaNHL10) and confirmed the interaction usingin-plantaco-immunoprecipitation and confocal microscopy co-localization analysis. We showed that the C-terminus of TaNHL10 is extracellular whilst the N-terminus was localized in the cytoplasm. Further analyses using yeast-two-hybrid and confocal microscopy co-localization showed that ToxA interacts with the C-terminal LEA2 extracellular domain of TaNHL10. Random mutagenesis was then used to identify a ToxA mutant, ToxAN109D, which was unable to interact with TaNHL10 in yeast-two-hybrid assays. Subsequent heterologous expression and purification of ToxAN109DinNicotiania benthamianarevealed that the mutated protein was unable to induce necrosis onTsn1-dominant wheat cultivars confirming that the interaction of ToxA with TaNHL10 is required to induce cell death. Collectively, these data advance our understanding on how ToxA induces cell death during infection and further highlights the importance of host cell surface interactions in necrotrophic pathosystems.
Tracing boundaries in Eastern Antarctica: Multi-scale drivers of soil microbial communities across the hyperarid Vestfold Hills
Authors: Zhang Eden , Czechowski Paul , Terauds Aleks , Wong Sin Yin , Chelliah Devan S. , Raven Mark , Tanaka Mark M. , Ferrari Belinda C.
Published: September 2021

Microorganisms are key to sustaining core ecosystem processes across terrestrial Antarctica but they are rarely considered in conservation frameworks. Whilst greater advocacy has been made towards the inclusion of microbial data in this context, there is still a need for better tools to quantify multispecies responses to environmental change. Here, we extend the scope of Gradient Forest modelling beyond macroorganisms and small datasets to the comprehensive polar soil microbiome encompassing >17, 000 sequence variants for bacteria, micro-eukarya and archaea throughout the hyperarid Vestfold Hills of Eastern Antarctica. Quantification of microbial diversity against 79 physiochemical variables revealed that whilst rank-order importance differed, predictors were broadly consistent between domains, with greatest sharing occurring between bacteria and micro-eukarya. Moisture was identified as the most robust predictor for shaping the regional soil microbiome, with highest compositional turnover or “splits” occurring within the 10 – 12 % moisture content range. Often the most responsive taxa were rarer lineages of bacteria and micro-eukarya with phototrophic and nutrient-cycling capacities such as Cyanobacteria (up to 61.81 % predictive capacity), Chlorophyta (62.17 %) and Ochrophyta (57.81 %). These taxa groups are thus at greater risk of biodiversity loss or gain to projected climate trajectories, which will inevitably disturb current ecosystem dynamics. Better understanding of these threshold tipping points will positively aid conservation efforts across Eastern Antarctica. Furthermore, the successful implementation of an improved Gradient Forest model also presents an exciting opportunity to broaden its use on microbial systems globally.
A new class of cell wall-recycling L,D-carboxypeptidase determines β-lactam susceptibility and morphogenesis in Acinetobacter baumannii
Authors: Dai Yunfei , Pinedo Victor , Tang Amy Y. , Cava Felipe , Geisinger Edward
Published: September 2021

The hospital-acquired pathogen Acinetobacter baumannii possesses a complex cell envelope that is key to its multidrug resistance and virulence. The bacterium, however, lacks many canonical enzymes that build the envelope in model organisms. Instead, A. baumannii contains a number of poorly annotated proteins that may allow alternative mechanisms of envelope biogenesis. We demonstrated previously that one of these unusual proteins, ElsL, is required for cell elongation and for withstanding antibiotics that attack the septal cell wall. Curiously, ElsL is composed of a leaderless YkuD-family domain usually found in secreted, cell-wall-modifying L,D-transpeptidases (LDTs). Here, we show that, rather than being an LDT, ElsL is actually a new class of cytoplasmic L,D-carboxypeptidase (LDC) that provides a critical step in cell-wall recycling previously thought to be missing from A. baumannii. Absence of ElsL impairs cell wall integrity, elongation, and intrinsic resistance due to buildup of murein tetrapeptide precursors, toxicity of which is bypassed by preventing muropeptide recycling. Multiple pathways in the cell become sites of vulnerability when ElsL is inactivated, including L,D-crosslink formation, cell division, and outer membrane lipid homoeostasis, reflecting its pleiotropic influence on cell envelope physiology. We thus reveal a novel class of cell-wall-recycling LDC critical to growth and homeostasis of A. baumannii and likely many other bacteria.

Importance

To grow efficiently, resist antibiotics, and control the immune response, bacteria recycle parts of their cell wall. A key step in the typical recycling pathway is the reuse of cell wall peptides by an enzyme known as an LDC. Acinetobacter baumannii, an “urgent-threat” pathogen causing drug-resistant sepsis in hospitals, was previously thought to lack this enzymatic activity due to absence of a known LDC homolog. Here, we show that A. baumannii possesses this activity in the form of an enzyme class not previously associated with cell wall recycling. Absence of this protein intoxicates and weakens the A. baumannii cell envelope in multiple ways due to the accumulation of dead-end intermediates. Several other organisms of importance to health and disease encode homologs of the A. baumannii enzyme. This work thus reveals an unappreciated mechanism of cell wall recycling, manipulation of which may contribute to enhanced treatments targeting the bacterial envelope.
The Influence of Model Violation on Phylogenetic Inference: A Simulation Study
Authors: Naser-Khdour Suha , Minh Bui Quang , Lanfear Robert
Published: September 2021

Abstract

Phylogenetic inference typically assumes that the data has evolved under Stationary, Reversible and Homogeneous (SRH) conditions. Many empirical and simulation studies have shown that assuming SRH conditions can lead to significant errors in phylogenetic inference when the data violates these assumptions. Yet, many simulation studies focused on extreme non-SRH conditions that represent worst-case scenarios and not the average empirical dataset. In this study, we simulate datasets under various degrees of non-SRH conditions using empirically derived parameters to mimic real data and examine the effects of incorrectly assuming SRH conditions on inferring phylogenies. Our results show that maximum likelihood inference is generally quite robust to a wide range of SRH model violations but is inaccurate under extreme convergent evolution.
Validation of the Panbio™ COVID-19 Antigen Rapid Test (Abbott) to screen for SARS-CoV-2 infection in Sint Maarten: a diagnostic accuracy study
Authors: King Colin , Weever Eva Lista-de , Henry Maria , Steingrover Radjin , Fleming Chérina , Panneflek Richard , van Kampen Sanne
Published: September 2021

Objectives

Control of the pandemic has required countries to look for other forms of tests besides the gold standard real-time polymerase chain reaction (RT-PCR). Rapid antigen tests (RAT), though less sensitive than RT-PCR, offer the possibility of rapid, inexpensive and early detection of the most infectious COVID-19 cases. Only very few studies have assessed the performance of the Abbott Panbio COVID-19 RAT among asymptomatic people or in Latin America. This study set out to validate this test among people attending the public test street in Sint Maarten, Dutch Caribbean.

Methods

People of all ages were recruited from the public COVID-19 test street regardless of COVID-19 symptoms. They received a nasopharyngeal swab for the Abbott Panbio COVID-19 RAT and the RT-PCR Qtower. Diagnostic accuracy of the RAT was compared to the RT-PCR among the overall study population and for subgroups with/without symptoms, with/without close contact and different Ct values.

Results

Using a RT-PCR Ct cut-off value of <33, 119 out of 1,411 people (8.4%) tested positive for SARS-CoV-2. Most were asymptomatic (59%). The overall sensitivity and specificity of the RAT was 84% (95% CI 76.2-90.1) and 99.9% (95% CI 99.6-100) respectively. The sensitivity reduced to 67.6% (95% CI: 49.5%, 82.6%) among people without symptoms, regardless of whether they were in close contact with a known COVID-19 case. Sensitivity reduced considerably with a Ct cut-off value of <35.

Conclusions

The Abbott Panbio RAT is a valid and cheaper alternative to RT-PCR when used on symptomatic individuals among the general population. However, among asymptomatic people it should not be used as a stand-alone test and negative results should be confirmed with RT-PCR.
Outcomes of gastrointestinal fistulas: Results from a multi-continent, multi-national, multi-center cohort
Authors: Márquez Humberto Arenas , Turcios Correia María Isabel , García Juan Francisco , Prado Roberto Anaya , Vergara Arturo , Garnica Jorge Luis , Cacho Alejandra , Guerra Daniel , Navarrete Miguel Mendoza , Porbén Sergio Santana
Published: September 2021

Rationale

Gastrointestinal fistulas (GIF) represent a severe and potentially lethal complication of the hospital surgical activity. However, evidences are lacking about prognosis and outcomes of GIF in Latin America (LATAM) hospitals.

Objective

To describe the prognosis and outcomes GIF in LATAM hospitals.

Study design

Prospective, longitudinal, cohort-type study. The cohort fostered three cross-sectional examinations: First examination: On admission of the patient in the study; Second examination: Thirty days later; and Third (and last) examination: Sixty days after patient’s admission.

Study serie

One hundred seventy-seven patients (Males: 58.2 %; Average age: 51.0 ± 16.7 years; Ages ≥ 60 years: 36.2 %) diagnosed with, and assisted for, GIF (ECF: Enterocutaneous: 64.9 % vs. EAF: Enteroathmospheric: 35.1 %) in 76 LATAM hospitals (13 countries) and Europe (4).

Methods

Condition (Alive vs. Deceased) and hospital status (Hospitalized vs. Discharged) of the patient, and the GFI patency (Closed vs. Non closed) were recorded in each of the cohort’s examination. Indicators of GFI prognosis thus constructed were correlated demographical, sanitary, surgical and nutritional characteristics of the patients

Results

On conclusion of the study indicators of GIF prognosis behaved as follows: Mortality:14.7 %; Prolonged hospitalization: 47.4 %; Spontaneous closure of GIF: 36.2 %. Type of GIF influenced upon patient’s survival: ECF: 87.0 % vs. EAF: 82.3 % (Δ = +4.7 %; χ2 = 6.787; p < 0.05). In each examination of the cohort, the number of surviving subjects was always greater among those with ECF: After 30 days: ECF: 92.1 % vs. EAF: 83.9 % (Δ = +8.2 %); After 60 days: ECF: 98.1 % vs. EAF: 90.4 % (Δ = +7.7 %; χ2 = 13.764; p < 0.05). On the other hand, hospital stay was prolonged in the subjects of elective surgery (Elective surgery: 61.4 % vs. Emergency surgery: 38.3 % (Δ = +23.1 %; χ2 = 9.064; p < 0.05) and those with a reduced calf circumference (χ2 = 12.655; p < 0.05). Location of the fistula also influenced upon prolongation of hospital stay (χ2 = 7.817; p < 0.05).

Conclusions

Type of GIF influences upon survival of the patient. On the hand, hospital stay was dependent upon type of surgery previously performed, location of the fistula, and calf circumference value on admission in the study serie.
A framework for evaluating edited cell libraries created by massively parallel genome engineering
Authors: Cawley Simon , Abbate Eric , Abraham Christopher G. , Alvarez Steven , Barber Mathew , Bolte Scott , Bruand Jocelyne , Church Deanna M. , Davis Clint , Estes Matthew , Federowicz Stephen , Fox Richard , Gander Miles W. , Garst Andrew D. , Gencer Gozde , Halweg-Edwards Andrea L. , Hardenbol Paul , Hraha Thomas , Jain Surbhi , Johnson Charlie , Juneau Kara , Krishnamurthy Nandini , Lambert Shea , Leland Bryan , Pearson Francesca , Ray J. Christian J. , Sanada Chad D. , Shaver Timothy M. , Shepherd Tyson R. , Shorenstein Joshua , Spindler Eileen C. , Struble Craig A. , Swat Maciej H. , Tanner Stephen , Tian Tian , Wishart Ken , Graige Michael S.
Published: September 2021

Genome engineering methodologies are transforming biological research and discovery. Approaches based on CRISPR technology have been broadly adopted and there is growing interest in the generation of massively parallel edited cell libraries. Comparing the libraries generated by these varying approaches is challenging and researchers lack a common framework for defining and assessing the characteristics of these libraries. Here we describe a framework for evaluating massively parallel libraries of edited genomes based on established methods for sampling complex populations. We define specific attributes and metrics that are informative for describing a complex cell library and provide examples for estimating these values. We also connect this analysis to generic phenotyping approaches, using either pooled (typically via a selection assay) or isolate (often referred to as screening) phenotyping approaches. We approach this from the context of creating massively parallel, precisely edited libraries with one edit per cell, though the approach holds for other types of modifications, including libraries containing multiple edits per cell (combinatorial editing). This framework is a critical component for evaluating and comparing new technologies as well as understanding how a massively parallel edited cell library will perform in a given phenotyping approach.
PcGCE is a potent elicitor of defense responses in aspen
Authors: Donev Evgeniy N. , Derba-Maceluch Marta , Liu Xiao-Kun , Bwanika Henri Colyn , Dobrowolska Izabela , Thapa Mohit , Leśniewska Joanna , Šimura Jan , Tsai Alex Yi-Lin , Boström Dan , Kleczkowski Leszek A. , Eriksson Maria E. , Ljung Karin , Master Emma R. , Mellerowicz Ewa J.
Published: September 2021

Using microbial enzymes in transgenesis is a powerful means to introduce new functionalities in plants. Glucuronoyl esterase (GCE) is a microbial enzyme hydrolyzing the ester bond between lignin and 4-O-methyl-α-D-glucuronic acid present as a side chain of glucuronoxylan. This bond mediates lignin-carbohydrate complex (LCC) formation, considered as crucial factor of lignocellulose recalcitrance to saccharification. Previous studies showed that hybrid aspen (Populus tremula L. x tremuloides Michx.) constitutively expressing Phanerochaete carnosa Burt GCE (PcGCE) had better efficiency of cellulose-to-glucose conversion but were stunned and had lower cellulose content indicating that more studies are needed to design strategy for deployment of this enzyme in planta. Here we report that the transgenic plants exhibit premature leaf senescence, increased accumulation of calcium oxalate crystals, tyloses and necrotic lesions and have strongly activated immune defense reactions as revealed by their altered profiles of transcriptomes, metabolomes and hormones in the leaves. To elucidate if these effects are triggered by damage-associated molecular patterns (DAMPs) or by PcGCE protein perceived as a pathogen-associated molecular pattern (PAMP), we ectopically expressed in aspen an enzymatically inactive PcGCES217A. The mutated PcGCE induced similar growth retardation, leaf necrosis and premature senescence as the active one, providing evidence that PcGCE protein is recognized as PAMP. Transcriptomics analysis of young expanding leaves of 35S:PcGCE plants identified several candidates for receptors of PcGCE, which were not expressed in developing wood tissues. Grafting experiments showed that PcGCE transcripts are not cell-to-cell mobile and that PcGCE expressing leaves augment systemic responses. In agreement, expressing PcGCE in developing wood by using the wood-specific promoter (WP), avoided all off-target effects. Moreover, WP:PcGCE lines grew better than control plants providing evidence that this strategy can be used in transgenic crops dedicated for biorefinery.
Inhibition of Bruton’s tyrosine kinase activity attenuates trauma-induced multiple organ dysfunction in rats
Authors: Patel Nikita M , Oliveira Filipe RMB , Ramos Hanna Pillmann , Aimaretti Eleonora , Alves Gustavo Ferreira , Coldewey Sina M , Collino Massimo , Sordi Regina , Thiemermann Christoph
Published: September 2021

Objective

The aim of this study was to investigate (a) the potential of the Bruton’s tyrosine kinase (BTK) inhibitors (BTKi) acalabrutinib and fenebrutinib to reduce multiple organ dysfunction syndrome (MODS) in acute and chronic hemorrhagic shock (HS) rat models and (b) whether treatment with either acalabrutinib or fenebrutinib attenuates BTK, NF-κB and NLRP3 activation in HS.

Background

The MODS caused by an excessive systemic inflammatory response following trauma is associated with a high morbidity and mortality. The protein BTK is known to play a role in the activation of the NLRP3 inflammasome, which is a key component of the innate inflammatory response. However, its role in trauma-hemorrhage is unknown.

Methods

Acute and chronic HS rat models were performed to determine the influence of acalabrutinib or fenebrutinib on MODS. The activation of BTK, NF-κB and NLRP3 pathways were analyzed by western blot in the kidney.

Results

We demonstrated that (a) HS caused organ injury and/or dysfunction and hypotension (post resuscitation) in rats, while (b) treatment of HS-rats with either acalabrutinib or fenebrutinib attenuated the organ injury and dysfunction in acute and chronic HS models and (c) reduced the activation of BTK, NF-κB and NLRP3 pathways in the kidney.

Conclusion

Our results point to a role of BTK in the pathophysiology of organ injury and dysfunction caused by trauma/hemorrhage and indicate that BTK inhibitors may be repurposed as a potential therapeutic approach for MODS after trauma and/or hemorrhage.

MINI-ABSTRACT

This study evaluated the role of Bruton’s tyrosine kinase (BTK) in trauma/hemorrhage. Patients with trauma had elevated gene expression of BTK. The BTK inhibitors acalabrutinib (irreversible) and fenebrutinib (reversible) attenuated the trauma-induced multiple organ dysfunction in rats with hemorrhagic shock, indicating that BTK could be a potential therapeutic target.
Gene regulation by DNA methylation is contingent on chromatin accessibility during transgenerational plasticity in the purple sea urchin
Authors: Bogan Samuel N , Strader Marie E , Hofmann Gretchen E
Published: September 2021

Epigenetic processes are proposed to contribute to phenotypic plasticity. In invertebrates, DNA methylation commonly varies across environments and can correlate or causally associate with phenotype, but its role in transcriptional responses to the environment remains unclear. Maternal environments experienced by the sea urchin Strongylocentrotus purpuratus induce 3 – 6x greater differential CpG methylation in offspring larvae relative to larval developmental environments, suggesting a role for DNA methylation in transgenerational plasticity (TGP). However, a negligible association has been observed between differentially methylated and differentially expressed genes. What gene regulatory roles does invertebrate DNA methylation possess under environmental change, if any? We quantified DNA methylation and gene expression in S. purpuratus larvae exposed to different ecologically relevant conditions during gametogenesis (maternal conditioning) or embryogenesis (developmental conditioning). We modeled differential gene expression and differential splicing under maternal conditioning as functions of DNA methylation, incorporating variables for genomic feature and chromatin accessibility. We detected significant interactions between differential methylation, chromatin accessibility, and genic architecture associated with differential expression and splicing. Observed transcriptional responses to maternal conditioning were also 4 – 13x more likely when accounting for interactions between methylation and chromatin accessibility. Our results provide evidence that DNA methylation possesses multiple functional roles during TGP in S. purpuratus, but its effects are contingent upon other genomic and epigenomic states. Singularly unpredictive of transcription, DNA methylation is likely one cog in the epigenomic machinery contributing to environmental responses and phenotypic plasticity in S. purpuratus and other invertebrates.
Modulation of sensory perception by hydrogen peroxide enables Caenorhabditis elegans to find a niche that provides both food and protection from hydrogen peroxide
Authors: Schiffer Jodie A. , Stumbur Stephanie V. , Seyedolmohadesin Maedeh , Xu Yuyan , Serkin William T. , McGowan Natalie G , Banjo Oluwatosin , Torkashvand Mahdi , Lin Albert , Hosea Ciara N. , Assié Adrien , Samuel Buck S. , O’Donnell Michael P. , Venkatachalam Vivek , Apfeld Javier
Published: September 2021

Hydrogen peroxide (H2O2) is the most common chemical threat that organisms face. Here, we show that H2O2 alters the bacterial food preference of Caenorhabditis elegans, enabling the nematodes to find a safe environment with food. H2O2 induces the nematodes to leave food patches of laboratory and microbiome bacteria when those bacterial communities have insufficient H2O2-degrading capacity. The nematode’s behavior is directed by H2O2-sensing neurons that promote escape from H2O2 and by bacteria-sensing neurons that promote attraction to bacteria. However, the input for H2O2-sensing neurons is removed by bacterial H2O2-degrading enzymes and the bacteria-sensing neurons’ perception of bacteria is prevented by H2O2. The resulting cross-attenuation provides a general mechanism that ensures the nematode’s behavior is faithful to the lethal threat of hydrogen peroxide, increasing the nematode’s chances of finding a niche that provides both food and protection from hydrogen peroxide.
Ratiometric RNA labeling allows dynamic multiplexed analysis of gene circuits in single cells
Authors: Xu Shuhui , Li Kai , Ma Liang , Zhang Jianhan , Yoon Shinae , Elowitz Michael B. , Lin Yihan
Published: September 2021

Biological processes are highly dynamic and are regulated by genes that connect with one and another, forming regulatory circuits and networks. Understanding how gene regulatory circuits operate dynamically requires monitoring the expression of multiple genes in the same cell. However, it is limited by the relatively few distinguishable fluorescent proteins. Here, we developed a multiplexed real-time transcriptional imaging method based on two RNA stem-loop binding proteins, and employed it to analyze the temporal dynamics of synthetic gene circuits. By incorporating different ratios of MS2 and PP7 stem-loops, we were able to monitor the real-time nascent transcriptional activities of up to five genes in the same cell using only two fluorescent proteins. Applying this multiplexing capability to synthetic linear or branched gene regulatory cascades revealed that propagation of transcriptional dynamics is enhanced by non-stationary dynamics and is dictated by the slowest regulatory branch in the presence of combinatorial regulation. Mathematical modeling provided further insight into temporal multi-gene interactions and helped to understand potential challenges in regulatory inference using snapshot single-cell data. Ratiometric multiplexing should scale exponentially with additional labelling channels, providing a way to track the dynamics of larger circuits.
Multiple molecular events underlie stochastic switching between two heritable cell states in a eukaryotic system
Authors: Ziv Naomi , Brenes Lucas R. , Johnson Alexander
Published: September 2021

Eukaryotic transcriptional networks are often large and contain several levels of feedback regulation. Many of these networks have the ability to generate and maintain several distinct transcriptional states across multiple cell divisions and to switch between them. In certain instances, switching between cell states is stochastic, occurring in a small subset of cells of an isogenic population in a seemingly homogenous environment. Given the scarcity and unpredictability of switching in these cases, investigating the determining molecular events is challenging. White-opaque switching in the fungal species Candida albicans is an example of stably inherited cell states that are determined by a complex transcriptional network and can serve as an experimentally accessible model system to study characteristics important for stochastic cell fate switching in eukaryotes. In standard lab media, genetically identical cells maintain their cellular identity (either “white” or “opaque”) through thousands of cell divisions and switching between the states is rare. By isolating populations of white or opaque cells, previous studies have elucidated the many differences between the two stable cell states and identified a set of transcriptional regulators needed for cell type switching. Yet little is known about the molecular events that determine the rare, stochastic switching events that occur in single cells. We use microfluidics combined with fluorescent reporters to directly observe rare switching events between the white and opaque states. We investigate the stochastic nature of switching by beginning with white cells and monitoring the activation of Wor1, a master regulator and marker for the opaque state, in single cells and throughout cell pedigrees. Our results indicate that switching requires two stochastic steps; first an event occurs that predisposes a lineage of cells to switch. In the second step, some but not all, of those predisposed cells rapidly express high levels of Wor1 and commit to the opaque state. To further understand the rapid rise in Wor1, we used a synthetic inducible system in Saccharomyces cerevisiae into which a controllable C. albicans Wor1 and a reporter for its transcriptional control region have been introduced. We document that Wor1 positive autoregulation is highly cooperative (Hill coefficient > 3), leading to rapid activation and producing an “all or none” rather than a graded response. Taken together, our results suggest that reaching a threshold level of a master regulator is sufficient to drive cell type switching in single cells and that an earlier molecular event increases the probability of reaching that threshold in certain small lineages of cells. Quantitative molecular analysis of the white-opaque circuit can serve as a model for the general understanding of complex circuits.
Prevalence of viral frequency-dependent infection in coastal marine prokaryotes revealed using monthly time series virome analysis
Authors: Tominaga Kento , Ogawa-Haruki Nana , Nishimura Yosuke , Watai Hiroyasu , Yamamoto Keigo , Ogata Hiroyuki , Yoshida Takashi
Published: September 2021

Viruses infecting marine prokaryotes have large impacts on the diversity and dynamics of their hosts. Model systems suggest viral infection is frequency-dependent and constrained by the virus-host encounter rate. However, it is unclear whether the frequency-dependent infection is pervasive among the abundant prokaryotic populations with different growth strategies (i.e. r-strategy and K-strategy). To address this question, we performed a comparison of prokaryotic and viral communities using 16S rRNA amplicon and virome sequencing based on samples collected monthly for two years at a Japanese coastal site, Osaka Bay. Concurrent seasonal shifts observed in prokaryotic and viral community dynamics indicated that abundances of viruses correlated with that of their predicted host phyla (or classes). Co-occurrence network analysis between abundant prokaryotes and viruses revealed 6 423 co-occurring pairs, suggesting a tight coupling of host and viral abundances and their “one to many” correspondence. Although dominant K-strategist like species, such as SAR11, showed few co-occurring viruses, a fast succession of their viruses suggests viruses infecting these populations changed continuously. Our results suggest the frequency-dependent viral infection prevailed in coastal marine prokaryotes regardless of host taxa and growth strategy.
A population based expression atlas provides insights into disease resistance and other physiological traits in cassava (Manihot esculentaCrantz)
Authors: Ogbonna Alex C. , Ramu Punna , Williams Esuma , Nandudu Leah , Morales Nicolas , Powell Adrian , Kawuki Robert , Bauchet Guillaume , Jannink Jean-Luc , Mueller Lukas A.
Published: September 2021

Cassava, a food security crop in Africa, is grown throughout the tropics and subtropics. Although cassava can provide high productivity in suboptimal conditions, the yield in Africa is substantially lower than in other geographies. The yield gap is attributable to many challenges faced by cassava in Africa, including susceptibility to diseases and poor soil conditions. In this study, we carried out 3’RNA sequencing on 150 accessions from the National Crops Resources Research Institute, Ugandan for 5 tissue types, providing population-based transcriptomics resources to the research community in a web-based queryable cassava expression atlas. Differential expression and weighted gene co-expression network analysis were performed to detect 8,820 significantly differentially expressed genes (DEGs), revealing similarity in expression patterns between tissue types and the clustering of detected DEGs into 18 gene modules. As a confirmation of data quality, differential expression and pathway analysis targeting cassava mosaic disease (CMD) identified 27 genes observed in the plant-pathogen interaction pathway, several previously identified CMD resistance genes and two peroxidase family proteins different from theCMD2gene. Present research work represents a novel resource towards understanding complex traits at expression and molecular levels for the development of resistant and high-yielding cassava varieties, as exemplified with CMD.
Personalizing the control law of an upper-limb exoskeleton using EMG signal
Authors: Treussart Benjamin , Caron Remi , Geffard Franck , Marin Frederic , Vignais Nicolas
Published: September 2021

Implementing an intuitive control law for an upper-limb exoskeleton dedicated to force augmentation is a challenging issue in the field of human-robot collaboration. The goal of this study is to adapt an EMG-based control system to a user based on individual caracteristics. To this aim, a method has been designed to tune the parameters of control using objective criteria, improving user’s feedback. The user’s response time is used as an objective value to adapt the gain of the controller. The proposed approach was tested on 10 participants during a lifting task. Two different conditions have been used to control the exoskeleton: with a generic gain and with a personalized gain. EMG signals was captured on five muscles to evaluate the efficiency of the conditions and the user’s adaptation. Results showed a statistically significant reduction of mean muscle activity of the deltoid between the beginning and the end of each situation (28.6 ± 13.5% to 17.2 ± 7.3% of Relative Maximal Contraction for the generic gain and from 24.9 ± 8.5% to 18.0 ± 6.8% of Relative Maximal Contraction for the personalized gain). When focusing on the first assisted movements, the personalized gain induced a mean activity of the deltoïd significantly lower (29.0 ± 8.0% of Relative Maximal Contraction and 37.4 ± 9.5% of Relative Maximal Contraction, respectively). Subjective evaluation showed that the system with a personalized gain was perceived as more intuitive, and required less concentration when compared to the system with a generic gain.
Engineering CAR-NK cells to secrete IL15 sustains their anti-AML functionality, but is associated with systemic toxicities
Authors: Christodoulou Ilias , Ho Won Jin , Marple Andrew , Ravich Jonas W. , Tam Ada , Rahnama Ruyan , Fearnow Adam , Rietberg Cambrynne , Yanik Sean , Solomou Elena E. , Varadhan Ravi , Koldobskiy Michael A. , Bonifant Challice L.
Published: September 2021

Background

The prognosis of patients with recurrent/refractory acute myelogenous leukemia (AML) remains poor and cell-based immunotherapies hold promise to improve outcomes. NK cells can elicit an anti-leukemic response via a repertoire of activating receptors that bind AML surface ligands. NK cell adoptive transfer is safe but thus far has shown limited anti-AML efficacy. Here, we aimed to overcome this limitation by engineering NK cells to express chimeric antigen receptors (CARs) to boost their anti-AML activity, and interleukin-15 (IL15) to enhance their persistence.

Methods

We characterized in detail NK cell populations expressing a panel of AML (CD123)-specific CARs and/or IL15 in vitro and in AML xenograft models.

Results

CARs with 2B4.ζ or 4-1BB.ζ signaling domains demonstrated greater cell surface expression and endowed NK cells with improved anti-AML activity in vitro. Initial in vivo testing revealed that only 2B4.ζ CAR-NK cells had improved anti-AML activity in comparison to untransduced (UTD) and 4-1BB.ζ CAR-NK cells. However, the benefit was transient due to limited CAR-NK cell persistence. Transgenic expression of secretory (s)IL15 in 2B4.ζ CAR and UTD NK cells improved their effector function in the setting of chronic antigen simulation in vitro. Multiparameter flow analysis after chronic antigen exposure identified the expansion of unique NK cell subsets. 2B4.ζ/sIL15 CAR and sIL15 NK cells maintained an overall activated NK cell phenotype. This was confirmed by transcriptomic analysis, which revealed a highly proliferative and activated signature in these NK cell groups. In vivo, 2B4.ζ/sIL15 CAR-NK cells had potent anti-AML activity in one model, while 2B4.ζ/sIL15 CAR and sIL15 NK cells induced lethal toxicity in a second model.

Conclusion

Transgenic expression of CD123-CARs and sIL15 enabled NK cells to function in the setting of chronic antigen exposure but was associated with systemic toxicities. Thus, our study provides the impetus to explore inducible and controllable expression systems to provide cytokine signals to AML-specific CAR-NK cells before embarking on early phase clinical testing.

Abstract Figure

In Brief

Secretion of IL15 from anti-CD123.2B4.CD3ζ CAR-NK cells enhances their activation and killing potential against Acute Myelogenous Leukemia, but carries the risk of severe systemic toxicity.
Novel microRNA-455-3p mouse models to study Alzheimer’s disease pathogenesis
Authors: Kumar Subodh , Mortan Hallie , Sawant Neha , Orlov Erika , Bunquin Lloyd , Pradeepkiran Jangampalli Adi , Reddy P. Hemachandra
Published: September 2021

MicroRNA-455-3p is one of the highly conserved miRNAs involved in several human diseases but newly explored by our lab in Alzheimer’s disease (AD). Our past studies unveiled the biomarker and therapeutic potentials of miR-455-3p in AD. Our in vitro study exhibited the protective role of miR-455-3p against AD toxicities in reducing full-length APP and amyloid-β (Aβ) protein levels, and also reducing defective mitochondrial biogenesis, impaired mitochondrial dynamics and synaptic deficiencies. Next, we sought to determine the essential roles of miR-455-3p in AD using mouse models. Therefore, for the first time we generated both transgenic (TG) and knockout (KO) mouse models of miR-455-3p. We determined the positive and negative effects of miR-455-3p on mice cognitive function, mitochondrial biogenesis, mitochondrial dynamics, mitochondrial number & length, dendritic spine density, synapse numbers and synaptic activity in 12-month-old miR-455-3p TG and KO mice. MiR-455-3p TG mice lived 5 months longer than wild-type (WT) mice, whereas KO mice lived 4 months shorter than their WT counter parts. Morris water maze test showed improved cognitive behavior, spatial learning and memory in miR-455-3p TG mice relative to age-matched WT mice and miR-455-3p KO mice. Further, mitochondrial biogenesis, dynamics and synaptic activities were enhanced in miR-455-3p TG mice, while these were reduced in KO mice. Overall, miR-455-3p TG mice displayed protective effects and miR-455-3p KO mice exhibited deleterious effects in relation to AD pathogenesis. Both mouse models could be ideal research tools to understand the molecular mechanism of miR-455-3p in AD and other human diseases.
Towards a system-level causative knowledge of pollinator communities
Authors: Saavedra Serguei , Bartomeus Ignasi , Godoy Oscar , Rohr Rudolf P. , Zu Penguan
Published: September 2021

Pollination plays a central role both in the maintenance of biodiversity and in crop production. However, habitat loss, pesticides, invasive species, and larger environmental fluctuations are contributing to a dramatic decline of numerous pollinators world-wide. This has increased the need for interventions to protect the composition, functioning, and dynamics of pollinator communities. Yet, how to make these interventions successful at the system level remains extremely challenging due to the complex nature of species interactions and the various unknown or unmeasured confounding ecological factors. Here, we propose that this knowledge can be derived by following a probabilistic causal analysis of pollinator communities. This analysis implies the inference of interventional expectations from the integration of observational and synthetic data. We propose that such synthetic data can be generated using theoretical models that can enable the tractability and scalability of unseen confounding ecological factors affecting the behavior of pollinator communities. We discuss a road map for how this probabilistic causal analysis can be accomplished to increase our system-level causative knowledge of natural communities.
Concurrent tau pathologies in frontotemporal lobar degeneration with TDP-43 pathology
Authors: Koga Shunsuke , Zhou Xiaolai , Murakami Aya , De Castro Cristhoper Fernandez , Baker Matthew C. , Rademakers Rosa , Dickson Dennis W.
Published: September 2021

Aims

Accumulating evidence suggests that patients with frontotemporal lobar degeneration (FTLD) can have pathologic accumulation of multiple proteins, including tau and TDP-43. This study aimed to determine the frequency and characteristics of concurrent tau pathology in FTLD with TDP-43 pathology (FTLD-TDP).

Methods

The study included 146 autopsy-confirmed cases of FTLD-TDP and 55 cases of FTLD-TDP with motor neuron disease (FTLD-MND). Sections from the basal forebrain were screened for tau pathology with phospho-tau immunohistochemistry. For cases with tau pathology on the screening section, additional brain sections were studied to establish a diagnosis. Genetic analysis of C9ORF72, GRN, and MAPT was performed on select cases.

Results

Among 201 cases, we found 72 cases (36%) with primary age-related tauopathy (PART), 85 (42%) with aging-related tau astrogliopathy (ARTAG), 45 (22%) with argyrophilic grain disease (AGD), and 2 cases (1%) with corticobasal degeneration (CBD). Patients with ARTAG or AGD were significantly older than those without these comorbidities. One of the patients with FTLD-TDP and CBD had C9ORF72 mutation and relatively mild tau pathology, consistent with incidental CBD.

Conclusion

The coexistence of TDP-43 and tau pathologies was relatively common, particularly PART and ARTAG. Although rare, individual patients with FTLD can have multiple concurrent proteinopathies. The absence of TDP-43-positive astrocytic plaques may suggest that CBD and FTLD-TDP were independent disease processes in the two patients with both tau and TDP-43 pathologies. It remains to be determined if mixed cases represent a unique disease process or two concurrent disease processes in an individual.
The structural basis of huntingtin (Htt) fibril polymorphism, revealed by cryo-EM of exon 1 Htt fibrils
Authors: Nazarov Sergey , Chiki Anass , Boudeffa Driss , Lashuel Hilal A.
Published: September 2021

The lack of detailed insight into the structure of aggregates formed by the huntingtin protein has hampered efforts to develop therapeutics and diagnostics targeting pathology formation in the brain of patients with Huntington’s disease. To address this knowledge gap, we investigated the structural properties of in vitro generated fibrils from exon1 of the huntingtin protein by electron cryo-microscopy and single-particle analysis. We show that wildtype and mutant exon1 of the huntingtin protein form non-helical fibrils with a polygultamine amyloid core composed of β-hairpins with unique characteristics that have not been previously observed with other amyloid filaments. The stacks of β-hairpins form long planar β- sheets (protofilaments) with variable stacking angle and occasional out-of-register state of individual β-hairpins. These features and the propensity of protofilament to undergo lateral association results in a high degree of fibril polymorphism, including fibrils composed of varying numbers of protofilaments. Our results also represent the first direct observation of how the flanking domains are organized around the polyglutamine core of the fibril and provide insight into how they might affect huntingtin fibril structure, polymorphism, and stacking of β-hairpins within its core structure. Removal of the first 17 amino acids at the N-terminus resulted in surprising intra-fibril structural heterogeneity and reduced fibril’s propensity to lateral associations. Overall, this work provides valuable insights that could guide future mechanistic studies to elucidate the sequence and structural determinants of huntingtin aggregation, as well as cryo-EM and structural studies of fibrils derived from huntingtin proteins and other disease-associated polyglutamine-containing proteins.