bluuebunny/crossref_metadata_2025 huggingface dataset:
Whole genome phylogeny of Cyanobacteria documents a distinct evolutionary trajectory of marine picocyanobacteria
Authors: Strunecký Otakar , Wachtlová Markéta , Koblížek Michal
Published: May 2021

During their 2.7 Gyr long evolution cyanobacteria radiated into a large number of different lineages. To better understand the evolution of cyanobacteria we compared a whole genome phylogenetic tree using 1,047 concatenated single copy orthologues ofProchlorococcus marinusMIT9313 obtained from 93 reference prokaryotic species with traditional phylogenies inferred from 16S rRNA gene and 109 genes found in all used genomes. In contrast to the traditional phylogeny, our genome wide analysis shows a split between picocyanobacteria encompassing all marineProchlorococcusspecies, marine “Synechococcus” species, and members of freshwaterCyanobiumgenus (altogether referred here as the “PSC” clade) and the remaining groups of extant cyanobacteria. To determine the influence of the horizontal gene transfer on the overall tree topology we removed the 374 genes identified as potentially transferred genes. A newly calculated tree utilizing the remaining 673 genes displayed the same topology as the former whole genome tree with the PSC clade as a basal group of all cyanobacteria. The picocyanobacteria also exhibited distinctly larger similarity to reference bacteria’s cell and genome size, carboxyzome architecture, various metabolic pathways, and chlorophyll synthesis than to the other cyanobacteria. Potentially horizontally transferred genes were found in connected chains throughout fundamental metabolic pathways suggesting that evolution of these genes was severely limited and or brought to a standstill. The environment related genes connected with metabolism of nitrogen, sulfur, and scarce seawater metals were more conserved in PSC group as they were already best tuned for its poor and stable environment. Other genes were found predominantly sequentially static as they were already accurately adapted with rare beneficial mutations. The PSC clade thus represents an isolated cyanobacterial lineage which followed a distinct evolutionary trajectory driven by its specific environment.
Microscopic characterization of local strain field in healing tissue in the central third defect of mouse patellar tendon at early-phase of healing
Authors: Maeda Eijiro , Kuronayagi Kaname , Matsumoto Takeo
Published: May 2021

Tendons exhibit a hierarchical collagen structure, wherein higher-level components, such as collagen fibres and fascicles, are elongated, slid, and rotated during macroscopic stretching. These mechanical behaviours of collagen fibres play important roles in stimulating tenocytes, imposing stretching, compression, and shear deformation. It was hypothesised that a lack of local fibre behaviours in healing tendon tissue may result in a limited application of mechanical stimuli to cells within the tissue, leading to incomplete recovery of tissue structure and functions in regenerated tendons. Therefore, the present study aimed to measure the microscopic strain field in the healing tendon tissue. A central third defect was created in the patellar tendon of mice, and the regenerated tissue in the defect was examined by tensile testing, collagen fibre analysis, and local strain measurement using confocal microscopy at 3 and 6 weeks after surgery. Healing tissue at 3 weeks exhibited a significantly lower strength and disorganised collagen fibre structure compared with the normal tendon. These characteristics at 6 weeks remained significantly different from those of the normal tendon. Moreover, the magnitude of local shear strain in the healing tissue under 4% tissue strain was significantly smaller than that in the normal tendon. Differences in the local strain field may be reflected in the cell nuclear shape and possibly the amount of mechanical stimuli applied to the cells during tendon deformation. Accordingly, restoration of a normal local mechanical environment in the healing tissue may be key to a better healing outcome of tendon injury.
Inhibitory Gating of Coincidence-Dependent Sensory Binding in Secondary Auditory Cortex
Authors: Kline Amber M. , Aponte Destinee A. , Tsukano Hiroaki , Giovannucci Andrea , Kato Hiroyuki K
Published: May 2021

Integration of multi-frequency sounds into a unified perceptual object is critical for recognizing syllables in speech. This “feature binding” relies on the precise synchrony of each component’s onset timing, but little is known regarding its neural correlates. We find that multi-frequency sounds prevalent in vocalizations, specifically harmonics, preferentially activate the mouse secondary auditory cortex (A2), whose response deteriorates with shifts in component onset timings. The temporal window for harmonics integration in A2 was broadened by inactivation of somatostatin-expressing interneurons (SOM cells), but not parvalbumin-expressing interneurons (PV cells). Importantly, A2 has functionally connected subnetworks of neurons encoding harmonic, but not inharmonic sounds. These subnetworks are stable across days and exist prior to experimental harmonics exposure, suggesting their formation during development. Furthermore, A2 inactivation impairs performance in a discrimination task for coincident harmonics. Together, we propose A2 as a locus for harmonic integration, which may form the circuit basis for vocal processing.
Experimentally unsupervised deconvolution for light-sheet microscopy with propagation-invariant beams
Authors: Wijesinghe Philip , Corsetti Stella , Chow Darren J.X. , Sakata Shuzo , Dunning Kylie R. , Dholakia Kishan
Published: May 2021

Deconvolution is a challenging inverse problem, particularly in techniques that employ complex engineered point-spread functions, such as microscopy with propagation-invariant beams. Here, we present a deep learning method for deconvolution that, in lieu of end-to-end training with ground truths, is trained using known physics of the imaging system. Specifically, we train a generative adversarial network with images generated with the known point-spread function of the system, and combine this with unpaired experimental data that preserves perceptual content. Our method rapidly and robustly deconvolves and superresolves microscopy images, demonstrating a two-fold improvement in image contrast to conventional deconvolution methods. In contrast to common end-to-end networks that often require 1,000–10,000s paired images, our method is experimentally unsupervised and can be trained solely on a few hundred regions of interest. We demonstrate its performance on light-sheet microscopy with propagation-invariant Airy beams, including in calibration beads, oocytes, preimplantation embryos, and excised brain tissue, as well as illustrate its utility for Bessel-beam LSM. This method aims to democratise learned methods for deconvolution, as it does not require data acquisition outwith the conventional imaging protocol.
Rational inattention in mice
Authors: Grujic Nikola , Brus Jeroen , Burdakov Denis , Polania Rafael
Published: May 2021

Behavior exhibited by humans and other organisms is generally inconsistent and biased, and thus is often labeled irrational. However, the origins of this seemingly suboptimal behavior remain elusive. We developed a behavioral task and normative framework to reveal how organisms should allocate their limited processing resources such that there is an advantage to being imprecise and biased for a given metabolic investment that guarantees maximal utility. We found that mice act as rational-inattentive agents by adaptively allocating their sensory resources in a way that maximizes reward consumption in novel stimulus-reward association environments. Surprisingly, perception to commonly occurring stimuli was relatively imprecise, however this apparent statistical fallacy implies “awareness” and efficient adaptation to their neurocognitive limitations. Interestingly, distributional reinforcement learning mechanisms efficiently regulate sensory precision via top-down normalization. These findings establish a neurobehavioral foundation for how organisms efficiently perceive and adapt to environmental states of the world within the constraints imposed by neurobiology.
The BH3-only protein NOXA serves as an independent predictor of breast cancer patient survival and defines susceptibility to microtubule targeting agents
Authors: Karbon Gerlinde , Haschka Manuel D. , Hackl Hubert , Soratroi Claudia , Rocamora-Reverte Lourdes , Parson Walther , Fiegl Heidelinde , Villunger Andreas
Published: May 2021

Breast cancer (BC) treatment frequently involves microtubule-targeting agents (MTAs), such as paclitaxel, that arrest cells in mitosis. Sensitivity to MTAs is defined by a subset of pro- and anti-apoptotic BCL2 family proteins controlling mitochondrial apoptosis. Here, we aimed to determine their prognostic value in primary tumour samples from 92 BC patients. Our analysis identified high NOXA/PMAIP mRNA expression levels as an independent prognostic marker for improved relapse-free survival (RFS) and overall survival (OS) in multivariate analysis in BC patients, independent of their molecular subtype. Analysis of available TCGA datasets of 1060 BC patients confirmed our results and added a clear predictive value of NOXA mRNA levels for patients who received MTA-based therapy. In this TCGA cohort, 122 patients received MTA-treatment and high NOXA mRNA levels correlated with their progression-free interval (PFI) and OS. Our follow-up analyses in a panel of BC cell lines of different molecular subtypes identified NOXA protein expression as a key determinant of paclitaxel sensitivity in triple-negative breast cancer (TNBC) cells. Moreover, we noted highest additive effects between paclitaxel and chemical inhibition of BCLX, but not BCL2 or MCL1, documenting dependence of TNBC cells on BCLX for survival and paclitaxel sensitivity, defined by NOXA expression levels.
Predicting Composition of Genetic Circuits with Resource Competition: Demand and Sensitivity
Authors: McBride Cameron D. , Del Vecchio Domitilla
Published: May 2021

The design of genetic circuits typically relies on characterization of constituent modules in isolation to predict the behavior of modules’ composition. However, it has been shown that the behavior of a genetic module changes when other modules are in the cell due to competition for shared resources. In order to engineer multi-module circuits that behave as intended, it is thus necessary to predict changes in the behavior of a genetic module when other modules load cellular resources. Here, we introduce two characteristics of circuit modules: the demand for cellular resources and the sensitivity to resource loading. When both are known for every genetic module in a circuit library, they can be used to predict any module’s behavior upon addition of any other module to the cell. We develop an experimental approach to measure both characteristics for any circuit module using a resource sensor module. Using the measured resource demand and sensitivity for each module in a library, the outputs of the modules can be accurately predicted when they are inserted in the cell in arbitrary combinations. These resource competition characteristics may be used to inform the design of genetic circuits that perform as predicted despite resource competition.
Female mice exhibit a more sensitive automated squint response to pain induced by CGRP and amylin
Authors: Rea Brandon J. , Sowers Levi P. , Davison Abigail L. , Fairbanks Aaron M. , Wattiez Anne-Sophie , Poolman Pieter , Kardon Randy H. , Russo Andrew F.
Published: May 2021

We developed an automated squint assay using both black C57BL/6J and white CD1 mice that measured the interpalpebral fissure area between the upper and lower eyelids as an objective quantification of pain. In C57BL/6J mice, we observed a squint response to increasing doses of a migraine trigger, the neuropeptide CGRP, including a significant response in female mice at a dose below detection by the manual grimace scale. Using the automated software, both C57BL/6J and CD1 mice lacked a detectable photic blink response. The CGRP-related peptide amylin induced squinting behavior in female mice, but not males. These data demonstrate that an automated squint assay can be used as an objective, real-time continuous-scale measure of pain that provides higher precision and real-time analysis compared to manual grimace assessments.
Expression of putative effectors of different Xylella fastidiosa subspecies/strains reveals recognition and defense activation in various model plants
Authors: Sertedakis Matthaios , Kotsaridis Konstantinos , Tsakiri Dimitra , Dominguez-Ferreras Ana , Ntoukakis Vardis , Sarris Panagiotis F.
Published: May 2021

The re-emergence of Gram-negative bacterium Xylella fastidiosa in Europe in 2013 impelled the scientific community to discover novel strategies for crop protection. The wide host range of Xylella indicates the existence of yet not characterized pathogenic mechanisms to overcome plant defenses. The recent uprising accuracy of a variety of bioinformatics tools, with the ability to predict the function of putative microbial protein represent a useful approach for understanding which of these proteins are associated with pathogens virulence. In this study we collected a number of putative effectors from two X. fastidiosa strains: Temecula1 and CoDiRo and the subspecies (ssp.) Sandyi Ann-1. We designed an in-planta Agrobacterium based expression system that drives the expressed proteins to the cell apoplast, in order to investigate their ability to activate defense in various model plants. Furthermore, we organized the resulted proteins according to their sequential and structural similarities via the I-TASSER online tool. We identified that various X. fastidiosa proteins were able to differentially elicit cell death-like phenotypes in Nicotiana tabacum, N. sylvestris and N. benthamiana. These proteins are members of different enzymatic groups: a) hydrolases/hydrolases inhibitors, b) serine proteases and c) metal transferases. Collectively, we identified structurally similar proteins that were able to differentially elicit cell death-like phenotypes in different cultivars of the same species. Our findings provide the bases for further studies on the mechanisms that underlie host-defense activation by X. fastidiosa putative effectors, as well as, pathogens adaptation in susceptible hosts.
Lack of Cas13a inhibition by anti-CRISPR proteins from Leptotrichia prophages
Authors: Meeske Alexander J , Johnson Matthew C. , Hille Logan T. , Kleinstiver Benjamin P. , Bondy-Denomy Joseph
Published: May 2021

CRISPR systems are prokaryotic adaptive immune systems that use RNA-guided Cas nucleases to recognize and destroy foreign genetic elements, like bacteriophages and plasmids. To overcome CRISPR immunity, phages have evolved diverse families of anti-CRISPR proteins (Acrs), each of which inhibits the nucleic acid binding or cleavage activities of specific Cas protein families. Recently, Lin et al. (2020) described the discovery and characterization of 7 different Acr families (AcrVIA1-7) that inhibit type VI-A CRISPR systems, which use the nuclease Cas13a to perform RNA-guided RNA cleavage. In this Matters Arising article, we detail several inconsistencies that question the results reported in the Lin et al. (2020) study. These include inaccurate bioinformatics analyses, as well as reported experiments involving bacterial strains that are impossible to construct. The authors were unable to provide their published strains with which we might reproduce their experiments. We independently tested the Acr sequences described in Lin et al. (2020) in two different Cas13 inhibition assays, but could not detect anti-CRISPR activity. Taken together, our data and analyses prompt us to question the claim that AcrVIA1-7 reported in Lin et al. are bona fide type VI anti-CRISPR proteins.
Identifying and Classifying Goals For Scientific Knowledge
Authors: Boguslav Mayla R. , Salem Nourah M. , White Elizabeth K. , Leach Sonia M. , Hunter Lawrence E.
Published: May 2021

Motivation

Science progresses by posing good questions, yet work in biomedical text mining has not focused on them much. We propose a novel idea for biomedical natural language processing: identifying and characterizing thequestionsstated in the biomedical literature. Formally, the task is to identify and characterizeignorance statements, statements where scientific knowledge is missing or incomplete. The creation of such technology could have many significant impacts, from the training of PhD students to ranking publications and prioritizing funding based on particular questions of interest. The work presented here is intended as the first step towards these goals.

Results

We present a novel ignorance taxonomy driven by the role ignorance statements play in the research, identifying specific goals for future scientific knowledge. Using this taxonomy and reliable annotation guidelines (inter-annotator agreement above 80%), we created a gold standard ignorance corpus of 60 full-text documents from the prenatal nutrition literature with over 10,000 annotations and used it to train classifiers that achieved over 0.80 F1 scores.

Availability

Corpus and source code freely available for download athttps://github.com/UCDenver-ccp/Ignorance-Question-Work. The source code is implemented in Python.

Contact

Mayla.Boguslav@CUAnshcutz.edu
Genetic Variation as a Long-Distance Modulator of RAD21 Expression in Humans
Authors: Schierding William , Horsfield Julia A. , O’Sullivan Justin
Published: May 2021

Mutations and changes in expression in RAD21 are common across cancers types and outside of cancer can result in cohesinopathy. As such, exploration of variants that modify RAD21 enhancer activity, across the genome, may also provide insights into mechanisms by which distinct variants impact healthy human development and disease. We searched 42,953,834 genomic variants for a spatial-eQTL association with the transcription of RAD21. We identified 123 significant associations (FDR < 0.05), which are local (cis) or long-distance (trans) regulators of RAD21 expression. The 123 variants co-regulate a further seven genes, enriched for having Sp2 transcription factor binding sites in their promoter regions. The Sp2 transcription factor and six of the seven genes had previously been associated with cancer onset, progression, and metastasis. Our results suggest that genome-wide variation in non-coding regions impacts on RAD21 transcript levels in addition to other genes, which then could impact on oncogenesis and the process of ubiquitination. This identification of distant co-regulation of oncogenes represents a strategy for discovery of novel genetic regions which impact cancer onset and a potential for diagnostics.
Fitness Landscapes Reveal Context-dependent Benefits of Oviposition Choice
Authors: Ravi Kumar Vrinda , Agavekar Gaurav , Agashe Deepa
Published: May 2021

Resource choice behaviour has enormous fitness consequences and can drive niche expansion. However, individual behavioural choices are often mediated by context, determined by past experience. Are such context-dependent behaviours adaptive? Using Tribolium castaneum (the red flour beetle), we demonstrate that context-dependent oviposition choice reflects distinct, context-specific local fitness peaks. Manipulating female egg allocation in a habitat containing optimal and suboptimal resource patches, we measured offspring fitness to generate fitness landscapes as a function of all possible oviposition behaviours (i.e., combinations of fecundity and resource preference). Females from different age and competition contexts exhibit distinct behaviours which optimize different fitness components that are linked in a tradeoff. With increasing age and prior exposure to competition, they produce few but fast-developing offspring that are advantageous under high resource competition. In contrast, young naïve females produce many slow-developing offspring, beneficial under weak competition. Systematically mapping complete context-dependent fitness landscapes is thus critical to infer behavioural optimality and offers predictive power in novel contexts.

Preprint available at - https://www.biorxiv.org/content/10.1101/2021.05.27.445916v1.full

Citation - Vrinda Ravi Kumar, Gaurav Agavekar, Deepa Agashe; bioRxiv 2021.05.27.445916; doi: https://doi.org/10.1101/2021.05.27.445916
Production of human translation-competent lysates using dual centrifugation
Authors: Gurzeler Lukas-Adrian , Ziegelmüller Jana , Mühlemann Oliver , Karousis Evangelos D.
Published: May 2021

Protein synthesis is a central process in gene expression and the development of efficient in vitro translation systems has been the focus of scientific efforts for decades. The production of translation-competent lysates originating from human cells or tissues remains challenging, mainly due to the variability of cell lysis conditions. Here we present a robust and fast method based on dual centrifugation that allows for detergent-free cell lysis under controlled mechanical forces. We optimized the lysate preparation to yield cytoplasm-enriched extracts from human cells that efficiently translate mRNAs in a cap-dependent as well as in an IRES-mediated way. Reduction of the phosphorylation state of eIF2α using recombinant GADD34 and 2-aminopurine considerably boosts the protein output, reinforcing the potential of this method to produce recombinant proteins from human lysates.
Distinct Neural Representations of Decision Uncertainty in Metacognition and Mentalizing
Authors: Jiang Shaohan , Wang Sidong , Wan Xiaohong
Published: May 2021

Metacognition and mentalizing are both associated with meta-level mental state representations. Specifically, metacognition refers to monitoring one’s own cognitive processes, while mentalizing refers to monitoring others’ cognitive processes. However, this self-other dichotomy is insufficient to delineate the two high-level mental processes. We here used functional magnetic resonance imaging (fMRI) to systematically investigate the neural representations of different levels of decision uncertainty in monitoring different targets (the current self, the past self, and others) performing a perceptual decision-making task. Our results reveal diverse formats of intrinsic mental state representations of decision uncertainty in mentalizing, separate from the associations with external information. External information was commonly represented in the right inferior parietal lobe (IPL) across the mentalizing tasks. However, the meta-level mental states of decision uncertainty attributed to others were uniquely represented in the dorsomedial prefrontal cortex (dmPFC), rather than the temporoparietal junction (TPJ) that also equivalently represented the object-level mental states of decision inaccuracy attributed to others. Further, the object-level and meta-level mental states of decision uncertainty, when attributed to the past self, were represented in the precuneus and the lateral frontopolar cortex (lFPC), respectively. In contrast, the dorsal anterior cingulate cortex (dACC) consistently represented both decision uncertainty in metacognition and estimate uncertainty during monitoring the different mentalizing processes, but not the inferred decision uncertainty in mentalizing. Hence, our findings identify neural signatures to clearly delineate metacognition and mentalizing and further imply distinct neural computations on the mental states of decision uncertainty during metacognition and mentalizing.
Integrating digital and field surveillance to complement efforts to manage epidemic diseases of livestock: African swine fever as a case study
Authors: Tizzani Michele , Muñoz-Gómez Violeta , De Nardi Marco , Paolotti Daniela , Muñoz Olga , Ceschi Piera , Viltrop Arvo , Capua Ilaria
Published: May 2021

The SARS-CoV-2 pandemic has unveiled the importance of stakeholders and ordinary citizens in managing infectious disease emergencies. Efficient management of infectious diseases requires a top-down approach which must be complemented with a bottom-up response to be effective. Here we investigate a novel approach to surveillance for transboundary animal diseases using African Swine fever as a model. We were able to collect data at a population level on information-seeking behavior and at a local level through a targeted questionnaire-based survey to relevant stakeholders such as farmers and veterinary authorities. Our study shows how information-seeking behavior and resulting public attention during an epidemic, can be addressed through novel data streams from digital platforms such as Wikipedia. We also bring evidence on how field surveys aimed at local workers (e.g. farmers) and public authorities remain a crucial tool to assess more in-depth preparedness and awareness among front-line actors. We conclude that they should be used in combination to maximize the outcome of surveillance and prevention activities for selected transboundary animal diseases.
Abiotic factors modulate interspecies competition mediated by the type VI secretion system effectors inVibrio cholerae
Authors: Tang Ming-Xuan , Pei Tong-Tong , Wang Zeng-Hang , Luo Han , Wang Xing-Yu , Dong Tao G.
Published: May 2021

Vibrio cholerae, the etiological pathogen of cholera, relies on its type VI secretion system (T6SS) as an effective weapon to survive in highly competitive communities. The anti-bacterial and anti-eukaryotic functions of T6SS depend on its secreted effectors that target multiple essential cellular processes. However, the mechanisms that account for effector diversity and different effectiveness during interspecies competition remain elusive. Here, we report that environmental cations and temperature play a key role in dictating effector-mediated competition ofVibrio cholerae. We found thatV. choleraecould employ its cell-wall-targeting effector TseH to outcompete the otherwise resistantEscherichia coliand theV. choleraeimmunity deletion mutantΔtsiHwhen Ca2+and Mg2+were supplemented. TheE. coliΔphoQmutant was more sensitive to TseH-mediated killing during competition, suggesting the metal-sensing PhoPQ two-component system is protective toE. colifrom TseH activity. Using transcriptome analysis, we found multiple stress response systems, including acid stress response, oxidative stress response, and osmotic stress response, were activated inE. coliexpressing TseH in comparison withE. coliexpressing the inactive mutant TseHH64A. The membrane-targeting lipase effector TseL also exhibited reduced killing againstE. coliwhen divalent cations were removed. In addition, competition analysis ofE. coliwithV. choleraesingle-effector active strains reveals a temperature-dependent susceptibility ofE. colito effectors, VasX, VgrG3, and TseL. These findings suggest that abiotic factors, thatV. choleraefrequently encounters in natural habitats, play a crucial role in dictating the competitive fitness conferred by the type VI secretion system in complex multispecies communities.
hei-tag: a highly efficient tag to boost targeted genome editing
Authors: Thumberger Thomas , Tavhelidse Tinatini , Gutierrez-Triana Jose Arturo , Medert Rebekka , Cornean Alex , Welz Bettina , Freichel Marc , Wittbrodt Joachim
Published: May 2021

Precise, targeted genome editing by CRISPR/Cas9 is key for basic research and translational approaches in model and non-model systems1. While active in all species tested so far, editing efficiencies still leave room for improvement. To reach its target, the bacterial Cas9 needs to be efficiently shuttled into the nucleus as attempted by fusion of nuclear localization signals (NLSs) to the Cas9 protein2. Additional domains such as FLAG- or myc-tags are added for immediate detection or straight-forward purification3. To avoid steric hinderance impacting on activity, amino acid linkers are employed connecting Cas9 and additional domains.

We present the ‘hei-tag (high efficiency-tag)’, boosting the activity of the wide variety of CRISPR/Cas genome editing tools. The addition of the hei-tag to Cas9 or a C-to-T base editor dramatically enhances the respective targeting efficiency in model systems ranging from fish to mammals, including tissue culture applications. This allows to instantly upgrade existing and potentially highly adapted systems as well as establish novel highly efficient tools.
The 3-phosphoinositide-dependent protein kinase 1 is an essential upstream activator of protein kinase A in malaria parasites
Authors: Hitz Eva , Wiedemar Natalie , Passecker Armin , Brancucci Nicolas M. B. , Vakonakis Ioannis , Mäser Pascal , Voss Till S.
Published: May 2021

Cyclic AMP (cAMP) signalling is crucial for the propagation of asexual malaria blood stage parasites. Recent work on Plasmodium falciparum demonstrated that phosphorylation of the invasion ligand AMA1 by the catalytic subunit of cAMP-dependent protein kinase A (PfPKAc) is an essential step during parasite invasion into red blood cells. However, the exact mechanisms regulating PfPKAc activity are only partially understood and PfPKAc function has not been extensively studied in gametocytes, the sexual blood stage forms that are essential for malaria transmission. By studying a conditional PfPKAc knockdown mutant, we confirm the essential role for PfPKAc in erythrocyte invasion and demonstrate that PfPKAc is involved in regulating gametocyte deformability. Interestingly, we observed that the conditional overexpression of PfPKAc also caused a profound lethal phenotype by preventing intra-erythrocytic parasite multiplication. Whole genome sequencing of parasites selected to tolerate increased PfPKAc expression levels identified missense mutations exclusively in the gene encoding the putative parasite orthologue of 3-phosphoinositide-dependent protein kinase-1 (PfPDK1). Using targeted mutagenesis, we show that PfPDK1 is essential for PfPKAc activation, most likely by phosphorylating T189 in the PfPKAc activation loop. In summary, our results corroborate the importance of tight regulation of PfPKA signalling for parasite survival and identify PfPDK1 as a crucial upstream regulator in this pathway and potential new drug target.
Adaptive anti-tumor immunity is orchestrated by a population of CCL5-producing tissue-resident NK cells
Authors: Kirchhammer Nicole , Trefny Marcel P , Natoli Marina , Brücher Dominik , Smith Sheena N , Werner Franziska , Koch Victoria , Schreiner David , Bartoszek Ewelina , Buchi Mélanie , Schmid Markus , Breu Daniel , Hartmann K Patricia , Zaytseva Polina , Thommen Daniela S , Läubli Heinz , Böttcher Jan P , Stanczak Michal A , Kashyap Abhishek S , Plückthun Andreas , Zippelius Alfred
Published: May 2021

T cell-directed cancer immunotherapy often fails to generate lasting tumor control. Harnessing additional effectors of the immune response against tumors may strengthen the clinical benefit of immunotherapies. Here, we demonstrate that therapeutic targeting of the IFNγ-IL-12 pathway relies on the ability of a population of tissue-resident NK (trNK) cells to orchestrate an anti-tumor microenvironment. Particularly, utilizing an engineered adenoviral platform, we show that paracrine IL-12 enhances functional DC-CD8 T cell interactions to generate adaptive anti-tumor immunity. This effect depends on the abundance of trNK cells and specifically their capacity to produce the cDC1-chemoattractant CCL5. Failure to respond to IL-12 and other IFNγ-inducing therapies such as immune checkpoint blockade in tumors with low trNK cell infiltration could be overcome by intra-tumoral delivery of CCL5. Our findings reveal a novel barrier for T cell-focused therapies and offer mechanistic insights into how T cell-NK cell-DC crosstalk can be enhanced to promote anti-tumor immunity and overcome resistance.

Significance

We identified the lack of CCL5-producing, tissue-resident NK (trNK) cells as a barrier to T cell-focused therapies. While IL-12 induces anti-tumoral DC-T cell crosstalk in trNK cellrichtumors, resistance to IL-12 or anti-PD-1 in trNK cellpoortumors can be overcome by the additional delivery of CCL5.
A deep-learning-based toolbox for Automated Limb Motion Analysis (ALMA) in murine models of neurological disorders
Authors: Aljovic Almir , Zhao Shuqing , Chahin Maryam , de la Rosa del Val Clara , Van Steenbergen Valerie , Kerschensteiner Martin , Bareyre Florence M
Published: May 2021

In neuroscience research, the refined analysis of rodent locomotion is complex and cumbersome, and access to the technique is limited because of the necessity for expensive equipment. In this study, we implemented a new deep-learning-based toolbox for Automated Limb Motion Analysis (ALMA) that requires only basic behavioral equipment and an inexpensive camera. The ALMA toolbox enables the unbiased and comprehensive analyses of locomotor kinematics and paw placement and can be applied to neurological conditions affecting the brain and spinal cord. We demonstrated that the ALMA toolbox can (1) robustly track the evolution of locomotor deficits after spinal cord injury, (2) sensitively detect locomotor abnormalities after traumatic brain injury, and (3) correctly predict disease onset in a multiple sclerosis model. We, therefore, established a broadly applicable automated and standardized approach that requires minimal financial and time commitments to facilitate the comprehensive analysis of locomotion in rodent disease models.
Oligodendrocyte precursor cells guide the migration of cortical interneurons by unidirectional contact repulsion
Authors: Lepiemme Fanny , Mazzucchelli Gabriel , Silva Carla G. , Nguyen Laurent
Published: May 2021

The cerebral cortex is built by neural cells that migrate away from their birthplace. In the forebrain, ventrally-derived oligodendrocyte precursor cells (vOPCs) travel tangentially together with cortical interneurons (cINs) to reach the cortex. After birth, vOPCs form transient synapses with cINs before engaging later into myelination. Here we tested whether these populations interact during embryogenesis while migrating. By coupling histological analysis of mouse genetic models with live imaging, we showed that, while responding to the chemokine Cxcl12, vOPCs and cINs occupy mutually-exclusive forebrain territories. Moreover, vOPCs depletion selectively disrupts the migration and distribution of cINs. At the cellular level, we found that by promoting unidirectional contact-repulsion (UCoRe) of cINs, vOPCs steer their migration away from blood vessels and contribute to their allocation to proper migratory streams. UCoRe is thus an efficient strategy to spatially control the competition for a shared chemoattractant, thereby allowing cINs to reach proper cortical territories.
Genetic tools for the stable overexpression of circular RNAs
Authors: Mecozzi Nicol , Nenci Arianna , Vera Olga , Falzone Aimee , DeNicola Gina M. , Karreth Florian A.
Published: May 2021

Circular RNAs (circRNAs) are a class of non-coding RNAs that feature a covalently closed ring structure formed through backsplicing. circRNAs are broadly expressed and contribute to biological processes through a variety of functions. Standard gain-of-function and loss-of-function approaches to study gene functions have significant limitations when studying circRNAs. Overexpression studies in particular suffer from the lack of efficient genetic tools. While mammalian expression plasmids enable transient overexpression of circRNAs in cultured cells, most cell biological studies require long-term ectopic expression. Here we report the development and characterization of genetic tools enabling stable circRNA overexpressionin vitroandin vivo. We demonstrated that circRNA expression constructs can be delivered to cultured cells via transposons, whereas lentiviral vectors have limited utility for the delivery of circRNA constructs. We further showed that circRNA transposons can be supplied to mouse livers via hydrodynamic tail vein injection, resulting in ectopic circRNA expression in a hepatocellular carcinoma mouse model. Furthermore, we generated genetically engineered mice harboring circRNA expression constructs. We demonstrate that this approach enables constitutive, global circRNA overexpression as well as inducible circRNA expression directed specifically to melanocytes in a melanoma mouse model. Overall, these tools expand the genetic toolkit available for the functional characterization of circRNAs of interest.
Late-in-life treadmill-training rejuvenates autophagy, protein aggregate clearance, and function in mouse hearts
Authors: Cho Jae Min , Ly Kellsey , Ramous Caroline , Thompson Lauren , Hansen Michele , Mattera Maria Sara de Lima Coutinho , Pires Karla Maria , Ferhat Maroua , Whitehead Kevin J , Carter Kandis , Buffolo Márcio , Ghosh Rajeshwary , Park Seul-Ki , Boudina Sihem , Symons J David
Published: May 2021

ABSTRACT AND KEYWORDS

There is evidence for a progressive decline of protein quality control mechanisms during the process of cardiac aging. This enables the accumulation of protein aggregates and damaged organelles that contribute to age-associated cardiac dysfunction. Macroautophagy (referred to as autophagy) is the process by which post-mitotic cells such as cardiomyocytes clear defective proteins and organelles. We hypothesized that late-in-life exercise training improves autophagy, protein aggregate clearance, and function that is otherwise dysregulated in hearts from old vs adult mice. As expected, 24-month old male C57BL/6J mice (old) exhibited : (i) repressed autophagosome formation and protein aggregate accumulation in the heart; (ii) systolic and diastolic dysfunction; and (iii) reduced exercise capacity, vs. 8-month old (adult) mice (all p< .05). Separate cohorts of 21 month old mice completed a 3-month progressive resistance treadmill-running program (old-ETR) that improved (all < .05) : (i) body composition; (ii) exercise capacity; and (iii) soleus muscle citrate synthase activity, vs. age-matched mice that did not train (old-SED). Importantly, (iv) protein expression of autophagy markers indicated trafficking of the autophagosome to the lysosome increased, (v) protein aggregate clearance improved, and (vi) overall function was enhanced (all p<0.05), in hearts from old-ETR vs. old- SED mice. Dietary maneuvers and pharmacological interventions shown to elevate basal autophagy are reported to mitigate / reverse age-associated cardiac dysfunction. Here we show the first evidence that a physiological intervention initiated late-in-life improves autophagic flux, protein aggregate clearance, and overall function in mouse hearts.
Predicting phenotypes from genetic, environment, management, and historical data using CNNs
Authors: Washburn Jacob D. , Cimen Emre , Ramstein Guillaume , Reeves Timothy , O’Briant Patrick , McLean Greg , Cooper Mark , Hammer Graeme , Buckler Edward S.
Published: May 2021

Predicting phenotypes from genetic (G), environmental (E), and management (M) conditions is a long-standing challenge with implications to agriculture, medicine, and conservation. Most methods reduce the factors in a dataset (feature engineering) in a subjective and potentially oversimplified manner. Convolutional Neural Networks (CNN) can overcome this by allowing the data itself to determine which factors are most important. CNN models were developed for predicting agronomic yield from a combination of replicated trials and historical yield survey data. The results were more accurate than standard methods when tested on heldout G, E, and M data (r=0.5 vs r=0.4), and performed slightly worse than standard methods when only G was held out (r=0.74 vs r=0.78). Pre-training on historical data increased accuracy by 1-36% compared to trial data alone. Saliency map analysis indicated the CNN has “learned” to prioritize many factors of known agricultural importance.
Metapopulations with habitat modification
Authors: Miller Zachary R. , Allesina Stefano
Published: May 2021

Across the tree of life, organisms modify their local environment, rendering it more or less hospitable for other species. Despite the ubiquity of these processes, simple models that can be used to develop intuitions about the consequences of widespread habitat modification are lacking. Here we extend the classic Levins’ metapopulation model to a setting where each of n species can colonize patches connected by dispersal, and when patches are vacated via local extinction, they retain a “memory” of the previous occupant—modeling habitat modification. While this model can exhibit a wide range of dynamics, we draw several overarching conclusions about the effects of modification and memory. In particular, we find that any number of species may potentially coexist, provided that each is at a disadvantage when colonizing patches vacated by a conspecific. This notion is made precise through a quantitative stability condition, which provides a way to unify and formalize existing conceptual models. We also show that when patch memory facilitates coexistence, it generically induces a positive relationship between diversity and robustness (tolerance of disturbance). Our simple model provides a portable, tractable framework for studying systems where species modify and react to a shared landscape.
An Efficient Method to Quantify Structural Distributions in Heterogeneous cryo-EM Datasets
Authors: Gu Hanlin , Wang Wei , Unarta Ilona Christy , Zeng Wenqi , Sheong Fu Kit , Cheung Peter Pak-Hang , Liu Song , Yao Yuan , Huang Xuhui
Published: May 2021

Cryogenic Electron Microscopy (cryo-EM) preserves the ensemble of protein conformations in solution and thus provide a promising way to characterize conformational changes underlying protein functions. However, it remains challenging for existing software to elucidate distributions of multiple conformations from a heterogeneous cryo-EM dataset. We developed a new algorithm: Linear Combinations of Template Conformations (LCTC) to obtain distributions of multiple conformations from cryo-EM datasets. LCTC assigns 2D images to the template 3D structures obtained by Multi-body Re-finement of RELION via a novel two-stage matching algorithm. Specifically, an initial rapid assignment of experimental 2D images to template 2D images was applied based on auto-correlation functions of image contours that can efficiently remove the majority of irrelevant 2D images. This is followed by pixel-pixel matching of images with fewer number of 2D images, which can accurately assign the 2D images to the template images. We validate the LCTC method by demonstrating that it can accurately reproduce the distributions of 3Thermus aquaticus(Taq) RNA polymerase (RNAP) structures with different degrees of clamp opening from a simulated cryo-EM dataset, in which the correct distributions are known. For this dataset, we also show that LCTC greatly outperforms clustering-based Manifold Embedding and Maximum Likelihood-based Multi-body Re-finement algorithms in terms of reproducing the structural distributions. Lastly, we also successfully applied LCTC to reveal the populations of various clamp-opening conformations from an experimentalEscherichia coliRNAP cryo-EM dataset. Source code is available athttps://github.com/ghl1995/LCTC.
Cooperativity of catalytic and lectin-like domain ofT. congolensetrans-sialidase modulates its catalytic activity
Authors: Waespy Mario , Gbem Thaddeus T. , Kumar Nilima Dinesh , Mani Shanmugam Solaiyappan , Rosenau Jana , Dietz Frank , Kelm Sørge
Published: May 2021

Trans-sialidases (TS) represent a multi-gene family of unusual enzymes, which catalyse the transfer of terminal sialic acids from sialoglycoconjugates to terminal galactose orN-acetylgalactosamine residues of oligosaccharides without the requirement of CMP-Neu5Ac, the activated Sia used by typical sialyltransferases. Most work on trypanosomal TS has been done on enzymatic activities of TS fromT. cruzi(causing Chagas disease in Latin America), subspecies ofT. brucei, (causing human sleeping sickness in Africa) andT. congolense(causing African Animal Trypanosomosis in livestock). Previously, we demonstrated thatT. congolenseTS (TconTS) lectin domain (LD) binds to several carbohydrates, such as 1,4-β-mannotriose.

To investigate the influence of TconTS-LD on enzyme activities, we firstly performedin silicoanalysis on structure models of TconTS enzymes. Findings strongly supports the potential of domain swaps between TconTS without structural disruptions of the enzymes overall topologies. Recombinant domain swapped TconTS1a/TS3 showed clear sialidase and sialic acid (Sia) transfer activities, when using fetuin and lactose as Sia donor and acceptor substrates, respectively. While Sia transfer activity remained unchanged from the level of TconTS1a, hydrolysis was drastically reduced. Presence of 1,4-β-mannotriose during TS reactions modulates enzyme activities favouring trans-sialylation over hydrolysis.

In summary, this study provides strong evidence that TconTS-LDs play pivotal roles in modulating enzyme activity and biological functions of these and possibly other TS, revising our fundamental understanding of TS modulation and diversity.
Comparative chromatin accessibility upon BDNF-induced neuronal activity delineates neuronal regulatory elements
Authors: Ibarra Ignacio L. , Ratnu Vikram S. , Gordillo Lucia , Hwang In-Young , Mariani Luca , Weinand Kathryn , Hammarén Henrik M. , Bulyk Martha L. , Savitski Mikhail M. , Zaugg Judith B. , Noh Kyung-Min
Published: May 2021

Neuronal activity induced by brain-derived neurotrophic factor (BDNF) triggers gene expression, which is crucial for neuronal survival, differentiation, synaptic plasticity, memory formation, and neurocognitive health. However, its role in chromatin regulation is unclear. Here, using temporal profiling of chromatin accessibility and transcription in mouse primary cortical neurons upon either BDNF stimulation or depolarization (KCl), we identify features that define BDNF-specific chromatin-to-gene expression programs. Enhancer activation is an early event in the regulatory control of BDNF-treated neurons, where the bZIP motif-binding Fos protein pioneered chromatin opening and cooperated with co-regulatory transcription factors (Homeobox, EGRs, and CTCF) to induce transcription. Deleting cis-regulatory sequences decreased BDNF-mediated Arc expression, a regulator of synaptic plasticity. BDNF-induced accessible regions are linked to preferential exon usage by neurodevelopmental disorder-related genes and heritability of neuronal complex traits, which were validated in human iPSC-derived neurons. Thus, we provide a comprehensive view of BDNF-mediated genome regulatory features using comparative genomic approaches to dissect mammalian neuronal activity.
Retrosplenial and hippocampal synchrony during retrieval of old memories in macaques
Authors: Hussin A.T. , Abbaspoor S. , Hoffman K.L.
Published: May 2021

To understand the neural activity behind recollections of the distant past, we recorded from the hippocampus and retrosplenial cortex of macaques as they retrieved year-old and newly-acquired object-scene associations. Year-old memoranda preferentially evoked retrosplenial sigma oscillations (10-15 Hz), and phase-locking between RSC and eye movements during retrieval. In contrast, gamma-band retrosplenial-hippocampal synchrony was stronger during retrieval of new items. Primate retrosplenial oscillations may therefore guide retrieval of visuospatial events long past.