bluuebunny/crossref_metadata_2025 huggingface dataset:
Chemogenetic manipulation of endogenous proteins in fission yeast using a self-localizing ligand-induced protein translocation (SLIPT) system
Authors: Nakamura Akinobu , Goto Yuhei , Sugiyama Hironori , Tsukiji Shinya , Aoki Kazuhiro
Published: July 2023

Cells sense extracellular stimuli through membrane receptors and process the information through an intracellular signaling network. Protein translocation triggers intracellular signaling, and techniques such as chemically induced dimerization (CID) have been used to manipulate signaling pathways by altering the subcellular localization of signaling molecules. However, in the fission yeastSchizosaccharomyces pombe, the commonly used FKBP-FRB system has technical limitations, and therefore perturbation tools with low cytotoxicity and high temporal resolution are needed. We here applied our recently developed self-localizing ligand-induced protein translocation (SLIPT) system toS. pombeand successfully perturbed several cell cycle-related proteins. The SLIPT system utilizes self-localizing ligands to recruit binding partners to specific subcellular compartments, such as the plasma membrane or nucleus. We optimzed the self-localizing ligands to maintain long-term recruitment of target molecules to the plasma membrane. By knocking in genes encoding the binding partners for self-localizing ligands, we observed changes in the localization of several endogenous molecules and found perturbations in the cell cycle and associated phenotypes. This study demonstrates the effectiveness of the SLIPT system as a chemogenetic tool for rapid perturbation of endogenous molecules inS. pombe, providing a valuable approach for studying intracellular signaling and cell cycle regulation with improved temporal resolution.
Inhibition of the YAP-MMB interaction and targeting NEK2 as potential therapeutic strategies for YAP-driven cancers
Authors: Jessen Marco , Gertzmann Dörthe , Liss Franziska , Zenk Franziska , Bähner Laura , Schöffler Victoria , Ade Carsten P. , von Eyss Björn , Gaubatz Stefan
Published: July 2023

YAP activation in cancer is linked to poor outcomes, making it an attractive therapeutic target. Previous research focused on blocking the interaction of YAP with TEAD transcription factors. Here, we took a different approach by disrupting YAP’s binding to the transcription factor B-MYB using MY-COMP, a fragment of B-MYB containing the YAP binding domain fused to a nuclear localization signal. MY-COMP induced cell cycle defects, nuclear abnormalities, and polyploidization. In an AKT and YAP-driven liver cancer model, MY-COMP significantly reduced liver tumorigenesis, highlighting the importance of the YAP-B-MYB interaction in tumor development. MY-COMP also perturbed the cell cycle progression of YAP-dependent uveal melanoma cells but not of YAP-independent cutaneous melanoma cell lines. It counteracted YAP-dependent expression of MMB-regulated cell cycle genes, explaining the observed effects. We also identified NIMA-related kinase (NEK2) as a downstream target of YAP and B-MYB, promoting YAP-driven transformation by facilitating centrosome clustering and inhibiting multipolar mitosis.
RRIkinDP: Direct Paths RNA-RNA Interaction Kinetics Model
Authors: Waldl Maria , Beckmann Irene K. , Will Sebastian , Hofacker Ivo L.
Published: July 2023

Interactions between RNAs are an essential mechanism in gene regulation. State-of-the-art tools predict RNA-RNA interactions based on thermodynamic stability but largely neglect kinetic effects. To overcome this limitation, we propose novel models of RNA-RNA interaction dynamics. On this basis we can improve our understanding of general principles that govern RNA-RNA interaction formation and improve target prediction tools.

While the dynamics of secondary structure formation of single RNAs have been successfully modeled using transition systems between conformations [2, 10], analogous approaches for RNA-RNA interaction quickly lead to infeasibly large systems. Therefore, we propose reducing the interaction system to the direct trajectories (shortest paths) from possible first contacts to full hybridization. This key idea enables studying general principles and relevant features of the interaction formation as well as model details; e.g. the relative speed of intra- and intermolecular folding.

Due to the design of our RNA kinetics model, features like energy barriers can be computed efficiently. In the future, this could enable refining genome-wide target predictions through kinetic criteria. Beyond these immediate practical improvements, it could shed light on general principles like the long-debated influence of the accessibility of the initial contact site.

RRIkinDP is is available on GitHub athttps://github.com/mwaldl/RRIkinDP.
A Compact and Effective Photon-Resolved Image Scanning Microscope
Authors: Tortarolo Giorgio , Zunino Alessandro , Piazza Simonluca , Donato Mattia , Zappone Sabrina , Pierzyńska-Mach Agnieszka , Castello Marco , Vicidomini Giuseppe
Published: July 2023

Fluorescence confocal laser-scanning microscopy (LSM) is one of the most popular tools for life science research. This popularity is expected to grow thanks to single-photon array detectors tailored for LSM. These detectors offer unique single-photon spatiotemporal information, opening new perspectives for gentle and quantitative super-resolution imaging. However, a flawless recording of this information poses significant challenges for the microscope data-acquisition system. Here, we present a data-acquisition module based on the digital frequency domain principle, able to record photons’ essential spatial and temporal features. We use this module to extend the capabilities of established imaging techniques based on single-photon avalanche diode (SPAD) array detectors, such as fluorescence lifetime image scanning microscopy. Furthermore, we use the module to introduce a robust multi-species approach encoding the fluorophore’s excitation spectra in the time domain. Lastly, we combine time-resolved stimulated emission depletion microscopy with image scanning microscopy, boosting spatial resolution. Our results demonstrate how a conventional fluorescence laser scanning microscope can transform into a simple, information-rich, super-resolved imaging system with the simple addition of a SPAD array detector with a tailored data acquisition system. We expected a blooming of advanced single-photon imaging techniques which effectively harness all the sample information encoded in each photon.
The previously uncharacterized RnpM (YlxR) protein modulates the activity of ribonuclease P inBacillus subtilis
Authors: Wicke Dennis , Neumann Piotr , Gößringer Markus , Chernev Aleksandar , Poehlein Anja , Daniel Rolf , Urlaub Henning , Hartmann Roland K. , Ficner Ralf , Stülke Jörg
Published: July 2023

Even thoughBacillus subtilisis one of the most studied organisms, no function has been identified for about 20% of its proteins. Among these unknown proteins are several RNA- and ribosome-binding proteins suggesting that they exert functions in cellular information processing. In this work, we have investigated the RNA-binding protein YlxR. This protein is widely conserved in bacteria and strongly constitutively expressed inB. subtilissuggesting an important function. We have identified the RNA subunit of the essential RNase P as the binding partner of YlxR. The main activity of RNase P is the processing of 5’ ends of pre-tRNAs.In vitroprocessing assays demonstrated that the presence of YlxR results in reduced RNase P activity. Chemical cross-linking studies followed byin silicodocking analysis and experiments with site-directed mutant proteins suggest that YlxR binds to the region of the RNase P RNA that is important for binding and cleavage of the pre-tRNA substrate. We conclude that the YlxR protein is a novel interaction partner of the RNA subunit of RNase P that serves to finetune RNase P activity to ensure appropriate amounts of mature tRNAs for translation. We rename the YlxR protein RnpM for RNase P modulator.
High-quality co-expression networks for accurate gene function predictions in the fungal cell factoryAspergillus nigerand beyond
Authors: Schäpe Paul , Starke Stephan , Schuetze Tabea , Basenko Evelina , Jung Sascha , Cairns Timothy , Meyer Vera
Published: July 2023

Co-expression networks have recently emerged as a useful approach for updating and improving gene annotation at a near-genome level. This is based on the hypothesis that function can be inferred by delineating transcriptional networks in which a gene of interest is embedded. In this study, we generated a co-expression network for the filamentous cell factoryAspergillus nigerfrom 128 RNA-seq experiments. We confirm that over 70% of the >14,000A. nigergenes are represented in this network and show that gene functions can be accurately predicted as evidenced by analysis of various control sub-networks. Our analyses further indicate that this RNA-seq co-expression network has a higher predictive power compared to the microarray co-expression network that we published in 2019. To demonstrate the potential of the new co-expression network to unveil complex and non-intuitive predictions for gene regulation phenomena, we provide here new insights into the temporal, spatial and metabolic expression profile that connects a secreted antifungal peptide with mycelial growth, asexual development, secondary metabolism and pectin degradation inA. niger. To empower biologists to generate or apply co-expression networks in the fungal kingdom and beyond, we also demonstrate that (i) high quality networks can be generated from only 32 transcriptional experiments; (ii) such low numbers of experiments can be safely compensated for by using higher thresholds for defining co-expression pairs; and (iii) a ‘safety in numbers’ rule applies, whereby experimental conditions have limited impacts on network content provided a certain number of experiments are included.
RecurrentFBXW7mutations bypass Wnt/β-catenin addiction in cancer
Authors: Zhong Zheng , Virshup David M.
Published: July 2023

Pathologic Wnt/β-catenin signaling drives various cancers, leading to multiple approaches to drug this pathway. Appropriate patient selection can maximize success of these interventions. Wnt ligand addiction is a druggable vulnerability inRNF43-mutant/RSPO-fusion cancers. However, pharmacologically targeting the biogenesis of Wnt ligands, e.g., with PORCN inhibitors, has shown mixed therapeutic responses, possibly due to tumor heterogeneity. Here we show that the tumor suppressorFBXW7is frequently mutated inRNF43-mutant/RSPO-fusion tumors, andFBXW7mutations cause intrinsic resistance to anti-Wnt therapies. Mechanistically, inactivation of FBXW7 stabilizes multiple oncoproteins including Cyclin E and MYC, and antagonizes the cytostatic effect of Wnt inhibitors. Moreover, althoughFBXW7mutations do not mitigate β-catenin degradation upon Wnt inhibition,FBXW7-mutantRNF43-mutant/RSPO-fusion cancers instead lose dependence on β-catenin signaling, accompanied by dedifferentiation and loss of lineage specificity. TheseFBXW7-mutant Wnt/β-catenin-independent tumors are susceptible to multi-CDK inhibition by dinaciclib. An in depth understanding of primary resistance to anti-Wnt/β-catenin therapies allows for more appropriate patient selection and use of alternative mechanism-based therapies.
Chromatin activity of IκBα mediates the exit from naïve pluripotency
Authors: Palma Luis G. , Álvarez-Villanueva Daniel , Maqueda María , Barrero Mercedes , Iglesias Arnau , Bertran Joan , Álvarez-Errico Damiana , García-Prieto Carlos A. , Ballaré Cecilia , Rodriguez-Cortez Virginia , Bueno Clara , Vidal August , Villanueva Alberto , Menéndez Pablo , Stik Gregoire , Di Croce Luciano , Payer Bernhard , Esteller Manel , Espinosa Lluís , Bigas Anna
Published: July 2023

Maintenance of pluripotency is a multifactorial process in which NF-κB is a negative regulator. Our previous work identified a chromatin role for IκBα, the master regulator of NF-κB signaling, that is critical for the proper regulation of various tissue stem cells. Here, we found that IκBα accumulates specifically in the chromatin fraction of pluripotent embryonic stem cells. IκBα depletion does not affect NF-kB-dependent transcription, but causes a profound epigenetic rewiring in pluripotent stem cells, including alterations in H3K27me3, a histone mark catalyzed by Polycomb repression complex 2. Chromatin changes induced by IκBα depletion affect a subset of pluripotency genes and are associated with altered gene transcription. At the cellular level, IκBα-deficient embryonic stem cells are arrested in a naive pluripotency state when cultured in serum/LIF conditions and fail to exit pluripotency under differentiation conditions. By constructing separation-of-function mutants, we show that the effects of IκBα in regulating stem cell pluripotency are NF-κB-independent, but mainly rely on its chromatin-related function. Taken together, our results reveal a novel mechanism by which IκBα participates in the regulation of the pluripotent state of embryonic stem cells and shed light on the interplay between inflammatory signals and the regulation of pluripotency.
Enhancing genetically engineeredEscherichia colibioreporters for the detection of buried TNT-based landmines
Authors: Amiel Eden
Published: July 2023

TNT-based buried landmines are explosive devices consisting of mainly 2,4,6-trinitrotoluene (2,4,6-TNT), but also a 2,4-dinitrotoluene (2,4-DNT) impurity. It is reported that some 2,4-DNT impurity vapors slowly leak through land mine covers and into the soil above them, making this compound a potential indicator for the presence of buried landmines. There are several approaches to identify the presence of chemicals in the environment, one of which is based on a microbial detection system. This approach was previously used to address the challenge of remotely identifying buried landmines by applying genetically engineered microbial whole cell biosensors capable of sensing trace signatures of the explosive compound 2,4,6-TNT and its degradation product, 2,4-DNT. Upon exposure to 2,4-DNT, the genetically engineered bacterial reporter strains, containing a genetic fusion between a sensing element for 2,4-DNT (yqjFgene promoter) and a fluorescence/ bioluminescence reporting element (GFP/luxCDABEgenes), generate a measurable dose-dependent signal within the whole cell biosensor system. This biological component of a comprehensive remote sensing solution was significantly improved after several rounds of random mutagenesis to theyqjFpromoter region using a prone-to-error PCR and selection of the best mutants. In the present project further significant improvements were obtained in signal intensity, response time, and in the limit of detection, following an additional round of random mutagenesis. Two new mutants, named A10 and C5, were identified. The new mutants present high induction levels both in terms of Ratio (sample/control) and delta (sample-control), making these strains more amenable to remote detection. To isolate a more sensitive mutant, the 3rd round of random mutagenesis library (about 1000 strains) was re-exposed to lower concentration of 2,4-DNT, but no better candidate was isolated. Possible alternative approaches to lower the detection threshold of the system, such as the use of permeability mutants, are now being tested.
A disinhibitory circuit mechanism explains a general principle of peak performance during mid-level arousal
Authors: Beerendonk Lola , Mejías Jorge F. , Nuiten Stijn A. , de Gee Jan Willem , Fahrenfort Johannes J. , van Gaal Simon
Published: July 2023

Perceptual decision-making is highly dependent on the momentary arousal state of the brain, which fluctuates over time on a scale of hours, minutes, and even seconds. The textbook relationship between momentary arousal and task performance is captured by an inverted U-shape, as put forward in the Yerkes-Dodson law (Yerkes and Dodson, 1908). This law suggests optimal performance at moderate levels of arousal, and impaired performance at low or high arousal levels. However, despite its popularity, the evidence for this relationship in humans is mixed at best. Here, we use pupil-indexed arousal and performance data from various perceptual decision-making tasks to provide converging evidence for the inverted U-shaped relationship between spontaneous arousal fluctuations and performance across different decision types (discrimination, detection) and sensory modalities (visual, auditory). To further understand this relationship, we built a neurobiologically plausible mechanistic model and show that it is possible to reproduce our findings by incorporating two types of interneurons that are both modulated by an arousal signal. The model architecture produces two dynamical regimes under the influence of arousal: one regime in which performance increases with arousal, and another regime in which performance decreases with arousal, together forming an inverted U-shaped arousal-performance relationship. We conclude that the inverted U-shaped arousal-performance relationship is a general and robust property of sensory processing. It might be brought about by the influence of arousal on two types of interneurons that together act as a disinhibitory pathway for the neural populations that encode the available sensory evidence used for the decision.

Significance statement

When people are repeatedly performing the exact same task, their performance fluctuates over time. One of the causes of this behavioral variability is spontaneous fluctuations in arousal. According to the Yerkes-Dodson law, task performance is optimal at moderate levels of species’ arousal, with impaired performance at very low or high arousal levels. However, until now, the evidence supporting this law as a general mechanism in human decision-making is mixed, and a neural mechanism that may explain the inverted U-shaped arousal-performance relationship is lacking. We show that the Yerkes-Dodson law is a general law that holds for human observers across decision-making tasks and settings. Furthermore, we present a simple and neurobiologically plausible mechanistic model that can explain its existence.
Centrosome amplification primes for apoptosis and favors the response to chemotherapy in ovarian cancer beyond multipolar divisions
Authors: Edwards Frances , Fantozzi Giulia , Simon Anthony Y. , Morretton Jean-Philippe , Herbette Aurelie , Tijhuis Andrea E. , Wardenaar Rene , Foulane Stacy , Gemble Simon , Spierings Diana C.J. , Foijer Floris , Mariani Odette , Vincent-Salomon Anne , Roman Sergio Roman , Sastre-Garau Xavier , Goundiam Oumou , Basto Renata
Published: July 2023

Centrosome amplification is a feature of cancer cells associated with chromosome instability and invasiveness. Enhancing chromosome instability and subsequent cancer cell death via centrosome unclustering and multipolar divisions is an aimed-for therapeutic approach. Here we show that centrosome amplification favors responses to conventional chemotherapy independently of multipolar divisions and chromosome instability. We perform single-cell live imaging of chemotherapy responses in epithelial ovarian cancer cell lines and observe increased cell death when centrosome amplification is induced. By correlating cell fate with mitotic behaviors, we show that enhanced cell death occurs independently of chromosome instability. We identify that cells with centrosome amplification are primed for apoptosis. We show they are dependent on the apoptotic inhibitor BCL-XL, and that this is not a consequence of mitotic stresses associated with centrosome amplification. Given the multiple mechanisms that promote chemotherapy responses in cells with centrosome amplification, we assess such a relationship in an epithelial ovarian cancer patient cohort. We show that high centrosome numbers associate with improved chemotherapy responses and longer overall survival. Our work identifies apoptotic priming as a clinically relevant consequence of centrosome amplification, expanding our understanding of this pleiotropic cancer cell feature.
Predicting real-life creativity using resting state electroencephalography
Authors: Chhade Fatima , Tabbal Judie , Paban Véronique , Auffret Manon , Hassan Mahmoud , Vérin Marc
Published: July 2023

Neuroscience research has shown that specific functional brain patterns can be related to creativity during multiple tasks but also at rest. Nevertheless, the electrophysiological correlates of a highly creative brain remain largely unexplored. This study aims to uncover resting-state networks related to real-life creativity using high-density electroencephalography (HD-EEG) and to test whether the strength of functional connectivity within these networks could predict individual creativity. We acquired resting-state HD-EEG data from 90 participants who completed a creativity questionnaire. We then employed connectome-based predictive modeling; a machine-learning technique that predicts behavioral measures from brain connectivity features. Using a support vector regression, our results revealed functional connectivity patterns related to high and low creativity in the gamma frequency band. In leave-one-out cross-validation, the combined model of high and low creativity networks predicted creativity scores with very good accuracy (r= 0.34, p= 0.0009). Furthermore, the model’s predictive power was established by an external validation on an independent dataset (N= 41), where we found a statistically significant relationship between the observed and predicted creativity scores (r= 0.37, p= 0.01). These findings reveal large-scale networks that could predict individual real-life creativity at rest, providing a crucial foundation for developing EEG network-based markers of creativity.
Laminar pattern of adolescent development changes in working memory neuronal activity
Authors: Zhu Junda , Hammond Benjamin M. , Zhou Xin Maizie , Constantinidis Christos
Published: July 2023

Adolescent development is characterized by an improvement in cognitive abilities, such as working memory. Neurophysiological recordings in a non-human primate model of adolescence have revealed changes in neural activity that mirror improvement in behavior, including higher firing rate during the delay intervals of working memory tasks. The laminar distribution of these changes is unknown. By some accounts, persistent activity is more pronounced in superficial layers, so we sought to determine whether changes are most pronounced there. We therefore analyzed neurophysiological recordings from neurons recorded in the young and adult stage, at different cortical depths. Superficial layers exhibited increased baseline firing rate in the adult stage. Unexpectedly, changes in persistent activity were most pronounced in the middle layers. Finally, improved discriminability of stimulus location was most evident in the deeper layers. These results reveal the laminar pattern of neural activity maturation that is associated with cognitive improvement.

NEW AND NOTEWORTHY

Structural brain changes are evident during adolescent development particularly in the cortical thickness of the prefrontal cortex, at a time when working memory ability increases markedly. The depth distribution of neurophysiological changes during adolescence is not known. Here we show that neurophysiological changes are not confined to superficial layers, which have most often been implicated in the maintenance of working memory. Contrary to expectations, greatest changes were evident in intermediate layers of the prefrontal cortex.
Conserved atypical cadherin, Fat2, regulates axon terminal organization in the developingDrosophilaolfactory sensory neurons
Authors: Vien Khanh M. , Duan Qichen , Yeung Chun , Volkan Pelin Cayirlioglu
Published: July 2023

A unique and defining characteristic of the olfactory sensory circuit is its functionally organized topographic map, which requires widely dispersed olfactory sensory neurons with the same identity to converge their axons into one neuropil, a class-specific glomerulus. Understanding the process of how neuronal identity confers circuit organization is a major endeavor in the field of neurobiology due to its intricate connection to neurodegeneration and neuronal dysfunction. In the olfactory system, many aspects of circuit organization, from axon guidance to synaptic matching, are regulated by a variety of cell surface proteins (for example Robo/Slit and Toll receptors). In this paper, we’ve identified a novel atypical cadherin protein, Fat2 (also known as Kugelei), as a regulator of class-specific axon organization. Fat2 is expressed in olfactory receptor neurons (ORNs) and local interneurons (LNs) within the olfactory circuits, but little to no expression is found in projection neurons (PNs). Fat2 expression levels vary in a neuronal class-specific manner and peak during pupal development. Infat2null mutants, ORN axon terminals belonging to different ORN classes present with varying phenotypic severity with the highestfat2expressing classes being most severely affected. In the most extreme cases,fat2mutations lead to ORN degeneration. We then show evidence that suggests Fat2 intracellular domain is necessary for Fat2 function in ORN axon organization. Within the developmental context, Fat2 is required starting at early stages of olfactory circuit development specifically for precise axon retraction to further condense class-specific glomeruli. We’ve also shown that PNs’ and LNs’ expression of Fat2 likely does not contribute to ORN organization1. Lastly, we narrow down potential Fat2 intracellular domain interactors, APC family proteins (Adenomatous polyposis coli) anddop(Drop out), that likely orchestrate the cytoskeletal remodeling required for axon retraction during protoglomerular development. Altogether, we provide a foundational understanding of how Fat2 functions in olfactory circuit organization and implicate the critical role of axon retraction during glomerular maturation.
Activation of Autoreactive Lymphocytes in the Lung by STING Gain-of-function Mutation Radioresistant Cells
Authors: Gao Kevin MingJie , Nündel Kerstin , Chiang Kristy , Yin Xihui , Utz Paul J. , Fitzgerald Kate , Marshak-Rothstein Ann
Published: July 2023

Gain-of-function mutations in the dsDNA sensing adaptor STING lead to a severe autoinflammatory syndrome known as STING-associated vasculopathy with onset in Infancy (SAVI). SAVI patients develop interstitial lung disease (ILD) and commonly produce anti-nuclear antibodies (ANAs), indicative of concomitant autoimmunity. Mice heterozygous for the most common SAVI mutation, V154M (VM), also develop ILD, triggered by nonhematopoietic VM cells, but exhibit severe peripheral lymphopenia, low serum Ig titers and fail to produce autoantibodies. In contrast, we now show that lethally irradiated VM mice reconstituted with WT stem cells (WT→VM chimeras) develop ANAs and lung-reactive autoantibodies associated with accumulation of activated lymphocytes and formation of germinal centers in lung tissues. Moreover, when splenocytes from WT→VM chimeras were adoptively transferred into unmanipulated Rag1-/-mice, donor T cells accumulated in the lung. Overall, these findings demonstrate that expression of the VM mutation in non-hematopoietic cells can promote the activation of immunocompetent autoreactive lymphocytes.

Chimeric mice expressing STING only in non-hematopoietic cells develop systemic and lung directed autoimmunity which recapitulates what is seen in pediatric patients with SAVI disease.
Targeted protein modification with an antibody-based system
Authors: Rimon Oded , Konc Juraj , Ali Montader , Roy Chowdhury Vaidehi , Černauskienė Inga , Sormanni Pietro , Bernardes Gonçalo J. L. , Vendruscolo Michele
Published: July 2023

The chemical modification of proteins is one of the major mechanisms used to regulate the properties and functions of these macromolecules in the cell. It is therefore of great interest to develop tools to exploit this type of modifications for applications in molecular biology, medicine and biotechnology. Here we present a method of using antibodies to perform post-translational covalent modifications of endogenous proteins in complex environments by exploiting proximity-driven chemistry. The method is based on the ability of antibodies to hold a weakly reactive group close to its intended site of reaction by binding the target protein on a nearby epitope. We characterise this approach by modifying the green fluorescent protein in increasingly complex environments, and illustrate its applicability by targeting the disease-associated protein beta-2 microglobulin.
A CRISPR screen of HIV dependency factors revealsCCNT1is non-essential in T cells but required for HIV-1 reactivation from latency
Authors: Hafer Terry L , Felton Abby , Delgado Yennifer , Srinivasan Harini , Emerman Michael
Published: July 2023

We sought to explore the hypothesis that host factors required for HIV-1 replication also play a role in latency reversal. Using a CRISPR gene library of putative HIV dependency factors, we performed a screen to identify genes required for latency reactivation. We identified several HIV-1 dependency factors that play a key role in HIV-1 latency reactivation includingELL,UBE2M,TBL1XR1,HDAC3,AMBRA1, andALYREF. Knockout of Cyclin T1 (CCNT1), a component of the P-TEFb complex important for transcription elongation, was the top hit in the screen and had the largest effect on HIV latency reversal with a wide variety of latency reversal agents. Moreover,CCNT1knockout prevents latency reactivation in a primary CD4+ T cell model of HIV latency without affecting activation of these cells. RNA sequencing data showed that CCNT1 regulates HIV-1 proviral genes to a larger extent than any other host gene and had no significant effects on RNA transcripts in primary T cells after activation. We conclude that CCNT1 function is redundant in T cells but is absolutely required for HIV latency reversal.
Deep mutational scanning and machine learning uncover antimicrobial peptide features driving membrane selectivity
Authors: Randall Justin R. , Vieira Luiz C. , Wilke Claus O. , Davies Bryan W.
Published: July 2023

Antimicrobial peptides commonly act by disrupting bacterial membranes, but also frequently damage mammalian membranes. Deciphering the rules governing membrane selectivity is critical to understanding their function and enabling their therapeutic use. Past attempts to decipher these rules have failed because they cannot interrogate adequate peptide sequence variation. To overcome this problem, we develop deep mutational surface localized antimicrobial display (dmSLAY), which reveals comprehensive positional residue importance and flexibility across an antimicrobial peptide sequence. We apply dmSLAY to Protegrin-1, a potent yet toxic antimicrobial peptide, and identify thousands of sequence variants that positively or negatively influence its antibacterial activity. Further analysis reveals that avoiding large aromatic residues and eliminating disulfide bound cysteine pairs while maintaining membrane bound secondary structure greatly improves Protegrin-1 bacterial specificity. Moreover, dmSLAY datasets enable machine learning to expand our analysis to include over 5.7 million sequence variants and reveal full Protegrin-1 mutational profiles driving either bacterial or mammalian membrane specificity. Our results describe an innovative, high-throughput approach for elucidating antimicrobial peptide sequence-structure-function relationships which can inform synthetic peptide-based drug design.
Characterization of covalent inhibitors that disrupt the interaction between the tandem SH2 domains of SYK and FCER1G phospho-ITAM
Authors: Bashore Frances M. , Katis Vittorio L. , Du Yuhong , Sikdar Arunima , Wang Dongxue , Bradshaw William J. , Rygiel Karolina A. , Leisner Tina M. , Chalk Rod , Mishra Swati , Williams Andrew C. , Gileadi Opher , Brennan Paul E. , Wiley Jesse C. , Gockley Jake , Cary Gregory A. , Carter Gregory W. , Young Jessica E. , Pearce Kenneth H. , Fu Haian ,   the Emory-Sage-SGC TREAT-AD Center, Axtman Alison D.
Published: July 2023

RNA sequencing and genetic data support spleen tyrosine kinase (SYK) and high affinity immunoglobulin epsilon receptor subunit gamma (FCER1G) as putative targets to be modulated for Alzheimer’s disease (AD) therapy. FCER1G is a component of Fc receptor complexes that contain an immunoreceptor tyrosine-based activation motif (ITAM). SYK interacts with the Fc receptor by binding to doubly phosphorylated ITAM (p-ITAM) via its two tandem SH2 domains (SYK-tSH2). Interaction of the FCER1G p-ITAM with SYK-tSH2 enables SYK activation via phosphorylation. Since SYK activation is reported to exacerbate AD pathology, we hypothesized that disruption of this interaction would be beneficial for AD patients. Herein, we developed biochemical and biophysical assays to enable the discovery of small molecules that perturb the interaction between the FCER1G p-ITAM and SYK-tSH2. We identified two distinct chemotypes using a high-throughput screen (HTS) and orthogonally assessed their binding. Both chemotypes covalently modify SYK-tSH2 and inhibit its interaction with FCER1G p-ITAM.
An inhibitor/anti-inhibitor system controls the activity of lytic transglycosylase MltF inPseudomonas aeruginosa
Authors: Wang Michelle , Ma Sheya Xiao , Darwin Andrew J.
Published: July 2023

Most bacterial cell envelopes contain a cell wall layer made of peptidoglycan. The synthesis of new peptidoglycan is critical for cell growth, division and morphogenesis, and is also coordinated with peptidoglycan hydrolysis to accommodate the new material. However, the enzymes that cleave peptidoglycan must be carefully controlled to avoid autolysis. In recent years, some control mechanisms have begun to emerge, although there are many more questions than answers for how most cell wall hydrolases are regulated. Here, we report a novel cell wall hydrolase control mechanism inPseudomonas aeruginosa, which we discovered during our characterization of a mutant sensitive to the overproduction of a secretin protein. The mutation affected an uncharacterized Sel1-like repeat protein encoded by the PA3978 locus. In addition to the secretin-sensitivity phenotype, PA3978 disruption also increased resistance to a β-lactam antibiotic used in the clinic.In vivoandin vitroanalysis revealed that PA3978 binds to the catalytic domain of the lytic transglycosylase MltF and inhibits its activity. ΔPA3978 mutant phenotypes were suppressed by deletingmltF, consistent with them having been caused by elevated MltF activity. We also discovered another interaction partner of PA3978 encoded by the PA5502 locus. The phenotypes of a ΔPA5502 mutant suggested that PA5502 interferes with the inhibitory function of PA3978 towards MltF, and we confirmed that activity for PA5502in vitro. Therefore, PA3978 and PA5502 form an inhibitor/anti-inhibitor system that controls MltF activity. We propose to name these proteins Ilt (inhibitor of lytic transglycosylase) and Lii (lytic transglycosylase inhibitor, inhibitor).

IMPORTANCE

A peptidoglycan cell wall is an essential component of almost all bacterial cell envelopes, which determines cell shape and prevents osmotic rupture. Antibiotics that interfere with peptidoglycan synthesis have been one of the most important treatments for bacterial infections. Peptidoglycan must also be hydrolyzed to incorporate new material for cell growth and division, and to help accommodate important envelope-spanning systems. However, the enzymes that hydrolyze peptidoglycan must be carefully controlled to prevent autolysis. Exactly how this control is achieved is poorly understood in most cases, but is a highly active area of current research. Identifying hydrolase control mechanisms has the potential to provide new targets for therapeutic intervention. The work here reports the important discovery of a novel inhibitor/anti-nhibitor system that controls the activity of a cell wall hydrolase in the human pathogenPseudomonas aeruginosa, and which also affects resistance to an antibiotic used in the clinic.
Intrinsic neural timescales attenuate information transfer along the uni-transmodal hierarchy
Authors: Çatal Yasir , Wolman Angelika , Abbasi Samira , Northoff Georg
Published: July 2023

The brain’s intrinsic timescales are organized in a hierarchy with shorter timescales in sensory regions and longer ones in associative regions. This timescale hierarchy overlaps with the timing demands of sensory information. Our question was how does this timescale hierarchy affect information transfer. We used a model of the timescale hierarchy based on connected excitatory and inhibitory populations across the cortex. We found that a hierarchy of information transfer follows the hierarchy of timescales with higher information transfer in sensory areas while it is lower in associative regions. Probing the effect of changes in timescale hierarchy on information transfer, we changed various model parameters which all, through, the loss of hierarchy, induced increased information transfer. Finally, the steepness of the timescale hierarchy relates negatively to total information transfer. Human MEG data confirmed our results. In sum, we demonstrate a key role of the brain’s timescale hierarchy in mediating information transfer.
Network Statistics of the Whole-Brain Connectome ofDrosophila
Authors: Lin Albert , Yang Runzhe , Dorkenwald Sven , Matsliah Arie , Sterling Amy R. , Schlegel Philipp , Yu Szi-chieh , McKellar Claire E. , Costa Marta , Eichler Katharina , Bates Alexander Shakeel , Eckstein Nils , Funke Jan , Jefferis Gregory S.X.E. , Murthy Mala
Published: July 2023

Brains comprise complex networks of neurons and connections. Network analysis applied to the wiring diagrams of brains can offer insights into how brains support computations and regulate information flow. The completion of the first whole-brain connectome of an adultDrosophila, the largest connectome to date, containing 130,000 neurons and millions of connections, offers an unprecedented opportunity to analyze its network properties and topological features. To gain insights into local connectivity, we computed the prevalence of two- and three-node network motifs, examined their strengths and neurotransmitter compositions, and compared these topological metrics with wiring diagrams of other animals. We discovered that the network of the fly brain displays rich club organization, with a large population (30% percent of the connectome) of highly connected neurons. We identified subsets of rich club neurons that may serve as integrators or broadcasters of signals. Finally, we examined subnetworks based on 78 anatomically defined brain regions or neuropils. These data products are shared within the FlyWire Codex and will serve as a foundation for models and experiments exploring the relationship between neural activity and anatomical structure.
Daytime experiences shape neural activity and dream content in the sleeping brain
Authors: Kumral Deniz , Palmieri Jessica , Gais Steffen , Schönauer Monika
Published: July 2023

Learning-related brain activity patterns are replayed during sleep, and memories of recent experiences appear in our dreams. The connection between these phenomena, however, remains unclear. We investigated whether memory reinstatement during sleep contributes to dreaming. Participants listened to audiobooks before falling asleep. We could determine which audiobook they had studied based on dream reports collected during the night. Audiobook content was also reinstated at the neural level, in high-density EEG recordings. Brain activity during rapid eye movement sleep, particularly in the high-frequency beta range, carried information about the audiobook and simultaneously benefitted memory retention. Crucially, when the learning condition was manifest in neural activity, it also emerged in dreams. Reprocessing of daytime experiences during sleep thus shapes our brain activity, our dreams, and our memories.
How spokespeople help or hurt business through crisis messaging: experiments testing the roles of narratives, non-narratives and counterargument
Authors: Clementson David , Page Tyler
Published: July 2023

Purpose

When an audience mentally counterargues a spokesperson, the message is backfiring. In such cases, audience members are practically persuading themselves to take the opposite position advocated by the spokesperson. Yet spokespeople who are professional persuaders serving corporations often seem to instill counterargument. This paper examines the role of counterargument as the conduit through which a spokesperson's different message types affect a company during a crisis. The authors explore the paradox of spokespeople's (in)effectiveness by testing divides in research drawn from normative crisis communication theory, narrative persuasion theory and the theory of reporting bias.

Design/methodology/approach

Two controlled, randomized experiments are reported. Participants (total N = 828) watch video clips of media interviews of a company spokesperson fielding questions about a scandal.

Findings

In the first study, non-narrative information most effectively bolsters purchase intentions and reduces negative word-of-mouth. The effect is mediated by decreased counterargument. The second study replicates the results concerning on-topic narratives compared with spinning, while on-topic narratives and non-narratives perform equally well.

Originality/value

This study addresses conflicts between two distinct traditions of theory as well as between normative crisis communication and its frequent practice. Reducing counterargument matters in the context of non-narrative persuasion, and non-narratives can perform at least as well as narratives in crisis communication.
Making sense of health in PE: conceptions of health among Swedish physical education teachers
Authors: Korp Peter , Quennerstedt Mikael , Barker Dean , Johansson Anna
Published: July 2023

Purpose

Over the last couple of decades, health has become a central part of the subject content in physical education (PE) curricula in many countries. As a result, issues of health have been foregrounded much more clearly in the teaching of PE. The aim of this study was to explore how Swedish PE teachers make sense of health in relation to their teaching practices. This was done through investigating conceptions and theories about health in the teachers' descriptions of their teaching practices.

Design/methodology/approach

The data analyzed in this paper were collected through focus group and individual interviews with PE teachers in the grades 7–9 within compulsory schools in Sweden. The data were analyzed using thematic analysis.

Findings

Four dominant themes were identified in the data: 1) Health as a healthy attitude, 2) Health as a functional ability, 3) Health as fitness, 4) Health as mental wellbeing. There is a clear impact from healthism and obesity discourses on the teachers' accounts of health, but there is also an impact from holistic views and approaches to health. The authors contend that teachers should be explicit in what they mean by health in relation to what they teach, how they teach and why they teach health in a certain way.

Originality/value

The knowledge produced by this study is crucial since teachers' assumptions regarding health affect the subject content (what), the pedagogies (how), as well as the reasons (why) they teach health and therefore what students learn regarding health.
Impact of workplace incivility and emotional exhaustion on job outcomes – a study of the higher education sector
Authors: Parray Zahoor Ahmad , Islam Shahbaz ul , Shah Tanveer Ahmad
Published: July 2023

Purpose

The main goal of this research study is to look at the mediating effect of emotional exhaustion in the association between workplace incivility and job outcomes (job stress, job satisfaction and employee turnover intentions) in the higher education sector.

Design/methodology/approach

The authors gathered field data from individuals working in the Higher Education Sector of Jammu and Kashmir to test the proposed study paradigm. A total of 550 respondents reported their perceptions of workplace incivility, emotional exhaustion and job-related outcomes (job stress, job satisfaction and employee turnover intention) at Time 1 in the Kashmir division and Time 2 in the Jammu division of Jammu and Kashmir via a two-wave data collection design.

Findings

The findings supported the hypothesized relationships, demonstrating that emotional exhaustion acts as a mediator between workplace incivility and employee job outcomes (job stress, employee job satisfaction and employee intention to leave).

Research limitations/implications

The research was undertaken in Jammu and Kashmir's higher education sectors. As a result, the findings may not apply to other sectors because workplace incivility may be regarded differently in different sectors.

Practical implications

The findings of this research study will assist organizations and practitioners in comprehending the significance of workplace incivility and emotional exhaustion, as well as how they positively impact job-related outcomes (employee job stress, turnover intention) and negatively on job satisfaction.

Originality/value

This research study added to the existing Self-Determination Theory model developed by Deci and associates (2017) by incorporating Emotional Exhaustion, and workplace incivility as autonomous intrinsic and workplace context factors respectively into the SDT model to study work behaviors in terms of job outcomes. This study adds to existing knowledge on SDTs by suggesting and testing emotional exhaustion as a mechanism for determining the impact of workplace incivility on employee job outcomes.

Contribution to impact
Operations of cost-effective charities: a qualitative study
Authors: Shevchenko Anton , Hajmohammad Sara , Pagell Mark
Published: July 2023

Purpose

People donate to charities with the aim of improving society. Yet, many charities fail to use donations efficiently or have ineffective interventions. The authors explore the strategic operational priorities and processes that enable charities to efficiently implement their interventions and have a positive impact on society.

Design/methodology/approach

The authors first review the literature on charities to gain a deeper understanding of the current state of knowledge on charity operations. The authors then employ the lens of paradox theory and perform a qualitative investigation of six case studies to explore various aspects of the operations of charities that are known for being cost-effective.

Findings

The authors reveal how the strategic operational decisions of charities, as well as the processes they implement, help them resolve the tensions arising from the cost-effectiveness paradox. The authors show that cost-effective charities make strategic operational decisions that help maintain two diverging priorities: prioritizing the status quo and prioritizing change in how they deliver value. Another set of strategic decisions helps balance these two diverging priorities. The authors then show how these charities create and then maintain cost-effective operations.

Originality/value

The authors address recent calls for research on non-profit organizations in the field of operations management. To authors’ knowledge, it is the first in-depth study of exemplary charity operations. The results can be used by charity executives as a benchmarking tool when they develop and implement their charitable interventions and by government agencies and potential donors when they select charities for their donations. Finally, the results should have implications for other organizations trying to have a positive societal impact.