bluuebunny/crossref_metadata_2025_split huggingface dataset:
Discovery of hypercompact epigenetic modulators for persistent CRISPR-mediated gene activation
Authors: Carosso Giovanni A. , Yeo Robin W. , Gainous T. Blair , Jawaid M. Zaki , Yang Xiao , Kim James Y.S. , Jadhav Kavita , Juan-Sing Nina , Boregowda Siddaraju V. , Cutillas Vincent , Qi Lei Stanley , l’Hortet Alexandra Collin de , Daley Timothy P. , Hart Daniel O. 
Published: June 2023

AbstractProgrammable epigenetic modulators provide a powerful toolkit for controlling gene expression in novel therapeutic applications, but recent discovery efforts have primarily selected for potency of effect rather than contextual robustness or durability thereof. Current CRISPR-based tools are further limited by large cargo sizes that impede clinical delivery and, in gene activation contexts, by brief activity windows that preclude transient, single-dose strategies such as lipid nanoparticle (LNP) delivery. To address these limitations, we perform high-throughput screening to discover novel classes of transcriptional modulators derived from thousands of human, viral, and archaeal proteomes. We identify high-potency activators capable of mitotically stable gene activation in a multitude of cellular contexts and leverage machine learning models to rationally engineer variants with improved activities. In liver and T-cells, novel hypercompact activators (64 to 98 amino acids) derived from vIRF2 core domain (vCD) achieve superior potency and durable activation lasting weeks beyond the current large activators (∼five-fold larger). In a humanized mouse model, we target a human hypercholesterolemia susceptibility gene and achieve activation persisting five weeks after a single dose by LNP delivery. Our discovery pipeline provides a predictive rubric for the development of contextually robust, potent, and persistent activators of compact size, broadly advancing the therapeutic potential of epigenetic gene activation.
Disordered regions and folded modules in CAF-1 promote histone deposition inS. pombe
Authors: Ouasti Fouad , Audin Maxime , Freon Karine , Quivy Jean-Pierre , Tachekort Mehdi , Cesard Elizabeth , Thureau Aurélien , Ropars Virginie , Varela Paloma F. , Moal Gwenaelle , Amadou Ibrahim Soumana , Uryga Aleksandra , Legrand Pierre , Andreani Jessica , Guerois Raphael , Almouzni Geneviève , Lambert Sarah , Ochsenbein Francoise 
Published: June 2023

AbstractGenome and epigenome integrity in eukaryotes depends on the proper coupling of histone deposition with DNA synthesis. This process relies on the evolutionary conserved histone chaperone CAF-1 for which the links between structure and functions are still a puzzle. While studies of theS. cerevisiaeCAF-1 complex enabled to propose a model for the histone deposition mechanism, we still lack a framework to demonstrate its generality and in particular, how its interaction with the polymerase accessory factor PCNA is operating. Here, we reconstituted a completeSpCAF-1 from fission yeast. We characterized its dynamic structure using NMR, SAXS and molecular modeling together within vitroandin vivofunctional studies on rationally designed interaction mutants. Importantly, we identify the unfolded nature of the acidic domain which folds up when binding to histones. We also show how the long KER helix mediates DNA binding and stimulatesSpCAF-1 association with PCNA. Our study highlights how the organization of CAF-1 comprising both disordered regions and folded modules enables the dynamics of multiple interactions to promote synthesis-coupled histone deposition essential for its DNA replication, heterochromatin maintenance, and genome stability functions.
Temporal composition of the cervicovaginal microbiome associates with hrHPV infection outcomes in a longitudinal study
Authors: Molina Mariano A. , Leenders William P. J. , Huynen Martijn A. , Melchers Willem J. G. , Andralojc Karolina M. 
Published: June 2023

AbstractPersistent infections with high-risk human papillomavirus (hrHPV) can cause cervical squamous intraepithelial lesions (SIL) that may progress to cancer. The cervicovaginal microbiome (CVM) correlates with SIL, but the temporal composition of the CVM after hrHPV infections has not been fully clarified. To determine the association between the CVM composition and infection outcome, we applied high-resolution microbiome profiling using the circular probes-based RNA sequencing technology on a longitudinal cohort of cervical smears obtained from 141 hrHPV DNA-positive women with normal cytology at first visit, of whom 51 were diagnosed by cytology with SIL six months later. Here we show that women with a microbial community characterized by low diversity and highLactobacillus crispatusabundance exhibit low risk to SIL development at both visits, while women with a microbial community characterized by high diversity andLactobacillusdepletion at first visit have a higher risk of developing SIL. At the level of individual species we observed that an increased abundance forGardnerella vaginalisandAtopobium vaginaeassociate with SIL outcomes at both visits. These species together withDialister micraerophilusshowed a moderate discriminatory power for hrHPV infection progression. Our results suggest that the CVM can potentially be used as a biomarker for cervical disease and SIL development after hrHPV infection diagnosis with implications on cervical cancer prevention strategies and treatment of SIL.
Beyond antibiotic resistance: thewhiB7transcription factor coordinates an adaptive response to alanine starvation in mycobacteria
Authors: Poulton Nicholas C. , DeJesus Michael A. , Munsamy-Govender Vanisha , Roberts Cameron G. , Azadian Zachary A. , Bosch Barbara , Lin Karl Matthew , Li Shuqi , Rock Jeremy M. 
Published: June 2023

ABSTRACTPathogenic mycobacteria are a significant cause of morbidity and mortality worldwide. These bacteria are highly intrinsically drug resistant, making infections challenging to treat. The conservedwhiB7stress response is a key contributor to mycobacterial intrinsic drug resistance. Although we have a comprehensive structural and biochemical understanding of WhiB7, the complex set of signals that activatewhiB7expression remain less clear. It is believed thatwhiB7expression is triggered by translational stalling in an upstream open reading frame (uORF) within thewhiB75’ leader, leading to antitermination and transcription into the downstreamwhiB7ORF. To define the signals that activatewhiB7, we employed a genome-wide CRISPRi epistasis screen and identified a diverse set of 150 mycobacterial genes whose inhibition results in constitutivewhiB7activation. Many of these genes encode amino acid biosynthetic enzymes, tRNAs, and tRNA synthetases, consistent with the proposed mechanism forwhiB7activation by translational stalling in the uORF. We show that the ability of thewhiB75’ regulatory region to sense amino acid starvation is determined by the coding sequence of the uORF. The uORF shows considerable sequence variation among different mycobacterial species, but it is universally and specifically enriched for alanine. Providing a potential rationalization for this enrichment, we find that while deprivation of many amino acids can activatewhiB7expression,whiB7specifically coordinates an adaptive response to alanine starvation by engaging in a feedback loop with the alanine biosynthetic enzyme,aspC. Our results provide a holistic understanding of the biological pathways that influencewhiB7activation and reveal an extended role for thewhiB7pathway in mycobacterial physiology, beyond its canonical function in antibiotic resistance. These results have important implications for the design of combination drug treatments to avoidwhiB7activation, as well as help explain the conservation of this stress response across a wide range of pathogenic and environmental mycobacteria.
RewiringSaccharomyces cerevisiaemetabolism for optimised Taxol® precursors production
Authors: Nowrouzi Behnaz , Torres-Montero Pablo , Kerkhoven Eduard J. , Martínez José L. , Rios-Solis Leonardo 
Published: June 2023

AbstractSaccharomyces cerevisiaehas been recognised as a convenient host for the production of early precursors to the Taxol® anticancer drug. Recent studies have highlighted the harmful impact of oxidative stress as a result of the activity of Taxol® first cytochrome P450-reductase enzymes (Taxusspp. CYP725A4-POR). Here, we evolved a new oxidative stress-tolerant yeast strain on galactose, which led to a three-fold higher titre of the CYP725A4 enzyme substrate, taxadiene. We comprehensively analysed the performance of the evolved and parent strain in galactose-limited chemostat cultures before and during oxidative stress induction. Integrating the transcriptomics and metabolite profiling data in an enzyme-constrained genome scale model enabled a more accurate prediction of changes that occurred to biological pathways as a response to/consequence of evolution and oxidative stress. The analyses showed a better performance of the evolved strain with improved respiration and reduced overflow metabolites production. The strain was robust to re-introduction of the oxidative stress, potentially due to the cross-protection mechanism, which contributed to likely better heme, flavin and NADPH availability for an optimal expression ofCYP725A4andPORin yeast. The increased level of taxadiene production has potentially occurred due to the antioxidant properties of taxadiene or as a mechanism to overcome the toxicity of geranylgeranyl diphosphate, the precursor to taxadiene synthase.HighlightsThe antioxidant properties of taxadiene promotes its production inSaccharomyces cerevisiaeS. cerevisiaeALE on H2O2and galactose regulates Flavin, iron and NADPH metabolism as well as carbon and protein recycling pathways through cross-protection and anticipation mechanismsAbstract FigureFigure 1.Graphical abstract of the study.Figure was created with BioRender.com.
Genetic determinants of virulence and extensive drug resistance inPseudomonas aeruginosaPPA14 isolated from eggplant rhizosphere
Authors: Ambreetha Sakthivel , Parshatd Govindasamy , Castellanos Christian , Narasimhan Giri , Balachandar Dananjeyan , Cickovski Trevor , Mathee Kalai 
Published: June 2023

AbstractPseudomonas aeruginosais one of the Priority Level I critical pathogens that are least sensitive to antibiotics and can cause fatal hospital-acquired infections. This bacterium is predominantly present in the agricultural ecosystem. However, there are very limited studies on health threats associated withP. aeruginosastrains flourishing in edible plants. Previously, we isolated and characterized 18P. aeruginosastrains from vegetable plants directly harvested from the farms. In the current work, it has been hypothesized that plant-associatedP. aeruginosaharbors genetic determinants for virulence and resistance. To test this hypothesis,in vitroresistome profiles of the plant-associatedP. aeruginosastrains were assessed based on the Kirby-Bauer disk diffusion method. Hierarchical clustering analysis was done to identify the plant-associated strains that are phenotypically similar to clinical isolates. An eggplant-associated strain, PPA14, that exhibited high virulence and extensivein vitroresistance against eight antibiotic classes was selected for complete genome analyses. The PPA14 genome was sequenced using the Solexa-Illumina and Oxford-Nanopore platforms, assembled, and annotated. The presence of virulence-related and antibiotic resistance (ABR) genes were predicted using the ABRicate tool and validated based on standard reference databases such as VFDB, NCBI AMRFinderPlus, MEGARes, CARD, and ResFinder. IslandViewer4 tool was used to predict the genes acquired through horizontal gene transfer. Additionally, comparative analyses of all the plant-associated and environmentalP. aeruginosagenomes characterized so far were done using the Roary tool. The PPA14 genome size was 6.72 Mbp, encoding 6315 open reading frames. The genome harbored 49 ABR genes, including those coding for multiple families of efflux pumps that collectively confer resistance against 11 antibiotic classes. In addition, we detected 225 virulence-related genes, 83 genomic islands, and 235 unique genes in the PPA14 genome. Over 4% of the PPA14 genome is devoted to conferring virulence and extensive drug resistance. Our report highlights the health threat associated with an eggplant-associatedP. aeruginosa.
CISD3 is required for Complex I function, mitochondrial integrity, and skeletal muscle maintenance
Authors: Marjault Henri-Baptiste , Karmi Ola , Rowland Linda , Nguyen Thi Thao , Grant DeAna , Manrique-Acevedo Camila , Nechushtai Rachel , Mittler Ron 
Published: June 2023

ABSTRACTMitochondria play a central role in muscle metabolism and function. In skeletal muscles, a unique family of iron-sulfur proteins, termed CISD proteins, support mitochondrial function. The abundance of these proteins declines with aging leading to muscle degeneration. Although the function of the outer mitochondrial proteins CISD1 and CISD2 has been defined, the role of the inner mitochondrial protein CISD3, is currently unknown. Here we show that CISD3 deficiency in mice results in muscle atrophy that shares proteomic features with Duchenne Muscular Dystrophy. We further reveal that CISD3 deficiency impairs the function and structure of skeletal muscle mitochondria, and that CISD3 interacts with, and donates its clusters to, Complex I respiratory chain subunit NDUFV2. These findings reveal that CISD3 is important for supporting the biogenesis and function of Complex I, essential for muscle maintenance and function. Interventions that target CISD3 could therefore impact muscle degeneration syndromes, aging, and related conditions.
Sialylation of EGFR by ST6GAL1 induces receptor activation and modulates trafficking dynamics
Authors: Ankenbauer Katherine E. , Rao Tejeshwar C. , Mattheyses Alexa L. , Bellis Susan L. 
Published: June 2023

AbstractAberrant glycosylation is a hallmark of a cancer cell. One prevalent alteration is an enrichment in α2,6-linked sialylation of N-glycosylated proteins, a modification directed by the ST6GAL1 sialyltransferase. ST6GAL1 is upregulated in many malignancies including ovarian cancer. Prior studies have shown that the addition of α2,6 sialic acid to the Epidermal Growth Factor Receptor (EGFR) activates this receptor, although the mechanism was largely unknown. To investigate the role of ST6GAL1 in EGFR activation, ST6GAL1 was overexpressed in the OV4 ovarian cancer line, which lacks endogenous ST6GAL1, or knocked down in the OVCAR-3 and OVCAR-5 ovarian cancer lines, which have robust ST6GAL1 expression. Cells with high expression of ST6GAL1 displayed increased activation of EGFR and its downstream signaling targets, AKT and NFκB. Using biochemical and microscopy approaches, including Total Internal Reflection Fluorescence (TIRF) microscopy, we determined that the α2,6 sialylation of EGFR promoted its dimerization and higher order oligomerization. Additionally, ST6GAL1 activity was found to modulate EGFR trafficking dynamics following EGF-induced receptor activation. Specifically, EGFR sialylation enhanced receptor recycling to the cell surface following activation while simultaneously inhibiting lysosomal degradation. 3D widefield deconvolution microscopy confirmed that in cells with high ST6GAL1 expression, EGFR exhibited greater co-localization with Rab11 recycling endosomes and reduced co-localization with LAMP1-positive lysosomes. Collectively, our findings highlight a novel mechanism by which α2,6 sialylation promotes EGFR signaling by facilitating receptor oligomerization and recycling.
Rewiring capsule production by CRISPRi-based genetic oscillators demonstrates a functional role of phenotypic variation in pneumococcal-host interactions
Authors: Rueff Anne-Stéphanie , Raaphorst Renske van , Aggarwal Surya , Santos-Moreno Javier , Laloux Géraldine , Schaerli Yolanda , Weiser Jeffrey N. , Veening Jan-Willem 
Published: June 2023

AbstractPhenotypic variation is the phenomenon in which clonal cells display different traits even under identical environmental conditions. This plasticity is thought to be important for processes including bacterial virulence1–8, but direct evidence for its relevance is often lacking. For instance, variation in capsule production in the human pathogenStreptococcus pneumoniaehas been linked to different clinical outcomes9–14, but the exact relationship between variation and pathogenesis is not well understood due to complex natural regulation15–20. In this study, we used synthetic oscillatory gene regulatory networks (GRNs) based on CRISPR interference together with live cell microscopy and cell tracking within microfluidics devices to mimic and test the biological function of bacterial phenotypic variation. We provide a universally applicable approach for engineering intricate GRNs using only two components: dCas9 and extended sgRNAs (ext-sgRNAs). Our findings demonstrate that variation in capsule production is beneficial for pneumococcal fitness in traits associated with pathogenesis providing conclusive evidence for this longstanding question.
Actin and microtubules position stress granules
Authors: Böddeker Thomas J. , Rusch Anja , Leeners Keno , Murrell Michael P. , Dufresne Eric R. 
Published: June 2023

AbstractMembraneless organelles, composed of protein and nucleic acids, alter the biochemical and physical landscape of the cell. While specific membraneless organelles are found in stereotypical locations, little is known about the physical mechanisms that guide their positioning. Here, we investigate how stress granules, a type of cytoplasmic membraneless organelle, establish their stereotypical perinuclear positioning. We find that actin and microtubules play complementary roles. Lamellar actin confines stress granules, and its retrograde flow drives them toward the cell center. Microtubules, in turn, adhere to stress granules through capillary interactions, which tend to concentrate stress granules in micro-tubule rich regions near the nucleus. Similar physical mechanisms are likely to play a role in the positioning of other membraneless organelles.
Filament structure and subcellular organization of the bacterial intermediate filament-like protein crescentin
Authors: Liu Yue , van den Ent Fusinita , Löwe Jan 
Published: June 2023

ABSTRACTThe coiled coil protein crescentin is required for the crescent shape of the freshwater bacteriumCaulobacter crescentus(vibrioides). Crescentin forms a filamentous structure on the inner, concave side of the curved cells. It shares features with eukaryotic intermediate filament (IF) proteins, such as its ability to form filamentsin vitro, the protein’s length, sequence comparisons and the presence of a coiled coil discontinuity called the “stutter”. Here, we have used electron cryomicroscopy (cryo-EM) to determine the structure of the full-length protein and its filament, exploiting a crescentin-specific nanobody. The filament is formed by two strands, related by two-fold symmetry that each consist of two dimers, resulting in an octameric assembly. Crescentin subunits form longitudinal contacts head-to-head and tail-to-tail, making the entire filament non-polar. Usingin vivosite-directed cysteine crosslinking we demonstrated that contacts observed in thein vitrofilament structure exist in cells. Electron cryotomography (cryo-ET) of cells expressing crescentin showed filaments on the concave side of the curved cells, close to the inner membrane, where they form a band. Comparison of our crescentin filament structure with current models of IF proteins and their filaments revealed similar coiled coil dimer formation as well as an absence of overall polarity. IF proteins form head-to-tail longitudinal contacts in contrast to crescentin and hence several inter-dimer contacts in IFs have no equivalents in crescentin filaments. Our work supports the idea that intermediate filament-like proteins achieve their shared polymerization and mechanical properties through a variety of filament architectures.SIGNIFICANCE STATEMENTCrescentin is a coiled coil protein that is required for the crescent cell shape of bacteria such asCaulobacter crescentus. Crescentin shares biochemical and cytoskeletal properties with intermediate filament (IF) proteins, which form the third major class of cytoskeletal proteins in eukaryotes. To better understand the relationship between crescentin and IF proteins, and the filaments they form, we have determined the three-dimensional structure of crescentin filaments by cryo-EM. This revealed the full-length structure of the parallel coiled coil dimer of crescentin and how dimers come together laterally and longitudinally, to form a non-polar, octameric filament. Differences in filament architecture highlight the versatility of intermediate filament-like proteins across the tree of life.
The cost of foreign ownership: Voluntary sustainability reporting and financial performance in an emerging economy
Authors: Liou Ru-Shiun , Ting Pi-Hui , Chen Ying-Yu 
Published: June 2023

PurposeMany emerging economy firms are under foreign owners' pressure to embrace the challenges of addressing corporate social responsibility (CSR) and consider adopting sustainability initiatives. However, it is not clear how foreign ownership plays a role to enable or inhibit these emerging economy firms from translating sustainability initiatives into improved financial performance. Utilizing neo-institutional theory, the authors argue that emerging economy firms that voluntarily report sustainability gain legitimacy in the eyes of shareholders and improve stock market performance. However, emerging economy firms may not have the resources to reconcile the internal stakeholders' various legitimacy requirements to promote sustainability practices, resulting in a negative association with accounting performance. Foreign ownership attenuates the relationship between sustainability reporting and firm performance due to the different legitimacy requirements in foreign markets.Design/methodology/approachTo test the study’s hypotheses, the authors collected and analyzed a large sample of publicly listed firms between 2010 and 2016 in Taiwan where the types of foreign ownership include foreign trust funds, foreign financial institutions and other foreign legal entities. Regression analyses were conducted to investigate whether the firms that report their sustainable practices have better financial performance, including stock market performance and accounting performance. Additionally, a three-step procedure was employed to address the endogeneity issue with a binary explanatory variable.FindingsThe positive stock market reaction to the emerging economy firms' voluntary sustainability reporting supports legitimacy gained among investors. By contrast, sustainability reporting has a negative association with accounting performance due to the difficulty of reconciling different legitimacy requirements among various stakeholders in emerging economies. Further, foreign ownership, particularly the trust fund, exhibits a negative moderating effect on the relationship between sustainability reporting in aligning corporate practices with sustainable development goals (SDGs) and the company's stock market performance.Originality/valueBy examining the less tested contingent role played by foreign ownership in the emerging economy firms' sustainability reporting, the authors provide insights into the influence exerted by different types of foreign ownership on firms' financial performances beyond previous studies that focus on family ownership, state ownership, or managerial ownership in emerging economies. The findings shed light on corporate sustainability strategy and foreign direct investment policies for an emerging economy.