bluuebunny/crossref_metadata_2025_split huggingface dataset:
Paused Polymerase synergises with transcription factors and RNA to enhance the transcriptional potential of promoters
Authors: Mann Rajat , Soota Deepanshu , Das Ayan , Podh Nitesh Kumar , Mehta Gunjan , Notani Dimple 
Published: October 2024

AbstractRegulated pausing of RNA polymerase II (Pol II) is essential for enabling rapid and coordinated transcriptional responses to signalling cues. Pausing also contributes to the formation of nucleosome-free regions with the help of chromatin remodellers. However, if these nucleosome-free regions engage with transcription factors to stimulate the transcription potential of paused promoters is not known. In this study, we demonstrate that ligand-induced estrogen receptor-alpha (ERα) binding is stabilized at Pol II-paused sites. This stabilization results from an increased dwell time of ERα on chromatin, as revealed by single molecule tracking (SMT) experiments. Notably, short chromatin-associated RNAs generated by the paused Pol II contribute to enhancing ERα binding at paused promoters. We also observe that pausing increases histone H3K27 acetylation (H3K27ac) levels, which primes paused promoters for robust transcriptional activation upon release. Collectively, these findings suggest that paused Pol II plays a central role in enhancing transcription factor binding through an RNA-dependent mechanism. This, in turn, results in a more vigorous transcriptional response following pausing release, thus contributing to the fine-tuning of ERα-mediated gene regulation.
Wnt specifically induces FZD5/8 endocytosis and degradation and the involvement of RSPO-ZNRF3/RNF43 and DVL
Authors: Luo Dong , Zheng Jing , Lv Shuning , Sheng Ren , Chen Maorong , He Xi , Zhang Xinjun 
Published: October 2024

AbstractFrizzled (FZD) proteins are the principal receptors of the Wnt signaling pathway. However, whether Wnt ligands induce FZD endocytosis and degradation remains elusive. The transmembrane E3 ubiquitin ligases ZNRF3 and RNF43 promote the endocytosis and degradation of FZD receptors to inhibit Wnt signaling, and their function is antagonized by R-spondin (RSPO) proteins. However, the dependency of RSPO-ZNRF3/RNF43-mediated FZD endocytosis and degradation on Wnt stimulation, as well as the specificity of this degradation for different FZD, remains unclear. Here, we demonstrated that Wnt specifically induces FZD5/8 endocytosis and degradation in a ZNRF3/RNF43-dependent manner. ZNRF3/RNF43 selectively targets FZD5/8 for degradation upon Wnt stimulation. RSPO1 enhances Wnt signaling by specifically stabilizing FZD5/8. Wnt promotes the interaction between FZD5 and RNF43. We further demonstrated that DVL proteins promote ligand-independent endocytosis of FZD but are dispensable for Wnt-induced FZD5/8 endocytosis and degradation. Our results reveal a novel negative regulatory mechanism of Wnt signaling at the receptor level and illuminate the mechanism by which RSPO-ZNRF3/RNF43 regulates Wnt signaling, which may provide new insights into regenerative medicine and cancer therapy.
Functional differentiation of Human Dental Pulp Stem Cells into neuron-like cells exhibiting electrophysiological activity
Authors: Pardo-Rodríguez B. , Baraibar A.M. , Manero-Roig I. , Luzuriaga J. , Salvador-Moya J. , Polo Y. , Basanta-Torres R. , Unda F. , Mato S. , Ibarretxe G. , Pineda J.R. 
Published: October 2024

AbstractBackgroundHuman dental pulp stem cells (hDPSCs) constitute a promising alternative for central nervous system (CNS) cell therapy. Unlike other human stem cells, hDPSCs can be differentiated, without genetic modification, to neural cells that secrete neuroprotective factors. However, a better understanding of their real capacity to give rise to functional neurons and integrate into synaptic networks is still needed. For that,ex vivodifferentiation protocols must be refined, especially to avoid the use of fetal animal serum.MethodsIn this study, we sought to improve existing differentiation protocols for obtaining functional neuron-like cells from hDPSCs. We compared the effects of the absence or presence of fetal serum during the initial expansion phase as a step prior to switching cultures to neurodifferentiation media. We improved hDPSC neurodifferentiation by adding retinoic acid (RA) and potassium chloride (KCl) pulses for 21 or 60 days and characterized the results by immunofluorescence, digital morphometric analysis, RT-qPCR and electrophysiology.ResultsWe found that neural markers like Nestin, GFAP, S100β and p75NTRwere expressed differently in neurodifferentiated hDPSC cultures depending on the presence or absence of serum during the initial cell expansion phase. In addition, hDPSCs previously grown as spheroids in serum-free medium exhibitedin vitroexpression of neuronal markers such as doublecortin (DCX), neuronal nuclear antigen (NeuN), Ankyrin-G and MAP2 after neurodifferentiation. Presynaptic vGLUT2, Synapsin-I, and excitatory glutamatergic and inhibitory GABAergic postsynaptic scaffold proteins and receptor subunits were also present in these neurodifferentiated hDPSCs. Treatment with KCl and RA increased the amount of both voltage-gated Na+and K+channel subunits in neurodifferentiated hDPSCs at the transcript level. Consistently, these cells displayed voltage-dependent K+and TTX-sensitive Na+currents as well as spontaneous electrophysiological activity and repetitive neuronal action potentials with a full baseline potential recovery.ConclusionOur study demonstrates, for the first time, that hDPSCs can be differentiated to neuronal-like cells that display functional excitability and thus evidence the potential of these easily accessible human stem cells for nerve tissue engineering. Our results highlight the importance of choosing an appropriate culture protocol to successfully neurodifferentiate hDPSCs.
Epigenetic modulation of RIPK3 by Transglutaminase 2-dependent serotonylation of H3K4me3 affects necroptosis
Authors: Vecchio Alessio , Colasuonno Fiorella , Bellanca Veronica , Pitolli Consuelo , Occhigrossi Luca , Ciccarone Fabio , D’Eletto Manuela , Sano Federica Di , Pagliarini Vittoria , Sette Claudio , Piacentini Mauro , Rossin Federica 
Published: October 2024

ABSTRACTThe receptor interacting protein kinase 3 (RIPK 3) is the main player in the activation of necroptosis, a pro-inflammatory regulated cell death modality induced by many different stimuli. RIPK3 is epigenetically regulated by DNA methylation and can be expressed when its promoter is associated with H3K4me3 histone. In this study, we show that Transglutaminase 2 protein (TG2) is necessary to induce necroptosis pathway allowing the expression ofRipk3gene. Indeed, cells lacking TG2 show a strong downregulation ofRipk3gene and are resistant to necroptotic stimuli. TG2 is known to promote the serotonylation of H3K4me3 histone (H3K4me3Q5ser) regulating in this way the target gene expression. Interestingly, we find that TG2 interacts with both histones H3K4me3 and H3K4me3Q5ser and these post-translational modifications are associated with theRipk3promoter only in presence of TG2. In addition, the absence of the H3K4me3Q5ser, in cells lacking TG2, is correlated withRipk3gene methylation. Altogether, these results indicate that RIPK3 expression requires TG2 mediated serotonylation of H3K4me3 to preventRipk3promoter methylation, thus favouring its expression and necroptosis induction.HIGHLIGHTS- Transglutaminase 2 protein (TG2) is necessary to induce necroptosis.- Cells lacking TG2 show a strong downregulation ofRipk3gene.- TG2 allows Ripk3 gene expression by promoting the serotonylation on H3K4me3 histone.SYNOPSIS- TG2 serotonylates histone H3K4me3, associated toRipk3promoter, favoring its expression.- TG2 dependent serotonylation of histone H3K4me3 prevents DNA methylation.- TG2 favours the expression of Ripk3 making cells susceptible to necroptosis.- In the absence of TG2, neither serotonylation nor methylation occur on histone H3, associated to Ripk3 promoter.- In the absence of TG2, Ripk3 promoter is methylated, thus blocking the activation of necroptosis.
SNAI2 cooperates with MEK1/2 and HDACs to suppress BIM- and BMF-dependent apoptosis inTERTpromoter mutant cancers
Authors: Tandon Amol , Stern Josh 
Published: October 2024

AbstractCancers withTERTpromoter mutations (TPM) display elevated RAS pathway signaling and mesenchymal traits, and associate with lower patient survival rates. We examined whether RAS pathway signaling in TPM cancers cooperates with mesenchymal features to drive resistance to apoptosis. We observed that RAS pathway signaling in TPM cancers inhibited apoptosis by downregulating the pro-apoptotic protein BIM. By using inhibitors of MEK1/2 kinases, we rescued the ability of TPM cancer cells to undergo apoptosis, which may have implications for targeted therapies. To further capitalize on this rescue, we explored combination treatments to drive apoptotic cell death. Treatment with the pan-BCL2 inhibitor, navitoclax (NX), in combination with MEK inhibition, significantly increased apoptosis, indicating that these cells are capable of undergoing intrinsic apoptosis, with BIM likely playing a critical role. Further, we found that transcriptional reprogramming of the mesenchymal state of TPM cancers using histone deacetylase inhibitors (HDACi) resulted in a synergistic increase in apoptosis, contingent upon BIM de-repression. Notably, the cause of this apoptosis appeared to be independent of DNA damage. The suppression of the mesenchymal transcription factor SNAI2, which has known roles in recruiting HDACs to silence gene expression, amplified apoptosis. Mechanistically, knockdown of SNAI2 impaired the cellular DNA repair leading to elevated basal levels of phosphorylated H2AX. Our findings show that TPM cancers exhibit specific small molecule vulnerabilities, driven by the convergence of RAS-MEK signaling and impaired HDAC regulation dependent on pro-apoptotic BH3-only proteins. Based on our findings, we propose that stratifying cancers based on TPM may identify a subset of tumors that are responsive to innovative combinations of inhibitors targeting these axes.
Systemic effects of cystic fibrosis transmembrane conductance regulator (CFTR) modulators on the blood proteome
Authors: Fentker Kerstin , Kirchner Marieluise , Ziehm Matthias , Niquet Sylvia , Popp Oliver , Duerr Julia , Schaupp Laura , Roehmel Jobst , Thee Stephanie , Hämmerling Susanne , Sommerburg Olaf , Stahl Mirjam , Graeber Simon Y. , Mall Marcus A. , Mertins Philipp 
Published: October 2024

SummaryCystic fibrosis (CF), resulting from a dysfunction in the cystic fibrosis transmembrane conductance regulator (CFTR), affects multiple organs through mucus obstruction and differences in secretion. The CFTR modulator drug combination elexacaftor/tezacaftor/ivacaftor (ELX/TEZ/IVA, ETI) has markedly improved clinical symptoms, but its broader molecular and systemic effects remain to be fully elucidated.We employed mass spectrometry-based proteomics to compare the blood proteomes of CF patients treated with the earlier, less effective lumacaftor/ivacaftor (LUM/IVA) combination against those receiving the more potent ELX/TEZ/IVA therapy. Our analysis revealed both specific and common pharmacodynamic signatures associated with inflammation and metabolic processes under each treatment regimen. Notably, ELX/TEZ/IVA therapy exhibited more consistent alterations across patients that were directed towards profiles observed in healthy individuals.Furthermore, by comparing sputum and blood proteomes of ELX/TEZ/IVA treated patients we identified counter-directional changes in the pulmonary surfactant-associated protein B, SFTPB, a potential biomarker of lung tissue repair, which also correlated with lung function improvements.This study provides a comprehensive resource that enhances our understanding of CFTR modulator-driven proteome alterations, offering insights to both systemic and local protein regulation in CF. Our findings indicate that ELX/TEZ/IVA promotes broader systemic health improvements, providing critical insights that could shape future therapeutic strategies in CF.
Multiplexed phosphoproteomics of low cell numbers using SPARCE
Authors: Gaizley Emily J. , Chen Xiuyuan , Bhamra Amandeep , Surinova Silvia 
Published: October 2024

AbstractUnderstanding cellular diversity and disease mechanisms requires global analysis of proteins and their modifications. While next-generation sequencing has advanced our understanding of cellular heterogeneity, it fails to capture downstream signalling networks. Ultrasensitive mass spectrometry-based proteomics enables unbiased protein-level analysis of low cell numbers, down to single cells. However, phosphoproteomics remains limited to high-input samples due to sample losses and poor reaction efficiencies associated with processing low cell numbers. Isobaric stable isotope labelling is a promising approach for reproducible and accurate quantification of low abundant phosphopeptides. Here, we introduce SPARCE (StreamlinedPhosphoproteomicAnalysis ofRareCElls) for multiplexed phosphoproteomic analysis of low cell numbers. SPARCE integrates cell isolation, water-based lysis, on-tip TMT labelling, and phosphopeptide enrichment. SPARCE outperforms traditional methods by enhancing labelling efficiency and phosphoproteome coverage. To demonstrate the utility of SPARCE, we analysed four patient-derived glioblastoma stem cell lines, reliably quantifying phosphosite changes from 1,000 FACS-sorted cells. This workflow expands the possibilities for signalling analysis of rare cell populations.
Monitoring lysosomal catabolism: a sensitive probe for assessing targeted lysosomal degradation of extracellular proteins
Authors: Holly Ryan W. , Hu Yi-Zhen , Valsan Vaishnavi Beena , Kougentakis Christos , Petronikolou Nektaria , Chen Tao , Li Diana , Chen Aimei , Rank Steve , Doran Kyle H. , Turtle Eric , Langsdorf Christopher , Shurtleff Matthew J. 
Published: October 2024

AbstractExtracellular targeted protein degradation (eTPD) is an emerging therapeutic field. The Lysosome targeting chimera (LYTAC) is a therapeutic modality that promotes degradation of extracellular drivers of disease in the lysosome. While widely available pH-sensitive probes may report on lysosome delivery, these probes do not necessarily report on the enzymatically active functional state of the lysosome. We report the development and application of a sensitive fluorescent probe, LysoLight Deep Red, to monitor catabolism of internalized proteins in the lysosome based on cleavage by cathepsin proteases. We demonstrate the application of Lysolight Deep Red to monitor the catabolic fate of therapeutic monoclonal antibodies, ASGPR-targeted LYTAC therapeutics and LYTAC targets in immortalized cell lines and in primary human hepatocytes.
An efflux pump family distributed across plant commensal bacteria conditions host- and organ-specific detoxification of a host-specific glucosinolate
Authors: Russ Dor , Fitzpatrick Connor R. , Saha Chinmay , Law Theresa F. , Jones Corbin D. , Kliebenstein Daniel J. , Dangl Jeffery L. 
Published: October 2024

AbstractIn nature, plants recruit a diverse microbial community, the plant microbiome, that is distinct from the surrounding soil community1–4. To understand the forces that shape the plant microbiome we need to characterize the microbial traits that contribute to plant colonization. We used barcoded mutant libraries to identify bacterial genes that contribute to the colonization of a monocot and a eudicot host5–7. We show that plant colonization is influenced by dozens of genes. While some of these colonization genes were shared between the two host plant species, most were highly specific, benefiting the colonization of a single host and organ. We characterized an efflux pump that specifically contributes to Arabidopsis shoot colonization. This efflux pump is prevalent across Pseudomonadota genomes, yet benefits the bacterial association with only a small subset ofArabidopsis thalianaaccessions. Leveraging genomic diversity withinArabidopsis thaliana, we confirmed that specific glucosinolate breakdown products are detoxified by this family of efflux pumps. The broad prevalence of this efflux pump family suggests that its members contribute to protection of commensal bacteria from collateral damage of plant glucosinolate-based defense responses to herbivores and necrotrophic pathogens.
Tissue-Like Multicellular Development Triggered by Mechanical Compression in Archaea
Authors: Rados Theopi , Leland Olivia S. , Escudeiro Pedro , Mallon John , Andre Katherine , Caspy Ido , von Kügelgen Andriko , Stolovicki Elad , Nguyen Sinead , Patop Inés Lucía , Rangel Thiberio , Kadener Sebastian , Renner Lars D. , Thiel Vera , Soen Yoav , Bharat Tanmay A.M. , Alva Vikram , Bisson Alex 
Published: October 2024

AbstractThe advent of clonal multicellularity is a critical evolutionary milestone, seen often in eukaryotes, rarely in bacteria, and only once observed in archaea. We show that uniaxial compression induces clonal multicellularity in haloarchaea, forming tissue-like structures. These archaeal tissues are mechanically and molecularly distinct from their unicellular lifestyle, mimicking several eukaryotic features. Archaeal tissues undergo a multinucleate stage followed by tubulin-independent cellularization, orchestrated by active membrane tension at a critical cell size. After cellularization, tissue junction elasticity become akin to animal tissues and give rise to two cell types—radial peribasal and central apicobasal cells— with distinct actin and protein glycosylation polarity patterns. Our findings highlight the potential convergent evolution of a biophysical mechanism in the emergence of multicellular systems across domains of life.
Shock wave propagation in a real gas with or without gravitational field in the presence of magnetic field and monochromatic radiation via group invariance method
Authors: Nath G. , Upadhyay P. 
Published: October 2024

PurposeThis article aims to find the similarity solutions for the one-dimensional motion of spherical symmetric shock wave in non-ideal gas influenced by the azimuthal magnetic field and monochromatic radiation in the presence or absence of gravitational field. This paper also aims to study the effects of physical parameters on the strength of shock wave, and on the flow variables in the flow-field region behind the shock front.Design/methodology/approachThe Roche model is used to describe the gravitational field effects due to a massive nucleus at the point of symmetry. To derive the similarity solutions, the Lie group symmetry method has been used. Also, the numerical solutions to the present problem are obtained by using Rung–Kutta method of the fourth order with the use of Mathematica software. The effects of variation in the parameter of non-idealness of the gas, the gravitation parameter, the strength of the ambient magnetic field and the adiabatic index of the gas on the shock wave, and on the flow variables is discussed. A comparative study between with and without gravitational field is also, made.FindingsFor different choices of the arbitrary constants that appeared in the solution of infinitesimal generators, we have obtained seven distinct cases of similarity solutions. In the absence of the gravitational field, the similarity solution exists to the power and exponential law shock paths, but in the presence of gravitational field, the similarity solution exists to the power law shock path case only. In the absence of gravitational field, the shock strength is enhanced in the exponential law shock path case in comparison to the power law shock path case. It is found that the shock wave decays with an increase in the value of the adiabatic exponent, the strength of magnetic field, non-idealness of the gas or gravitational parameter.Research limitations/implicationsThe consideration of medium under the influence of gravitational field due to a heavy nucleus at the center and presence of magnetic field decrease the shock strength. This result may be helpful in designing space vehicle and jet engine.Practical implicationsThe result of the present study may be used in the analysis of data from the measurements by space craft in the solar wind and in neighborhood of the Earth’s magnetosphere.Social implicationsThe obtained results may be used for mankind.Originality/valueThe study of spherical shock wave propagation influenced by monochromatic radiation and azimuthal magnetic field in a non-ideal gas with or without gravitational field has yet to be discussed by any authors by using the Lie group symmetry method. In this article, we have discussed all possible cases of similarity solutions using the Lie group symmetry method, which is not studied by anyone as known to us.
Adoption challenges of building information modelling (BIM) and off-site construction (OSC) in healthcare construction: are they fellow sufferers?
Authors: Tan Tan , Mills Grant , Ma Xiaolin , Papadonikolaki Eleni 
Published: October 2024

PurposeOff-Site Construction (OSC) has received much government and public attention during and after COVID. Building Information Modelling (BIM) is an initiative discussed widely to promote OSC implementation. Although many policy promotions have been published, there are many challenges to implementing BIM and OSC in real life and questions of whether they really offer value to healthcare design professionals. This research aims to investigate BIM and OSC to understand their commonalities and differences of challenges by collecting empirical evidence from China’s healthcare construction.Design/methodology/approachThis exploratory research adopted a mixed method with a questionnaire survey and interviews. A total of 261 questionnaires were received (with 183 valid), followed by 31 semi-structured interviews.FindingsThis research reveals that although both OSC and BIM face similar adoption challenges and suspicious attitudes in real-life projects, their challenges’ connotations and reasons are different. OSC faces scepticism for its customisation costs and technical constraints, while BIM is seen as limited in utility and complex to integrate. Highlighting these as socio-technical challenges, the research advocates for an integrated framework to effectively implement OSC and BIM, addressing both technical and collaborative needs in healthcare construction.Originality/valueThis research examines OSC and BIM within the context of healthcare construction, a focus that is relatively underexplored. The research provides a juxtaposition of the perceived and practical challenges of adopting these technologies, revealing a gap between the industry’s expectations and the current capabilities of OSC and BIM, thereby contributing to the development of modern methods of design in healthcare.