bluuebunny/crossref_metadata_2025_split huggingface dataset:
Small transcriptional differences among cell clones lead to distinct NF-κB dynamics
Authors: Kizilirmak Cise , Monteleone Emanuele , García-Manteiga José Manuel , Brambilla Francesca , Agresti Alessandra , Bianchi Marco E. , Zambrano Samuel 
Published: December 2021

ABSTRACTTranscription factor dynamics is fundamental to determine the activation of accurate transcriptional programs and yet is heterogeneous at single-cell level, even between very similar cells. We asked how such heterogeneity emerges for the nuclear factor κB (NF-κB), whose dynamics have been reported to cover a wide spectrum of behaviors, including persistent, oscillatory and weak activation. We found that clonal populations of immortalized fibroblasts derived from a single mouse embryo display robustly distinct dynamics upon tumor necrosis factor α (TNF-α) stimulation, which give rise to differences in the transcription of NF-κB targets. Notably, standard transcriptomic analyses indicate that the clones differ mostly in transcriptional programs related with development, but not in TNF-α signaling. However, by combining transcriptomics data and simulations we show how the expression levels of genes coding for proteins of the signaling cascade determine the differences in early NF-κB activation; differences in the expression of IκBα determine differences in its persistence and oscillatory behavior. The same analysis predicts inter-clonal differences in the NF-κB response to IL-1β. We propose that small (less than twofold) differences at transcript level can lead to distinct transcription factor dynamics in cells within homogeneous cell populations, and all the more so among different cell types.
Altricial bird early-stage embryos express the molecular ‘machinery’ to respond to maternal thyroid hormone cues
Authors: Ruuskanen Suvi , Hukkanen Mikaela , Garcin Natacha , Cossin-Sevrin Nina , Hsu Bin-Yan , Stier Antoine 
Published: December 2021

AbstractMaternal hormones, such as thyroid hormones transferred to embryos and eggs, are key signalling pathways to mediate maternal effects. To be able to respond to maternal cues, embryos must express key molecular ‘machinery’ of the hormone pathways, such as enzymes and receptors. While altricial birds begin thyroid hormone (TH) production only at/after hatching, experimental evidence suggests that their phenotype can be influenced by maternal THs deposited in the egg. However, it is not understood, how and when altricial birds express genes in the TH-pathway. For the first time, we measured the expression of key TH-pathway genes in altricial embryos, using two common altricial ecological model species (pied flycatcher, Ficedula hypoleuca and blue tit Cyanistes caeruleus). Deiodinase DIO1 gene expression could not be reliably confirmed in either species, but deiodinase enzyme DIO2 and DIO3 genes were expressed in both species. Given that DIO2 coverts T4 to biologically active T3, and DIO3 mostly T3 to inactive forms of thyroid hormones, our results suggest that embryos may modulate maternal signals. Thyroid hormone receptor (THRA and THRB) and monocarboxyl membrane transporter gene (SLC15A2) were also expressed, enabling TH-responses. Our results suggest that early altricial embryos may be able to respond and potentially modulate maternal signals conveyed by thyroid hormones.
Large Scale Discovery of Microbial Fibrillar Adhesins and Identification of Novel Members of Adhesive Domain Families
Authors: Monzon Vivian , Bateman Alex 
Published: December 2021

AbstractFibrillar adhesins are bacterial cell surface proteins that mediate interactions with the environment including host cells during colonisation or other bacteria during biofilm formation. These proteins are characterised by a stalk that projects the adhesive domain closer to the binding target. Fibrillar adhesins evolve quickly and thus can be difficult to computationally identify, yet they represent an important component for understanding bacterial host interactions.To detect novel fibrillar adhesins we developed a random forest prediction approach based on common characteristics we identified for this protein class. We applied this approach to Firmicute and Actinobacterial proteomes, yielding over 6,500 confidently predicted fibrillar adhesins. To verify the approach we investigated predicted fibrillar adhesins that lacked a known adhesive domain. Based on these proteins, we identified 24 sequence clusters representing potential novel members of adhesive domain families. We used AlphaFold to verify that 15 clusters showed structural similarity to known adhesive domains such as the TED domain.Overall our study has made a significant contribution to the number of known fibrillar adhesins and has enabled us to identify novel members of adhesive domain families involved in the bacterial pathogenesis.ImportanceFibrillar adhesins are a class of bacterial cell surface proteins that enable bacteria to interact with their environment. We developed a Machine Learning approach to identify fibrillar adhesins and applied this classification approach on the Firmicutes and Actinobacteria Reference Proteomes. This method allowed us to detect a high number of novel fibrillar adhesins, and also novel members of adhesive domain families. To confirm our predictions of these potential adhesin protein domains, we predicted their structure using the AlphaFold tool.
HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform
Authors: Friedl Marie-Sophie , Djakovic Lara , Kluge Michael , Hennig Thomas , Whisnant Adam W. , Backes Simone , Dölken Lars , Friedel Caroline C. 
Published: December 2021

AbstractThe herpes simplex virus 1 (HSV-1) virion host shut-off (vhs) protein cleaves both cellular and viral mRNAs by a translation-initiation-dependent mechanism. Here, we show thatvhs-mediated degradation of mRNAs leads to an accumulation of circular RNAs (circRNAs) relative to linear mRNAs during HSV-1 infection. Strikingly, we found that circular splicing of the long isoform (NEAT1_2) of the nuclear paraspeckle assembly transcript 1 (NEAT1) was massively induced during HSV-1 infection. In contrast to other circRNAs, induction of the NEAT1_2 circRNA was independent ofvhsand occurred while NEAT1_2 was still bound to the chromatin. This was associated with induction of linear splicing of NEAT1_2 both within and downstream of the circRNA. NEAT1_2 splicing was absent in uninfected cells but can be induced by ectopic co-expression of the HSV-1 immediate-early proteins ICP22 and ICP27. Interestingly, NEAT1_2 circular and linear splicing was also up-regulated in influenza A virus (IAV) infection but absent in stress conditions, which disrupt transcription termination similar to but not by the same mechanisms as HSV-1 and IAV infection. Large-scale analysis of published RNA-seq data uncovered induction of NEAT1_2 splicing in cancer cells upon inhibition or knockdown of cyclin-dependent kinase 7 (CDK7) or the MED1 subunit of the Mediator complex phosphorylated by CDK7. Finally, CDK7 inhibition also disrupted transcription termination, highlighting a possible link between disruption of transcription termination and NEAT1_2 splicing.
Structural insights into distinct signaling profiles of the μOR activated by diverse agonists
Authors: Qu Qianhui , Huang Weijiao , Aydin Deniz , Paggi Joseph M. , Seven Alpay B. , Wang Haoqing , Chakraborty Soumen , Che Tao , DiBerto Jeffrey F. , Robertson Michael J. , Inoue Asuka , Roth Bryan L. , Majumdar Susruta , Dror Ron O. , Kobilka Brian K. , Skiniotis Georgios 
Published: December 2021

AbstractDrugs targeting the G protein-coupled μ-opioid receptor (μOR) are the most effective analgesics available but are also associated with fatal respiratory depression. While some partial opioid agonists appear to be safer than full agonists, the signaling pathways responsible for respiratory depression have yet to be elucidated. Here we investigated the structural and mechanistic basis of action of lofentanil (LFT) and mitragynine pseudoindoxyl (MP), two μOR agonists with different safety profiles. LFT, one of the most potent and lethal opioids, and MP, a derivative from the kratom plant with reduced respiratory depression in animal studies at equianalgesic doses, exhibited markedly different signaling efficacy profiles for G protein subtype activation and recruitment of β-arrestins. Cryo-EM structures of the μOR-Gi1 complex with MP (2.5Å) and LFT (3.2Å) revealed that the two ligands engage distinct sub-pockets, and molecular dynamics (MD) simulations showed additional differences in the binding site that propagate to the intracellular side of the receptor where G proteins and β-arrestins bind. While MP favors the precise G protein-bound active state observed in the cryo-EM structures, LFT favors a distinct active state. These results highlight how drugs engaging different parts of the μOR orthosteric pocket can lead to distinct signaling outcomes.
Rapid and robust directed differentiation of mouse epiblast stem cells into definitive endoderm and forebrain organoids
Authors: Medina-Cano Daniel , Corrigan Emily K. , Glenn Rachel A. , Islam Mohammed T. , Lin Yuan , Kim Juliet , Cho Hyunwoo , Vierbuchen Thomas 
Published: December 2021

ABSTRACTDirected differentiation of pluripotent stem cells (PSCs) is a powerful model system for deconstructing embryonic development. Although mice are the most advanced mammalian model system for genetic studies of embryonic development, state-of-the-art protocols for directed differentiation of mouse PSCs into defined lineages require additional steps and generate target cell types with lower purity than analogous protocols for human PSCs, limiting their application as models for mechanistic studies of development. Here, we examine the potential of mouse epiblast stem cells (EpiSCs) cultured in media containing Wnt pathway inhibitors as a starting point for directed differentiation. As a proof-of-concept, we focused our efforts on two specific cell/tissue types that have proven difficult to generate efficiently and reproducibly from mouse embryonic stem cells: definitive endoderm and neural organoids. We present new protocols for rapid generation of nearly pure definitive endoderm and forebrain-patterned neural organoids that model the development of prethalamic and hippocampal neurons. These differentiation models present new possibilities for combining mouse genetic tools within vitrodifferentiation to characterize molecular and cellular mechanisms of embryonic development.SUMMARY STATEMENTNew optimized protocols for directed differentiation of mouse epiblast stem cells into definitive endoderm and forebrain-patterned organoids.
Spatiotemporal transcriptomics reveals pathogenesis of viral myocarditis
Authors: Mantri Madhav , Hinchman Meleana M. , McKellar David W. , Wang Michael F. Z. , Cross Shaun T. , Parker John S. L. , De Vlaminck Iwijn 
Published: December 2021

ABSTRACTA significant fraction of sudden death in children and young adults is due to myocarditis, an inflammatory disease of the heart, most often caused by viral infection. Here we used integrated single-cell and spatial transcriptomics to create a high-resolution, spatially resolved map of reovirus-induced myocarditis in neonatal murine hearts. We assayed hearts collected at three timepoints after reovirus infection and studied the temporal, spatial, and cellular heterogeneity of host-virus interactions. We further assayed the intestine, the primary site of reovirus infection to establish a full chronology of molecular events that ultimately lead to myocarditis. We implemented targeted enrichment of viral transcripts to establish the cellular targets of the virus in the intestine and the heart. Our data give insight into the cell-type specificity of innate immune responses, and into the transcriptional states of inflamed cardiac cells in reovirus-infected heart. We find that inflamed endothelial cells recruit cytotoxic T cells and undergo pyroptosis in the myocarditic tissue. Analyses of spatially restricted gene expression in myocarditic regions and the border zone around those regions identified immune-mediated cell-type specific injury and stress responses. Overall, we observe a dynamic and complex network of cellular phenotypes and cell-cell interactions associated with viral myocarditis.
Neural Oscillatory Characteristics of Feedback Associated Activity in Globus Pallidus Interna
Authors: Choubdar Hadi , Mahdavi Mahdi , Rostami Zahra , Zabeh Erfan , Gillies Martin J. , Green Alexander L. , Aziz Tipu Z. , Lashgari Reza 
Published: December 2021

AbstractNeural oscillatory activities in basal ganglia have prominent roles in cognitive processes on local and global scales. However, the characteristics of high frequency oscillatory activities during cognitive tasks have not been extensively explored in human Globus Pallidus internus (GPi). This study aimed to investigate amplitude and interhemispheric coupling of bilateral GPi high gamma bursts in dystonia and Parkinson’s Disease (PD) patients, in on and off medication states, after feedback during the Intra-Extra-Dimension shift (IED) task. Bilateral GPi Local Field Potentials (LFP) activity was recorded via externalized DBS electrodes during the IED task. Inter hemisphere phase synchrony was assessed using Inter-Site Phase Clustering (ISPC). Transient high gamma activity (∼100-150Hz) was observed immediately after feedback in the dystonia patient. Moreover, these bursts were phase synchronous between left and right GPis with an antiphase clustering of phase differences. In contrast, no synchronous high gamma activity was detected in the PD patient with or without dopamine administration. The off-med PD patient displayed enhanced low frequency clusters ameliorated by medication in the on-med state. Furthermore, an increased low frequency activity was observed after feedback of incorrect trials in both disease states. The current study provides a rare report of antiphase homotopic synchrony in human GPi, potentially related to incorporating and processing feedback information. The absence of these activities in off and on-med PD indicates the potential presence of impaired medication independent circuits related to feedback processing. Together, these findings are helpful in pointing to the potential role of GPi’s synchronized high frequency activity in cognitive tasks and feedback information processing.
High-Speed Low-Light In Vivo Two-Photon Voltage Imaging of Large Neuronal Populations
Authors: Platisa Jelena , Ye Xin , Ahrens Allison M. , Liu Chang , Chen Ichun Anderson , Davison Ian G. , Tian Lei , Pieribone Vincent A. , Chen Jerry L. 
Published: December 2021

SUMMARYMonitoring spiking activity across large neuronal populations at behaviorally relevant timescales is critical for understanding neural circuit function. Unlike calcium imaging, voltage imaging requires kilohertz sampling rates which reduces fluorescence detection to near shot noise levels. High-photon flux excitation can overcome photon-limited shot noise but photo-bleaching and photo-damage restricts the number and duration of simultaneously imaged neurons. We investigated an alternative approach aimed at low two-photon flux, voltage imaging below the shot noise limit. This framework involved developing: a positive-going voltage indicator with improved spike detection (SpikeyGi); an ultra-fast two-photon microscope for kilohertz frame-rate imaging across a 0.4×0.4mm2 field of view, and; a self-supervised denoising algorithm (DeepVID) for inferring fluorescence from shot-noise limited signals. Through these combined advances, we achieved simultaneous high-speed, deep-tissue imaging of more than one hundred densely-labeled neurons over one hour in awake behaving mice. This demonstrates a scalable approach for voltage imaging across increasing neuronal populations.
Revealing a hidden intermediate of rotatory catalysis with X-ray crystallography and Molecular simulations
Authors: Shekhar Mrinal , Gupta Chitrak , Suzuki Kano , Murata Takeshi , Singharoy Abhishek 
Published: December 2021

ABSTRACTThe mechanism of rotatory catalysis in ATP-hydrolyzing molecular motors remain an unresolved puzzle in biological energy transfer. Notwith standing the wealth of available biochemical and structural information inferred from years of experiments, knowledge on how the coupling between the chemical and mechanical steps within motors enforces directional rotatory movements remains fragmentary. Even more contentious is to pinpoint the rate-limiting step of a multi-step rotation process. Here, using Vacuolar or V1-type hexameric ATPase as an exemplary rotational motor, we present a model of the complete 4-step conformational cycle involved in rotatory catalysis. First, using X-ray crystallography a new intermediate or ‘dwell’ is identified, which enables the release of an inorganic phosphate (or Pi) after ATP hydrolysis. Using molecular dynamics simulations, this new dwell is placed in a sequence with three other crystal structures to derive a putative cyclic rotation path. Free-energy simulations are employed to estimate the rate of the hexameric protein transformations, and delineate allosteric effects that allow new reactant ATP entry only after hydrolysis product exit. An analysis of transfer entropy brings to light how the sidechain-level interactions transcend into larger-scale reorganizations, highlighting the role of the ubiquitous arginine-finger residues in coupling chemical and mechanical information. Inspection of all known rates encompassing the 4-step rotation mechanism implicates overcoming of the ADP interactions with V1-ATPase to be the rate-limiting step of motor action.
Interpretable and Generalizable Attention-Based Model for Predicting Drug-Target Interaction Using 3D Structure of Protein Binding Sites: SARS-CoV-2 Case Study and in-Lab Validation
Authors: Yazdani-Jahromi Mehdi , Yousefi Niloofar , Tayebi Aida , Garibay Ozlem Ozmen , Seal Sudipta , Kolanthai Elayaraja , Neal Craig J. 
Published: December 2021

AbstractIn this study, we introduce and implement an interpretable graph-based deep learning prediction model, which utilizes protein binding sites along with self-attention to learn which protein binding sites interact with a given ligand. Our proposed model enables interpretability by identifying the protein binding sites that contribute the most towards the Drug-Target Interaction. Results on three benchmark datasets show improved performance compared to previous graph-based models. More significantly, unlike previous studies our model performance remains close to the optimal performance when tested with new proteins (ie., high generalizablity). Through multidisciplinary collaboration, we further experimentally evaluate the practical potential of our proposed approach. To achieve this, we first computationally predict binding interaction of some candidate compounds with a target protein, then experimentally validate the binding interactions for these pairs in the laboratory. The high agreement between the computationally-predicted and experimentally-observed (measured) DTIs illustrates the potential of our method as an effective pre-screening tool in drug re-purposing applications.
Differential Histone-DNA Interactions Dictate Nucleosome Recognition of the Pioneer Transcription Factor Sox
Authors: Ozden Burcu , Boopathi Ramachandran , Barlas Ayşe Berçin , Lone Imtiaz N. , Bednar Jan , Petosa Carlo , Kale Seyit , Hamiche Ali , Angelov Dimitar , Dimitrov Stefan , Karaca Ezgi 
Published: December 2021

ABSTRACTPioneer transcription factors (PTFs) have the remarkable ability to directly bind to chromatin for stimulating vital cellular processes. In this work, we dissect the universal binding mode of Sox PTF by combining extensive molecular simulations and DNA footprinting techniques. As a result, we show that when Sox consensus DNA is located at the solvent-facing DNA strand, Sox binds to the compact nucleosome without imposing any significant conformational changes. We also reveal that the basespecific Sox:DNA interactions (base reading) and the Sox-induced DNA changes (shape reading) are concurrently required for the sequence-specific DNA recognition. Among different nucleosomal positions, such a specific reading mechanism is satisfied solely at superhelical location 2 (SHL2). While SHL2 acts transparently to Sox binding, SHL4 permits only shape reading, and SHL0 (dyad) allows no reading mechanism. These findings demonstrate for the first time that Sox-based nucleosome recognition is essentially guided by the distinct histone-DNA interactions, permitting varying degrees of DNA flexibility.
A high-throughput amplicon screen for somatic UBA1 variants in Cytopenic and Giant Cell Arteritis cohorts
Authors: Poulter James A. , Khan Alesia , Martin Stephen , Grey Mark , Andjelic Bosko , Nga Emma , Triantafyllopoulou Diana I.D. , Evans Paul , Sorensen Louise , Mackie Sarah L. ,   UKGCA Consortium, Morgan Ann W. , Cargo Catherine , Savic Sinisa 
Published: December 2021

AbstractSomatic mutations in the gene encoding the major E1 ubiquitin ligase, UBA1, were recently identified as a cause of VEXAS, a late-onset acquired auto-inflammatory syndrome. Differential diagnoses for patients subsequently found to have VEXAS include relapsing polychondritis, Sweet’s syndrome, myelodysplastic syndrome (MDS), giant cell arteritis (GCA) and undifferentiated systemic autoinflammatory disease (uSAID). We therefore sought to screen DNA from individuals with a non-diagnostic cytopenia or GCA, for known VEXAS-associated mutations. To this end, we developed a multiplexed UBA1 amplicon sequencing assay, allowing quick screening of large cohorts while also providing sufficient sequencing depth to identify somatic mutations to an allele frequency <1%. Using this assay, we screened genomic DNA from 612 males diagnosed with GCA, and bone marrow DNA from 1,055 cases with an undiagnosed cytopenia. No GCA cases were found to have UBA1 mutations, however 4 different mutations in the cytopenic cohort were identified in 7 individuals. Furthermore, we describe a female case identified in the screen with a UBA1 mutation and all VEXAS-associated phenotypes, but without Monosomy X. Our study suggests that, despite the overlap in clinical features, VEXAS is rarely misdiagnosed as GCA, but identified in 1.0% of males with an undiagnosed cytopenia. The identification of a UBA1 variant in a female case adds further evidence that VEXAS should not be ruled out as a differential diagnosis in females with VEXAS-like symptoms.Key points-Mutations in UBA1 exon 3 have been associated with VEXAS syndrome-UBA1 exon 3 was screened in 1650 patients with cytopenia or GCA by amplicon sequencing.-6 males were identified from the non-diagnostic cytopenia cohort (1.0%) with UBA1 mutations.-A female with a somatic UBA1 mutation was identified without Monosomy X