bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dysregulation of thede novoproteome accompanies pathology progression in the APP/PS1 mouse model of Alzheimer’s disease
Authors: Elder Megan K. , Erdjument-Bromage Hediye , Oliveira Mauricio M. , Mamcarz Maggie , Neubert Thomas A. , Klann Eric 
Published: August 2020

AbstractAlzheimer’s disease (AD) is an age-related neurodegenerative disorder, but neuropathological changes in AD begin years before memory impairment. Investigation of the early molecular abnormalities in AD might offer innovative opportunities to target memory impairment prior to its onset. Decreased protein synthesis plays a fundamental role in AD, yet the consequences for cellular function remain unknown. We hypothesize that alterations in thede novoproteome drive early metabolic changes in the hippocampus that persist throughout AD progression. Using a combinatorial amino acid tagging approach to selectively label newly synthesized proteins, we found that thede novoproteome is disturbed in APP/PS1 mice prior to pathology development, affecting the synthesis of multiple components of synaptic, lysosomal and mitochondrial pathways. Furthermore, the synthesis of large clusters of ribosomal subunits were affected throughout neuropathology development in these mice. Our data suggest that large-scale changes in protein synthesis could underlie cellular dysfunction in AD.
Routine sub-2.5 Å cryo-EM structure determination of B-family G protein-coupled receptors
Authors: Danev Radostin , Belousoff Matthew , Liang Yi-Lynn , Zhang Xin , Wootten Denise , Sexton Patrick M. 
Published: August 2020

AbstractCryo-electron microscopy (cryo-EM) experienced game-changing hardware and software advances about a decade ago. Since then, there have been gradual and steady improvements in experimental and data analysis methods. Nonetheless, structural analysis of nonsymmetric membrane proteins, such as G protein-coupled receptors (GPCRs), remains challenging. Their relatively low molecular weight and obstruction by the micelle/nanodisc result in marginal signal levels, which combined with the intrinsic flexibility of such complexes creates difficult structural study scenarios. Pushing the performance limits of cryo-EM requires careful optimization of all experimental aspects. To this end, it is necessary to build quantitative knowledge of the effect each parameter has on the outcome. Here, we present in-depth analysis of the influence of the main cryo-EM experimental factors on the performance for GPCR structure determination. We used a tandem experiment approach that combined real-world structural studies with parameter testing. We quantified the effects of using a Volta phase plate, zero-loss energy filtering, objective lens aperture, defocus magnitude, total exposure, and grid type. Through such systematic optimization of the experimental conditions, it has been possible to routinely determine class B1 GPCR structures at resolutions better than 2.5 Å. The improved fidelity of such maps helps to build higher confidence atomic models and will be crucial for the future expansion of cryo-EM into the structure-based drug design domain. The optimization guidelines drafted here are not limited to GPCRs and can be applied directly for the study of other challenging membrane protein targets.
Evolution of dynamical networks enhances catalysis in a designer enzyme
Authors: Bunzel H. Adrian , Anderson J. L. Ross , Hilvert Donald , Arcus Vickery L. , van der Kamp Marc W. , Mulholland Adrian J. 
Published: August 2020

AbstractActivation heat capacity is emerging as a crucial factor in enzyme thermoadaptation, as shown by non-Arrhenius behaviour of many natural enzymes1,2. However, its physical origin and relationship to evolution of catalytic activity remain uncertain. Here, we show that directed evolution of a computationally designed Kemp eliminase introduces dynamical changes that give rise to an activation heat capacity absent in the original design3. Extensive molecular dynamics simulations show that evolution results in the closure of solvent exposed loops and better packing of the active site with transition state stabilising residues. Remarkably, these changes give rise to a correlated dynamical network involving the transition state and large parts of the protein. This network tightens the transition state ensemble, which induces an activation heat capacity and thereby nonlinearity in the temperature dependence. Our results have implications for understanding enzyme evolution (e.g. in explaining the role of distal mutations and evolutionary tuning of dynamical responses) and suggest that integrating dynamics with design and evolution will accelerate the development of efficient novel enzymes.
Automated analysis of rabbit knee calcified cartilage morphology using micro-computed tomography and deep learning segmentation
Authors: Rytky Santeri J. O. , Huang Lingwei , Tanska Petri , Tiulpin Aleksei , Panfilov Egor , Herzog Walter , Korhonen Rami K. , Saarakkala Simo , Finnilä Mikko A. J. 
Published: August 2020

AbstractPurposeOnly little is known how calcified cartilage (CC) structure changes during exercise, aging and disease. CC thickness (CC.Th) can be analyzed using conventional histological sections. Micro-computed tomography (μCT) allows for three-dimensional (3D) imaging of mineralized tissues, however, the segmentation between bone and CC is challenging. Here, we present state-of-the-art deep learning segmentation for μCT images to enable assessment of CC morphology.MethodsSixteen knees from twelve New Zealand White rabbits were dissected into osteochondral samples from six anatomical regions: lateral and medial femoral condyles, lateral and medial tibial plateaus, femoral groove and patella (n = 96). Samples were imaged with μCT and processed for conventional histology. Manually segmented CC from the histology and reconstructed μCT images was used as the gold standard to train segmentation models with different encoder-decoder architectures. The models with the greatest out-of-fold evaluation Dice score were used for automated CC.Th analysis. Subsequently, the automated CC.Th analysis was compared across a total of 24 regions, co-registered between the imaging modalities, using Pearson correlation and Bland-Altman analyses. Finally, the anatomical variation in CC.Th was assessed via a Linear Mixed Model analysis.ResultsThe best segmentation models yielded average Dice scores of 0.891 and 0.807 for histology and μCT segmentation, respectively. The correlation between the co-registered regions across the modalities was strong (r = 0.897). The Bland-Altman analysis yielded a bias of 21.9 μm and a standard deviation of 21.5 μm between the methods. Finally, both methods could separate the CC morphology between the patella, femoral, and tibial regions (p < 0.001).ConclusionThe presented method allows for ex vivo 3D assessment of CC.Th in an automated and non-destructive manner. We demonstrated its utility by quantifying CC.Th in different anatomical regions. CC.Th was the thickest in the patella and the thinnest in the tibial plateau.Graphical abstractWe present a μCT-based method with deep learning segmentation for analyzing calcified cartilage thickness (CC.Th). The method is compared throughout the study against conventional histology. The comparison against co-registered regions yielded a strong Pearson correlation (r = 0.90). Both methods were able to separate the CC.Th properties between tibia, femur, and patella.
Recurrent human papillomavirus-related head and neck cancer undergoes metabolic re-programming and is driven by oxidative phosphorylation
Authors: Vyas Avani , Harbison R. Alex , Faden Daniel , Kubik Mark , Palmer Drake , Zhang Qing , Osmanbeyoglu Hatice U. , Méndez Eduardo , Duvvuri Umamaheswar 
Published: August 2020

AbstractHuman papillomavirus (HPV) infection drives the development of some head and neck cancer squamous cell carcinomas (HNSC). This disease is rapidly increasing in incidence worldwide. Although these tumors are sensitive to treatment, ~10% of patients fail therapy. However, the mechanisms that underlie treatment failure remain unclear. Here, we show that the oxidative phosphorylation (OXPHOS) pathway is enriched in recurrent HPV-associated HNSC and may contribute to treatment failure. Nrf2-enriched HNSC samples from the Cancer Genome Atlas with enrichment in OXPHOS, fatty acid metabolism, Myc, Mtor, ROS, and glycolytic signaling networks exhibited worse survival. HPV-positive HNSC cells demonstrated sensitivity to the OXPHOS inhibitor, IACS-010759, in a Nrf2-dependent manner. Further, using murine xenograft models, we identified Nrf2 as a driver of tumor growth. Mechanistically, Nrf2 drives ROS and mitochondrial respiration, and Nrf2 is a critical regulator of redox homeostasis that can be crippled by disruption of OXPHOS. Nrf2 also mediated cisplatin sensitivity in endogenously overexpressing primary HPV-related HNSC cells. Cisplatin treatment demonstrated Nrf2-dependent synergy with OXPHOS inhibition. These results unveil a paradigm shifting translational target harnessing Nrf2-mediated metabolic reprogramming in HPV-related HNSC.
Hemodynamic responses link individual differences in informational masking to the vicinity of superior temporal gyrus
Authors: Zhang Min , Alamatsaz Nima , Ihlefeld Antje 
Published: August 2020

AbstractSuppressing unwanted background sound is crucial for aural communication. A particularly disruptive type of background sound, informational masking (IM), often interferes in social settings. However, IM mechanisms are incompletely understood. At present, IM is identified operationally: when a target should be audible, based on suprathreshold target/masker energy ratios, yet cannot be heard because perceptually similar background sound interferes. Here, functional near infrared spectroscopy recordings show that task-evoked blood oxygenation changes near the superior temporal gyrus (STG) covary with behavioral speech detection performance for high-IM but not low-IM background sound, suggesting that the STG is part of an IM-dependent network. Moreover, listeners who are more vulnerable to IM show increased hemodynamic recruitment near STG. In contrast, task-evoked responses near another auditory region of cortex, the caudal inferior frontal sulcus (cIFS), do not predict behavioral sensitivity, suggesting that the cIFS belongs to an IM-independent network. Results are consistent with the idea that cortical gating shapes individual vulnerability to IM.
Extensive age-dependent loss of antibody diversity in naturally short-lived turquoise killifish
Authors: Bradshaw William J , Poeschla Michael , Placzek Aleksandra , Kean Samuel , Valenzano Dario Riccardo 
Published: August 2020

AbstractAging individuals exhibit a pervasive decline in adaptive immune function, with important implications for health and lifespan. Previous studies have found a pervasive loss of immune-repertoire diversity in human peripheral blood during aging; however, little is known about repertoire aging in other immune compartments, or in species other than humans. Here, we perform the first study of immune-repertoire aging in an emerging model of vertebrate aging, the African turquoise killifish (Nothobranchius furzeri). Despite their extremely short lifespans, these killifish exhibit complex and individualized heavy-chain repertoires, with a generative process capable of producing millions of distinct productive sequences. Whole-body killifish repertoires decline rapidly in within-individual diversity with age, while between-individual variability increases. Large, expanded B-cell clones exhibit far greater diversity loss with age than small clones, suggesting important differences in how age affects different B cell populations. The immune repertoires of isolated intestinal samples exhibit especially dramatic age-related diversity loss, related to an elevated prevalence of expanded clones. Lower intestinal repertoire diversity was also associated with transcriptomic signatures of reduced B-cell activity, supporting a functional role for diversity changes in killifish immunosenescence. Our results highlight important differences in systemic vs. organ-specific aging dynamics in the adaptive immune system.Significance StatementImmune function declines dramatically with age, with important consequences for health in the elderly. Changes in the diversity and composition of the body’s antibody repertoire are an important contributor to this age-related dysfunction. However, studying repertoire aging at high spatial and temporal resolution is hampered by the long lifespans of humans and other vertebrate models, and the difficulty of acquiring human samples other than peripheral blood. Here, we characterize the aging of the antibody repertoire in a naturally short-lived vertebrate, the turquoise killifish(Nothobranchius furzeri).We found that aging killifish undergo rapid loss of repertoire diversity in both whole-body and intestinal samples within four months of hatching. Loss of repertoire diversity in the gut was also associated with broader gene-expression changes indicative of reduced B-cell activity. As such, killifish represent a powerful laboratory model to study adaptive immune aging.
Combining repetition suppression and pattern analysis provides new insights into the role of M1 and parietal areas in skilled sequential actions
Authors: Berlot Eva , Popp Nicola J. , Grafton Scott T. , Diedrichsen Jörn 
Published: August 2020

AbstractHow does the brain change during learning? In functional magnetic resonance imaging studies, both multivariate pattern analysis and repetition suppression (RS) have been used to detect changes in neuronal representations. In the context of motor sequence learning, the two techniques have provided discrepant findings: pattern analysis showed that only premotor and parietal regions, but not primary motor cortex (M1), develop a representation of trained sequences. In contrast, RS suggested trained sequence representations in all these regions. Here we applied both analysis techniques to a 5-week finger sequence training study, in which participants executed each sequence twice before switching to a different sequence. Both RS and pattern analysis indicated learning-related changes for parietal areas, but only RS showed a difference between trained and untrained sequences in M1. A more fine-grained analysis, however, revealed that the RS effect in M1 reflects a fundamentally different process than in parietal areas. On the first execution, M1 represents especially the first finger of each sequence, likely reflecting preparatory processes. This effect dramatically reduces during the second execution. In contrast, parietal areas represent the identity of a sequence, and this representation stays relatively stable on the second execution. These results suggest that the RS effect does not reflect a trained sequence representation in M1, but rather a preparatory signal for movement initiation. More generally, our study demonstrates that across regions RS can reflect different representational changes in the neuronal population code, emphasizing the importance of combining pattern analysis and RS techniques.Significance statementPrevious studies using pattern analysis have suggested that primary motor cortex (M1) does not represent learnt sequential actions. However, a study using repetition suppression (RS) has reported M1 changes during motor sequence learning. Combining both techniques, we first replicate the discrepancy between them – with learning-related changes in M1 in RS, but not pattern dissimilarities. We further analysed the representational changes with repetition, and found that the RS effects differ across regions. M1’s activity represents the starting finger of the sequence, an effect that vanishes with repetition. In contrast, activity patterns in parietal areas exhibit sequence dependency, which persists with repetition. These results demonstrate the importance of combining RS and pattern analysis to understand the function of brain regions.
The receptor binding domain of SARS-CoV-2 spike is the key target of neutralizing antibody in human polyclonal sera
Authors: Steffen Tara L. , Stone E. Taylor , Hassert Mariah , Geerling Elizabeth , Grimberg Brian T. , Espino Ana M. , Pantoja Petraleigh , Climent Consuelo , Hoft Daniel F. , George Sarah L. , Sariol Carlos A. , Pinto Amelia K. , Brien James D. 
Published: August 2020

AbstractNatural infection of SARS-CoV-2 in humans leads to the development of a strong neutralizing antibody response, however the immunodominant targets of the polyclonal neutralizing antibody response are still unknown. Here, we functionally define the role SARS-CoV-2 spike plays as a target of the human neutralizing antibody response. In this study, we identify the spike protein subunits that contain antigenic determinants and examine the neutralization capacity of polyclonal sera from a cohort of patients that tested qRT-PCR-positive for SARS-CoV-2. Using an ELISA format, we assessed binding of human sera to spike subunit 1 (S1), spike subunit 2 (S2) and the receptor binding domain (RBD) of spike. To functionally identify the key target of neutralizing antibody, we depleted sera of subunit-specific antibodies to determine the contribution of these individual subunits to the antigen-specific neutralizing antibody response. We show that epitopes within RBD are the target of a majority of the neutralizing antibodies in the human polyclonal antibody response. These data provide critical information for vaccine development and development of sensitive and specific serological testing.
Regulation of extracellular matrix assembly and structure by hybrid M1/M2 macrophages
Authors: Witherel Claire E. , Sao Kimheak , Brisson Becky K. , Han Biao , Volk Susan W. , Petrie Ryan J. , Han Lin , Spiller Kara L. 
Published: August 2020

AbstractAberrant extracellular matrix (ECM) assembly surrounding implanted biomaterials is the hallmark of the foreign body response, in which implants become encapsulated in thick fibrous tissue that prevents their proper function. While macrophages are known regulators of fibroblast behavior, how their phenotype influences ECM assembly and the progression of the foreign body response is poorly understood. In this study, we usedin vitromodels with physiologically relevant macrophage phenotypes, as well as controlled release of macrophage-modulating cytokines from gelatin hydrogels implanted subcutaneouslyin vivoto investigate the role of macrophages in ECM assembly. Primary human macrophages were polarized to four distinct phenotypes, which have each been associated with fibrosis, including pro-inflammatory M1, pro-healing M2, and a hybrid M1/M2, generated by exposing macrophages to M1- and M2-promoting stimuli simultaneously. Additionally, macrophages were first polarized to M1 and then to M2 (M1→M2) to generate a phenotype typically observed during normal wound healing. Human dermal fibroblasts that were cultured in macrophage-conditioned media upregulated numerous genes involved in regulation of ECM assembly, especially in M2-conditioned media. Hybrid M1/M2 macrophage-conditioned media caused fibroblasts to produce a matrix with thicker and less aligned fibers, while M2 macrophage-conditioned media caused the formation of a more aligned matrix with thinner fibers. Gelatin methacrylate hydrogels containing interleukin-4 (IL4) and IL13-loaded poly(lactic-co-glycolic acid) (PLGA) microparticles were designed to promote the M2 phenotype in a murine subcutaneousin vivomodel. NanoString multiplex gene expression analysis of hydrogel explants showed that hydrogels with and without drug caused markers of both M1 and M2 phenotypes to be highly expressed, but the release of IL4+IL13 promoted upregulation of M2 markers and genes associated with regulation of ECM assembly, such asCol5a1andCol6a1. Biochemical analysis and second harmonic generation microscopy showed that the release of IL4+IL13 increased total sulfated glycosaminoglycan content and decreased fibril alignment, which is typically associated with less fibrotic tissue. Together, these results show that hybrid M1/M2 macrophages regulate ECM assembly, and that shifting the balance towards M2 may promote architectural and compositional changes in ECM with enhanced potential for downstream remodeling.
Cyclophilin A is a mitochondrial factor that forms antiapoptotic complexes with p23
Authors: Daneri-Becerra Cristina , Valeiras Brenda , Lagadari Mariana , Galigniana Mario D. 
Published: August 2020

AbstractCyclophilin A (CyPA) is an abundant and ubiquitously expressed protein belonging to the immunophilin family that has intrinsic peptidyl-prolyl-(cis/trans)-isomerase enzymatic activity. In addition to mediating the immunosuppressive effects of the drug cyclosporine A, CyPA is involved in multiple cellular processes such as protein folding, intracellular trafficking, signal transduction, and transcriptional regulation. Because CyPA is also a molecular chaperone, its expression is induced by several stressor agents and is a highly abundant protein in cancer cells. In this study, it is demonstrated that in several cell types and at least in murine liver, a significant pool of this immunophilin is primarily an intramitochondrial factor that migrates to the nucleus upon the onset of stress. It is also shown that CyPA has antiapoptotic action. Importantly, the capability of CyPA to form complexes with the small acidic cochaperone p23 is proven, this interaction being independent of the usual association of p23 with the heat-shock protein of 90-kDa, Hsp90. Furthermore, it is demonstrated that the CyPA•p23 complex enhances the antiapoptotic response of the cell, suggesting that both proteins form a functional unit whose high level of expression plays a significant role in cell survival.
Analysis of the oxidative stress regulon identifiessoxSas a genetic target for resistance reversal in multi-drug resistantKlebsiella pneumoniae
Authors: Anes João , Dever Katherine , Eshwar Athmanya , Nguyen Scott , Cao Yu , Sivasankaran Sathesh K , Sakalauskaitė Sandra , Lehner Angelika , Devineau Stéphanie , Daugelavičius Rimantas , Fanning Séamus , Srikumar Shabarinath 
Published: August 2020

AbstractIn bacteria, the defense system deployd to counter oxidative stress is orchestrated by three transcriptional factors – SoxS, SoxR, and OxyR. Although the regulon that these factors control is known in many bacteria, similar data is not available forKlebsiella pneumoniae. To address this data gap, oxidative stress was artificially induced inK. pneumoniaeMGH 78578 using paraquat and the corresponding oxidative stress regulon recorded using RNA-seq. ThesoxSgene was significantly induced during oxidative stress and a knock-out mutant was constructed, to explore its functionality. The wild-type and mutant were grown in the presence of paraquat and subjected to RNA-seq to elucidate thesoxSregulon inK. pneumoniaeMGH78578. Genes that are commonly regulated both in the oxidative stress regulon andsoxSregulon were identified and denoted as the ‘oxidative SoxS regulon’ – these included a stringent group of genes specifically regulated by SoxS. Efflux pump encoding genes such asacrAB-tolC, acrE, and global regulators such asmarRABwere identified as part of this regulon. Consequently, the isogenicsoxSmutant was found to exhibit a reduction in the minimum bactericidal concentration against tetracycline compared to that of the wild type. Impaired efflux activity, allowing tetracycline to be accumulated in the cytoplasm to bactericidal levels, was further evaluated using a tetraphenylphosphonium (TPP+) accumulation assay. ThesoxSmutant was also susceptible to tetracyclinein vivo, in a zebrafish embryo model. We conclude that thesoxSgene could be considered as a genetic target against which an inhibitor could be developed in the future and used in combinatorial therapy with tetracycline to combat infections associated with multi-drug resistantK. pneumoniae.