bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dynamic competition between SARS-CoV-2 NSP1 and mRNA on the human ribosome inhibits translation initiation
Authors: Lapointe Christopher P. , Grosely Rosslyn , Johnson Alex G. , Wang Jinfan , Fernández Israel S. , Puglisi Joseph D. 
Published: August 2020

ABSTRACTSARS-CoV-2 recently emerged as a human pathogen and is the causative agent of the COVID-19 pandemic. A molecular framework of how the virus manipulates host cellular machinery to facilitate infection remains unclear. Here, we focus on SARS-CoV-2 NSP1, which is proposed to be a virulence factor that inhibits protein synthesis by directly binding the human ribosome. Using extract-based and reconstitution experiments, we demonstrate that NSP1 inhibits translation initiation on model human and SARS-CoV-2 mRNAs. NSP1 also specifically binds to the small (40S) ribosomal subunit, which is required for translation inhibition. Using single-molecule fluorescence assays to monitor NSP1–40S subunit binding in real time, we demonstrate that eukaryotic translation initiation factors (eIFs) modulate the interaction: NSP1 rapidly and stably associates with most ribosomal pre-initiation complexes in the absence of mRNA, with particular enhancement and inhibition by eIF1 and eIF3j, respectively. Using model mRNAs and an inter-ribosomal-subunit FRET signal, we elucidate that NSP1 competes with RNA segments downstream of the start codon to bind the 40S subunit and that the protein is unable to associate rapidly with 80S ribosomes assembled on an mRNA. Collectively, our findings support a model where NSP1 associates with the open head conformation of the 40S subunit to inhibit an early step of translation, by preventing accommodation of mRNA within the entry channel.SIGNIFICANCE STATEMENTSARS-CoV-2 is the causative agent of the COVID-19 pandemic. A molecular framework for how SARS-CoV-2 manipulates host cellular machinery to facilitate infection is needed. Here, we integrate biochemical and single-molecule strategies to reveal molecular insight into how NSP1 from SARS-CoV-2 inhibits translation initiation. NSP1 directly binds to the small (40S) subunit of the human ribosome, which is modulated by human initiation factors. Further, NSP1 and mRNA compete with each other to bind the ribosome. Our findings suggest that the presence of NSP1 on the small ribosomal subunit prevents proper accommodation of the mRNA. How this competition disrupts the many steps of translation initiation is an important target for future studies.
Secondary metabolites of Bacillus subtilis impact soil-derived semi-synthetic bacterial community assembly
Authors: Kiesewalter Heiko T. , Lozano-Andrade Carlos N. , Strube Mikael L. , Kovács Ákos T. 
Published: August 2020

AbstractSecondary metabolites provide Bacillus subtilis with increased competitiveness towards other microorganisms. In particular nonribosomal peptides (NRPs) have an enormous antimicrobial potential by causing cell lysis, perforation of fungal membranes, enzyme inhibition or disruption of bacterial protein synthesis. This knowledge was primarily acquired in vitro when B. subtilis was competing with other microbial monocultures. However, our understanding of the true ecological role of these small molecules is limited.In this study, we have established soil-derived semi-synthetic mock communities containing 13 main genera and supplemented them with B. subtilis P5_B1 WT, its NRP deficient strain sfp or single NRP mutants incapable of producing surfactin, plipastatin or bacillaene. 16S amplicon sequencing revealed that the invasion of NRP-producing B. subtilis strains had no major impact on the bacterial communities. Still, the abundances of the two genera Lysinibacillus and Viridibacillus were reduced. Interestingly, this effect was diminished in communities supplemented with the NRPs deficient strain. Growth profiling of Lysinibacillus fusiformis M5 exposed to either spent media of the B. subtilis strains or pure surfactin indicates the sensitivity of this strain towards the biosurfactant surfactin.Our study provides a more in-depth insight into the influence of B. subtilis NRPs on semi-synthetic bacterial communities and helps to understand their ecological role.
Biocontrol ofAspergillus nigerin 3D-lung cell tissues by oxalotrophic bacteria
Authors: Palmieri Fabio , Palmieri Ilona , Noormamode Nourine , Estoppey Aislinn , Ishak M. Omar , Kelliher Julia M. , Vallat Armelle , Iyer Rashi , Bindschedler Saskia , Davenport Karen , Chain Patrick S. G. , Harris Jennifer Foster , Junier Pilar 
Published: August 2020

AbstractAspergillusfungi are opportunistic pathogens that affect a large number of people worldwide. Many aspects ofAspergillusspp. pathogenesis toward humans are known, but their ability to enhance their infectious potential by manipulating the environmental pH of its host has not been considered yet. In this study, we tested the hypothesis that by producing oxalic acid,Aspergillus nigercan manipulate pH during lung infection and thus, interfering with this process could limit pathogenicity. To test this hypothesis, we co-culturedA. nigerwith oxalotrophic bacteria in increasingly complex testing systems (Petri dishes and 3D-cell cultures systems). Inin vitrotests, oxalotrophic bacteria limit oxalic acid production and suppressed the pH shift induced byA. niger. In 3D-cell cultures (Transwells® and Bronchioles-on-a-chip),A. nigeralso modified pH, Ca2+and oxalic acid concentrations. Co-inoculation with as little as 10 cells of the oxalatrophic bacterium strongly inhibited the germination and development ofA. nigerand returned each of the three parameters to the baseline physiological values of uninfected cells. This biocontrol interaction between oxalotrophic bacteria and oxalate-producingA. nigercould represent a paradigm shift in the fight against opportunistic fungal pathogens, where the host environment is rendered less permissive to fungal development.
Phase separation by the Sterile Alpha Motif of Polyhomeotic compartmentalizes Polycomb Group proteins and enhances their activity
Authors: Seif Elias , Kang Jin Joo , Sasseville Charles , Senkovitch Olga , Kaltashov Alexander , Boulier Elodie L. , Kapur Ibani , Kim Chongwoo A. , Francis Nicole J. 
Published: August 2020

AbstractPolycomb Group (PcG) proteins organize chromatin at multiple scales to regulate gene expression. A conserved Sterile Alpha Motif (SAM) in thePolycombRepressiveComplex 1 (PRC1) subunit Polyhomeotic (Ph) is important for chromatin compaction and large-scale chromatin organization. Like many SAMs, Ph SAM forms helical head to tail polymers, and SAM-SAM interactions between chromatin-bound Ph/PRC1 are believed to compact chromatin and mediate long-range interactions. To understand mechanistically how this occurs, we analyzed the effects of Ph SAM on chromatinin vitro. We find that incubation of chromatin or DNA with a truncated Ph protein containing the SAM results in formation of concentrated, phase-separated condensates. Condensate formation depends on Ph SAM, and is enhanced by but not strictly dependent on, its polymerization activity. Ph SAM-dependent condensates can recruit PRC1 from extracts and enhance PRC1 ubiquitin ligase activity towards histone H2A. Overexpression of Ph with an intact SAM increases ubiquitylated H2A in cells. Thus, phase separation is an activity of the SAM, which, in the context of Ph, can mediate large-scale compaction of chromatin into biochemical compartments that facilitate histone modification.
Mass Spectrometry Imaging of N-Glycans Reveals Racial Discrepancies in Low Grade Prostate Tumors
Authors: Conroy Lindsey R. , Young Lyndsay E.A. , Stanback Alexandra E. , Austin Grant L. , Liu Jinpeng , Liu Jinze , Allison Derek B. , Sun Ramon C. 
Published: August 2020

AbstractProstate cancer is the most common cancer in men worldwide. Despite its prevalence, there is a critical knowledge gap regarding the underlining molecular events that result in higher incidence and mortality rate in Black men. Identifying molecular features that separate racial disparities is a critical step in prostate cancer research that could lead to predictive biomarkers and personalized therapy. N-linked glycosylation is a co-translational event during protein folding that modulates a myriad of cellular processes. Recently, aberrant N-linked glycosylation has been reported in prostate cancers. However, the full clinical implications of dysregulated glycosylation in prostate cancer has yet to be explored. Herein, we performed high-throughput matrix-assisted laser desorption ionization mass spectrometry analysis to characterize the N-glycan profile from tissue microarrays of over 100 patient tumors with over 10 years of follow up data. We identified several species of N-glycans that were profoundly different between low grade prostate tumors resected from White and Black patients. Further, these glycans predict opposing overall survival between White and Black patients with prostate cancer. These data suggest differential N-linked glycosylation underline the racial disparity of prostate cancer prognosis. Our study highlights the potential applications of MALDI-MSI for digital pathology and biomarker to study racial disparity of prostate cancer patients.Graphical Abstract
Maladaptive myelination promotes epileptogenesis in absence epilepsy
Authors: Knowles Juliet K. , Soane Caroline , Frost Eleanor , Tam Lydia T. , Fraga Danielle , Xu Haojun , Batra Ankita , Ni Lijun , Villar Katlin , Saucedo Tristan , Huguenard John , Monje Michelle 
Published: August 2020

SummaryNeuronal activity can influence the generation of new oligodendrocytes (oligodendrogenesis) and myelination. In health, this is an adaptive process that can increase synchrony within distributed neuronal networks and contribute to cognitive function. We hypothesized that in seizure disorders, aberrant neuronal activity may promote maladaptive myelination that contributes to pathogenesis. Absence epilepsy is a disease defined by increasingly frequent behavioral arrest seizures over time, thought to be due to thalamocortical network hypersynchrony. We tested the hypothesis that activity-dependent myelination resulting from absence seizures promotes epileptogenesis. Using two distinct models of absence epilepsy, Wag/Rij rats andScn8a+/mutmice, we found increased oligodendrogenesis and myelination specifically within the absence seizure network. These changes are evident only after seizure onset in both models and are prevented with pharmacological inhibition of seizures. Genetic blockade of activity-dependent myelination during epileptogenesis markedly decreased seizure frequency in theScn8a+/mutmouse model of absence epilepsy. Taken together, these findings indicate that activity-dependent myelination driven by absence seizures contributes to seizure kindling during epileptogenesis.
Naphthalene monoimide derivative ameliorates amyloid burden and cognitive decline in a transgenic mouse model of Alzheimer’s disease
Authors: Samanta Sourav , Rajasekhar Kolla , Ramesh Madhu , Murugan N. Arul , Alam Shadab , Shah Devanshi , Clement James P , Govindaraju Thimmaiah 
Published: August 2020

ABSTRACTAlzheimer’s disease (AD) is a major neurodegenerative disorder and the leading cause of dementia worldwide. Predominantly, misfolding and aggregation of amyloid-β (Aβ) peptides associated with multifaceted toxicity is the neuropathological hallmark of AD pathogenesis and thus, primary therapeutic target to ameliorate neuronal toxicity and cognitive deficits. Herein, we report the design, synthesis and evaluation of small molecule inhibitors with naphthalene monoimide scaffold to amelioratein vitroandin vivoamyloid induced neurotoxicity. The detailed studies established TGR63 as the lead candidate to rescue neuronal cells from amyloid toxicity. Thein silicostudies showed disruption of salt bridges and intermolecular hydrogen bonding interactions within Aβ42 fibrils by the interaction of TGR63, causing destabilization of Aβ42 assembly. Remarkably, TGR63 treatment showed a significant reduction in cortical and hippocampal amyloid burden in the progressive stages of APP/PS1 AD mice brain. Various behavioral tests demonstrated rescued cognitive deficits. The excellent biocompatibility, BBB permeability and therapeutic efficacy to reduce amyloid burden make TGR63 a promising candidate for the treatment of AD.
Virucidal and antiviral activity of astodrimer sodium against SARS-CoV-2 in vitro
Authors: Paull Jeremy R.A. , Heery Graham P. , Bobardt Michael D. , Castellarnau Alex , Luscombe Carolyn A. , Fairley Jacinth K. , Gallay Philippe A. 
Published: August 2020

AbstractAn effective response to the ongoing coronavirus disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) will involve a range of complementary preventive modalities. The current studies were conducted to evaluate the in vitro SARS-CoV-2 antiviral and virucidal activity of astodrimer sodium, a dendrimer with broad spectrum antimicrobial activity, including against enveloped viruses in in vitro and in vivo models, that is marketed for antiviral and antibacterial applications. We report that astodrimer sodium inhibits replication of SARS-CoV-2 in Vero E6 and Calu-3 cells, with 50% effective concentrations (EC50) for i) reducing virus-induced cytopathic effect of 0.002 to 0.012 mg/mL in Vero E6 cells, and ii) infectious virus release by plaque assay of 0.019 to 0.031 mg/mL in Vero E6 cells and 0.031 to 0.045 mg/mL in Calu-3 cells. The selectivity index (SI) in these assays was as high as 2197. Astodrimer sodium was also virucidal, reducing SARS-CoV-2 infectivity by >99.9% (>3 log10) within 1 minute of exposure, and up to >99.999% (>5 log10) shown at astodrimer sodium concentrations of 10 to 30 mg/mL in Vero E6 and Calu-3 cell lines. Astodrimer sodium also inhibited infection in a primary human airway epithelial cell line. The data were similar for all investigations and were consistent with the potent antiviral and virucidal activity of astodrimer sodium being due to inhibition of virus-host cell interactions, as previously demonstrated for other viruses. Further studies will confirm if astodrimer sodium binds to SARS-CoV-2 spike protein and physically blocks initial attachment of the virus to the host cell. Given the in vitro effectiveness and significantly high SI, astodrimer sodium warrants further investigation for potential as a nasally administered or inhaled antiviral agent for SARS-CoV-2 prevention and treatment applications.
A simple, high-throughput method of protein and label removal from extracellular vesicle samples
Authors: Welsh Joshua A. , Killingsworth Bryce , Kepley Julia , Traynor Tim , McKinnon Kathy , Savage Jason , Appel Deven , Aldape Kenneth , Camphausen Kevin , Berzofsky Jay A. , Ivanov Alexander R. , Ghiran Ionita H. , Jones Jennifer C. 
Published: August 2020

AbstractEvidence continues to increase of the clinical utility extracellular vesicles (EVs) can provide as translational biomarkers. While a wide variety of EV isolation and purification methods have been implemented, few techniques are high-throughput and scalable for removing excess fluorescent reagents (e.g. dyes, antibodies). EVs are too small to be recovered from routine cell-processing procedures, such as filtration or centrifugation. The lack of suitable methods for removing unbound labels, especially in optical assays, is a major roadblock to accurate EV phenotyping and utilization of EV assays in a translational or clinical setting. Therefore, we developed a method for using a multi-modal resin, referred to as EV-Clean, to remove unbound labels from EV samples, and we demonstrate improvement in flow cytometric EV analysis with the use of this EV-Clean method.
Qualitative and quantitative bioanalytical methods validation of aristolochic acid DNA adducts and application in human formalin-fixed paraffin-embedded hepatocellular carcinoma tissues
Authors: xin Wang , jinping Song , yapping Dong , linghua Hu , xiaoying Chen , fangfang Yang , haiyang Qi , xinming Qi , wen Wen , shuzhen Chen , guozhen Xing , jin Ren 
Published: August 2020

AbstractMutation signature of aristolochic acid (AA) found in urothelial or hepatocellular carcinoma causes public concern about the cancer risk of AA. In contrast, direct evidence based on the reliable bioanalytical method for the exposure of AA is still lacking and not universal. Here, we strictly complied with the qualitative and quantitative guidance for forensic toxicological analysis: In the sample preprocessing, DNA from formalin-fixed and paraffin-embedded (FFPE) tissues was digested to single nucleotide by a series of enzymes with 70% enzymatic digestion efficiency. After protein precipitation, the samples were submitted to an ABI6500+ mass spectrometer for LC-MS/MS analysis. Ion pairs 543.2/427.2 and 543.2/395.2 of dA-AAI were selected from 5 ion pairs due to their higher LC-MS/MS response. Both these ion pairs have excellent selectivity and specificity in rat liver DNA matrix, and a linear regression range from 5 pg/mL to 200 pg/mL with the best fit and determination coefficient (r) greater than 0.99. The intra and inter batch accuracy and precision of these two ion pairs are also acceptable with less than 15% variation. The total recovery for ion pair 543.2/427.2 and 543.2/395.2 of dA-AAI was 90.06% and 90.76%, respectively. Our method has a minor matrix effect and good stability under different temperature and time conditions. With signal to noise ratio ≥ 3, 2 ion pairs (< 50 % relative abundance variation), the lower limit of quantification (LLOQ) of our method is set to 5 pg/mL(∼3.6 AAI-DNA adducts per 108DNA bases). By using this validated bioanalytical method of dA-AAI, 165 human HCC FFPE tissues were analyzed, the total ratio of samples with peak543.2/427.2is 17.0% (28/165), with peak543.2/395.2is 9.09% (15/165) which yields the total ratio of samples combined peak543.2/427.2and peak543.2/395.2is 7.27% (12/165). Two samples are higher than 5 pg/mL under the qualitative requirements. In conclusion, we first reported a fully validated methods to analyze the DNA adducts level of aristolochic acid, which could be qualitatively and quantitatively applied to the investigation of AA exposure in the human and other species.
Feasibility and promise of circulating tumor DNA analysis in dogs with naturally-occurring sarcoma
Authors: Favaro Patricia Filippsen , Stewart Samuel D. , McDonald Bradon R. , Cawley Jacob , Contente-Cuomo Tania , Wong Shukmei , Hendricks William P.D. , Trent Jeffrey M. , Khanna Chand , Murtaza Muhammed 
Published: August 2020

AbstractComparative studies of naturally-occurring canine cancers have provided new insight into many areas of cancer research. The inclusion of pet dogs in the development and validation of circulating tumor DNA (ctDNA) diagnostics may be uniquely informative for human translation for many reasons, including: high incidence of certain spontaneous cancers, repeated access to blood and tumor from the same individuals during the course of disease progression, and molecular heterogeneity of naturally-occurring cancers in dogs. Here, we present a feasibility study of ctDNA analysis performed in 9 healthy dogs and 39 dogs with either benign or malignant splenic tumors (hemangiosarcoma) using shallow whole genome sequencing (sWGS) of cell-free DNA. To enable detection and quantification of ctDNA using sWGS, we adapted two informatic approaches and compared their performance for the canine genome. At presentation, mean ctDNA tumor fraction in dogs with malignant splenic tumors was 11.2%, significantly higher than dogs with benign lesions (3.2%; p 0.001), achieving an AUC of 0.84. ctDNA tumor fraction was 14.3% and 9.0% in dogs with metastatic and localized disease, respectively although this difference was not statistically significant (p 0.227). In paired analysis, ctDNA fraction decreased from 11.0% to 7.9% after resection of malignant tumors (p 0.047). Our results demonstrate that ctDNA analysis is feasible in dogs with hemangiosarcoma using a cost-effective approach such as sWGS. Future studies are underway to validate these findings, and further evaluate the role of ctDNA to assess burden of disease and treatment response during drug development.
Centriole-less pericentriolar material serves as a microtubule organizing center at the base ofC. eleganssensory cilia
Authors: Magescas Jérémy , Eskinazi Sani , Tran Michael V. , Feldman Jessica L. 
Published: August 2020

SummaryDuring mitosis in animal cells, the centrosome acts as a microtubule organizing center (MTOC) to assemble the mitotic spindle. MTOC function at the centrosome is driven by proteins within the pericentriolar material (PCM), however the molecular complexity of the PCM makes it difficult to differentiate the proteins required for MTOC activity from other centrosomal functions. We used the natural spatial separation of PCM proteins during mitotic exit to identify a minimal module of proteins required for centrosomal MTOC function inC. elegans. Using tissue specific degradation, we show that SPD-5, the functional homolog of CDK5RAP2, is essential for embryonic mitosis while SPD-2/CEP192 and PCMD-1, which are essential in the zygote, are dispensable. Surprisingly, although the centriole is known to be degraded in the ciliated sensory neurons inC. elegans[1-3], we find evidence for “centriole-less PCM” at the base of cilia and use this structure as a minimal testbed to dissect centrosomal MTOC function. Super-resolution imaging revealed that this PCM inserts inside the lumen of the ciliary axoneme and directly nucleates the assembly of dendritic microtubules towards the cell body. Tissue-specific degradation in ciliated sensory neurons revealed a role for SPD-5 and the conserved microtubule nucleator γ-TuRC, but not SPD-2 or PCMD-1, in MTOC function at centriole-less PCM. This MTOC function was in the absence of regulation by mitotic kinases, highlighting the intrinsic ability of these proteins to drive microtubule growth and organization and further supporting a model that SPD-5 is the primary driver of MTOC function at the PCM.
PeptideMind – applying machine learning algorithms to assess replicate quality in shotgun proteomic data
Authors: Handler D.C.L. , Haynes P.A. 
Published: August 2020

AbstractAssessment of replicate quality is an important process for any shotgun proteomics experiment. One fundamental question in proteomics data analysis is whether any specific replicates in a set of analyses are biasing the downstream comparative quantitation. In this paper, we present an experimental method to address such a concern. PeptideMind uses a series of clustering Machine Learning algorithms to assess outliers when comparing proteomics data from two states with six replicates each. The program is a JVM native application written in the Kotlin language with Python sub-process calls to scikit-learn. By permuting the six data replicates provided into four hundred triplet non redundant pairwise comparisons, PeptideMind determines if any one replicate is biasing the downstream quantitation of the states. In addition, PeptideMind generates useful visual representations of the spread of the significance measures, allowing researchers a rapid, effective way to monitor the quality of those identified proteins found to be differentially expressed between sample states.