bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mineralized collagen scaffolds fabricated with amniotic membrane matrix increase osteogenesis under inflammatory conditions
Authors: Dewey Marley J. , Johnson Eileen M. , Slater Simona T. , Milner Derek J. , Wheeler Matthew B. , Harley Brendan A.C. 
Published: January 2020

ABSTRACTDefects in craniofacial bones occur congenitally, after high-energy impacts, and during the course of treatment for stroke and cancer. These injuries are difficult to heal due to the overwhelming size of the injury area and the inflammatory environment surrounding the injury. Significant inflammatory response after injury may greatly inhibit regenerative healing. We have developed mineralized collagen scaffolds that can induce osteogenic differentiation and matrix biosynthesis in the absence of osteogenic media or supplemental proteins. The amniotic membrane is derived from placentas and has been recently investigated as an extracellular matrix to prevent chronic inflammation. Herein, we hypothesized that a mineralized collagen-amnion composite scaffold could increase osteogenic activity in the presence of inflammatory cytokines. We report mechanical properties of a mineralized collagen-amnion scaffold and investigated osteogenic differentiation and mineral deposition of porcine adipose derived stem cells within these scaffolds as a function of inflammatory challenge. Incorporation of amniotic membrane matrix promotes osteogenesis similarly to un-modified mineralized collagen scaffolds, and increases in mineralized collagen-amnion scaffolds under inflammatory challenge. Together, these findings suggest that a mineralized collagen-amnion scaffold may provide a beneficial environment to aid craniomaxillofacial bone repair, especially in the course of defects presenting significant inflammatory complications.
Estimation of Full-Length TprK Diversity inTreponema pallidumsubspeciespallidum
Authors: Addetia Amin , Lin Michelle , Phung Quynh , Xie Hong , Huang Meei-Li , Ciccarese Giulia , Dal Conte Ivano , Cusini Marco , Drago Francesco , Giacani Lorenzo , Greninger Alexander L. 
Published: January 2020

AbstractImmune evasion and disease progression ofTreponema pallidumsubspeciespallidumare associated with sequence diversity in the hypervariable, putative outer membrane protein TprK. Previous attempts to study variation within TprK have sequenced at depths insufficient to fully appreciate the hypervariable nature of the protein, failed to establish linkage between the protein’s 7 variable regions, or were conducted on strains passed through rabbits. As a consequence, a complete profiling oftprKduring infection in the human host is still lacking. Furthermore, prior studies examining howT. pallidumuses its repertoire of genomic donor sites to generate diversity within the V regions of thetprKalso yielded a partial understanding of this process, due to the limited number oftprKalleles examined. In this study, we used short- and long-read deep sequencing to directly characterize full-lengthtprKalleles fromT. pallidumcollected from early lesions of patients attending two STD clinics in Italy. Our data, combined with recent data available on ChineseT. pallidumstrains, show the near complete absence of overlap in TprK sequences among the 41 strains profiled to date. Moreover, our data allowed us to redefine the boundaries oftprKV regions, identify 55 donor sites, and estimate the total number of TprK variants thatT. pallidumcan potentially generate. Altogether, our results support howT. pallidumTprK antigenic variation system is an unsurmountable obstacle for the human immune system to naturally achieve infection eradication, and reiterate the importance of this mechanism for pathogen persistence in the host.ImportanceSyphilis continues to be a significant public health issue in both low- and high-income nations, including the United States, where the number of infectious syphilis cases has increased dramatically over the past five years.T. pallidum, the causative agent of syphilis, encodes an outer membrane protein TprK that undergoes segmental gene conversion to constantly create new sequences. We performed deep TprK profiling to understand full-length TprK diversity inT. pallidum-positive clinical specimens and compared these to all samples for which TprK deep sequencing is available. We found almost no overlap in TprK sequences between different patients. We further estimate that the total baseline junctional diversity of full-length TprK rivals that of current estimates of the human adaptive immune system. These data underscore the immunoevasive ability of TprK that allowsT. pallidumto establish lifelong infection.
Divided Attention in Perception: A Unified Analysis of Dual-Task Deficits and Congruency Effects
Authors: Palmer John , White Alex L. , Moore Cathleen M. , Boynton Geoffrey M. 
Published: January 2020

AbstractHow well can one perceive simultaneous stimuli at two widely spaced visual locations? Are the stimuli processed independently? If not, does the dependency affect perception, disrupt signals in later stages, or both? To address these questions, we measured effects of divided attention using a dual-task paradigm with stimuli presented in noise on either side of fixation. This paradigm was applied to detecting Gabor patches and to the semantic categorization of words. We measureddual-task deficitswhich are a decline in mean performance for a dual task compared to a single task. There was such a deficit for categorizing two words but relatively little deficit for detecting two Gabors. We also measuredcongruency effectswhich are when performance at one location depends on whether the stimulus at the other location requires the same response. There was such a congruency effect for detecting two Gabors but relatively little congruency effect for categorizing two words. Further experiments were consistent with the dual-task deficit in word categorization being perceptual, but the congruency effect in Gabor detection being due to later processes. Results of additional experiments showed that the congruency effect was consistent with either graded selection errors or with all-or-none selection followed by graded interactive processing. To answer our opening question: for Gabor detection, perceptual processes were largely independent but later processes caused congruency effects; for word categorization, perceptual processes had capacity limits but even in combination with later processes caused relatively little congruency effects. In summary, there was evidence for two different kinds of dependency. Such complementary dependencies are inconsistent with theories of divided attention that depend on a single dependency such as a single resource or single source of interactive processing.
Designing Effective small interfering RNA for Post-Transcriptional Silencing of Human GREM1: A Comprehensive Bioinformatics Approach
Authors: Ebenezer Asiedu 
Published: January 2020

ABSTRACTHuman gremlin-1 is a physiologically versatile signaling molecule that has been associated with several human diseases including cancer. The ability of gremlin-1 to induce fibrosis in organs and transduce angiogenesis makes it a target for cancer therapy. RNAi-based therapy has proven to be very efficient and specific in tumor growth inhibition. The efficacy and specificity of siRNA-mediated gene silencing depends on the designing approaches. Here, empirical guidelines for siRNA design and comprehensive target site availability analysis were used to select effective siRNA from a plethora of potential candidates designed using several computation algorithms. Then, the selected siRNA candidates were subjected to stringent similarity searches in order to obtain siRNA candidates with reduced off-target effects (high specificity). The best candidates were compared to experimentally successful gremlin-1 siRNAs in order to predict the silencing potency of the selected siRNAs. siRNA-6 (sense strand: 5’-CCAAGAAAUUCACUACCAU-3’), siRNA-7 (sense strand: 5’-CCAUGAUGGUCACACUCAA-3’) and siRNA-47 (sense strand: 5’-GGCCCAGCACAAUGACUCA-3’) were predicted to be highly effective siRNA candidates for gremlin-1 silencing. These siRNAs can be considered for RNAi-based therapy because off-target effects are predicted to be minimal.
Enhancing CO2-valorization usingClostridium autoethanogenumfor sustainable fuel and chemicals production
Authors: Heffernan James K. , Valgepea Kaspar , de Souza Pinto Lemgruber Renato , Casini Isabella , Plan Manuel , Tappel Ryan , Simpson Sean D. , Köpke Michael , Nielsen Lars K. , Marcellin Esteban 
Published: January 2020

AbstractAcetogenic bacteria can convert waste gases into fuels and chemicals. Design of bioprocesses for waste carbon valorization requires quantification of steady-state carbon flows. Here, steady-state quantification of autotrophic chemostats containingClostridium autoethanogenumgrown on CO2and H2revealed that captured carbon (460 ± 80 mmol/gDCW/day) had a significant distribution to ethanol (54 ± 3 mol% with a 2.4 ± 0.3 g/L titer). We were impressed with this initial result, but also observed limitations to biomass concentration and growth rate. Metabolic modelling predicted culture performance and indicated significant metabolic adjustments when compared to fermentation with CO as the carbon source. Moreover, modelling highlighted flux to pyruvate, and subsequently reduced ferredoxin, as a target for improving CO2and H2fermentation. Supplementation with a small amount of CO enabled co-utilisation with CO2, and enhanced CO2fermentation performance significantly, while maintaining an industrially relevant product profile. Additionally, the highest specific flux through the Wood-Ljungdahl pathway was observed during co-utilization of CO2and CO. Furthermore, the addition of CO led to superior CO2-valorizing characteristics (9.7 ± 0.4 g/L ethanol with a 66 ± 2 mol% distribution, and 540 ± 20 mmol CO2/gDCW/day). Similar industrial processes are commercial or currently being scaled up, indicating CO-supplemented CO2and H2fermentation has high potential for sustainable fuel and chemical production. This work also provides a reference dataset to advance our understanding of CO2gas fermentation, which can contribute to mitigating climate change.
Analysis of cell-associated DENV RNA by oligo(dT) primed 5’ capture scRNAseq
Authors: Sanborn Mark , Li Tao , Victor Kaitlin , Siegfried Hayden , Fung Christian , Rothman Alan L. , Srikiatkhachorn Anon , Fernandez Stefan , Ellison Damon , Jarman Richard G. , Friberg Heather , Berry Irina Maljkovic , Currier Jeffrey R. , Waickman Adam T 
Published: January 2020

ABSTRACTDengue is one of the most widespread vector-borne viral diseases in the world. However, the size, heterogeneity, and temporal dynamics of the cell-associated viral reservoir during acute dengue virus (DENV) infection remains unclear. In this study, we analyzed cells infectedin vitrowith DENV and PBMC from an individual experiencing a natural DENV infection utilizing 5’ capture single cell RNA sequencing (scRNAseq). Both positive- and negative-sense DENV RNA was detected in reactions containing either an oligo(dT) primer alone, or in reactions supplemented with a DENV-specific primer. The addition of a DENV-specific primer did not increase the total amount of DENV RNA captured or the fraction of cells identified as containing DENV RNA. However, inclusion of a DENV-specific cDNA primer did increase the viral genome coverage immediately 5’ to the primer binding site. Furthermore, while the majority of intracellular DENV sequence captured in this analysis mapped to the 5’ end of the viral genome, distinct patterns of enhanced coverage within the DENV polyprotein coding region were observed. The 5’ capture scRNAseq analysis of PBMC not only recapitulated previously published reports by detecting virally infected memory and naïve B cells, but also identified cell-associated genomic variants not observed in contemporaneous serum samples. These results demonstrate that oligo(dT) primed 5’ capture scRNAseq can detect DENV RNA and quantify virus-infected cells in physiologically relevant conditions, and provides insight into viral sequence variability within infected cells.IMPORTANCEDengue is one of the most widespread vector-borne viral diseases in the world. However, it is still unclear which cells harbor virus during DENV infection, and how viral reservoirs in serum and infected cells are related. These results demonstrate for the first time that intracellular DENV RNA can be identified and infected cells quantified by 5’ capture scRNAseq. This strategy provides a significantly greater throughput and higher sensitivity than previously published methods and has the potential to provide additional information on the genomic heterogeneity of intracellular DENV RNA.
Rapid and scalable profiling of nascent RNA with fastGRO
Authors: Barbieri Elisa , Hill Connor , Quesnel-Vallieres Mathieu , Barash Yoseph , Gardini Alessandro 
Published: January 2020

AbstractGenome-wide profiling of nascent RNA has become a fundamental tool to study transcription regulation. Over the past decade, next-generation sequencing has fostered development of a handful of techniques (i.e. GRO-seq, PRO-seq, TT-seq and NET-seq) that map unprocessed transcripts originating from both the coding and the noncoding portion of the genome. Unlike steady-state RNA sequencing, nascent RNA profiling mirrors the real-time activity of RNA Polymerases and provides an accurate readout of transcriptome-wide variations that occur during short time frames (i.e. response to external stimuli or rapid metabolic changes). Some species of nuclear RNAs, albeit functional, have a short half-life and can only be accurately gauged by nascent RNA techniques (i.e. lincRNAs and eRNAs). Furthermore, these techniques capture uncapped post-cleavage RNA at termination sites or promoter-associated antisense RNAs, providing a unique insight into RNAPII dynamics and processivity.Here we present a run-on assay with 4s-UTP labelling, followed by reversible biotinylation and affinity purification via streptavidin. Our protocol allows streamlined sample preparation within less than 3 days. We named the technique fastGRO (fast Global Run-On). We show that fastGRO is highly reproducible and yields a more complete and extensive coverage of nascent RNA than comparable techniques. Importantly, we demonstrate that fastGRO is scalable and can be performed with as few as 0.5×10^6 cells.
The dendritic spatial code: branch-specific place tuning and its experience-dependent decoupling
Authors: Rashid Shannon K. , Pedrosa Victor , Dufour Martial A. , Moore Jason J. , Chavlis Spyridon , Delatorre Rodrigo G. , Poirazi Panayiota , Clopath Claudia , Basu Jayeeta 
Published: January 2020

AbstractDendrites of pyramidal neurons integrate different sensory inputs, and non-linear dendritic computations drive feature selective tuning and plasticity. Yet little is known about how dendrites themselves represent the environment, the degree to which they are coupled to their soma, and how that coupling is sculpted with experience. In order to answer these questions, we developed a novel preparation in which we image soma and connected dendrites in a single plane across days usingin vivotwo-photon microscopy. Using this preparation, we monitored spatially tuned activity in area CA3 of the hippocampus in head-fixed mice running on a linear track. We identified “place dendrites”, which can stably and precisely represent both familiar and novel spatial environments. Dendrites could display place tuning independent of their connected soma and even their sister dendritic branches, the first evidence for branch-specific tuning in the hippocampus. In a familiar environment, spatially tuned somata were more decoupled from their dendrites as compared to non-tuned somata. This relationship was absent in a novel environment, suggesting an experience dependent selective gating of dendritic spatial inputs. We then built a data-driven multicompartment computational model that could capture the experimentally observed correlations. Our model predicts that place cells exhibiting branch-specific tuning have more flexible place fields, while neurons with homogenous or co-tuned dendritic branches have higher place field stability. These findings demonstrate that spatial representation is organized in a branch-specific manner within dendrites of hippocampal pyramidal cells. Further, spatial inputs from dendrites to soma are selectively and dynamically gated in an experience-dependent manner, endowing both flexibility and stability to the cognitive map of space.One sentence summaryHippocampal pyramidal cells show branch-specific tuning for different place fields, and their coupling to their soma changes with experience of an environment.
The allosteric control mechanism of bacterial glycogen biosynthesis disclosed by cryoEM
Authors: Cifuente Javier O. , Comino Natalia , D’Angelo Cecilia , Marina Alberto , Gil-Carton David , Albesa-Jové David , Guerin Marcelo E. 
Published: January 2020

ABSTRACTGlycogen and starch are the major carbon and energy reserve polysaccharides in nature, providing living organisms with a survival advantage. The evolution of the enzymatic machinery responsible for the biosynthesis and degradation of such polysaccharides, led the development of mechanisms to control the assembly and disassembly rate, to store and recover glucose according to cell energy demands. The tetrameric enzyme ADP-glucose pyrophosphorylase (AGPase) catalyzes and regulates the initial step in the biosynthesis of both α-polyglucans. Importantly, AGPase displays cooperativity and allosteric regulation by sensing metabolites from the cell energy flux. The understanding of the allosteric signal transduction mechanisms in AGPase arises as a long-standing challenge. In this work, we disclose the cryoEM structures of the paradigmatic homotetrameric AGPase fromEscherichia coli(EcAGPase), in complex with either positive or negative physiological allosteric regulators, FBP and AMP respectively, both at 3.0 Å resolution. Strikingly, the structures reveal that FBP binds deeply into the allosteric cleft and overlaps the AMP site. As a consequence, FBP promotes a concerted conformational switch of a regulatory loop, RL2, from a ‘locked’ to a ‘free’ state, modulating ATP binding and activating the enzyme. This notion is strongly supported by our complementary biophysical and bioinformatics evidence, and a careful analysis of vast enzyme kinetics data on single-point mutants ofEcAGPase. The cryoEM structures uncover the residue interaction networks (RIN) between the allosteric and the catalytic components of the enzyme, providing unique details on how the signaling information is transmitted across the tetramer, from which cooperativity emerges. Altogether, the conformational states visualized by cryoEM reveal the regulatory mechanism ofEcAGPase, laying the foundations to understand the allosteric control of bacterial glycogen biosynthesis at the molecular level of detail.
NF90 Modulates Processing of a Subset of Human Pri-miRNAs
Authors: Grasso Giuseppa , Higuchi Takuma , Barbier Jérôme , Helsmoortel Marion , Lorenzi Claudio , Sanchez Gabriel , Bello Maxime , Ritchie William , Sakamoto Shuji , Kiernan Rosemary 
Published: January 2020

ABSTRACTMicroRNAs are predicted to regulate the expression of more than 60% of mammalian genes and play fundamental roles in most biological processes. Deregulation of miRNA expression is a hallmark of most cancers and further investigation of mechanisms controlling miRNA biogenesis is needed. The dsRNA-binding protein, NF90 has been shown to act as a competitor of Microprocessor for a limited number of pri-miRNAs. Here, we show that NF90 has a more widespread effect on pri-miRNA biogenesis than previously thought. Genome-wide approaches revealed that NF90 is associated with the stem region of 38 pri-miRNAs, in a manner that is largely exclusive of Microprocessor. Following loss of NF90, 22 NF90-bound pri-miRNAs showed increased abundance of mature miRNA products. NF90-targeted pri-miRNAs are highly stable, having a lower free energy and fewer mismatches compared to all pri-miRNAs. Mutations leading to less stable structures reduced NF90 binding while increasing pri-miRNA stability led to acquisition of NF90 association, as determined by RNA EMSA. NF90-bound and modulated pri-miRNAs are embedded in introns of host genes and expression of several host genes is concomitantly modulated. These data suggest that NF90 controls the processing of a subset of highly stable, intronic miRNAs.
Bacterial FtsZ induces mitochondrial fission in human cells
Authors: Spier Anna , Sachse Martin , Tham Nam To , Matondo Mariette , Cossart Pascale , Stavru Fabrizia 
Published: January 2020

AbstractMitochondria are key eukaryotic organelles that evolved from an intracellular bacterium, in a process involving bacterial genome rearrangement and streamlining. As mitochondria cannot formde novo, their biogenesis relies on growth and division. In human cells, mitochondrial division plays an important role in processes as diverse as mtDNA distribution, mitochondrial transport and quality control. Consequently, defects in mitochondrial division have been associated with a wide range of human pathologies. While several protists have retained key components of the bacterial division machinery, none have been detected in human mitochondria, where the dynamin-related protein Drp1, a cytosolic GTPase is recruited to the mitochondrial outer membrane, forming helical oligomers that constrict and divide mitochondria. Here, we created a human codon optimized version of FtsZ, the central component of the bacterial division machinery, and fused it to a mitochondrial targeting sequence. Upon expression in human cells, mt-FtsZ was imported into the mitochondrial matrix, specifically localizing at fission sites prior to Drp1 and significantly increasing mitochondrial fission levels. Our data suggests that human mitochondria have an internal, matrix-localized fission machinery, whose structure is sufficiently conserved as to accommodate bacterial FtsZ. We identified interaction partners of mt-FtsZ, and show that expression of PGAM5, FAM210, SFXN3 and MTCH1 induced mitochondrial fission. Our results thus represent an innovative approach for the discovery of novel critical mitochondrial fission components.
METTL4 catalyzes m6Am methylation inU2 snRNAto regulate pre-mRNA splicing
Authors: Goh Yeek Teck , Koh Casslynn W. Q. , Sim Donald Yuhui , Roca Xavier , Goh W. S. Sho 
Published: January 2020

AbstractN6-methylation of 2’-O-methyladenosine (Am) in RNA occurs in eukaryotic cells to generateN6,2’-O-dimethyladenosine (m6Am). Identification of the methyltransferase responsible for m6Am catalysis has accelerated studies on the function of m6Am in RNA processing. While m6Am is generally found in the first transcribed nucleotide of mRNAs, the modification is also found internally withinU2 snRNA. However, the writer required for catalyzing internal m6Am formation had remained elusive. By sequencing transcriptome-wide RNA methylation at single-base-resolution, we identified human METTL4 as the writer that directly methylates Am atU2 snRNAposition 30 into m6Am. We found that METTL4 localizes to the nucleus and its conserved methyltransferase catalytic site is required forU2 snRNAmethylation. By sequencing human cells with overexpressedMettl4, we determined METTL4’sin vivotarget RNA motif specificity. In the absence ofMettl4in human cells,U2 snRNAlacks m6Am thereby affecting a subset of splicing events that exhibit specific features such as overall 3’ splice-site weakness with certain motif positions more affected than others. This study establishes that METTL4 methylation ofU2 snRNAregulates splicing of specific pre-mRNA transcripts.