bluuebunny/crossref_metadata_2025_split huggingface dataset:
Label-free amino acid identification forde novoprotein sequencing via tRNA charging and current blockade in a nanopore
Authors: Sampath G. 
Published: June 2020

AbstractA label-free procedure to identify single amino acids (AAs) for protein sequencing is developed in theory and simulated in part. A terminal AA cleaved from a protein/peptide, a tRNA, its cognate amino-acyl tRNA synthetase (AARS), and adenosine triphosphate (ATP) are brought together in a container where tRNA, if cognate, gets charged with AA and adenosine monophosphate (AMP) is released. The released AMP (and any free AA and ATP molecules) filters into thecischamber of an adjoining electrolytic cell (e-cell) from where they pass through a nanopore into thetranschamber. Addition of NaOH to the container deacylates the tRNA if it is charged. The resulting free AA passes into thecischamber of the e-cell, translocates intotrans, and causes a current blockade; AA is immediately known from the identity of the tRNA (the two are cognate). If the tRNA is not charged there is no AA bound to it so AA remains unidentified. In this approach there is no need to distinguish among the 20 AAs by blockade size; it suffices to distinguish blockades from noise: thus a high-precision analog measurement has been transformed into a low-precision binary one. Identification is accurate because of tRNA superspecificity (the tRNA charging error rate is < 1/350); parallel execution with 20 different tRNAs can identify AA in one cycle. This is ade novomethod in which no prior information about the protein is used or needed.
Quantifying adaptive evolution and the effects of natural selection across the Norway spruce genome
Authors: Wang Xi , Ingvarsson Pär K 
Published: June 2020

AbstractDetecting natural selection is one of the major goals of evolutionary genomics. Here, we sequence whole genomes of 34Picea abiesindividuals and quantify the amount of selection across the genome. Using an estimate of the distribution of fitness effects, we show that negative selection is very limited in coding regions, while positive selection is rare in coding regions but very strong in non-coding regions, suggesting the great importance of regulatory changes in evolution of Norway spruce. Additionally, we found a positive correlation between adaptive rate with recombination rate and a negative correlation between adaptive rate and gene density, suggesting a widespread influence from Hill-Robertson interference to efficiency of protein adaptation inP. abies. Finally, the distinct population statistics between genomic regions under either positive or balancing selection with that under neutral regions indicated impact from selection to genomic architecture of Norway spruce. Further gene ontology enrichment analysis for genes located in regions identified as undergoing either positive or long-term balancing selection also highlighted specific molecular functions and biological processes in that appear to be targets of selection in Norway spruce.
The Set Increment with Limited Views Encoding Ratio (SILVER) Method for Optimizing Radial Sampling of Dynamic MRI
Authors: Schauman S. Sophie , Okell Thomas W. , Chiew Mark 
Published: June 2020

AbstractPurposeTo present and assess a method for choosing a fixed increment between spokes in radially sampled MRI that results in higher SNR than other radial sampling increments.Theory and MethodsSampling uniformity contributes to image SNR when reconstructed using linear methods. Thus, for a radial trajectory, uniformly spaced sampling is ideal. However, uniform sampling lacks post-acquisition reconstruction flexibility, which is often needed in dynamic imaging. Golden ratio-based increments are often used for this purpose. The method presented here, Set Increment with Limited Views Encoding Ratio (SILVER), optimizes sampling uniformity when a limited number of temporal resolutions are required. With SILVER, an optimization algorithm finds the angular increment that provides the highest uniformity for a pre-defined set of reconstruction window sizes. SILVER was tested over multiple sets and assessed in terms of uniformity, noise amplification, and SNR both in simulations and in acquisitions of a phantom and healthy volunteers.ResultsThe proposed algorithm produced trajectories that, for the optimized window sizes, had higher uniformity and lower image noise than golden ratio sampling both in a simulated single-coil system and in a multi-coil system, assessed using simulation, phantom, and in vivo experiments. The noise in SILVER optimized trajectories was comparable to uniformly distributed spokes whilst retaining flexibility in reconstruction at multiple temporal resolutions. In a resting state fMRI experiment, tSNR increases at different spatial and temporal resolutions ranged from 21-72%.ConclusionSILVER is a simple addition to any sequence currently using golden ratio sampling and significantly increases sampling efficiency and SNR.
Direct Supercritical Angle Localization Microscopy for Nanometer 3D Superresolution
Authors: Dasgupta Anindita , Deschamps Joran , Matti Ulf , Hübner Uwe , Becker Jan , Strauss Sebastian , Jungmann Ralf , Heintzmann Rainer , Ries Jonas 
Published: June 2020

Abstract3D single molecule localization microscopy (SMLM) is an emerging superresolution method for structural cell biology, as it allows probing precise positions of proteins in cellular structures. Supercritical angle fluorescence strongly depends on the z-position of the fluorophore and can be used for z localization in a method called supercritical angle localization microscopy (SALM). Here, we realize the full potential of SALM by directly splitting supercritical and undercritical emission, using an ultra-high NA objective, and applying new fitting routines to extract precise intensities of single emitters, resulting in a four-fold improved z-resolution compared to the state of the art. We demonstrate nanometer isotropic localization precision on DNA origami structures, and on clathrin coated vesicles and microtubules in cells, illustrating the potential of SALM for cell biology.
Facilitating candidate gene discovery in an emerging model plant lineage: Transcriptomic and genomic resources forThalictrum(Ranunculaceae)
Authors: Arias Tatiana , Riaño-Pachón Diego Mauricio , Di Stilio Verónica S. 
Published: June 2020

ABSTRACTThe plant genusThalictrumis a representative of the order Ranunculales (a sister lineage to all other Eudicots) with diverse floral morphologies, encompassing four sexual systems and two pollination modes. Previous studies suggest multiple transitions from insect to wind pollination within this genus, in association with polyploidy and unisexual flowers, but the underlying genes remain unknown. We generated a draft reference genome forThalictrum thalictroides, a representative of a clade with ancestral floral traits (diploidy, hermaphroditism, and insect pollination) and a model for functional studies. To facilitate candidate gene discovery in flowers with different sexual and pollination systems we also generated floral transcriptomes ofT. thalictroidesand of wind-pollinated, andromonoecious (staminate and hermaphroditic flowers on the same plant)T. hernandezii.TheT. thalictroidesdraft genome assembly consisted of 44,860 contigs (N50=12,761 bp. and 243 Mbp. total length) and contained 84.5% conserved embryophyte single-copy genes. Floral transcriptomes from Illumina sequencing andde novoassembly contained representatives of most eukaryotic core genes (approximately 80%), with most of their genes falling into common orthologous groups (orthogroups). Simple Sequence Repeat (SSR) motifs were also identified, which together with the single-copy genes constitute a resource for population-level or phylogenetic studies. Finally, to validate the utility of these resources, putative candidate genes were identified for the different floral morphologies using stepwise dataset comparisons. In conclusion, we present genomic and transcriptomic resources forThalictrum, including the first genome ofT. thalictroidesand potential candidate genes for flowers with distinct sexual and pollination systems.
Crossing the Blood-Brain-Barrier: A bifunctional liposome for BDNF gene delivery – A Pilot Study
Authors: Diniz Danielle M. , Franze Silvia , Homberg Judith R. 
Published: June 2020

AbstractTo achieve their therapeutic effect on the brain, molecules need to pass the blood-brain-barrier (BBB). Many pharmacological treatments of neuropathologies encounter the BBB as a barrier, hindering their effective use. Pharmaceutical nanotechnology based on optimal physicochemical features and taking advantage of naturally occurring permeability mechanisms, nanocarriers such as liposomes offer an attractive alternative to allow drug delivery across the BBB. Liposomes are spherical bilayer lipid-based nanocapsules that can load hydrophilic molecules in their inner compartment and on their outer surface can be functionally modified by peptides, antibodies and polyethyleneglycol (PEG). When composed of cationic lipids, liposomes can serve as gene delivery devices, encapsulating and protecting genetic material from degradation and promoting nonviral cell transfection. In this study, we aimed to develop a liposomal formulation to encapsulate a plasmid harbouring brain-derived neurotrophic factor (BDNF) and infuse these liposomes via the peripheral bloodstream into the brain. To this end, liposomes were tagged with PEG, transferrin, and arginine and characterized regarding their physical properties, such as particle size, zeta-potential and polydispersity index (PDI). Moreover, we selected liposomes preparations for plasmid DNA (pDNA) encapsulation and checked for loading efficiency, in vitro cell uptake, and transfection. The preliminary results from this pilot study revealed that we were able to replicate the liposomes synthesis described in literature, achieving compatible size, charge, PDI, and loading efficiency. However, we could not properly determine whether the conjugation of the surface ligands transferrin and arginine to PEG worked and whether they were attached to the surface of the liposomes. Additionally, we were not able to see transfection in SH-SY5Y cells after 24 or 48 hours of incubation with the pDNA loaded liposomes. In conclusion, we synthesized liposomes encapsulation pBDNF, however, further research will be necessary to address the complete physicochemical characterization of the liposomes. Furthermore, preclinical studies will be helpful to verify transfection efficiency, cytotoxicity, and in the future, safe delivery of BDNF through the BBB.
Interferon-Induced Transmembrane Protein 3 Blocks Fusion of Diverse Enveloped Viruses by Locally Altering Mechanical Properties of Cell Membranes
Authors: Guo Xiangyang , Steinkühler Jan , Marin Mariana , Li Xiang , Lu Wuyuan , Dimova Rumiana , Melikyan Gregory B. 
Published: June 2020

AbstractInterferon-induced transmembrane protein 3 (IFITM3) potently inhibits entry of diverse enveloped viruses by trapping the viral fusion at a hemifusion stage, but the underlying mechanism remains unclear. Here, we show that recombinant IFITM3 reconstituted into lipid vesicles induces negative membrane curvature and that this effect maps to its small amphipathic helix (AH). We demonstrate that AH: (i) partitions into lipid-disordered domains where IAV fusion occurs, (ii) induces negative membrane curvature, and (iii) increases lipid order and membrane stiffness. Additionally, replacing the IFITM3 AH with AH from an unrelated protein did not compromise its antiviral activity. These effects on membrane properties correlate with the fusion-inhibitory activity, as exogenous addition of AH to insulin-producing cells reduces glucose-stimulated insulin secretion. Our results thus imply that IFITM3 inhibits the transition from hemifusion to full fusion by imposing an unfavorable membrane curvature and increasing the order and stiffness of the cytoplasmic leaflet of endosomal membranes.
DiMSum: an error model and pipeline for analyzing deep mutational scanning data and diagnosing common experimental pathologies
Authors: Faure Andre J. , Schmiedel Jörn M. , Baeza-Centurion Pablo , Lehner Ben 
Published: June 2020

AbstractDeep mutational scanning (DMS) enables multiplexed measurement of the effects of thousands of variants of proteins, RNAs and regulatory elements. Here, we present a customizable pipeline – DiMSum – that represents an end-to-end solution for obtaining variant fitness and error estimates from raw sequencing data. A key innovation of DiMSum is the use of an interpretable error model that captures the main sources of variability arising in DMS workflows, outperforming previous methods. DiMSum is available as an R/Bioconda package and provides summary reports to help researchers diagnose common DMS pathologies and take remedial steps in their analyses.
Bi-compartmentalized stem cell organization of the corneal limbal niche
Authors: Farrelly Olivia , Suzuki-Horiuchi Yoko , Brewster Megan , Kuri Paola , Huang Sixia , Rice Gabriella , Xu Jianming , Dentchev Tzvete , Lee Vivian , Rompolas Panteleimon 
Published: June 2020

AbstractStem cells exist in precise locations within tissues, yet how their organization supports tissue architecture and function is poorly understood. The limbus is the presumptive stem cell niche of the corneal epithelium. Here, we visualize the live limbus and track the activity of single stem cells in their native environment by 2-photon microscopy. We identify previously unknown niche compartments and show that long implicated slow-cycling cells form separate lineages in the outer limbus, with only local clonal dynamics. Instead, we find distinct stem cells in the pericorneal limbus to be required for corneal regeneration. Unbiased photolabeling captures their progeny exiting the niche, then moving centripetally in unison before undergoing terminal differentiation. This study demonstrates how a compartmentalized stem cell organization coordinates tissue regeneration.One Sentence SummaryIn vivo live imaging of the regenerating cornea reveals distinct stem cell activities in the limbal niche
Follicle Stem Cells (FSCs) in the Drosophila ovary; a critique of published studies defining the number, location and behavior of FSCs
Authors: Kalderon Daniel , Melamed David , Reilein Amy 
Published: June 2020

SummaryA paper by Reilein et al., (2017) presented several key new insights into the behavior of adult Follicle Stem Cells (FSCs) in the Drosophila ovary, including overwhelming evidence that each ovariole hosts a large number of FSCs (about 14-16) maintained by population asymmetry (Reilein et al., 2017), rather than just two FSCs, dividing with largely individually asymmetric outcomes, as originally proposed (Margolis and Spradling, 1995; Nystul and Spradling, 2007). Here we provide further discussion asserting the merits of the conclusions of Reilein et al., (2017) and the deficiencies in the contrary assertions recently presented by Fadiga and Nystul (Fadiga and Nystul, 2019). The principles that we discuss here, particularly with regard to lineage tracing and population asymmetry, are common to the investigation of most types of adult stem cell and should therefore be instructive and of interest to investigators studying any type of adult stem cell. The improved understanding of FSC numbers, location and behavior afforded by Reilein et al., (2017) and Reilein et al., (2018) can only provide a firm foundation for future progress once they are widely appreciated and seen to be resistant to challenge, as described in detail here.
A targeted next generation sequencing approach to develop robust, genotype-specific mutation profiles and uncover novel variants inSaccharomyces cerevisiae
Authors: Lamb Natalie A. , Bard Jonathan , Buck Michael J. , Surtees Jennifer A. 
Published: June 2020

ABSTRACTDistinct mutation signatures arise from environmental exposures and/or from defects in metabolic pathways that promote genome stability. The presence of a particular mutation signature in a cell or a tumor can therefore predict the underlying mechanism of mutagenesis, which, in practice, may be clinically important. These insults to the genome often alter dNTP pools, which itself impacts replication fidelity. Therefore, the impact of altered dNTP pools should be considered when making mechanistic predictions based on mutation signatures. We developed a targeted deep-sequencing approach on theCAN1gene inSaccharomyces cerevisiaeto define information-rich mutational profiles associated with distinctrnr1backgrounds that alter replication fidelity by elevating dNTP levels.. The mutation spectra ofrnr1Y285Fandrnr1Y285Aalleles were characterized previously; our analysis was consistent with this prior work but the sequencing depth achieved in our study allowed a significantly more robust and nuanced computational analysis of the variants observed, generating profiles that integrated information about mutation spectra, position effects, and sequence context. This approach revealed novel, genotype-specific mutation profiles in the presence of even modest changes in dNTP pools. Furthermore, we identified broader sequence contexts and specific nucleotide motifs that influenced variant profiles in differentrnr1backgrounds, which allowed us to make specific mechanistic predictions about the impact of altered dNTP pools on replication fidelity.
ESCRT recruitment by the inner nuclear membrane protein Heh1 is regulated by Hub1-mediated alternative splicing
Authors: Capella Matías , Caballero Lucía Martín , Pfander Boris , Braun Sigurd , Jentsch Stefan 
Published: June 2020

AbstractMisassembled nuclear pore complexes (NPCs) are removed by sealing off the surrounding nuclear envelope (NE), which is mediated by members of the ESCRT (endosomalsortingcomplexesrequired fortransport) machinery. Recruitment of ESCRT proteins to the NE is mediated by the interaction between the ESCRT member Chm7 and the inner nuclear membrane protein Heh1, which belongs to the conserved LEM family. Increased ESCRT recruitment results in excessive membrane scission at damage sites but its regulation remains poorly understood. Here, we show that Hub1-mediated alternative splicing ofHEH1pre-mRNA, resulting into its shorter form Heh1-S, is critical for the integrity of the NE. ESCRT-III mutants lacking Hub1 or Heh1-S display severe growth defects and accumulate improperly assembled NPCs. This depends on the interaction of Chm7 with the conserved MSC domain only present in the longer spliced variant Heh1-L. Heh1 variants assemble into heterodimers and we demonstrate that a unique splice segment in Heh1-S suppresses growth defects associated with uncontrolled interaction between Heh1-L and Chm7. Together, our findings reveal that Hub1-mediated splicing generates Heh1-S to regulate ESCRT recruitment to the nuclear envelope.Summary statementHeh1-S, the Hub1-mediated spliced version ofHEH1pre-mRNA, contributes to nuclear envelope maintenance by preventing excessive recruitment of Chm7.
A single cell atlas of the healthy breast tissues reveal clinically relevant clusters of breast epithelial cells
Authors: Bhat-Nakshatri Poornima , Gao Hongyu , McGuire Patrick C. , Xuei Xiaoling , Sheng Liu , Wan Jun , Liu Yunlong , Althouse Sandra K. , Colter Austyn , Sandusky George , Storniolo Anna Maria , Nakshatri Harikrishna 
Published: June 2020

SummarySingle cell RNA sequencing is an evolving field to elucidate cellular architecture of adult organs. Using normal breast tissues from healthy volunteers and a rapid procurement/processing/sequencing protocol, 13 breast epithelial cell clusters were identified. Approximately 90% of breast cancers were enriched for cell-of-origin signatures derived from differentiated luminal clusters and two minor luminal progenitor clusters. Expression of cell cycle and chromosome segregation-related genes were higher in one of the minor clusters and breast tumors with this cluster signature displayed the highest mutation rate and poor outcome. We identified TBX3 and PDK4 as genes co-expressed with estrogen receptor (ER) in the normal breasts and their expression analyses in >550 breast cancers enabled prognostically relevant cell-of-origin based subclassification of ER+ breast cancers.SignificanceThis study elucidates different epithelial cell types of the normal breasts and identifies a minor subpopulation of cells from which the majority of breast cancers may originate. This observation should help to develop methods to characterize breast tumors based on cell-of-origin. Although it was suggested that intrinsic subtypes of breast cancers have distinct cells of origins, this study suggests multiple cell-of-origin for an intrinsic subtype of breast cancer, including for hormone responsive breast cancers. Cell-of-origin signatures allowed survival-associated subclassification of intrinsic subtypes. Critically, this normal breast cell atlas would allow for the classification of genes differentially expressed in a breast tumor compared to normal breast due to the cell-of-origin of tumor and those that are acquired due to genomic aberrations.
Tsetse salivary glycoproteins are modified with paucimannosidicN-glycans, are recognised by C-type lectins and bind to trypanosomes
Authors: Kozak Radoslaw P. , Mondragon-Shem Karina , Williams Christopher , Rose Clair , Perally Samirah , Caljon Guy , Van Den Abbeele Jan , Wongtrakul-Kish Katherine , Gardner Richard , Spencer Daniel , Lehane Michael J. , Acosta-Serrano Alvaro 
Published: June 2020

AbstractAfrican sleeping sickness is caused byTrypanosoma brucei,a parasite transmitted by the bite of a tsetse fly. Trypanosome infection induces a severe transcriptional downregulation of tsetse genes encoding for salivary proteins, which reduces its anti-hemostatic and anti-clotting properties. To better understand trypanosome transmission and the possible role of glycans in insect bloodfeeding, we characterized theN-glycome of tsetse saliva glycoproteins. Tsetse salivaryN-glycans were enzymatically released, tagged with either 2-aminobenzamide (2-AB) or procainamide, and analyzed by HILIC-UHPLC-FLR coupled online with positive-ion ESI-LC-MS/MS. We found that theN-glycan profiles ofT. brucei-infected and naïve tsetse salivary glycoproteins are almost identical, consisting mainly (>50%) of highly processed Man3GlcNAc2 in addition to several other paucimannose, high mannose, and few hybrid-type glycans. In overlay assays, these sugars were differentially recognized by the C-type lectins mannose receptor and DC-SIGN. We also show that salivary glycoproteins bind strongly to the surface of transmissible metacyclic trypanosomes. We suggest that although the repertoire of tsetse salivaryN-glycans does not change during a trypanosome infection, the interactions with mannosylated glycoproteins may influence parasite transmission into the vertebrate host.
The Evolution of Insect Metallothioneins
Authors: Luo Mei , Finet Cédric , Cong Haosu , Wei Hong-yi , Chung Henry 
Published: June 2020

ABSTRACTMetallothioneins (MTs) are a family of cysteine-rich metal-binding proteins that are important in the chelating and detoxification of toxic heavy metals. Until now, the short length and the low sequence complexity of MTs has hindered the possibility of any phylogenetic reconstruction, hampering the study of their evolution. To answer this longstanding question, we developed an iterative BLAST search pipeline that allowed us to build a unique dataset of more than 300 MT sequences in insects. By combining phylogenetics and synteny analysis, we reconstructed the evolutionary history of MTs in insects. We show that the MT content in insects has been shaped by lineage-specific tandem duplications from a single ancestral MT. Strikingly, we also uncovered a sixth MT,MtnF, in the model organismDrosophila melanogaster.MtnFevolves faster than other MTs and is characterized by a non-canonical length and higher cysteine content. Our methodological framework not only paves the way for future studies on heavy metal detoxification but also can allow us to identify other previously unidentified genes and other low complexity genomic features.