bluuebunny/crossref_metadata_2025_split huggingface dataset:
High-throughput identification of FLT3 wild-type and mutant kinase substrate preferences and application to design of sensitive in vitro kinase assay substrates
Authors: Perez Minervo , Blankenhorn John , Murray Kevin J. , Parker Laurie L. 
Published: October 2018

SummaryAcute myeloid leukemia (AML) is an aggressive disease that is characterized by abnormal increase of immature myeloblasts in blood and bone marrow. The FLT3 receptor tyrosine kinase plays an integral role in haematopoiesis, and one third of AML diagnoses exhibit gain-of-function mutations in FLT3, with the juxtamembrane domain internal tandem duplication (ITD) and the kinase domain D835Y variants observed most frequently. Few FLT3 substrates or phosphorylation sites are known, which limits insight into FLT3’s substrate preferences and makes assay design particularly challenging. We applied in vitro phosphorylation of a cell lysate digest (adaptation of the Kinase Assay Linked with Phosphoproteomics (KALIP) technique and similar methods) for high-throughput identification of substrates for three FLT3 variants (wild-type, ITD mutant, and D835Y mutant). Incorporation of identified substrate sequences as input into the KINATEST-ID substrate preference analysis and assay development pipeline facilitated the design of several peptide substrates that are phosphorylated efficiently by all three FLT3 kinase variants. These substrates could be used in assays to identify new FLT3 inhibitors that overcome resistant mutations to improve FLT3-positive AML treatment.
Mutations in Rv2983 as a novel determinant of resistance to nitroimidazole drugs in Mycobacterium tuberculosis
Authors: Rifat Dalin , Li Si-Yang , Ioerger Thomas , Lanoix Jean-Philippe , Lee Jin , Bashiri Ghader , Sacchettini James , Nuermberger Eric 
Published: October 2018

AbstractDelamanid represents one of two novel antimicrobial classes approved to treat tuberculosis in over 40 years. Pretomanid is another promising nitroimidazole pro-drug in clinical development. Characterization of the full spectrum of mutations conferring resistance to nitroimidazoles and their related phenotypes in Mycobacterium tuberculosis will inform development of suitable genotypic and phenotypic drug susceptibility tests. Here, we used a range of pretomanid doses to select pretomanid-resistant mutants in two pathologically distinct murine TB models. The frequency of spontaneous pretomanid resistance mutations was approximately 10−5 CFU. Pretomanid demonstrated dose-dependent bactericidal activity and selective amplification of resistant mutants. Whole genome sequencing of 161 resistant isolates from 47 mice revealed 99 unique mutations, 90% of which were found in 1 of 5 genes previously associated with nitroimidazole activation and resistance. The remaining 10% harbored isolated mutations in Rv2983. Complementing an Rv2983 mutant with a wild-type copy of Rv2983 restored wild-type susceptibility to pretomanid and delamanid, confirming that loss of Rv2983 function causes nitroimidazole resistance. By quantifying F420 and its precursor Fo in Mycobacterium smegmatis overexpressing Rv2983 and an M. tuberculosis Rv2983 mutant, we provide evidence that Rv2983 is necessary for F420 biosynthesis and nitroimidazole activation, perhaps as the guanylyltransferase CofC. F420H2-deficient mutants displayed hypersusceptibility to malachite green (MG), a selective decontaminant present in solid media used to isolate and propagate mycobacteria from clinical samples. The wide diversity of mutations causing high-level pretomanid resistance and MG hypersusceptibility of most mutants poses significant challenges to clinical detection of nitroimidazole resistance using either genotypic or phenotypic methods.SignificanceNitroimidazole pro-drugs represent a promising new class of anti-tuberculosis drugs. Reliable methods to assure nitroimidazole susceptibility are critical to assure their optimal use. Yet, the spectrum of nitroimidazole resistance mutations remains incompletely characterized. Using 161 pretomanid-resistant Mycobacterium tuberculosis isolates selected in pretomanid-treated mice, we discovered a novel resistance determinant, Rv2983, required for cofactor F420 biosynthesis and characterized the remarkable diversity of mutations in this and 5 other genes involved in nitroimidazole activation. We show that F420H2–deficient nitroimidazole-resistant mutants are hypersusceptible to the selective decontaminant malachite green used in solid media to isolate mycobacteria and may evade detection on such media. These results have important implications for development and clinical use of genotypic and phenotypic methods for nitroimidazole susceptibility testing.
Remodelling of pSK1 Family Plasmids and Enhanced Chlorhexidine Tolerance in Methicillin-ResistantStaphylococcus aureus
Authors: Baines Sarah L , Jensen Slade O , Firth Neville , da Silva Anders Gonçalves , Seemann Torsten , Carter Glen , Williamson Deborah A. , Howden Benjamin P , Stinear Timothy P 
Published: October 2018

AbstractStaphylococcus aureusis a significant human pathogen whose evolution and adaptation has been shaped in part by mobile genetic elements (MGEs), facilitating global spread of extensive antimicrobial resistance. However, our understanding of the evolutionary dynamics surrounding MGEs is incomplete, in particular how changes in the structure of multidrug-resistant (MDR) plasmids may influence important staphylococcal phenotypes. Here, we undertook a population-and functional-genomics study of 212 methicillin-resistantS. aureus(MRSA) ST239 isolates collected over 32 years to explore the evolution of the pSK1 family of MDR plasmids, illustrating how these plasmids have co-evolved with and contributed to the successful adaptation of this persistent MRSA lineage. Using complete genomes and temporal phylogenomics we reconstructed the evolution of the pSK1 family lineage from its emergence in the late 1970s, with multiple structural variants arising. Plasmid maintenance and stability was linked to IS256- and IS257-mediated chromosomal integration and disruption of plasmid replication machinery. Overlaying genomic comparisons with phenotypic susceptibility data for gentamicin and chlorhexidine, it appeared that pSK1 has contributed to enhanced resistance in ST239 MRSA through two mechanisms: (i) acquisition of plasmid-borne resistance mechanisms increasing rates of gentamicin resistance and reduced chlorhexidine susceptibility, and (ii) changes in plasmid configuration linked with further enhancement of chlorhexidine tolerance. While the exact mechanism of enhanced tolerance remains elusive, this research has uncovered a clear evolutionary response of ST239 MRSA to chlorhexidine, one which may contribute to the ongoing persistence and adaptation of this lineage within healthcare institutions.
Integrating Molecular Simulation and Experimental Data: A Bayesian/Maximum Entropy reweighting approach
Authors: Bottaro Sandro , Bengtsen Tone , Lindorff-Larsen Kresten 
Published: October 2018

AbstractWe describe a Bayesian/Maximum entropy (BME) procedure and software to construct a conformational ensemble of a biomolecular system by integrating molecular simulations and experimental data. First, an initial conformational ensemble is constructed using for example Molecular Dynamics or Monte Carlo simulations. Due to potential inaccuracies in the model and finite sampling effects, properties predicted from simulations may not agree with experimental data. In BME we use the experimental data to refine the simulation so that the new conformational ensemble has the following properties: (i) the calculated averages are close to the experimental values taking uncertainty into account and (ii) it maximizes the relative Shannon entropy with respect to the original simulation ensemble. The output of this procedure is a set of optimized weights that can be used to calculate arbitrary properties and distributions. Here, we provide a practical guide on how to obtain and use such weights, how to choose adjustable parameters and discuss shortcomings of the method.
RNA is a critical element for the sizing and the composition of phase-separated RNA-protein condensates
Authors: Navarro Marina Garcia-Jove , Kashida Shunnichi , Chouaib Racha , Souquere Sylvie , Pierron Gerard , Weil Dominique , Gueroui Zoher 
Published: October 2018

SummaryLiquid-liquid phase separation is thought to be a key organizing principle in eukaryotic cells to generate highly concentrated dynamic assemblies, such as the RNP granules. Numerous in vitro approaches have validated this model, yet a missing aspect is to take into consideration the complex molecular mixture and promiscuous interactions found in vivo. Here we report the versatile scaffold “ArtiG” to generate concentration-dependent RNA-protein condensates within living cells, as a bottom-up approach to study the impact of co-segregated endogenous components on phase separation. We demonstrate that intracellular RNA seeds the nucleation of the condensates, as it provides molecular cues to locally coordinate the formation of endogenous high order RNP assemblies. Interestingly, the co-segregation of intracellular components ultimately impacts the size of the phase-separated condensates. Thus, RNA arises as an architectural element that can influence the composition and the morphological outcome of the condensate phases in an intracellular context.
A completeCannabischromosome assembly and adaptive admixture for elevated cannabidiol (CBD) content
Authors: Grassa Christopher J. , Wenger Jonathan P. , Dabney Clemon , Poplawski Shane G. , Motley S. Timothy , Michael Todd P. , Schwartz C.J. , Weiblen George D. 
Published: October 2018

AbstractCannabishas been cultivated for millennia with distinct cultivars providing either fiber and grain or tetrahydrocannabinol. Recent demand for cannabidiol rather than tetrahydrocannabinol has favored the breeding of admixed cultivars with extremely high cannabidiol content. Despite several draftCannabisgenomes, the genomic structure ofcannabinoid synthaseloci has remained elusive. A genetic map derived from a tetrahydrocannabinol/cannabidiol segregating population and a complete chromosome assembly from a high-cannabidiol cultivar together resolve the linkage ofcannabidiolicandtetrahydrocannabinolic acid synthasegene clusters which are associated with transposable elements. High-cannabidiol cultivars appear to have been generated by integrating hemp-typecannabidiolic acid synthasegene clusters into a background of marijuana-type cannabis. Quantitative trait locus mapping suggests that overall drug potency, however, is associated with other genomic regions needing additional study.Resources available online at:http://cannabisgenome.orgSummaryA complete chromosome assembly and an ultra-high-density linkage map together identify the genetic mechanism responsible for the ratio of tetrahydrocannabinol (THC) to cannabidiol (CBD) in Cannabis cultivars, allowing paradigms for the evolution and inheritance of drug potency to be evaluated.
Extracellular Electron Uptake by TwoMethanosarcinaSpecies
Authors: Yee Mon Oo , Snoeyenbos-West Oona , Thamdrup Bo , Ottosen Lars DM , Rotaru Amelia-Elena 
Published: October 2018

AbstractDirect electron uptake by prokaryotes is a recently described mechanism with a potential application for energy and CO2storage into value added chemicals. Members of Methanosarcinales, an environmentally and biotechnologically relevant group of methanogens, were previously shown to retrieve electrons from an extracellular electrogenic partner performing Direct Interspecies Electron Transfer (DIET) and were therefore proposed to be electroactive. However, their intrinsic electroactivity has never been examined. In this study, we tested two methanogens belonging to the genusMethanosarcina, M. barkeriandM. horonobensis,regarding their ability to accept electrons directly from insoluble electron donors like other cells, conductive particles and electrodes. Both methanogens were able to retrieve electrons fromGeobacter metallireducensvia DIET. Furthermore, DIET was also stimulated upon addition of electrically conductive granular activated carbon (GAC) when each was co-cultured withG. metallireducens. However, when provided with a cathode poised at −400 mV (vs. SHE), onlyM. barkericould perform electromethanogenesis. In contrast, the strict hydrogenotrophic methanogen,Methanobacterium formicicum, did not produce methane regardless of the type of insoluble electron donor provided (Geobactercells, GAC or electrodes). A comparison of functional gene categories between the twoMethanosarcinashowed differences regarding energy metabolism, which could explain dissimilarities concerning electromethanogenesis at fixed potentials. We suggest that these dissimilarities are minimized in the presence of an electrogenic DIET partner (e.g.Geobacter), which can modulate its surface redox potentials by adjusting the expression of electroactive surface proteins.
The Cten signalling pathway stabilises Src protein to promote Epithelial-Mesenchymal Transition (EMT) in colorectal cancer
Authors: Asiri Abdulaziz , Toss Michael S , Pereira Raposo Teresa , Akhlaq Maham , Thorpe Hannah , Alfahed Abdulaziz , Asiri Abutaleb , Ilyas Mohammad 
Published: October 2018

AbstractCten is an oncogene which promotes epithelial-mesenchymal transition (EMT) in many signalling pathways. Having previously shown that Cten promotes EMT through Snail, we investigated whether Cten function could be mediated through Src (a known regulator of Snail).Cten levels were modulated by forced expression in colorectal cancer (CRC) cell lines with low Cten expression (HCT116 and RKO) and gene knockdown in a cell line with high Cten expression (SW620). In all cell lines, Cten was a positive regulator of Src expression. The functional importance of Src was tested by forcibly expressing Cten and simultaneously knocking down Src. This resulted in abrogation of Cten motility-inducing activity (cell migration, cell invasion, wound healing – each p<0.001) and abrogation of the promotion of colony formation by Cten (p<0.001) together with failure to induce the Cten targets - Snail and ROCK1. To complement these experiments, Cten expression was restored by forced expression in a subclone of SW620 in which the Cten gene had been deleted (SW620ΔCten). SW620ΔCten showed reduced expression of Src which increased following restoration of Cten by forced expression. In SW620ΔCten, restoration of Cten increased cell motility (cell migration, cell invasion, wound healing) and colony formation (each p<0.001) which were all lost if Src was concomitantly knocked down. Quantitative Reverse-Transcription PCR (qRT-PCR) showed that modulation of Cten had no effect on Src mRNA levels. However, a cycloheximide (CHX) pulse chase assay demonstrated stabilisation of Src protein by Cten. Finally, the expression of Cten and Src was tested in a series of 84 primary CRCs and there was significant correlation between Cten and Src expression (p=0.001).We conclude that Src is a novel and functionally important target of the Cten signalling pathway and that Cten protein causes post-transcriptional stabilisation of Src protein in order to promote EMT and possibly metastasis in CRC.
A new class of constitutively active super-enhancers is associated with fast recovery of 3D chromatin loops
Authors: Ryu Jayoung , Kim Hyunwoong , Yang Dongchan , Lee Andrew J. , Jung Inkyung 
Published: October 2018

AbstractSuper-enhancers or stretch enhancers are clusters of active enhancers that often coordinate cell-type specific gene regulation. However, little is known about the function of super-enhancers beyond gene regulation. In this study, through a comprehensive analysis of super-enhancers in 30 human cell/tissue types, we identified a new class of super-enhancers which are constitutively active across most cell/tissue types. These ‘common’ super-enhancers are associated with universally highly expressed genes in contrast to the canonical definition of super-enhancers that assert cell-type specific gene regulation. In addition, the genome sequence of these super-enhancers is highly conserved by evolution and among humans, advocating their universal function in genome regulation. Integrative analysis of 3D chromatin loops demonstrates that, in comparison to the cell-type specific super-enhancers, the cell-type common super-enhancers present a striking association with rapidly recovering loops. We propose that a new class of super-enhancers may play an important role in the early establishment of 3D chromatin structure.BackgroundSuper-enhancers or stretch enhancers are defined by a strong enrichment of mediators and transcription-regulating proteins, appearing to play a deterministic role in cellular identity by controlling the expression of cell-type specific genes[1, 2]. Previous studies have revealed the critical function of super-enhancers during development and differentiation[3]. The enrichment of disease-associated single nucleotide polymorphism (SNP) in super-enhancers compared to that of typical enhancers proposed a substantial link between super-enhancers and many complex human diseases[2]. In addition, a set of recent studies have proposed potential functions of super-enhancers in the extremely long-range chromatin communications and the establishment of 3D chromatin loops[4]. These results suggest a more universal role of super-enhancers in genome regulation apart from cell-type specific gene regulation, but little is known about the mechanisms underlying these various functions. To extend the current knowledge of super-enhancers and their biological roles, we conducted a comprehensive analysis of super-enhancer activities across 30 human cell/tissue types. Our analysis suggests that a substantial number of super-enhancers exhibits prevalent activities across cell-types in terms of H3K27ac signals, and that these non-canonical super-enhancers are involved in the formation of fast recovering chromatin loops.NoteA genome browser session has been set up for visualization of the super-enhancer domains described in the current study– https://genome.ucsc.edu/cgi-bin/hgTracks?hgS-doOtherUser=submit&hgS-otherUserName=abundantiavosliberabit&hgS-otherUserSessionName=SuperEnhancerDomain
Genome-wide data for effective conservation of manta and devil ray species
Authors: Hosegood Jane , Humble Emily , Ogden Rob , de Bruyn Mark , Creer Si , Stevens Guy , Abudaya Mohammed , Bassos-Hull Kim , Bonfil Ramon , Fernando Daniel , Foote Andrew D. , Hipperson Helen , Jabado Rima W. , Kaden Jennifer , Moazzam Muhammad , Peel Lauren , Pollett Stephen , Ponzo Alessandro , Poortvliet Marloes , Salah Jehad , Senn Helen , Stewart Joshua , Wintner Sabine , Carvalho Gary 
Published: October 2018

AbstractPractical biodiversity conservation relies on delineation of biologically meaningful units, particularly with respect to global conventions and regulatory frameworks. Traditional approaches have typically relied on morphological observation, resulting in artificially broad delineations and non-optimal species units for conservation. More recently, species delimitation methods have been revolutionised with High-Throughput Sequencing approaches, allowing study of diversity within species radiations using genome-wide data. The highly mobile elasmobranchs, manta and devil rays (Mobulaspp.), are threatened globally by targeted and bycatch fishing pressures resulting in recent protection under several global conventions. However, a lack of global data, morphological similarities, a succession of recent taxonomic changes and ineffectual traceability measures combine to impede development and implementation of a coherent and enforceable conservation strategy. Here, we generate genome-wide Single Nucleotide Polymorphism (SNP) data from among the most globally and taxonomically representative set of mobulid tissues. The resulting phylogeny and delimitation of species units represents the most comprehensive assessment of mobulid diversity with molecular data to date. We find a mismatch between current species classifications, and optimal species units for effective conservation. Specifically, we find robust evidence for an undescribed species of manta ray in the Gulf of Mexico and show that species recently synonymised are reproductively isolated. Further resolution is achieved at the population level, where cryptic diversity is detected in geographically distinct populations, and indicates potential for future traceability work determining regional location of catch. We estimate the optimal species tree and uncover substantial incomplete lineage sorting, where standing variation in extinct ancestral populations is identified as a driver of phylogenetic uncertainty, with further conservation implications. Our study provides a framework for molecular genetic species delimitation that is relevant to wide-ranging taxa of conservation concern, and highlights the potential for genomic data to support effective management, conservation and law enforcement strategies.
BRK Phosphorylates SMAD4 for proteasomal degradation and inhibits tumor suppressor FRK to control SNAIL, SLUG and metastatic potential
Authors: Miah Sayem , Banks Charles A. S. , Ogunbolude Yetunde , Bagu Edward T. , MacAusland-Berg Josh , Saraf Anita , Hattem Gaye , Kempf Cassandra G. , Sardiu Mihaela , Napper Scott , Florens Laurence , Lukong Kiven E. , Washburn Michael P. 
Published: October 2018

AbstractThe tumor-suppressing function of SMAD4 is frequently subverted during mammary tumorigenesis, leading to cancer growth, invasion, and metastasis. A long-standing concept is that SMAD4 is not regulated by phosphorylation but ubiquitination. Interestingly, our search for signaling pathways regulated by BRK, a non-receptor protein tyrosine kinase that is up-regulated in ∼80% of invasive ductal breast tumors, led us to discover that BRK competitively binds and phosphorylates SMAD4, and regulates TGF-β/ SMAD4 signaling pathway. A constitutively active BRK (BRK-Y447F), phosphorylates SMAD4 resulting in its recognition by the ubiquitin-proteasome system, which accelerates SMAD4 degradation. In agreement, we also observed an inverse protein expression pattern of BRK and SMAD4 in a panel of breast cancer cell lines and breast tumors. Activated BRK mediated degradation of SMAD4 causes the repression of tumor suppressor genes FRK that was associated with increased expression of mesenchymal markers and decreased cell adhesion ability. Thus, our data suggest that combination therapies targeting activated BRK signaling may have synergized the benefits in the treatment of SMAD4 repressed cancers. Therefore, our data propose that combination therapies which includes targeting activated BRK signaling may synergize the benefits in the treatment of SMAD4 deficient cancers.