bluuebunny/crossref_metadata_2025_split huggingface dataset:
Pervasive head-to-tail insertions of DNA templates mask desired CRISPR/Cas9-mediated genome editing events
Authors: Skryabin Boris V. , Gubar Leonid , Seeger Birte , Kaiser Helena , Stegemann Anja , Roth Johannes , Meuth Sven G. , Pavenstädt Hermann , Sherwood Joanna , Pap Thomas , Wedlich-Söldner Roland , Sunderkötter Cord , Schwartz Yuri B. , Brosius Juergen , Rozhdestvensky Timofey S. 
Published: March 2019

AbstractCRISPR/Cas9 mediated homology-directed DNA repair is the method of choice for precise gene editing in a wide range of model organisms, including mouse and human. Broad use by the biomedical community refined the method, making it more efficient and sequence specific. Nevertheless, the rapidly evolving technique still contains pitfalls. During the generation of six different conditional knock-out mouse models, we discovered that frequently (sometimes solely) homology-directed repair and/or non-homologous end-joining mechanisms caused multiple unwanted head-to-tail insertions of donor DNA templates. Disturbingly, conventionally applied PCR analysis—in most cases—failed to identify such multiple integration events, which led to a high rate of falsely claimed precisely edited alleles. We caution that comprehensive analysis of modified alleles is essential, and offer practical solutions to correctly identify precisely edited chromosomes.
The Neural Basis of Shared Preference Learning
Authors: Farmer Harry , Hertz Uri , Hamilton Antonia 
Published: March 2019

AbstractDuring our daily lives, we often learn about the similarity of the traits and preferences of others to our own and use that information during our social interactions. However, it is unclear how the brain represents similarity between the self and others. One possible mechanism is to track similarity to oneself regardless of the identity of the other (Similarity account); an alternative is to track each confederate in terms of consistency of the similarity to the self, with respect to the choices they have made before (consistency account). Our study combined fMRI and computational modelling of reinforcement learning (RL) to investigate the neural processes that underlie learning about preference similarity. Participants chose which of two pieces of artwork they preferred and saw the choices of one confederate who usually shared their preference and another who usually did not. We modelled neural activation with RL models based on the similarity and consistency accounts. Data showed more brain regions whose activity pattern fits with the consistency account, specifically, areas linked to reward and social cognition. Our findings suggest that impressions of other people can be calculated in a person-specific manner which assumes that each individual behaves consistently with their past choices.
The order of trait emergence in the evolution of cyanobacterial multicellularity
Authors: Hammerschmidt Katrin , Landan Giddy , Tria Fernando Domingues Kümmel , Alcorta Jaime , Dagan Tal 
Published: March 2019

AbstractThe transition from unicellular to multicellular organisms is one of the most significant events in the history of life. Key to this process is the emergence of Darwinian individuality at the higher level: groups must become single entities capable of reproduction for selection to shape their evolution. Evolutionary transitions in individuality are characterized by cooperation between the lower level entities and by division of labor. Theory suggests that division of labor may drive the transition to multicellularity by eliminating the trade-off between two incompatible processes that cannot be performed simultaneously in one cell. Here we examine the evolution of the most ancient multicellular transition known today, that of cyanobacteria, where we reconstruct the sequence of ecological and phenotypic trait evolution. Our results show that the prime driver of multicellularity in cyanobacteria was the expansion in metabolic capacity offered by nitrogen fixation, which was accompanied by the emergence of the filamentous morphology and succeeded by a reproductive life cycle. This was followed by the progression of multicellularity into higher complexity in the form of differentiated cells and patterned multicellularity.Significance StatementThe emergence of multicellularity is a major evolutionary transition. The oldest transition, that of cyanobacteria, happened more than 3 to 3.5 billion years ago. We find N2 fixation to be the prime driver of multicellularity in cyanobacteria. This innovation faced the challenge of incompatible metabolic processes since the N2 fixing enzyme (nitrogenase) is sensitive to oxygen, which is abundantly found in cyanobacteria cells performing photosynthesis. At the same time, N2-fixation conferred an adaptive benefit to the filamentous morphology as cells could divide their labour into performing either N2-fixation or photosynthesis. This was followed by the culmination of complex multicellularity in the form of differentiated cells and patterned multicellularity.
Genomic analysis reveals shared genes and pathways in human and canine angiosarcoma
Authors: Megquier Kate , Turner-Maier Jason , Swofford Ross , Kim Jong-Hyuk , Sarver Aaron L. , Wang Chao , Sakthikumar Sharadha , Johnson Jeremy , Koltookian Michele , Lewellen Mitzi , Scott Milcah C. , Graef Ashley J. , Borst Luke , Tonomura Noriko , Alfoldi Jessica , Painter Corrie , Thomas Rachael , Karlsson Elinor K. , Breen Matthew , Modiano Jaime F. , Elvers Ingegerd , Lindblad-Toh Kerstin 
Published: March 2019

AbstractAngiosarcoma is a highly aggressive cancer of blood vessel-forming cells with high fatality and few effective treatment options. It is both rare and heterogenous, making large, well powered genomic studies nearly impossible. In dogs, angiosarcoma is common, with breeds like the golden retriever carrying heritable genetic factors that put them at very high risk. If the clinical similarity of canine and human angiosarcoma reflects shared genomic etiology, dogs could be a critically needed model for advancing angiosarcoma research. We assessed the genomic landscape of canine angiosarcoma via whole exome sequencing (47 golden retriever angiosarcomas) and RNA sequencing (74 angiosarcomas from multiple breeds). The predominant mutational signature was the age-associated deamination of cytosine to thymine, and somatic coding mutations occurred most frequently in the tumor suppressorTP53(59.6% of cases) as well as two genes in the PI3K pathway: the oncogenePIK3CA(29.8%) and its regulatory subunitPIK3R1(8.5%). We compared the canine data to human data recently released by The Angiosarcoma Project, and found the same genes and many of the same pathways significantly enriched for somatic mutations, most notably protein kinases, glycoproteins, fibronectin Type III domains, EGF-like domains, and cell adhesion proteins such as cadherins. As in human angiosarcoma,CDKN2A/Bwas recurrently deleted andVEGFA, KDR, and KITrecurrently gained. Canine angiosarcoma closely models human angiosarcoma on a genomic level, and is a powerful tool for investigating the pathogenesis of this devastating disease.
Designing a novel multi-epitope T vaccine for “targeting protein for Xklp-2” (TPX2) in hepatocellular carcinoma based on immunoinformatics approach
Authors: Ghahremanifard Parisa , Afzali Farzaneh , Rostami Amin , Nayeri Zahra , Bambai Bijan , Minuchehr Zarrin 
Published: March 2019

AbstractHepatocellular carcinoma (HCC) is one of the leading cancer-related deaths worldwide. Recently, studies for HCC treatment are focused on cancer immunotherapy, particularly cancer vaccines, to complete and assist other therapies. TPX2 is a microtubule-associated protein necessary for cell division; therefore, alteration in its expression, especially up regulation, is associated with several human carcinomas such as HCC.In this study, immunoinformatics tools were used to design a rational multi-epitope T vaccine against TPX2 in HCC. Cytotoxic T lymphocytes (CTL) and Helper T lymphocytes (HTL) epitopes were predicted and Maltose-binding protein (MBP) was added to the construct as an adjuvant. Evaluation of vaccine properties was indicated that our construct is stable and immunogenic enough to induce relevant responses besides not being allergic. After predicting the tertiary structure and energy minimization, protein-protein docking was performed to calculate the free energy of possible interactions between the vaccine and toll-like receptor 4 (TLR4) to assure that simultaneous complementary responses would be activated by our construct. Finally, Codon optimization and in-silico cloning were performed to ensure the vaccine expression efficiency in the desired host.
Genetic determinants inSalmonella entericaserotype Typhimurium required for overcoming stressors in the host environment
Authors: Mandal Rabindra K. , Jiang Tieshan , Min Kwon Young 
Published: March 2019

AbstractSalmonella entericaserovar Typhimurium (S. Typhimurium), a non-typhoidalSalmonella(NTS), result in a range of diseases, including self-limiting gastroenteritis, bacteremia, enteric fever, and focal infections representing a major disease burden worldwide. There is still a significant portion ofSalmonellagenes whose functional basis to overcome host innate defense mechanisms, consequently causing disease in host, largely remains unknown. Here, we have applied a high-throughput transposon sequencing (Tn-seq) method to unveil the genetic factors required for the growth or survival of S. Typhimurium under various host stressors simulatedin vitro. A highly saturating Tn5 library ofS. Typhimurium 14028s was subjected to selection during growth in the presence of short chain fatty acid (100 mM propionate), osmotic stress (3% NaCl) or oxidative stress (1 mM H2O2) or survival in extreme acidic pH (30 min in pH3) or starvation (12 days in 1X PBS). We have identified an overlapping set of 339 conditionally essential genes (CEGs) required byS. Typhimurium to overcome these host insults. Interestingly, entire eight genes encoding F0F1-ATP synthase subunit proteins were required for fitness in all five stresses. Intriguingly, total 88 genes inSalmonellapathogenicity island (SPI), including SPI-1, SPI-2, SPI-3, SPI-5, SPI-6 and SPI-11 are also required for fitness under thein vitroconditions evaluated in this study. Additionally, by comparative analysis of the genes identified in this study and the genes previously shown to be required forin vivofitness, we identified novel genes (marBCT,envF,barA,hscA,rfaQ,rfbIand putative proteins STM14_1138, STM14_3334, STM14_4825, and STM_5184) that has compelling potential to be exploited as vaccine development and/or drug target to curb theSalmonellainfection.
Fear Learning Induces Long-Lasting Changes in Gene Expression and Pathway Specific Presynaptic Growth
Authors: Dillingham Blythe C. , Cameron Peter , Pieraut Simon , Cardozo Leonardo M. , Yoo Eun J. , Maximov Anton , Stowers Lisa , Mayford Mark 
Published: March 2019

AbstractThe stabilization or consolidation of long-term memories lasting more than a few hours requires new gene expression. While neural activity has been shown to induce expression of a variety of genes within several hours of learning, whether this leads to persistent changes in gene expression that lasts for days or weeks remains unclear. We developed a novel mouse line which expresses Cre recombinase in an inducible manner and used it to examine gene expression in learning-activated neurons of the medial prefrontal cortex (mPFC) one month following contextual fear conditioning. The mPFC is not required for the initial retrieval of contextual memory but becomes necessary after one month, suggesting a slowly developing plasticity. We found a variety of changes in gene expression in learning-activated neural ensembles that were specific to the mPFC. One group of transcriptional changes observed was the coordinated upregulation of presynaptic proteins suggesting a potential learning-induced elaboration of presynaptic terminals. We tested this idea by labeling the projections of mPFC neurons active during initial learning and found an increase in the number of terminals in neurons projecting to the basolateral amygdala at 1 month following training. These results suggest a presynaptic growth mechanism that could account for the enhanced role of the mPFC in fear memory retrieval at long time points after learning.
Protein kinase B controls Mycobacterium tuberculosis growth via phosphorylation of the global transcriptional regulator Lsr2
Authors: Alqaseer Kawther , Turapov Obolbek , Barthe Philippe , Jagatia Heena , De Visch Angélique , Roumestand Christian , Wegrzyn Malgorzata , Bartek Iona L. , Voskuil Martin I. , O’Hare Helen , Witney Adam A. , Cohen-Gonsaud Martin , Waddell Simon J. , Mukamolova Galina V. 
Published: March 2019

ABSTRACTMycobacterium tuberculosis is able to persist in the body through months of multi-drug therapy. Mycobacteria possess a wide range of regulatory proteins, including the essential protein kinase B (PknB), that control transitions between growth states. Here, we establish that depletion of PknB in replicating M. tuberculosis results in transcriptional adaptations that implicate the DNA-binding protein Lsr2 in coordinating these changes. We show that Lsr2 is phosphorylated by PknB, and that phosphorylation of Lsr2 at threonine 112 is important for M. tuberculosis growth and survival under hypoxic conditions. Fluorescence anisotropy and electrophoretic mobility shift assays demonstrate that phosphorylation reduces Lsr2 binding to DNA, and ChIP-sequencing confirms increased DNA binding of a phosphoablative (T112A) Lsr2 mutant in M. tuberculosis. Altered expression of target genes in T112A Lsr2 compared to wild type Lsr2 M. tuberculosis offers further evidence that phosphorylation mediates expression of the Lsr2 regulon. Structural studies reveal increased dynamics of the Lsr2 DNA binding domain from a T112D phosphomimetic Lsr2 mutant, providing a molecular basis for decreased DNA binding by phosphorylated Lsr2. Our findings suggest that, the essential protein kinase, PknB controls M. tuberculosis growth and adaptations to the changing host environment by phosphorylating the global transcriptional regulator Lsr2.
An atlas of anteriorhoxgene expression in the embryonic sea lamprey head:hox-code evolution in vertebrates
Authors: Parker Hugo J. , Bronner Marianne E. , Krumlauf Robb 
Published: March 2019

AbstractIn the hindbrain and the adjacent cranial neural crest (NC) cells of jawed vertebrates (gnathostomes), nested and segmentally-restricted domains ofHoxgene expression provide a combinatorialHox-code for specifying regional properties during head development. Extant jawless vertebrates, such as the sea lamprey(Petromyzon marinus),can provide insights into the evolution and diversification of thisHox-code in vertebrates. There is evidence for gnathostome-like spatial patterns ofHoxexpression in lamprey; however, the expression domains of the majority of lampreyhoxgenes from paralogy groups (PG) 1-4 are yet to be characterized, so it is unknown whether they are coupled to hindbrain segments (rhombomeres) and NC. In this study, we systematically describe the spatiotemporal expression of all 14 sea lampreyhoxgenes from PG1-PG4 in the developing hindbrain and pharynx to investigate the extent to which their expression conforms to the archetypal gnathostome hindbrain and pharyngealhox-codes. We find many similarities inHoxexpression between lamprey and gnathostome species, particularly in rhombomeric domains during hindbrain segmentation and in the cranial neural crest, enabling inference of aspects ofHoxexpression in the ancestral vertebrate embryonic head. These data are consistent with the idea that aHoxregulatory network underlying hindbrain segmentation is a pan vertebrate trait. We also reveal differences in hindbrain domains at later stages, as well as expression in the endostyle and in pharyngeal arch (PA) 1 mesoderm. Our analysis suggests that manyHoxexpression domains that are observed in extant gnathostomes were present in ancestral vertebrates but have been partitioned differently acrossHoxclusters in gnathostome and cyclostome lineages after duplication.
Functional and signaling characterization of the neutrophil FPR2 selective agonist Act-389949
Authors: Lind Simon , Sundqvist Martina , Holmdahl Rikard , Dahlgren Claes , Forsman Huamei , Olofsson Peter 
Published: March 2019

AbstractDespite the steadily increased numbers of formyl peptide receptor (FPR) ligands identified over the years, few have been characterized in studies using animal disease models and even less have entered clinical trials in human subjects. A small-molecule compound, Act-389949, was however recently tested in a phase I clinical trial and found to be safe and well tolerated in healthy human subjects. The desired anti-inflammatory property of Act-389949 was proposed to be mediated through FPR2, one of the FPRs expressed in neutrophils, but no basic characterization was included in the study. To gain more insights into FPR2 recognition of this first-in-class compound for future utility of the agonist, we have in this study determined the receptor preference and down-stream signaling characteristics induced by Act-389949 in human blood neutrophils isolated from healthy donors. Our data demonstrate that Act-389949 is an agonist for FPR2 that triggers functional/signaling repertoires comparable to what has been earlier described for other FPR2 agonists, including neutrophil chemotaxis, granule mobilization and activation of the NADPH-oxidase. In fact, Act-389949 was found to be as potent as the prototype FPR2 peptide agonist WKYMVM and had the advantage of being resistant to oxidation by the MPO-H2O2-halide derived oxidants, as compared to the sensitive WKYMVM. The down-stream signals generated by Act-389949 include an FPR2-dependent and Gαq-independent transient rise in intracellular Ca2+and recruitment of β-arrestin. In summary, our data show that Act-389949 serves as an excellent tool-compound for further dissection of FPR2-regulated activitiesin vitroandin vivo. Potent and stable FPR ligands such as Act-389949 may therefore be used to develop the next generation of FPR signaling regulating anti-inflammatory therapeutics.
Fast dose fractionation using ultra-short laser accelerated proton pulses can increase cancer cell mortality, which relies on functional PARP1 protein
Authors: Bayart E. , Flacco A. , Delmas O. , Pommarel L. , Levy D. , Cavallone M. , Megnin-Chanet F. , Deutsch E. , Malka V. 
Published: March 2019

AbstractRadiotherapy is a cornerstone of cancer management. The improvement of spatial dose distribution in the tumor volume by minimizing the dose deposited in the healthy tissues have been a major concern during the last decades. Temporal aspects of dose deposition are yet to be investigated. Laser-plasma-based particle accelerators are able to emit pulsed-proton beams at extremely high peak dose rates (~109Gy/s) during several nanoseconds. The impact of such dose rates on resistant glioblastoma cell lines, SF763 and U87-MG, was compared to conventionally accelerated protons and X-rays. No difference was observed in DNA double-strand breaks generation and cells killing. The variation of the repetition rate of the proton bunches produced an oscillation of the radio-induced cell susceptibility in HCT116 cells, which appeared to be related to the presence of the PARP1 protein and an efficient parylation process. Interestingly, when laser-driven proton bunches were applied at 0.5 Hz, survival of the radioresistant HCT116 p53−/−cells equaled that of its radiosensitive counterpart, HCT116 WT, which was also similar to cells treated with the PARP1 inhibitor Olaparib. Altogether, these results suggest that the application modality of ultrashort bunches of particles could provide a great therapeutic potential in radiotherapy.
β-Catenin tumour-suppressor activity depends on its ability to promote Pro-N-Cadherin maturation
Authors: Herrera Antonio , Menendez Anghara , Torroba Blanca , Pons Sebastian 
Published: March 2019

SUMMARYNeural stem cells (NSCs) form a pseudostratified, single-cell layered epithelium with a marked apico-basal polarity. In these cells, β-Catenin associates with classic cadherins in order to form the apical adherens junctions (AJs). We previously reported that oncogenic forms of β-Catenin (sβ-Catenin) maintain neural precursors as progenitors, while also enhancing their polarization and adhesiveness, thereby limiting their malignant potential. Here we show that β-Catenin can bind to phosphorylated Pro-N-Cadherin, promoting the excision of the propeptide and its maturation into N-Cadherin in the trans-Golgi network (TGN). Moreover, β-Catenin-assisted maturation of Pro-N-Cadherin is required for the formation of the AJs and for them to recruit other apical complex (AC) components like aPKC, and accordingly, to establish apico-basal polarity. Notably, we show that NSCs expressing unprocessed Pro-N-Cadherin invade the ventricle and they breach the basement membrane to invade the surrounding mesenchyme. Hence, we propose that the tumour-suppressor activity of sβ-Catenin depends on it promoting Pro-N-Cadherin processing.
An evolutionary-conserved redox regulatory mechanism in human Ser/Thr protein kinases
Authors: Byrne Dominic P. , Shrestha Safal , Kannan Natarajan , Eyers Patrick A. 
Published: March 2019

ABSTRACTReactive oxygen species (ROS) are products of oxygen metabolism, but are also recognized as endogenous physiological mediators of cellular signaling. Eukaryotic protein kinase (ePK) regulation occurs through reversible phosphorylation events in the flexible activation segment. In this study, we demonstrate that the catalytic phosphotransferase output from the mitotic Ser/Thr kinase Aurora A is also controlled by cysteine (Cys) oxidation. Reversible regulation occurs by direct modification of a conserved residue (Cys 290), which lies adjacent to Thr 288, the activating site of phosphorylation. Strikingly, redox modulation of the Cys 290-equivalent in other ePKs is predicted to be an underappreciated regulatory mechanism, since ~100 human Ser/Thr kinases possess a Cys at this position in the conserved activation loop. Using real-time enzyme assays, we confirm that the presence of the equivalent Cys residue is prognostic for redox-sensitivity amongst a cohort of human CAMK, AGC and AGC-like kinases, including AKT, AMPK, CAMK1, MAPKAP-K2/3 and SIK1-3. Our findings demonstrate that dominant Cys-based redox-switching in the activation segment represents an evolutionary-conserved mode of regulation for a significant subset of the human kinome. This finding has important implications for understanding physiological and pathological signaling responses to ROS, and emphasises the importance of multivalent activation segment regulation in ePKs.ONE-SENTENCE SUMMARYThe catalytic activity of Ser/Thr kinases is regulated through a conserved Cys-based redox mechanism.