bluuebunny/crossref_metadata_2025 huggingface dataset:
Single-cell resolution view of the transcriptional landscape of developingDrosophilaeye
Authors: Ejsmont Radoslaw Kamil , Houser Grace , Garcia Natalia Mora , Topp Sara Fonseca , Danda Natalia , Wong-Chung Agnes , Hassan Bassem A.
Published: September 2019

Faithful and reliable quantification of gene expression at a single-cell level is an outstanding challenge in developmental biology. Most existing approaches face a trade-off between the signal to noise ratio, resolution, and sensitivity. Here, we present a novel approach for in situ quantification of gene expression in a developing tissue. Our pipeline combines computational prediction of transcription factor targets, gene tagging, fluorescent reporter imaging, state-of-the-art image analysis, and automated cell-type identification. By applying this approach to identify the sequence of quantitative changes in gene expression which govern the development of theDrosophilaneural retina, we demonstrate the feasibility of our method. We analyze the targets of Atonal (Ato), a transcription factor that controls the transition from eye disc progenitor cell to photoreceptor neurons. We utilized recombineering and genomic engineering to tag all predicted Ato targets with novel transcriptional reporters. These reporters enable following the expression of both regulator and regulated genes to accurately quantify their expression levels in individual cells. Our complete computational pipeline identifies nuclei in the eye discs and detects different states of cells as they progress through differentiation. Based on detailed gene expression analysis, our technique revealed genes likely to be direct Ato targets and provided insight into how gene expression changes drive the specification of photoreceptors.
To infect or not to infect: molecular determinants of bacterial outer membrane vesicle internalization by host membranes
Authors: Jefferies Damien , Khalid Syma
Published: September 2019

Outer membrane vesicles (OMVs) are spherical liposomes that are secreted by almost all forms of Gram-negative bacteria. The nanospheres contribute to bacterial pathogenesis by trafficking molecular cargo from bacterial membranes to target cells at the host-pathogen interface. We have simulated the interaction of OMVs with host cell membranes to understand why OMV uptake depends on the length of constituent lipopolysaccharide macromolecules. Using coarse-grained molecular dynamics simulations, we show that lipopolysaccharide lipid length affects OMV shape at the host-pathogen interface: OMVs with long (smooth-type) lipopolysaccharide lipids retain their spherical shape when they interact with host cell membranes, whereas OMVs with shorter (rough-type) lipopolysaccharide lipids distort and spread over the host membrane surface. In addition, we show that OMVs preferentially coordinate domain-favoring ganglioside lipids within host membranes to enhance curvature and affect the local lipid composition. We predict that these differences in shape preservation affect OMV internalization on long timescales: spherical nanoparticles tend to be completely enveloped by host membranes, whereas low sphericity nanoparticles tend to remain on the surface of cells.
Structural and functional analysis of genes with potential involvement in resistance to coffee leaf rust: a functional marker based approach
Authors: Barka Geleta Dugassa , Caixeta Eveline Teixeira , Ferreira Sávio Siqueira , Zambolim Laércio
Published: September 2019

Physiology-based differentiation of SHgenes andHemileia vastatrixraces is the principal method employed for the characterization of coffee leaf rust resistance. Based on the gene-for-gene theory, nine major rust resistance genes (SH1-9) have been proposed. However, these genes have not been characterized at the molecular level. Consequently, the lack of molecular data regarding rust resistance genes or candidates is a major bottleneck in coffee breeding. To address this issue, we screened a BAC library with resistance gene analogs (RGAs), identified RGAs, characterized and explored for any SHrelated candidate genes. Herein, we report the identification and characterization of a gene (gene 11), which shares conserved sequences with other SHgenes and displays a characteristic polymorphic allele conferring different resistance phenotypes. Furthermore, comparative analysis of the two RGAs belonging to CC-NBS-LRR revealed more intense diversifying selection in tomato and grape genomes than in coffee. For the first time, the present study has unveiled novel insights into the molecular nature of the SHgenes, thereby opening new avenues for coffee rust resistance molecular breeding. The characterized candidate RGA is of particular importance for further biological function analysis in coffee.
Long-term potentiation prevents ketamine-induced aberrant neurophysiological dynamics in the hippocampus-prefrontal cortex pathway in vivo
Authors: Lopes-Aguiar Cleiton , Ruggiero Rafael N. , Rossignoli Matheus T. , de Miranda Esteves Ingrid , Santos José Eduardo Peixoto , Romcy-Pereira Rodrigo N. , Leite João P.
Published: September 2019

N-methyl-D-aspartate receptor (NMDAr) antagonists such as ketamine (KET) produce psychotic-like behavior in both humans and animal models. NMDAr hypofunction affects normal oscillatory dynamics and synaptic plasticity in key brain regions related with schizophrenia, particularly in the hippocampus and the prefrontal cortex. In contrast, long-term potentiation (LTP) induction is known to increase glutamatergic transmission. Thus, we hypothesized that LTP could mitigate the electrophysiological changes promoted by KET. We recorded HPC-PFC local field potentials and evoked responses in urethane anesthetized rats, before and after KET administration, preceded or not by LTP induction. Our results show that KET promotes an aberrant delta-high-gamma crossfrequency coupling in the PFC and an enhancement in HPC-PFC evoked responses. LTP induction prior to KET attenuates changes in synaptic efficiency and prevents the increase in cortical gamma amplitude comodulation. These findings are consistent with evidence that increased efficiency of glutamatergic receptors attenuates cognitive impairment in animal models of psychosis. Therefore, high-frequency stimulation in HPC may be a useful tool to better understand how to prevent NMDAr hypofunction effects on synaptic plasticity and oscillatory coordination in cortico-limbic circuits.
Systematical analysis reveals the novel function of Cyp2c29 in liver injury
Authors: Wang Qi , Tang Qin , Zhao Lijun , Zhang Qiong , Wu Yuxin , Hu Hui , Liu Lan-Lan , Liu Xiang , Zhu Yanhong , Guo An-Yuan , Yang Xiangliang
Published: September 2019

As a severe lethal cancer, hepatocellular carcinoma (HCC) usually originates from chronic liver injury and inflammation, in which the discovery of key genes is important for HCC prevention. Here, we analyzed the time serial (from 0 week to 30 weeks) transcriptome data of liver injury samples in diethylnitrosamine (DEN)-induced HCC mouse model. Through expression and function analyses, we identified that Cyp2c29 was a key gene continuously downregulated during liver injury. Overexpression of Cyp2c29 suppressed the NF-κB activation, proinflammatory cytokine production and hepatocyte proliferation by increasing its production 14,15-epoxyeicosatrienoic acid (14,15-EET). Furthermore, in vivo Cyp2c29 protected against liver inflammation in liver injury mice models by reversing the expression on functions of cell proliferation, metabolism and inflammation including suppressing NF-κB pathway and compensatory proliferation. CYP2C8 and CYP2C9, two human homologs of mouse Cyp2c29, were decreased in human HCC progression and positively correlated with HCC patient survival. Therefore, through systematical analysis and verification, we identified that Cyp2c29 is a novel gene in liver injury and inflammation, which may be a potential biomarker for HCC prevention and prognosis.
Co-ordinated Ras and Rac activity shapes macropinocytic cups and enables phagocytosis of geometrically diverse bacteria
Authors: Buckley Catherine M. , Pots Henderikus , Gueho Aurelie , Phillips Ben A. , Gilsbach Bernd , Traynor David , Nikolaev Anton , Soldati Thierry , Parnell Andrew J. , Kortholt Arjan , King Jason S.
Published: September 2019

Engulfment of extracellular material by phagocytosis or macropinocytosis depends on the ability of cells to generate specialised cup shaped protrusions. To effectively capture and internalise their targets, these cups are organised into a ring or ruffle of actin-driven protrusion encircling a non-protrusive interior domain. These functional domains depend on the combined activities of multiple Ras and Rho family small GTPases, but how their activities are integrated and differentially regulated over space and time is unknown. Here, we show that the amoeba Dictyostelium discoideum coordinates Ras and Rac activity using the multidomain protein RGBARG (RCC1, RhoGEF, BAR and RasGAP-containing protein). We find RGBARG uses a tripartite mechanism of Ras, Rac and phospholipid interactions to localise at the protruding edge and interface with the interior of both macropinocytic and phagocytic cups. There, RGBARG shapes the protrusion by driving Rac activation at the rim whilst suppressing expansion of the active Ras interior domain. Consequently, cells lacking RGBARG form enlarged, flat interior domains unable to generate large macropinosomes. During phagocytosis, we find that disruption of RGBARG causes a geometry-specific defect in engulfing rod-shaped bacteria and ellipsoidal beads. This demonstrates the importance of co-ordinating small GTPase activities during engulfment of more complex shapes and thus the full physiological range of microbes, and how this is achieved in a model professional phagocyte.
Dynamics of the 4D genome during lineage specification, differentiation and maturation in vivo
Authors: Oudelaar A. Marieke , Beagrie Robert A. , Gosden Matthew , de Ornellas Sara , Georgiades Emily , Kerry Jon , Hidalgo Daniel , Carrelha Joana , Shivalingam Arun , El-Sagheer Afaf H. , Telenius Jelena M. , Brown Tom , Buckle Veronica J. , Socolovsky Merav , Higgs Douglas R. , Hughes Jim R.
Published: September 2019

Mammalian gene expression patterns are controlled by regulatory elements, which interact within Topologically Associating Domains (TADs). The relationship between activation of regulatory elements, formation of structural chromatin interactions and gene expression during development is unclear. We developed Tiled-C, a low-input Chromosome Conformation Capture (3C) approach, to study chromatin architecture at high spatial and temporal resolution through in vivo mouse erythroid differentiation. Integrated analysis of matched chromatin accessibility and single-cell expression data shows that regulatory elements gradually become accessible within pre-existing TADs during early differentiation. This is followed by structural re-organization within the TAD and formation of specific contacts between enhancers and promoters. In contrast to previous reports, our high-resolution data show that these enhancer-promoter interactions are not established prior to gene expression, but formed gradually during differentiation, concomitant with progressive upregulation of gene activity. Together, these results provide new insight into the close, interdependent relationship between chromatin architecture and gene regulation during development.
An efficient CRISPR-based strategy to insert small and large fragments of DNA using short homology arms
Authors: Kanca Oguz , Zirin Jonathan , Garcia-Marques Jorge , Knight Shannon , Yang-Zhou Donghui , Amador Gabriel , Chung Hyunglok , Zuo Zhongyuan , Ma Liwen , He Yuchun , Lin Wen-Wen , Fang Ying , Ge Ming , Yamamoto Shinya , Schulze Karen L. , Hu Yanhui , Spradling Allan C. , Mohr Stephanie E. , Perrimon Norbert , Bellen Hugo J.
Published: September 2019

We previously reported a CRISPR-mediated knock-in strategy into introns ofDrosophilagenes, generating anattP-FRT-SA-T2A-GAL4-polyA-3XP3-EGFP-FRT-attPtransgenic library for multiple uses (Lee et al., 2018b). The method relied on double stranded DNA (dsDNA) homology donors with ∼1 kb homology arms. Here, we describe three new simpler ways to edit genes in flies. We create single stranded DNA (ssDNA) donors using PCR and add 100 nt of homology on each side of an integration cassette, followed by enzymatic removal of one strand. Using this method, we generated GFP-tagged proteins that mark organelles in S2 cells. We then describe two dsDNA methods using cheap synthesized donors flanked by 100 nt homology arms and gRNA target sites cloned into a plasmid. Upon injection, donor DNA (1 to 5 kb) is released from the plasmid by Cas9. The cassette integrates efficiently and preciselyin vivo. The approach is fast, cheap, and scalable.
PDS5 proteins regulate the length of axial elements and telomere integrity during male mouse meiosis
Authors: Viera Alberto , Berenguer Inés , Ruiz-Torres Miguel , Gómez Rocío , Guajardo Andrea , Barbero José Luis , Losada Ana , Suja José A.
Published: September 2019

Cohesin cofactors regulate the loading, maintenance and release of cohesin complexes from chromosomes during the mitotic cell cycle but little is known on their role during vertebrate meiosis. One such cofactor is PDS5, which exists in two versions in somatic and germline cells, PDS5A and PDS5B, with unclear functional specificity. Here we have analyzed their distribution and functions in mouse spermatocytes. We show that simultaneous elimination of PDS5A and PDS5B results in severe defects during prophase I while their individual depletion does not, suggesting a functional redundancy of the two factors. Shortened axial/lateral elements and a reduction of early recombination nodules are observed in the absence of both PDS5 proteins. Moreover, telomere integrity and their association to the nuclear envelope are severely compromised. As these defects occur without detectable reduction in chromosome-bound cohesin, we propose that the dynamic behavior of the complex, mediated by PDS5 proteins, is key for successful completion of meiotic prophase I.
DeepMAsED: Evaluating the quality of metagenomic assemblies
Authors: Rojas-Carulla Mateo , Ley Ruth E. , Schölkopf Bernhard , Youngblut Nicholas D.
Published: September 2019

Motivation/background

Methodological advances in metagenome assembly are rapidly increasing in the number of published metagenome assemblies. However, identifying misassemblies is challenging due to a lack of closely related reference genomes that can act as pseudo ground truth. Existing reference-free methods are no longer maintained, can make strong assumptions that may not hold across a diversity of research projects, and have not been validated on large scale metagenome assemblies.

Results

We present DeepMAsED, a deep learning approach for identifying misassembled contigs without the need for reference genomes. Moreover, we provide an in silico pipeline for generating large-scale, realistic metagenome assemblies for comprehensive model training and testing. DeepMAsED accuracy substantially exceeds the state-of-the-art when applied to large and complex metagenome assemblies. Our model estimates close to a 5% contig misassembly rate in two recent large-scale metagenome assembly publications.

Conclusions

DeepMAsED accurately identifies misassemblies in metagenome-assembled contigs from a broad diversity of bacteria and archaea without the need for reference genomes or strong modelling assumptions. Running DeepMAsED is straight-forward, as well as is model re-training with our dataset generation pipeline. Therefore, DeepMAsED is a flexible misassembly classifier that can be applied to a wide range of metagenome assembly projects.

Availability

DeepMAsED is available from GitHub at https://github.com/leylabmpi/DeepMAsED.
Optimization and quantitative evaluation of fluorescence in situ hybridization chain reaction in clarified fresh-frozen brain tissues
Authors: Kumar Vivek , Krolewski David M , Hebda-Bauer Elaine K , Parsegian Aram , Martin Brian , Foltz Mathew , Akil Huda , Watson Stanley
Published: September 2019

Transcript labelling in intact tissues using in situ hybridization chain reaction has potential to provide vital spatiotemporal information for molecular characterization of heterogeneous neuronal populations. However, it remains relatively unexplored in fresh-frozen brain which provides more flexible utilization than perfused tissue. In the present study, we optimized the combination of in situ hybridization chain reaction in fresh-frozen rodent brains and then evaluated the uniformity of neuronal labelling between two clearing methods, CLARITY and iDISCO+. We found that CLARITY yielded higher signal-to-noise ratios but more limited imaging depth and required longer clearing times, whereas, iDISCO+ resulted in better tissue clearing, greater imaging depth and a more uniform labelling of larger samples. Based on these results, we used iDISCO+-cleared fresh-frozen rodent brains to further validate this combination and map the expression of a few genes of interest pertaining to mood disorders. We then examined the potential of in situ hybridization chain reaction to label transcripts in cleared postmortem human brain tissues. The combination failed to produce adequate mRNA labelling in postmortem human cortical slices but produced visually adequate labeling in the cerebellum tissues. We next, investigated the multiplexing ability of in situ hybridization chain reaction in cleared tissues which revealed inconsistent fluorescence output depending upon the fluorophore conjugated to the hairpins. Finally, we applied our optimized protocol to assess the effect of glucocorticoid receptor overexpression on basal somatostatin expression in the mouse cortex. The constitutive glucocorticoid receptor overexpression resulted in lower number density of somatostatin-expressing neurons compared to wild type. Overall, the combination of in situ hybridization chain reaction with clearing methods especially iDISCO+ may find broad application in the transcript analysis in rodent studies but its limited use in postmortem human tissues can be improved by further optimizations.
Protein Tyrosine Phosphatase 1B Regulates MicroRNA-208b-Argonaute 2 Association and Thyroid Hormone Responsiveness in Cardiac Hypertrophy
Authors: Coulis Gérald , Shi Yanfen , Labbé David P. , Bergeron Alexandre , Sahmi Fatiha , Vinette Valérie , Karsenty Gérard , Allen Bruce G. , Tremblay Michel L. , Tardif Jean-Claude , Boivin Benoit
Published: September 2019

Elevated reactive oxygen species (ROS) production plays an important role in the pathogenesis of several diseases, including cardiac hypertrophy. While the regulation of diverse sources of ROS is well characterized in the heart, the redox-sensitive targets that contribute to redox signaling remain largely undefined. We now report that protein tyrosine phosphatase 1B (PTP1B) is reversibly oxidized and inactivated in hearts undergoing hypertrophy and that gene deletion of PTP1B in mouse hearts cause an hypertrophic phenotype that is critically exacerbated in mice subjected to pressure overload. Furthermore, we show that PTP1B dephosphorylates Tyr393on argonaute 2, a key component of the RNA-induced silencing complex, and sustains gene silencing in the heart. Our results indicate that PTP1B inactivation and argonaute 2 Tyr393phosphorylation specifically prevents argonaute 2 from interacting with miR-208b. Phosphorylation and inactivation of argonaute 2 in PTP1B cKO mice revealed a mechanism by which defective miR-208b-mediated repression of thyroid hormone receptor-associated protein 1 (THRAP1/MED13) contributes to thyroid hormone-mediated cardiac hypertrophy. In support of this conclusion, inhibiting the synthesis of triiodothyronine (T3), using propylthiouracil, rescued TAC-induced hypertrophy and improved myocardial contractility and systolic function in PTP1B cKO mice. Together, our data illustrate that PTP1B activity exerts a cardioprotective effect in the heart and that redox signaling is tightly linked to thyroid hormone responsiveness and to microRNA-mediated gene silencing in pathological hypertrophy.
Secondary origin, hybridization and sexual reproduction in a diploid- tetraploid contact zone of the facultative apomictic orchidZygopetalum mackayi
Authors: de Moura Yohans Alves , Alves-Pereira Alessandro , Silva Carla Cristina , de Souza Lívia Moura , de Souza Anete Pereira , Koehler Samantha
Published: September 2019

>Mixed-cytotype populations are ideal to understand polyploid establishment and diversification. We used the orchidZygopetalum mackayito understand how facultative apomictic reproduction relates to polyploidy. Sexual diploids and facultative apomictic tetraploids occur under distinct niches, with a contact zone where triploids occur. We hypothesized that facultative apomictic reproduction increases the fitness of tetraploids through reproductive interference between cytotypes. We predict patterns of genetic diversity of allopatric tetraploid populations to be significantly different from contact zone populations as a result of dominant apomictic reproduction in the later. We also describe the contact nature of diploids and tetraploids and the role of the intermediate triploids based on patterns of genetic structure within and among pure and mixed-cytotype populations.

>We designed eight microsatellite markers and genotyped 155 individuals from six populations resulting in 237 alleles. We described patterns of genetic diversity and structure within and among populations and cytotypes.

>Genotypic diversity is similarly high among all populations and cytotypes. Each cytotype emerged as a genetically cluster, combining individuals from different populations. Triploids clustered in an intermediate position between diploids andtetraploids.

>We rejected the hypothesis of reproductive interference between cytotypes ofZ. mackayi. Patterns of genetic diversity are incongruent with the occurrence of apomict reproduction in tetraploids. Mixed-cytotype populations originate from secondary contact and triploids are hybrids between diploids and tetraploids and act as a reproductive barrier. We suggest polyploidy rather than facultative apomixis explains higher fitness of tetraploids in this species and, therefore, eco-geographical patterns of distribution.
HELLS and PRDM9 form a Pioneer Complex to Open Chromatin at Meiotic Recombination Hotspots
Authors: Spruce Catrina , Dlamini Sibongakonke , Ananda Guruprasad , Bronkema Naomi , Tian Hui , Paigen Ken , Carter Gregory W. , Baker Christopher L
Published: September 2019

Chromatin barriers prevent spurious interactions between regulatory elements and DNA-binding proteins. One such barrier, whose mechanism for overcoming is poorly understood, is access to recombination hotspots during meiosis. Here we show that the chromatin remodeler HELLS and DNA-binding protein PRDM9 function together to open chromatin at hotspots and provide access for the DNA double-strand break (DSB) machinery. Recombination hotspots are decorated by a unique combination of histone modifications, not found at other regulatory elements. HELLS is recruited to hotspots by PRDM9, and is necessary for both histone modifications and DNA accessibility at hotspots. In male mice lacking HELLS, DSBs are retargeted to other sites of open chromatin, leading to germ cell death and sterility. Together, these data provide a model for hotspot activation where HELLS and PRDM9 function as a pioneer complex to create a unique epigenomic environment of open chromatin, permitting correct placement and repair of DSBs.
Direct-fit to nature: an evolutionary perspective on biological (and artificial) neural networks
Authors: Hasson Uri , Nastase Samuel A. , Goldstein Ariel
Published: September 2019

Evolution is a blind fitting process by which organisms, over generations, adapt to the niches of an ever-changing environment. Does the mammalian brain use similar brute-force fitting processes to learn how to perceive and act upon the world? Recent advances in training deep neural networks has exposed the power of optimizing millions of synaptic weights to map millions of observations along ecologically relevant objective functions. This class of models has dramatically outstripped simpler, more intuitive models, operating robustly in real-life contexts spanning perception, language, and action coordination. These models do not learn an explicit, human-interpretable representation of the underlying structure of the data; rather, they use local computations to interpolate over task-relevant manifolds in a high-dimensional parameter space. Furthermore, counterintuitively, over-parameterized models, similarly to evolutionary processes, can be simple and parsimonious as they provide a versatile, robust solution for learning a diverse set of functions. In contrast to traditional scientific models, where the ultimate goal is interpretability, over-parameterized models eschew interpretability in favor of solving real-life problems or tasks. We contend that over-parameterized blind fitting presents a radical challenge to many of the underlying assumptions and practices in computational neuroscience and cognitive psychology. At the same time, this shift in perspective informs longstanding debates and establishes unexpected links with evolution, ecological psychology, and artificial life.
Refining the composition of the Arabidopsis thaliana 80S cytosolic ribosome
Authors: Salih Karzan Jalal , Duncan Owen , Li Lei , Troesch Josua , Millar A. Harvey
Published: September 2019

The cytosolic 80S ribosome is composed of protein and RNA molecules and its function in protein synthesis is modulated through interaction with other cytosolic components. Defining the role of each of the proteins associated with ribosomes in plants is a major challenge which is hampered by difficulties in attribution of different proteins to roles in ribosome biogenesis, the mature cytosolic ribosome (r-proteins) or to the broader translatome associated with functioning ribosomes. Here we refined the core r-protein composition in plants by determining the abundance of proteins in low, partially and highly purified ribosomal samples from Arabidopsis thaliana cell cultures. To characterise this list of proteins further we determined their degradation (KD) and synthesis (KS) rate by progressive labelling with 15N combined with peptide mass spectrometry analysis. The turnover rates of 55 r-proteins, including 26 r-proteins from the 40S subunit and 29 r-proteins from the 60S subunit could be determined. Overall, ribosome proteins showed very similar KD and KS rates suggesting that half of the ribosome population is replaced every 3-4 days. Three proteins showed significantly shorter half-lives; ribosomal protein P0D (RPP0D) with a half-life of 0.5 days and RACK1b and c with half-lives of 1-1.4 days. The ribosomal RPP0D protein is a homolog of the human Mrt4 protein, a trans-acting factor in the assembly of the pre-60S particle, while RACK1 has known regulatory roles in cell function beyond its role as a 40S subunit. Our experiments also identified 58 proteins that are not from r-protein families but co-purify with ribosomes and co-express with r-proteins in Arabidopsis. Of this set, 26 were enriched more than 10-fold during ribosome purification. A number have known roles in translation or ribosome-association while others are newly proposed ribosome-associated factors in plants. This analysis provides a more robust understanding of Arabidopsis ribosome content, shows that most r-proteins turnover in unison in vivo, identifies a novel set of potential plant translatome components, and reveals how protein turnover can identify r-proteins involved in ribosome biogenesis or regulation in plants. Data are available via ProteomeXchange with identifier PXD012839.
Excessive assimilation of ammonium by plastidic glutamine synthetase is a major cause of ammonium toxicity in Arabidopsis thaliana
Authors: Hachiya Takushi , Inaba Jun , Wakazaki Mayumi , Sato Mayuko , Toyooka Kiminori , Miyagi Atsuko , Kawai-Yamada Maki , Kiba Takatoshi , Gojon Alain , Sakakibara Hitoshi
Published: September 2019

Plants use nitrate and ammonium in the soil as their main nitrogen sources. Recently, ammonium has attracted attention due to evidence suggesting that, in C3 species, an elevated CO2 environment inhibits nitrate assimilation. However, high concentrations of ammonium as the sole nitrogen source for plants causes impaired growth, i.e. ammonium toxicity. Although ammonium toxicity has been studied for a long time, the primary cause remains to be elucidated. Here, we show that ammonium assimilation in plastids rather than ammonium accumulation is a primary cause for toxicity. Our genetic screen of ammonium-tolerant Arabidopsis lines with enhanced shoot growth identified plastidic GLUTAMINE SYNTHETASE 2 (GLN2) as the causal gene. Our reciprocal grafting of wild-type and GLN2 or GLN1;2-deficient lines suggested that shoot GLN2 activity results in ammonium toxicity, whilst root GLN1;2 activity prevents it. With exposure to toxic levels of ammonium, the shoot GLN2 reaction produced an abundance of protons within cells, thereby elevating shoot acidity and stimulating expression of acidic stress-responsive genes. Application of an alkaline ammonia solution to the toxic ammonium medium efficiently alleviated the ammonium toxicity with a concomitant reduction in shoot acidity. Consequently, we conclude that a primary cause of ammonium toxicity is acidic stress in the shoot. This fundamental insight provides a framework for enhanced understanding of ammonium toxicity in plants.
cGMP dynamics underlie thermosensation in C. elegans
Authors: Aoki Ichiro , Shiota Makoto , Nakano Shunji , Mori Ikue
Published: September 2019

Animals sense ambient temperature so that they can adjust their behavior to the environment; they avoid noxious heat and coldness and stay within a survivable temperature range. C. elegans can sense temperature, memorize past cultivation temperature and navigate towards preferable temperature, for which a thermosensory neuron, AFD, is essential. AFD responds to temperature increase from the past cultivation temperature by increasing intracellular Ca2+ level. We aimed to reveal how AFD encodes and memorizes the information of temperature. Although cGMP synthesis is crucial for thermosensation by AFD, whether and how cGMP level temporally fluctuates in AFD remained elusive. We therefore monitored cGMP level in AFD and found that cGMP dynamically responded to temperature change in a manner dependent on past cultivation temperature. Given that cGMP dynamics is supposed to be upstream of Ca2+ dynamics, our results suggest that AFD’s memory is formed by simpler molecular mechanisms than previously expected from the Ca2+ dynamics. Moreover, we analyzed how guanylyl cyclases and phosphodiesterases, which synthesize and degrade cGMP, respectively, contributed to cGMP and Ca2+ dynamics and thermotaxis behavior.
Arabidopsis PPC2 is crucial for growth at low CO2 by involvement in photorespiratory metabolism and integration of ABI5
Authors: You Lei , Zhang Jumei , Li Long , Xiao Chuanlei , Feng Xinhua , Chen Shaoping , Guo Liang , Hu Honghong
Published: September 2019

Phosphoenolpyruvate carboxylase (PEPC) is a pivotal enzyme that plays a key role in photosynthetic CO2 fixation in C4. However, the function of C3 PEPCs and their roles at environmental CO2 changes are still limited. Here, we report the role of PPC2 in seedling growth at low CO2 by linking photorespiratory metabolism with primary metabolism and involvement of ABA and ABI5. Mutation of PPC2 caused seedling growth arrest, with reduced Fv/Fm, photosynthetic carbohydrates and ABA biosynthesis at low CO2. PPC2 is induced by low CO2 and the PEPC activity was greatly reduced in ppc2 leaves. Moreover, metabolic analyses showed the photorespiratory intermediates, glycine and serine, were greatly increased and primary metabolites were reduced. Application of sucrose, malate and ABA greatly rescued the growth arrest phenotype of ppc2. The expression of glycine/serine synthesis and metabolism related photorespiratory enzyme genes were decreased in ppc2 and regulated by ABI5 at low CO2 conditions. ppc2 and abi5 mature plants exhibited reduced A-Ci curves at relatively low CO2, which could be recovered by non-photorespiratory low oxygen conditions. ABI5 expression greatly rescued the growth arrest and A-Ci curves of ppc2 at low CO2. Our findings demonstrate the important role of C3 PEPCs in carbon fixation and metabolism.
Induced RPB1 depletion reveals a direct gene-specific control of RNA Polymerase III function by RNA Polymerase II
Authors: Gerber Alan , Ito Keiichi , Chu Chi-Shuen , Roeder Robert G.
Published: September 2019

Increasing evidence suggests that tRNA levels are dynamically and specifically regulated in response to internal and external cues to modulate the cellular translational program. However, the molecular players and the mechanisms regulating the gene-specific expression of tRNAs are still unknown. Using an inducible auxin-degron system to rapidly deplete RPB1 (the largest subunit of RNA Pol II) in living cells, we identified Pol II as a direct gene-specific regulator of tRNA transcription. Our data suggest that Pol II transcription robustly interferes with Pol III function at specific tRNA genes. This activity was further found to be essential for MAF1-mediated repression of a large set of tRNA genes during serum starvation, indicating that repression of tRNA genes by Pol II is dynamically regulated. Hence, Pol II plays a direct and central role in the gene-specific regulation of tRNA expression.
Cell type specific novel lincRNAs and circRNAs in the BLUEPRINT haematopoietic transcriptomes atlas
Authors: Grassi Luigi , Izuogu Osagie G. , Jorge Natasha A.N. , Seyres Denis , Bustamante Mariona , Burden Frances , Farrow Samantha , Farahi Neda , Martin Fergal J. , Frankish Adam , Mudge Jonathan M. , Kostadima Myrto , Petersen Romina , Lambourne John J. , Rowlston Sophia , Martin-Rendon Enca , Clarke Laura , Downes Kate , Estivill Xavier , Flicek Paul , Martens Joost H.A. , Yaspo Marie-Laure , Stunnenberg Hendrik G. , Ouwehand Willem H. , Passetti Fabio , Turro Ernest , Frontini Mattia
Published: September 2019

Transcriptional profiling of hematopoietic cell subpopulations has helped characterize the developmental stages of the hematopoietic system and the molecular basis of malignant and non-malignant blood diseases for the past three decades. The introduction of high-throughput RNA sequencing has increased knowledge of the full repertoire of RNA molecules in hematopoietic cells of different types, without relying on prior gene annotation. Here, we introduce the analysis of the BLUEPRINT consortium gene expression data for mature hematopoietic cells, comprising 90 total RNA and 32 small RNA sequencing experiments, from 27 different cell types. We used these data to describe the transcriptional profile of each we used guided transcriptome assembly to extend the annotation of the transcribed genome, which led to the identification of hundreds of novel non-coding RNA genes, which display a high degree of cell type specificity. We also characterized the expression of circular RNAs and found that these are also highly cell type specific. This resource refines the active transcriptional landscape of mature hematopoietic cells, highlights abundant genes and transcriptional isoforms for each cell type, and provides valuable data and visualisation tools for the scientific community working on hematological development and diseases.
Homeostatic maintenance and age-related functional decline in theDrosophilaear
Authors: Keder Alyona , Tardieu Camille , Malong Liza , Filia Anastasia , Kashkenbayeva Assel , Gale Jonathan E. , Lovett Mike , Jarman Andrew P. , Albert Joerg T.
Published: September 2019

The widespread loss of hearing is one of the major threats to future wellbeing in ageing human societies. Amongst its various forms, age-related hearing loss (ARHL) carries the vast bulk of the global disease burden. The causes for the terminal decline of auditory function, however, are as unknown as the mechanisms that maintain sensitive hearing before its breakdown. We here present an in-depth analysis of maintenance and ageing in the auditory system of the fruit flyDrosophila melanogaster. We show thatDrosophila, just like humans, display ARHL and that their auditory life span is homeostatically supported by a set of evolutionarily conserved transcription factors. The transcription factors Onecut (closest human orthologues: ONECUT2, ONECUT3), Optix (SIX3, SIX6), Worniu (SNAI2) and Amos (ATOH1, ATOH7, NEUROD1) emerged as key regulators acting upstream of core sensory genes, including components of the fly’s molecular machinery for auditory transduction and amplification.
Predicting intentional and unintentional task unrelated thought with EEG
Authors: Martel Adrien , Arvaneh Mahnaz , Robertson Ian , Dockree Paul
Published: September 2019

Our attention seldom remains on a singular activity, instead veering off into thoughts unrelated to the task at hand. Studies adopting a component process view of off-task thought have begun to identify the underlying mechanisms and associated electrophysiological correlates underlying ongoing thought. In the present study, we developed subject-independent classification algorithms based on electroencephalographic (EEG) markers to discriminate on-task vs off-task as well as intentional vs unintentional off-task thought. To that end, spatio-temporal and spectral features extracted from EEG activity prior to reports of ongoing thought during a test of sustained attention were ranked according to their discriminative power. Using data collected from 26 participants, average classification accuracies of 83.4% and 71.6% were achieved using a regularized linear model for on-task vs off-task and intentional vs unintentional off-task thought, respectively. Our results identified gamma oscillations as the most discriminative feature to distinguish on-task from off-task states, and alpha synchronization as the most prominent feature when off-task states are engaged in deliberately rather than when experienced as arising spontaneously. Our work represents the first successful attempt at reliably discriminating the degree of intentionality experienced during task-unrelated thought and highlights the importance of recognizing the heterogeneous nature of off-task states.
Human embryoid bodies model basal lamina assembly and muscular dystrophy
Authors: Nickolls Alec R. , Lee Michelle M. , Zukosky Kristen , Mallon Barbara S. , Bönnemann Carsten G.
Published: September 2019

The basal lamina is a specialized sheet of dense extracellular matrix (ECM), linked to the plasma membrane of specific cell types in their tissue context, that serves as a structural scaffold for organ genesis and maintenance. Disruption of the basal lamina and its functions is central to many disease processes, including cancer metastasis, kidney disease, eye disease, muscular dystrophies, and specific types of brain malformation. The latter three pathologies occur in the dystroglycanopathies, which are caused by dysfunction of the ECM receptor dystroglycan. However, opportunities to study the basal lamina in various human disease tissues are restricted due to its limited accessibility. Here, we report the generation of embryoid bodies from human induced pluripotent stem cells to model basal lamina formation. Embryoid bodies cultured via this protocol mimic pre-gastrulation embryonic development, consisting of an epithelial core surrounded by a basal lamina and a peripheral layer of ECM-secreting endoderm. In dystroglycanopathy patient embryoid bodies, electron and fluorescence microscopy revealed ultrastructural basal lamina defects and reduced ECM assembly. By starting from patient-derived cells, these results establish a method for the in vitro synthesis of patient-specific basal lamina and recapitulate disease-relevant ECM defects seen in muscular dystrophies. Finally, we applied this system to evaluate an experimental ribitol supplement therapy on genetically diverse dystroglycanopathy patient samples.
The Na—O bond in sodium fenamate
Authors: Krawczyk Marta S. , Majerz Irena
Published: September 2019

The one-dimensional polymeric structure of sodium diaquafenamate–water (1/1) was studied by X-ray diffraction. The sodium cation is coordinated to one oxygen atom of the carboxylate group and to four water oxygen atoms. To characterize the Na—O bonds, the quantum theory of atoms in molecules (QTAIM) and noncovalent interaction (NCI) approaches have been used. Both methods confirmed that the Na—O bonds are very weak, comparable with the weak N—H...O intramolecular hydrogen bond. The polymeric structure is stabilized by the interaction of the sodium cation with the surrounding water molecules.
A commensurately modulated crystal structure and the physical properties of a novel polymorph of the caesium manganese phosphate CsMnPO4
Authors: Bolotina Nadezhda , Yakubovich Olga , Shvanskaya Larisa , Dimitrova Olga , Volkov Anatoly , Vasiliev Alexander
Published: September 2019

A novel modification of the CsMnPO4 β-phase was achieved by hydrothermal synthesis at 553 K. The compound crystallizes in the monoclinic system with the basic unit-cell parameters a = 11.0699 (4), b = 11.0819 (6), c = 9.1106 (3) Å, γ = 119.480 (5)o; the modulation vectors are q1 = 0.4a* and q2 = 0.4b*. The structure was determined based on single-crystal X-ray diffraction data obtained from a pseudo-merohedral twin using a superspace approach in the (3 + 2)D symmetry group P11a(a1,b1,0)0(a2,b2,0)0 and refined to R = 0.083 for 10 266 reflections with I > 3σ(I). It is considered as a low-temperature polymorph of CsMnPO4 with the same UUUDDD-type layer topology built by MnO4 and PO4 tetrahedra, and stacked in a framework in the same manner as β-tridymite. Large open channels parallel to the [110] and [001] directions incorporate Cs atoms. All Cs atoms are distributed along the a
super = 55.35 (1) and b
super = 55.41 (1) axes of the large unit cell with pseudo periods of a
super/5 and b
super/5 which are broken mainly by the positions of oxygen atoms (orientation of Mn- and P-centered tetrahedra). The β-phase is discussed as a member of the morphotropic series of manganese phosphates with large cations of AMnPO4, where A = Cs, Rb, K and Ag. The title compound is an antiferromagnet with the Neel temperature T
N = 4.5 K.
Analytical description of nanowires. I. Regular cross sections for zincblende and diamond structures
Authors: König Dirk , Smith Sean C.
Published: September 2019

Semiconductor nanowires (NWires) experience stress and charge transfer from their environment and impurity atoms. In response, the environment of a NWire experiences a NWire stress response which may lead to propagated strain and a change in the shape and size of the NWire cross section. Here, geometric number series are deduced for zincblende- (zb-) and diamond-structured NWires of diameter d
Wire to obtain the numbers of NWire atoms N
Wire(d
Wire[i]), bonds between NWire atoms N
bnd(d
Wire[i]) and interface bonds N
IF(d
Wire[i]) for six high-symmetry zb NWires with the low-index faceting that occurs frequently in both bottom-up and top-down approaches of NWire processing. Along with these primary parameters, the specific lengths of interface facets, the cross-sectional widths and heights and the cross-sectional areas are presented. The fundamental insights into NWire structures revealed here offer a universal gauge and thus could enable major advancements in data interpretation and understanding of all zb- and diamond-structure-based NWires. This statement is underpinned with results from the literature on cross-section images from III–V core–shell NWire growth and on Si NWires undergoing self-limiting oxidation and etching. The massive breakdown of impurity doping due to self-purification is shown to occur for both Si NWires and Si nanocrystals (NCs) for a ratio of N
bnd/N
Wire = N
bnd/N
NC = 1.94 ± 0.01 using published experimental data.
The crystal structures and Hirshfeld surface analyses of four 3,5-diacetyl-2-methyl-2,3-dihydro-1,3,4-thiadiazol-2-yl derivatives
Authors: NizamMohideen M. , Syed Abuthahir S. , Viswanathan V. , Velmurugan D. , Karthik Ananth M.
Published: September 2019

The title compounds, 4-(5-acetamido-3-acetyl-2-methyl-2,3-dihydro-1,3,4-thiadiazol-2-yl)phenyl benzoate, C20H19N3O4S (I), 4-(5-acetamido-3-acetyl-2-methyl-2,3-dihydro-1,3,4-thiadiazol-2-yl)phenyl isobutyrate 0.25-hydrate, C17H21N3O4S·0.25H2O (II), 4-(5-acetamido-3-acetyl-2-methyl-2,3-dihydro-1,3,4-thiadiazol-2-yl)phenyl propionate, C16H19N3O4S (III) and 4-(5-acetamido-3-acetyl-2-methyl-2,3-dihydro-1,3,4-thiadiazol-2-yl)phenyl cinnamate chloroform hemisolvate, C22H21N3O4S·0.5CHCl3 (IV), all crystallize with two independent molecules (A and B) in the asymmetric unit in the triclinic P\overline{1} space group. Compound II crystallizes as a quaterhydrate, while compound IV crystallizes as a chloroform hemisolvate. In compounds I, II, III (molecules A and B) and IV (molecule A) the five-membered thiadiazole ring adopts an envelope conformation, with the tetrasubstituted C atom as the flap. In molecule B of IV this ring is flat (r.m.s. deviation 0.044 Å). The central benzene ring is in general almost normal to the mean plane of the thiadiazole ring in each molecule, with dihedral angles ranging from 75.8 (1) to 85.5 (2)°. In the crystals of all four compounds, the A and B molecules are linked via strong N—H...O hydrogen bonds and generate centrosymmetric four-membered R
4
4(28) ring motifs. There are C—H...O hydrogen bonds present in the crystals of all four compounds, and in I and II there are also C—H...π interactions present. The intermolecular contacts in the crystals of all four compounds were analysed using Hirshfeld surface analysis and two-dimensional fingerprint plots.
The structure and Hirshfeld surface analysis of the salt 3-methacrylamido-N,N,N-trimethylpropan-1-aminium 2-acrylamido-2-methylpropane-1-sulfonate
Authors: Wickramasinhage Ravindra N. , McAdam C. John , Hanton Lyall R. , Moratti Stephen C. , Simpson Jim
Published: September 2019

The title salt, C10H21N2O+·C7H12NO4S−, comprises a 3-methacrylamido-N,N,N-trimethylpropan-1-aminium cation and a 2-acrylamido-2-methylpropane-1-sulfonate anion. The salt crystallizes with two unique cation–anion pairs in the asymmetric unit of the orthorhombic unit cell. The crystal studied was an inversion twin with a 0.52 (4):0.48 (4) domain ratio. In the crystal, the cations and anions stack along the b-axis direction and are linked by an extensive series of N—H...O and C—H...O hydrogen bonds, forming a three-dimensional network. Hirshfeld surface analysis was carried out on both the asymmetric unit and the two individual salts. The contribution of interatomic contacts to the surfaces of the individual cations and anions are also compared.
Crystal structure of 4-bromo-N-[(3,6-di-tert-butyl-9H-carbazol-1-yl)methylidene]aniline
Authors: Kubono Koji , Matsumoto Taisuke , Taneda Masatsugu
Published: September 2019

In the title compound, C27H29BrN2, the carbazole ring system is essentially planar, with an r.m.s. deviation of 0.0781 (16) Å. An intramolecular N—H...N hydrogen bond forms an S(6) ring motif. One of the tert-butyl substituents shows rotational disorder over two sites with occupancies of 0.592 (3) and 0.408 (3). In the crystal, two molecules are associated into an inversion dimer through a pair of C—H...π interactions. The dimers are further linked by another pair of C—H...π interactions, forming a ribbon along the c-axis direction. A C—H...π interaction involving the minor disordered component and the carbazole ring system links the ribbons, generating a network sheet parallel to (100).