bluuebunny/crossref_metadata_2025 huggingface dataset:
Female Assamese macaques bias their affiliation to paternal and maternal kin
Authors: De Moor Delphine , Roos Christian , Ostner Julia , Schülke Oliver
Published: July 2019

Forming strong social bonds leads to higher reproductive success, increased longevity and/or increased infant survival in several mammal species. Given these adaptive benefits, understanding what determines partner preferences in social bonding is important. Maternal relatedness strongly predicts partner preference across many mammalian taxa. Although paternal and maternal kin share the same number of genes, and theoretically similar preferences would therefore be expected for paternal kin, the role of paternal relatedness has received relatively little attention. Here, we investigate the role of maternal and paternal relatedness for female bonding in Assamese macaques (Macaca assamensis), a species characterized by a relatively low male reproductive skew. We studied a wild population under natural conditions using extensive behavioural data and relatedness analyses based on pedigree reconstruction. We found stronger social bonds and more time spent grooming between maternal kin and paternal half-sisters compared to non-kin, with no preference of maternal over paternal kin. Paternally related and non-related dyads did not form stronger bonds when they had less close maternal kin available, however we would need a bigger sample size to confirm this. As expected given the low reproductive skew, bonds between paternal half-sisters closer in age were not stronger than between paternal half-sisters with larger age differences, suggesting that age similarity was not the mechanism by which paternally related individuals recognized each other. An alternative way through which paternal kin could get familiarized is mother- and/or father-mediated familiarity.
Targeting USP9x/SOX2 axis contributes to the anti-osteosarcoma effect of neogambogic acid
Authors: Chen Xiangyun , Zhang Xingming , Cai Haiyan , Yang Wupeng , Lei Hu , Xu Hanzhang , Wang Weiwei , Zhu Qi , Kang Jingwu , Yin Tong , Gu Wenli , Wu Ying-Li
Published: July 2019

SOX2 has been viewed as a critical oncoprotein in osteosarcoma. Emerging evidence show that inducing the degradation of transcription factors such as SOX2 is a promising strategy to make them druggable. Here, we show that neogambogic acid (NGA), an active ingredients in garcinia, significantly inhibited the proliferation of osteosarcoma cells with ubiquitin proteasome-mediated degradation of SOX2 in vitro and in vivo. We further identified USP9x as a bona fide deubiquitinase for SOX2 and NGA directly interacts with USP9x in cells. Moreover, knockdown of USP9x inhibited the proliferation and colony formation of osteosarcoma cells, which could be rescued by overexpression of SOX2. Consistent with this, knockdown of USP9x inhibited the proliferation of osteosarcoma cells in a xenograft mouse model. Collectively, we identify USP9x as the first deubiquitinating enzyme for controlling the stability of SOX2 and USP9x is a direct target for NGA. We propose that targeting the USP9x/SOX2 axis represents a novel strategy for the therapeutic of osteosarcoma and other SOX2 related cancers.
Regional oceanographic features and hydrothermal activity influence protist diversity and biogeography in the Okinawa Trough
Authors: Brisbin Margaret Mars , Conover Asa E. , Mitarai Satoshi
Published: July 2019

Microbial eukaryotes (protists) contribute substantially to ecological functioning in marine ecosystems, but factors shaping protist diversity, such as dispersal barriers and environmental selection, remain difficult to parse. Deep-sea water masses, which form geographic barriers, and hydrothermal vents, which represent isolated productivity hotspots, are ideal opportunities for studying the effects of dispersal barriers and environmental selection on protist communities. The Okinawa Trough, a deep, back-arc spreading basin, contains distinct water masses in the bottom waters of northern and southern regions and at least twenty-five active hydrothermal vents. In this study, we used metabarcoding to characterize protist communities from fourteen stations spanning the length of the Okinawa Trough, including three hydrothermal vent sites. Significant differences in community structure reflecting regional oceanography and water mass composition were present, indicating the importance of geographic factors in shaping protist communities. Protist communities in bottom waters affected by hydrothermal activity were significantly different from communities in other bottom waters, suggesting that environmental factors can be especially important in shaping community composition under specific conditions. Amplicon sequence variants that were enriched in hydrothermally influenced bottom waters largely derived from cosmopolitan protists that were present, but rare, in other near-bottom samples, thus highlighting the importance of the rare biosphere.
Polyanions provide selective control of APC/C interactions with the activator subunit
Authors: Mizrak Arda , Morgan David O.
Published: July 2019

Transient interactions between the Anaphase-Promoting Complex/Cyclosome (APC/C) and its activator subunit Cdc20 or Cdh1 generate oscillations in ubiquitination activity necessary to maintain the order of cell cycle events. Activator binds the APC/C with high affinity and exhibits negligible dissociation kinetics in vitro, and it is not clear how the rapid turnover of APC/C-activator complexes is achieved in vivo. Here, we describe a mechanism that controls APC/C-activator interactions based on the availability of substrates. We find that APC/C-activator dissociation is stimulated by abundant cellular polyanions such as nucleic acids and polyphosphate. Polyanions also interfere with the ubiquitination of low-affinity substrates. However, engagement with high-affinity substrates blocks the inhibitory effects of polyanions on activator binding and APC/C activity. This mechanism amplifies the effects of substrate affinity on APC/C function, stimulating processive ubiquitination of high-affinity substrates and suppressing ubiquitination of low-affinity substrates.
ΔSCOPE: A new method to quantify 3D biological structures and identify differences in zebrafish forebrain development
Authors: Schwartz Morgan S , Schnabl Jake , Litz Mackenzie P.H. , Baumer Benjamin S , Barresi Michael
Published: July 2019

Research in the life sciences has traditionally relied on the analysis of clear morphological phenotypes, which are often revealed using increasingly powerful microscopy techniques analyzed as maximum intensity projections (MIPs). However, as biology turns towards the analysis of more subtle phenotypes, MIPs and qualitative approaches are failing to adequately describe these phenotypes. To address these limitations and quantitatively analyze the three-dimensional (3D) spatial relationships of biological structures, we developed the computational method and program called ΔSCOPE (Changes in Spatial Cylindrical Coordinate Orientation using PCA Examination). Our approach uses the fluorescent signal distribution within a 3D data set and reorients the fluorescent signal to a relative biological reference structure. This approach enables quantification and statistical analysis of spatial relationships and signal density in 3D multichannel signals that are positioned around a well-defined structure contained in a reference channel. We validated the application of ΔSCOPE by analyzing normal axon and glial cell guidance in the zebrafish forebrain and by quantifying the commissural phenotypes associated with abnormal Slit guidance cue expression in the forebrain. Despite commissural phenotypes which display disruptions to the reference structure, ΔSCOPE was able to detect subtle, previously uncharacterized changes in zebrafish forebrain midline crossing axons and glia. This method has been developed as a user-friendly, open source program. We propose that ΔSCOPE is an innovative approach to advancing the state of image quantification in the field of high resolution microscopy, and that the techniques presented here are of broad applications to the life science field.
Autonomous emergence of connectivity assemblies via spike triplet interactions
Authors: Montangie Lisandro , Gjorgjieva Julijana
Published: July 2019

Non-random connectivity can emerge without structured external input driven by activity-dependent mechanisms of synaptic plasticity based on precise spiking patterns. Here we analyze the emergence of global structures in recurrent networks based on a triplet model of spike timing dependent plasticity (STDP) which depends on the interactions of three precisely-timed spikes and can describe plasticity experiments with varying spike frequency better than the classical pair-based STDP rule. We describe synaptic changes arising from emergent higher-order correlations, and investigate their influence on different connectivity motifs in the network. Our motif expansion framework reveals novel motif structures under the triplet STDP rule, which support the formation of bidirectional connections and loops in contrast to the classical pair-based STDP rule. Therefore, triplet STDP drives the spontaneous emergence of self-connected groups of neurons, or assemblies, proposed to represent functional units in neural circuits. Assembly formation has often been associated with plasticity driven by firing rates or external stimuli. We propose that assembly structure can emerge without the need for externally patterned inputs or assuming a symmetric pair-based STDP rule commonly assumed in previous studies. The emergence of non-random network structure under triplet STDP occurs through internally-generated higher-order correlations, which are ubiquitous in natural stimuli and neuronal spiking activity, and important for coding. We further demonstrate how neuromodulatory mechanisms that modulate the shape of triplet STDP or the synaptic transmission function differentially promote connectivity motifs underlying the emergence of assemblies, and quantify the differences using graph theoretic measures.
Dynamic measurement of cytosolic pH and [NO3-] uncovers the role of the vacuolar transporter AtCLCa in the control of cytosolic pH
Authors: Demes Elsa , Besse Laetitia , Satiat-Jeunemaitre Béatrice , Thomine Sébastien , De Angeli Alexis
Published: July 2019

Ion transporters are key players of cellular processes. The mechanistic properties of ion transporters have been well elucidated by biophysical methods. Meanwhile the understanding of their exact functions in the whole cell homeostasis is limited by the difficulty to monitor their activity in vivo. The development of biosensors to track subtle changes in intracellular parameters provides an invaluable key to tackle this challenging issue. Here, we adapted the use of a dual biosensor using guard cells as experimental model to visualize the impact on the cytosol of anion transport from intracellular compartments. To image the activity of AtCLCa, a vacuolar NO3-/H+ exchanger regulating stomata aperture in Arabidopsis thaliana, we expressed a genetically encoded biosensor, ClopHensor allowing monitoring the dynamics of cytosolic anion concentration and pH. We first show that ClopHensor is not only a Cl- but also a NO3- sensor. We were then able to unravel and quantify the variations of NO3- and pH in the cytosol. Our data show that AtCLCa activity modifies cytosolic pH and NO3-, demonstrating that the transport activity of a vacuolar exchanger has a profound impact on cytosolic homeostasis. We propose that a major function of this endomembrane transporter is to adjust cytosolic conditions to cellular needs. This opens a novel perspective on the function of intracellular transporters of the CLC family in eukaryotes: not only controlling the intra organelle lumen but also actively modifying cytosolic conditions.

Significance

Intracellular transporters are key actors in cell biological processes. Their disruption causes major physiological defects. The role of intracellular ion transporters is usually seen through an “intra organelle” lens, meanwhile their potential action on cytosolic ion homeostasis is still a black box. The case of a plant CLC is used as a model to uncover the missing link between the regulation of conditions inside the vacuole and inside the cytosol. The development of an original live imaging workflow to simultaneously measure pH and anion dynamics in the cytosol reveals the role of an Arabidopsis thaliana CLC, AtCLCa, in the modification of cytosolic pH. Our data highlight an unsuspected function of endomembrane transporters in the regulation of cytosolic pH.
Meta-analysis of longitudinal epigenome-wide association studies of military cohorts reveals multiple CpG sites associated with post-traumatic stress disorder
Authors: Snijders Clara , Maihofer Adam X. , Ratanatharathorn Andrew , Baker Dewleen G. , Boks Marco P. , Geuze Elbert , Jain Sonia , Kessler Ronald C. , Pishva Ehsan , Risbrough Victoria B. , Stein Murray B. , Ursano Robert J. , Vermetten Eric , Vinkers Christiaan H. ,   PGC PTDS EWAS Consortium, Smith Alicia K. , Uddin Monica , Rutten Bart P. F. , Nievergelt Caroline M.
Published: July 2019

Background

Epigenetic mechanisms have been suggested to play a role in the development of post-traumatic stress disorder (PTSD). Here, blood-derived DNA methylation data (HumanMethylation450 BeadChip) collected prior to and following combat exposure in three cohorts composed of male military members were combined to assess whether DNA methylation profiles are associated with the development of PTSD.

Methods

A total of 123 cases and 143 trauma-exposed controls were included. The Psychiatric Genomics Consortium (PGC) PTSD EWAS QC pipeline was used on all cohorts, and results were combined using a sample size weighted meta-analysis. We first combined two cohorts in a discovery stage (N=126 and 78), sought targeted replication in the third cohort (N=62) and then performed a meta-analysis of all three datasets.

Results

The discovery stage identified four CpG sites in which, conditional on pre-deployment DNA methylation, post-deployment DNA methylation was associated with PTSD status after adjustment for multiple comparisons. The most significant CpG (p = 1.0 × 10−08) was located on 5q31 and replicated in the third cohort. When combining all cohorts, this intergenic site remained most significant along with two CpGs located in MAD1L1 and HEXDC. Interestingly, the CpG site of MAD1L1 had an underlying single nucleotide polymorphism (SNP) which was located within the same LD block as a recently identified PTSD-associated SNP. Twelve differential methylated regions (DMRs) were also identified, one of which was located in MAD1L1 and four were situated in the human leukocyte antigen (HLA) region.

Conclusion

This study suggests that the development of PTSD is associated with distinct methylation patterns in several genomic positions and regions. Our most prominent finding points to the involvement of MAD1L1 which was previously associated with PTSD.
Integrating memories: Congruency and reactivation aid memory integration through reinstatement of prior knowledge
Authors: van Kesteren Marlieke T.R. , Rignanese Paul , Gianferrara Pierre G. , Krabbendam Lydia , Meeter Martijn
Published: July 2019

Building consistent knowledge schemas that organize information and guide future learning is of great importance in everyday life. Such knowledge building is suggested to occur through reinstatement of prior knowledge during new learning in stimulus-specific brain regions. This process is proposed to yield integration of new with old memories, supported by the medial prefrontal cortex (mPFC) and medial temporal lobe (MTL). Possibly as a consequence, congruency of new information with prior knowledge is known to enhance subsequent memory. Yet, it is unknown how reactivation and congruency interact to optimize memory integration processes that lead to knowledge schemas. To investigate this question, we here used an adapted AB-AC inference paradigm in combination with functional Magnetic Resonance Imaging (fMRI). Participants first studied an AB-association followed by an AC-association, so B (a scene) and C (an object) were indirectly linked through their common association with A (an unknown pseudoword). BC-associations were either congruent or incongruent with prior knowledge (e.g. a bathduck or a hammer in a bathroom), and participants were asked to report subjective reactivation strength for B while learning AC. Behaviorally, both the congruency and reactivation measures enhanced memory integration. In the brain, these behavioral effects related to univariate and multivariate parametric effects of congruency and reactivation on activity patterns in the MTL, mPFC, and Parahippocampal Place Area (PPA). Moreover, mPFC exhibited larger connectivity with the PPA for more congruent associations. These outcomes provide insights into the neural mechanisms underlying memory integration enhancement, which can be important for educational learning.

Significance statement

How does our brain build knowledge through integrating information that is learned at different periods in time? This question is important in everyday learning situations such as educational settings. Using an inference paradigm, we here set out to investigate how congruency with, and active reactivation of previously learned information affects memory integration processes in the brain. Both these factors were found to relate to activity in memory-related regions such as the medial prefrontal cortex (mPFC) and the hippocampus. Moreover, activity in the parahippocampal place area (PPA), assumed to reflect reinstatement of the previously learned associate, was found to predict subjective reactivation strength. These results show how we can moderate memory integration processes to enhance subsequent knowledge building.
The neural bases of resilient cognitive systems: Evidence of variable neuro-displacement in the semantic system
Authors: Jung JeYoung , Rice Grace E. , Lambon Ralph Matthew A.
Published: July 2019

The purpose of this study was to initiate exploration of an equally-important research goal: what are the neurocomputational mechanisms that make these cognitive systems “well engineered” and thus resilient across a range of performance demands and to mild levels of perturbation or even damage? We achieved this aim by investigating the neural dynamics of the semantic network with two task difficulty manipulations. We found that intrinsic resilience-related mechanisms were observed in both the domain-specific semantic representational system and the parallel executive control networks. Functional connectivity between these regions was also increased and these increases were related to better semantic task performance. Our results suggest that higher cognitive functions are made resilient by flexible, dynamic changes (variable neuro-displacement) across both domain-specific and multi-demand networks. Our findings provide strong evidence that the compensatory functional alterations in the impaired brain might reflect intrinsic mechanisms of a well-engineered neural system.
The Embodied Nature of Well-Timed Behavior
Authors: Safaie Mostafa , Jurado-Parras Maria-Teresa , Sarno Stefania , Louis Jordane , Karoutchi Corane , Petit Ludovic F. , Pasquet Matthieu O. , Eloy Christophe , Robbe David
Published: July 2019

How animals adapt their movements to take advantage of behaviorally-relevant time intervals is not well understood, especially in the supra-second timescale. It has been proposed that motor timing depends on the emergence of self-sustained dynamics across ensembles of neurons. Alternatively, evidence from operant conditioning suggests that animals can develop motor routines to adapt their behavior to fixed temporal constraints. But it is unclear whether animals can accurately time their behavior without the help of motor routines. To address this issue, we used a task in which rats, freely moving on a motorized treadmill, could obtain a reward if they approached it after a fixed interval. Most animals took advantage of the treadmill length and its moving direction to develop, by trial-and-error, a unique motor routine whose execution resulted in the precise timing of their reward approaches. Noticeably, when proficient animals occasionally failed to follow this routine, the timing of their reward approaches was systematically poor. In a second step, we trained naive animals in modified versions of the task specifically designed to prevent the development of this motor strategy. Compared to rats trained in the first protocol, these animals never reached a comparable level of timing accuracy. We conclude that motor timing critically depends on the ability of animals to develop motor routines adapted to the structure of their environment. Our work also suggests that self-sustained neuronal activity alone may not be sufficient to support motor timing, at least in the supra-second timescale.
Post-learning replay of hippocampal-striatal activity is biased by reward-prediction errors
Authors: Roscow Emma L. , Howe Timothy , Lepora Nathan F. , Jones Matthew W.
Published: July 2019

Neural activity encoding recent experiences is replayed during sleep and rest to promote consolidation of memories. However, precisely which features of experience influence replay prioritisation to optimise adaptive behaviour remains unclear. Here, we trained adult male rats on a novel maze-based reinforcement learning task designed to dissociate reward outcomes from reward-prediction errors. Four variations of a reinforcement learning model were fitted to the rats’ behaviour over multiple days. Behaviour was best predicted by a model incorporating replay biased by reward-prediction error, compared to the same model with no replay, random replay or reward-biased replay. Neural population recordings from the hippocampus and ventral striatum of rats trained in the task evidenced preferential reactivation of reward-prediction signals during post-task rest. These insights disentangle the influences of salience on replay, suggesting that reinforcement learning is tuned by post-learning replay biased by reward-prediction error, not by reward per se. This work therefore provides a behavioural and theoretical toolkit with which to measure and interpret the neural mechanisms linking replay and reinforcement learning.
Anisotropic ESCRT-III architecture governs helical membrane tube formation
Authors: von Filseck Joachim Moser , Barberi Luca , Talledge Nathaniel , Johnson Isabel , Frost Adam , Lenz Martin , Roux Aurélien
Published: July 2019

ESCRT-III proteins assemble into ubiquitous membrane-remodeling polymers during many cellular processes. Here we describe the structure of helical membrane tubes that are scaffolded by bundled ESCRT-III filaments. Cryo-ET reveals how the shape of the helical membrane tube arises from the assembly of distinct bundles of protein filaments that bind the membrane with different mean curvatures. Cryo-EM reveals how one of these ESCRT-III filaments engages the membrane tube through a novel interface. Mathematical modeling of the helical membrane tube suggests how its shape emerges from differences in membrane binding energy, positional rigidity, and membrane tension. Altogether, our findings support a model in which increasing the rigidity of ESCRT-III filaments through the assembly of multi-strands triggers buckling of the membrane.

One Sentence Summary

ESCRT-III heteropolymers deform membranes into helical tubes.
Distinct roles for innexin gap junctions and hemichannels in mechanosensation
Authors: Walker Denise S. , Schafer William R.
Published: July 2019

Mechanosensation is central to a wide range of functions, including tactile and pain perception, hearing, proprioception, and control of blood pressure, but identifying the molecules underlying mechanotransduction has proved challenging. In Caenorhabditis elegans, the avoidance response to gentle body touch is mediated by 6 touch receptor neurons (TRNs), and is dependent on MEC-4, a DEG/ENaC channel. We show that hemichannels containing the innexin protein UNC-7 are also essential for gentle touch in the TRNs, as well as harsh touch in both the TRNs and the PVD nociceptors. UNC-7 and MEC-4 do not colocalize, suggesting that their roles in mechanosensory transduction are independent. Heterologous expression of unc-7 in touch-insensitive chemosensory neurons confers ectopic touch sensitivity, indicating a direct role for UNC-7 hemichannels in mechanosensation. The unc-7 touch defect can be rescued by the homologous mouse gene Panx1 gene, thus, innexin/pannexin proteins may play broadly conserved roles in neuronal mechanotransduction.
The 20S proteasome activator PA28γ controls the compaction of chromatin
Authors: Fesquet Didier , Llères David , Grimaud Charlotte , Viganò Cristina , Méchali Francisca , Boulon Séverine , Coux Olivier , Bonne-Andrea Catherine , Baldin Véronique
Published: July 2019

PA28γ, a nuclear activator of the 20S proteasome, is involved in the degradation of several proteins regulating cell growth and proliferation and in the dynamics of various nuclear bodies, but its precise cellular functions remain unclear. Here, using a quantitative FLIM-FRET based microscopy assay monitoring close proximity between nucleosomes in living human cells, we show that PA28γ controls chromatin compaction. We find that its depletion induces a decompaction of pericentromeric heterochromatin, similarly to that observed upon the knockdown of HP1β, a key factor in heterochromatin structure. We show that PA28γ is present at HP1β-containing repetitive-DNA sequences abundant in heterochromatin and importantly, that HP1β on its own is unable to drive chromatin compaction without the presence of PA28γ. At the molecular level, we show that this novel function of PA28γ is independent of its stable interaction with the 20S proteasome, and most likely depends on its ability to maintain appropriate levels of H3K9me3 and H4K20me3, histone modifications that are both involved in heterochromatin formation. Overall, our results implicate PA28γ as a key factor involved in the higher order structuration of chromatin.
Balanced polymorphism at the Pgm-1 locus of the Pompeii worm Alvinella pompejana and its variant adaptability is only governed by two QE mutations at linked sites
Authors: Alexis Bioy , Anne-Sophie Le Port , Emeline Sabourin , Marie Verheye , Patrice Piccino , Baptiste Faure , Stéphane Hourdez , Jean Mary , Didier Jollivet
Published: July 2019

The polychaete Alvinella pompejana lives exclusively on the walls of deep-sea hydrothermal chimneys along the East Pacific Rise and, display specific adaptations to withstand high temperature and hypoxia associated with this highly variable habitat. Previous studies revealed the existence of a balanced polymorphism on the enzyme phosphoglucomutase associated with thermal variations where allozymes 90 and 100 exhibited different optimal activities and thermostabilities. The exploration of the mutational landscape of the phosphoglucomutase1 revealed the maintenance of four highly divergent allelic lineages encoding the three most frequent electromorphs over the worm’s geographic range. This polymorphism is only governed by two linked amino-acid replacements located in exon 3 (E155Q and E190Q). A two-niches model of selection with ‘cold’ and ‘hot’ conditions represents the most likely way for the long-term persistence of these isoforms. Using directed mutagenesis, overexpression of the three recombinant variants allowed us to test the additive effect of these two mutations on the biochemical properties of this enzyme. Results are coherent with those previously obtained from native proteins and reveal a thermodynamic trade-off between the protein thermostability and catalysis, which is likely to have maintained these functional phenotypes prior to the geographic separation of populations across the Equator, about 1.2 Mya.
Leveraging biobank-scale rare and common variant analyses to identify ASPHD1 as the main driver of reproductive traits in the 16p11.2 locus
Authors: Männik Katrin , Arbogast Thomas , Lepamets Maarja , Lepik Kaido , Pellaz Anna , Ademi Herta , Kupchinsky Zachary A , Ellegood Jacob , Attanasio Catia , Messina Andrea , Rotman Samuel , Martin-Brevet Sandra , Dubruc Estelle , Chrast Jacqueline , Lerch Jason P , Qiu Lily R , Laisk Triin ,   The 16p11.2 European Consortium,   The Simons VIP Consortium,   The eQTLGen Consortium, Henkelman R Mark , Jacquemont Sébastien , Herault Yann , Lindgren Cecilia M , Peterson Hedi , Stehle Jean Christophe , Katsanis Nicholas , Kutalik Zoltan , Nef Serge , Draganski Bogdan , Davis Erica E , Mägi Reedik , Reymond Alexandre
Published: July 2019

Whereas genome-wide association studies (GWAS) allowed identifying thousands of associations between variants and traits, their success rate in pinpointing causal genes has been disproportionately low. Here, we integrate biobank-scale phenotype data from carriers of a rare copy-number variant (CNV), Mendelian randomization and animal modeling to identify causative genes in a GWAS locus for age at menarche (AaM). We show that the dosage of the 16p11.2 BP4-BP5 interval is correlated positively with AaM in the UK and Estonian biobanks and 16p11.2 clinical cohorts, with a directionally consistent trend for pubertal onset in males. These correlations parallel an increase in reproductive tract disorders in both sexes. In support of these observations, 16p11.2 mouse models display perturbed pubertal onset and structurally altered reproductive organs that track with CNV dose. Further, we report a negative correlation between the 16p11.2 dosage and relative hypothalamic volume in both humans and mice, intimating a perturbation in the gonadotropin-releasing hormone (GnRH) axis. Two independent lines of evidence identified candidate causal genes for AaM; Mendelian randomization and agnostic dosage modulation of each 16p11.2 gene in zebrafish gnrh3:egfp models. ASPHD1, expressed predominantly in brain and pituitary gland, emerged as a major phenotype driver; and it is subject to modulation by KCTD13 to exacerbate GnRH neuron phenotype. Together, our data highlight the power of an interdisciplinary approach to elucidate disease etiologies underlying complex traits.
Extracellular adenosine enhances the ability of PMNs to killStreptococcus pneumoniaeby inhibiting IL-10 production
Authors: Siwapornchai Nalat , Lee James N. , Tchalla Essi Y. I. , Bhalla Manmeet , Yeoh Jun Hui , Roggensack Sara E. , Leong John M. , Bou Ghanem Elsa N.
Published: July 2019

PMNs are crucial for initial control ofStreptococcus pneumoniae(pneumococcus) lung infection; however, as the infection progresses their persistence in the lungs becomes detrimental. Here we explored why the anti-microbial efficacy of PMNs declines over the course of infection. We found that the progressive inability of PMNs to control infection correlated with phenotypic differences characterized by a decrease in CD73 expression, an enzyme required for production of extracellular adenosine (EAD). EAD production by CD73 was crucial for the ability of both murine and human PMNs to killS. pneumoniae.In exploring the mechanisms by which CD73 controlled PMN function, we found that CD73 mediated its anti-microbial activity by inhibiting IL-10 production. PMNs from wild type mice did not increase IL-10 production in response toS. pneumoniae,however, CD73-/-PMNs up-regulated IL-10 production upon pneumococcal infectionin vitroand during lung challenge. IL-10 inhibited the ability of wild type PMNs to kill pneumococci. Conversely, blocking IL-10 boosted the bactericidal activity of CD73-/-PMNs as well as host resistance of CD73-/-mice to pneumococcal pneumonia. CD73/IL-10 did not affect apoptosis, bacterial uptake and intracellular killing or production of anti-microbial Neutrophil Elastase and Myeloperoxidase. Rather, inhibition of IL-10 production by CD73 was important for optimal ROS production by PMNs. ROS contributed to PMN anti-microbial function as their removal or detoxification impaired the ability of PMNs to efficiently killS. pneumoniae. This study demonstrates that CD73 controls PMN anti-microbial phenotype duringS. pneumoniaeinfection.
Targeting dormant ovarian cancer cellsin vitroand in anin vivomodel of platinum resistance
Authors: Huang Zhiqing , Kondoh Eiji , Visco Zachary , Baba Tsukasa , Matsumura Noriomi , Dolan Emma , Whitaker Regina S. , Konishi Ikuo , Fujii Shingo , Berchuck Andrew , Murphy Susan K.
Published: July 2019

Objective

Ovarian cancer cells often existin vivoas multicellular spheroids. Spheroid formationin vitrohas been used to enrich for cancer stem cell populations from primary tumors. Such spheroids exhibit drug resistance and slow proliferation, suggesting involvement in disease recurrence. Our objectives were to characterize cancer spheroid phenotypes, determine gene expression profiles associated with spheroid forming capacity and to evaluate the responsiveness of spheroids to commonly used and novel therapeutic agents.

Methods

Tumorigenic potential was assessed using anchorage independent growth assays in 24 cell lines. Spheroids from cell lines (N=12) and from primary cancers (N=8) were grown on non-adherent tissue culture plates in serum-free media. Cell proliferation was measured using MTT assays and Ki67 immunostaining. Affymetrix HT U133A gene expression data was used to identify differentially expressed genes based on spheroid forming capacity. Matched monolayers and spheroids (N=7 pairs) were tested for response to cisplatin, paclitaxel and 7-hydroxystaurosporine (UCN-01) while mitochondrial inhibition was performed using oligomycin. Xenograft tumors from intraperitoneal injection of CAOV2-GFP/LUC ovarian cancer cells into nude mice were treated with carboplatin to reduce tumor burden followed by secondary treatment with carboplatin, UCN-01, or Oltipraz. Tumor formation and response was monitored using live imaging.

Results

Of 12 cell lines with increased anchorage-independent growth, 8 also formed spheroids under serum-free spheroid culture conditions. Spheroids showed reduced proliferation (p<0.0001) and Ki67 immunostaining (8% versus 87%) relative to monolayer cells. Spheroid forming capacity was associated with increased mitochondrial pathway activity (p ≤ 0.001). The mitochondrial inhibitors, UCN-01 and Oligomycin, demonstrated effectiveness against spheroids, while spheroids were refractory to cisplatin and paclitaxel. By livein vivoimaging, ovarian cancer xenograft tumors were reduced after primary treatment with carboplatin. Continued treatment with carboplatin was accompanied by an increase in tumor signal while there was little or no increase in tumor signal observed with subsequent treatment with UCN-01 or Oltipraz.

Conclusions

Our findings suggest that the mitochondrial pathway in spheroids may be an important therapeutic target in preventing disease recurrence.
Functional innovation in the evolution of the calcium-dependent system of the eukaryotic endoplasmic reticulum
Authors: Schäffer Daniel E. , Iyer Lakshminarayan M. , Burroughs A. Maxwell , Aravind L.
Published: July 2019

The origin of eukaryotes was marked by the emergence of several novel subcellular systems. One such is the calcium (Ca2+)-stores system of the endoplasmic reticulum, which profoundly influences diverse aspects of cellular function including signal transduction, motility, division, and biomineralization. We use comparative genomics and sensitive sequence and structure analyses to investigate the evolution of this system. Our findings reconstruct the core form of the Ca2+- stores system in the last eukaryotic common ancestor as having at least 15 proteins that constituted a basic system for facilitating both Ca2+flux across endomembranes and Ca2+-dependent signaling. We present evidence that the key EF-hand Ca2+-binding components had their origins in a likely bacterial symbiont other than the mitochondrial progenitor, whereas the protein phosphatase subunit of the ancestral calcineurin complex was likely inherited from the asgardarchaeal progenitor of the stem eukaryote. This further points to the potential origin of the eukaryotes in a Ca2+-rich biomineralized environment such as stromatolites. We further show that throughout eukaryotic evolution there were several acquisitions from bacteria of key components of the Ca2+-stores system, even though no prokaryotic lineage possesses a comparable system. Further, using quantitative measures derived from comparative genomics we show that there were several rounds of lineage-specific gene expansions, innovations of novel gene families, and gene losses correlated with biological innovation such as the biomineralized molluscan shells, coccolithophores, and animal motility. The burst of innovation of new genes in animals included the wolframin protein associated with Wolfram syndrome in humans. We show for the first time that it contains previously unidentified Sel1, EF-hand, and OB-fold domains, which might have key roles in its biochemistry.
Targeting Toxic Nuclear RNA Foci with CRISPR-Cas13 to Treat Myotonic Dystrophy
Authors: Poosala Pornthida , Lindley Sean R. , Anderson Kelly M. , Anderson Douglas M.
Published: July 2019

Human monogenetic diseases can arise from the aberrant expansion of tandem nucleotide repeat sequences, which when transcribed into RNA, can misfold and aggregate into toxic nuclear foci1. Nuclear retention of repeat-containing RNAs can disrupt their normal expression and induce widespread splicing defects by sequestering essential RNA binding proteins. Among the most prevalent of these disorders is myotonic dystrophy type 1 (DM1), a disease occurring from the expression of a noncoding CTG repeat expansion in the 3’UTR of the human dystrophia myotonica protein kinase (DMPK) gene2,3. Here we show that RNA-binding CRISPR-Cas13, with a robust non-classical nuclear localization signal, can be efficiently targeted to toxic nuclear RNA foci for either visualization or cleavage, tools we named hilightR and eraseR, respectively. HilightR combines catalytically dead Cas13b (dCas13b) with a fluorescent protein to directly visualize CUG repeat RNA foci in the nucleus of live cells, allowing for quantification of foci number and observation of foci dynamics. EraseR utilizes the intrinsic endoribonuclease activity of Cas13b, targeted to nuclear CUG repeat RNA, to disrupt nuclear foci. These studies demonstrate the potential for targeting toxic nuclear RNA foci directly with CRISPR-Cas13 for either the identification or treatment of DM1. The efficient and sequence programmable nature of CRISPR-Cas13 systems will allow for rapid targeting and manipulation of other human nuclear RNA disorders, without the associated risks of genome editing.
Dopamine in the basal amygdala signals salient somatosensory events during fear learning
Authors: Tang Wei , Kochubey Olexiy , Kintscher Michael , Schneggenburger Ralf
Published: July 2019

The amygdala is a brain area critical for the formation of threat memories. However, the nature of the teaching signal(s) that drive plasticity in the amygdala are still under debate. Here, we use optogenetic methods to investigate whether dopamine release in the amygdala contributes to fear learning. Antero- and retrograde labeling showed that a sparse, and relatively evenly distributed population of ventral tegmental area (VTA) neurons projects to the basal amygdala (BA). In-vivo optrode recordings in behaving mice showed that many VTA neurons, amongst them putative dopamine neurons, are excited by footshocks. Correspondingly, in-vivo fiber photometry of dopamine in the BA revealed robust dopamine concentration transients upon footshock presentation. Finally, silencing VTA dopamine neurons, or their axon terminals in the BA during the footshock, reduced the extent of threat memory retrieval one day later. Thus, VTA dopamine neurons projecting to the BA code for the saliency of the footshock event, and the resulting dopamine release in the BA facilitates threat memory formation.
Withdraw of prophylactic antimicrobials does not change the pigs’ resistome
Authors: Fernanda Loayza-Villa , Alejandro Torres , Lixin Zhang , Gabriel Trueba
Published: July 2019

The use of antimicrobials in the animal industry has increased the prevalence of antimicrobial resistant commensal bacteria in food products derived from animals, which could be associated with antimicrobial resistance in human pathogens. To reduce the influx of antibiotic resistant bacteria (and genes) to the human microbiota, restrictions on antimicrobials (in food animals) have been implemented in different countries. We investigated the impact of antimicrobial restriction in the frequency of antimicrobial resistant bacteria in pigs. No differences in antimicrobial resistance or antimicrobial resistance genes (richness or abundance) was found when we compared animals fed with and without antibiotics. Fitness costs of antimicrobial resistance in bacteria (in the field) seems to be overestimated.
Use of multivariable Mendelian randomization to address biases due to competing risk before recruitment
Authors: Schooling C Mary , Lopez Priscilla M , Yang Zhao , Zhao J V , Au Yeung SL , Huang Jian V
Published: July 2019

Background

Mendelian randomization (MR) provides unconfounded estimates. MR is open to selection bias particularly when the underlying sample is selected on surviving the genetically instrumented exposure and other conditions that share etiology with the outcome (competing risk before recruitment). Few methods to address this bias exist.

Methods

We use directed acyclic graphs to show this selection bias can be addressed by adjusting for common causes of survival and outcome. We use multivariable MR to obtain a corrected MR estimate, specifically, the effect of statin use on ischemic stroke, because statins affect survival and stroke typically occurs later in life than ischemic heart disease so is open to competing risk.

Results

In univariable MR the genetically instrumented effect of statin use on ischemic stroke was in a harmful direction in MEGASTROKE and the UK Biobank (odds ratio (OR) 1.33, 95% confidence interval (CI) 0.80 to 2.20). In multivariable MR adjusted for major causes of survival and ischemic stroke, (blood pressure, body mass index and smoking initiation) the effect of statin use on stroke in the UK Biobank was as expected (OR 0.81, 95% CI 0.68 to 0.98) with a Q-statistic indicating absence of genetic pleiotropy or selection bias, but not in MEGASTROKE.

Conclusion

MR studies concerning late onset chronic conditions with shared etiology based on samples recruited in later life need to be conceptualized within a mechanistic understanding, so as to any identify potential bias due to competing risk before recruitment, and to inform the analysis and interpretation.
The single-progenitor model as the unifying paradigm of squamous epithelial maintenance
Authors: Piedrafita Gabriel , Kostiou Vasiliki , Wabik Agnieszka , Colom Bartomeu , Fernandez-Antoran David , Herms Albert , Murai Kasumi , Hall Benjamin A , Jones Philip H
Published: July 2019

Adult tissues such as the epidermis of the skin and the epithelium lining the esophagus are continuously turned over throughout life. Cells are shed from the tissue surface and replaced by cell division. Yet, the cellular mechanisms that underpin these tissues homeostasis remain poorly established, having important implications for wound healing and carcinogenesis. Lineage tracing, in which of a cohort of proliferating cells and their descendants are genetically labelled in transgenic mice, has been used to study the fate behavior of the proliferating cells that maintain these tissues. However, based on this technique, distinct mutually irreconcilable models, differing in the implored number and hierarchy of proliferating cell types, have been proposed to explain homeostasis. To elucidate which of these conflicting scenarios should prevail, here we performed cell proliferation assays across multiple body sites in transgenic H2BGFP mouse epidermis and esophagus. Cell-cycle properties were then extracted from the H2BGFP dilution kinetics and adopted in a common analytic approach for a refined analysis of a new lineage-tracing experiment and eight published clonal data sets from esophagus and different skin territories. Our results show H2BGFP dilution profiles remained unimodal over time, indicating the absence of slow-cycling stem cells across all tissues analyzed. We find that despite using diverse genetic labelling approaches, all lineage-tracing data sets are consistent with tissues maintenance by a single population of proliferating cells. The outcome of a given division is unpredictable but, on average the likelihood of producing proliferating and differentiating cells is balanced, ensuring tissue homeostasis. The fate outcomes of sister cells are anticorrelated. We conclude a single cell population maintains squamous epithelial homeostasis.
Uncoupling sodium channel dimers rescues phenotype of pain-linked Nav1.7 mutation
Authors: Rühlmann Annika H. , Körner Jannis , Bebrivenski Nikolay , Detro-Dassen Silvia , Hautvast Petra , Benasolo Carène A. , Meents Jannis , Machtens Jan-Philipp , Schmalzing Günther , Lampert Angelika
Published: July 2019

The voltage-gated sodium channel Nav1.7 is essential for an adequate perception of painful stimuli. Its mutations cause various pain syndromes in human patients. The hNav1.7/A1632E mutation induces symptoms of erythromelalgia and paroxysmal extreme pain disorder (PEPD), and its main gating change is a strongly enhanced persistent current.

Using molecular simulations, we demonstrate that the disease causing persistent current of hNav1.7/A1632E is due to impaired binding of the IFM motif, thus affecting proper function of the recently proposed allosteric fast inactivation mechanism. By using native polyacrylamide gel electrophoresis (PAGE) gels, we show that hNav1.7 dimerizes. The disease-linked persistent current depends on the channel’s functional dimerization status: Using difopein, a 14-3-3 inhibitor known to uncouple dimerization of hNav1.5, we detect a significant decrease in hNav1.7/A1632E induced persistent currents.

Our work identifies that functional uncoupling of hNav1.7/A1632E dimers rescues the pain-causing molecular phenotype by interferes with an allosteric fast inactivation mechanism, which we link for the first time to channel dimerization. Our work supports the concept of sodium channel dimerization and reveals its relevance to human pain syndromes.
Quantitative, super-resolution localization of small RNAs with sRNA-PAINT
Authors: Huang Kun , Demirci Feray , Batish Mona , Meyers Blake C. , Caplan Jeffrey L.
Published: July 2019

Small RNAs are non-coding RNAs that play important roles in the lives of both animals and plants. They are 21- to 24-nt in length and around 10 nanometers (nm) in size. Their small size and high diversity have made it challenging to develop detection methods that have sufficient resolution and specificity to multiplex and quantify. We created a method, sRNA-PAINT, for the detection of small RNAs with 20 nm resolution by combining the super-resolution method, DNA-based points accumulation in nanoscale topography (DNA-PAINT), and the specificity of locked nucleic acid (LNA) probes for thein situdetection of multiple small RNAs. The method relies on designing probes to target small RNAs that combine DNA oligonucleotides (oligos) for PAINT with LNA-containing oligos for hybridization; therefore, we developed an online tool called “Vetting & Analysis of RNA forin situHybridization probes” (VARNISH) for probe design. Our method utilizes advances in DNA-PAINT methodologies, including qPAINT for quantification, and Exchange-PAINT for multiplexing. We demonstrated these capabilities of sRNA-PAINT by detecting and quantifying small RNAs in different cell layers of early developmental stage maize anthers that are important for male sexual reproduction.