bluuebunny/crossref_metadata_2025 huggingface dataset:
Genome-wide analysis identifies cis-acting elements regulating mRNA polyadenylation and translation during vertebrate oocyte maturation
Authors: Yang Fei , Wang Wei , Cetinbas Murat , Sadreyev Ruslan I. , Blower Michael D.
Published: July 2019

Changes in gene expression are required to orchestrate changes in cell state during development. Most cells change patterns of gene expression through transcriptional regulation. In contrast, oocytes are transcriptionally silent and use changes in mRNA poly-A tail length to control protein production. Poly-A tail length is positively correlated with translation activation during early development. However, it is not clear how poly-A tail changes affect mRNA translation at a during vertebrate oocyte maturation. We used Tail-seq and polyribosome analysis to measure poly-A tail and translational changes during oocyte maturation in Xenopus laevis. We identified large-scale poly-A and translational changes during oocyte maturation and found that poly-A tail changes precede translation changes. Additionally, we identified a family of U-rich sequence elements that are enriched near the polyadenylation signal of polyadenylated and translationally activated mRNAs. A modest density of U-rich elements was correlated with polyadenylation while a high density of U-rich elements was required to activate translation, showing that polyadenylation and translation activation can be uncoupled. Collectively, our data show that changes in mRNA polyadenylation are a key mechanism regulating protein expression during vertebrate oocyte maturation and that these changes are controlled by a spatial code of cis-acting sequence elements. Our results provide insight into mechanisms of translational control in oocytes and identify novel proteins important for the completion of meiosis.
Fungal seed pathogens of wild chili peppers possess multiple mechanisms to tolerate capsaicinoids
Authors: Adams Catharine A. , Zimmerman Kolea , Fenstermacher Kristi , Thompson Mitchell G. , Skyrud Will , Behie Scott , Pringle Anne
Published: July 2019

The wild chili pepper Capsicum chacoense produces the spicy defense compounds known as capsaicinoids, including capsaicin and dihydrocapsaicin, antagonistic to the growth of fungal pathogens. Compared to other microbes, fungi isolated from infected seeds of C. chacoense possess much higher tolerance to these spicy compounds, having their growth slowed, but not entirely inhibited. Previous research has shown capsaicinoids inhibit microbes by disrupting ATP production via the binding of NADH dehydrogenase in the Electron Transport Chain (ETC), throttling Oxidative Phosphorylation (OXPHOS). Capsaicinoids may also disrupt cell membranes. Here, we investigated capsaicinoid tolerance in fungal seed pathogens isolated from C. chacoense. We selected 16 fungal isolates from four Ascomycete genera (Alternaria, Colletotrichum, Fusarium and Phomopsis). Using relative growth rate as a readout for tolerance, fungi were challenged with ETC inhibitors to infer if fungi possess alternative respiratory enzymes, and if effects on the ETC fully explained inhibition by capsaicinoids. In all isolates, we found evidence for at least one alternative NADH dehydrogenase. In many isolates we also found evidence for an alternative oxidase. These data suggest wild plant pathogens may be a rich source of alternative respiratory enzymes. We further demonstrate these fungal isolates are capable of the breakdown of capsaicinoids. Lastly, we determine the OXPHOS theory weakly explains the primary mechanism by which dihydrocapsaicin slows fungal growth, but not capsaicin. Our findings suggest capsaicinoids likely disrupt membranes in addition to energy poisoning, with implications for microbiology and human health.

Importance

Plants make chemical compounds to protect themselves. For example, chili peppers produce the spicy compound capsaicin to inhibit animal feeding and pathogen damage. In humans, capsaicin binds to a membrane channel protein, creating the sensation of heat, while in microbes, capsaicin limits energy production by binding respiratory enzymes. However, some data suggest capsaicin also disrupts membranes. Here we studied fungal pathogens (Alternaria, Colletotrichum, Fusarium, and Phomopsis) isolated from a wild chili pepper, Capsicum chacoense. By measuring growth rate in the presence of antibiotics with known respiratory targets, we infer wild plant pathogens may be rich with alternative respiratory enzymes. A zone of clearance around the colonies, as well as LCMS data, further indicate these fungi can break down capsaicin. Lastly, the total inhibitory effect of capsaicin was not fully explained by its effect on respiratory enzymes. Our findings lend credence to studies proposing capsaicin may disrupt cell membranes, with implications for microbiology as well as human health.
A small membrane protein critical to both the offensive and defensive capabilities of Staphylococcus aureus
Authors: Duggan Seána , Laabei Maisem , Alnahari Alaa , O’Brien Eóin C. , Lacey Keenan A. , Bacon Leann , Heesom Kate , Fu Chih-Lung , Otto Michael , Skaar Eric , McLoughlin Rachel M. , Massey Ruth C.
Published: July 2019

Staphylococcus aureus is a major human pathogen, where the widespread emergence of antibiotic resistance is making infections more challenging to treat. Toxin induced tissue damage and resistance to the host’s immune system are well established as critical to its ability to cause disease. However, recent attempts to study S. aureus pathogenicity at a population level have revealed significant complexity and hierarchical levels of regulation. In an effort to better understand this we have identified and characterized a principle effector protein, MasA. The inactivation of this small highly-conserved membrane protein simultaneously disrupts toxin production and impairs S. aureus’ ability to resist several aspects of the innate immune system. These pleiotropic effects are mediated by both a change in the stability of the bacterial membrane and the dysregulation of iron homeostasis, which results in a significant impairment in the ability of S. aureus to cause infection in both a subcutaneous and a sepsis model of infection. That proteins with such major effects on pathogenicity remain unidentified in a bacterium as well studied as S. aureus demonstrates how incomplete our understanding of their ability to cause disease is, an issue that needs to be addressed if effective control and treatment strategies are to be developed.
Composite plants for a composite plant: An efficient protocol for root studies in the sunflower using composite plants approach
Authors: Parks Tyler , Yordanov Yordan S.
Published: July 2019

Sunflower (Helianthus annuus L.) is important oilseed crop in the world and the sunflower oil is prized for its’ exceptional quality and flavor. The recent availability of the sunflower genome can allow genome-wide characterization of genes and gene families. With plant transformation usually being the rate limiting step for gene functional studies of sunflower, composite plants can alleviate this bottleneck. Composite plants, produced using Agrobacterium rhizogenes, are plants with transgenic roots and wild type shoots. Composite plants offer benefits over creating fully transgenic plants, namely time and cost. Here we outlined the critical steps and parameters for a protocol for the sunflower composite plants production. We use more than a dozen genotypes and three constitutive promoters to validates the utility and efficiency of this protocol. Moreover, functional gene characterization by overexpression and RNAi silencing of a root related transcription factor HaLBD16 further emphasize the value of the system in the sunflower studies. With the protocol developed here an experiment can be carried out with efficiency and in only two months. This procedure adds to arsenal of approaches for the functional genetics/genomics in sunflower for characterization candidate genes involved in root development and stress adaptation.

Key message

Composite plants technique described here is fast and efficient approach for roots functional studies in sunflower.
Prophylactic efficacy of 5-HT4R agonists against stress
Authors: Chen Briana K. , Mendez-David Indira , Faye Charlene , Gardier Alain M. , David Denis J. , Denny Christine A.
Published: July 2019

Enhancing stress resilience could protect against stress-induced psychiatric disorders in at-risk populations. We and others have previously reported that (R,S)-ketamine acts as a resilience enhancing drug (e.g., prophylactic) against stress when administered 1 week before stress. While we have shown that the selective 5-hydroxytryptamine (5-HT) (serotonin) reuptake inhibitor (SSRI) fluoxetine (Flx) is ineffective as a prophylactic, it remains to be determined if other serotonergic drugs could be effective prophylactics. Here, we hypothesized that serotonin 4 receptor (5-HT4R) agonists could be prophylactic against fear, depressive-like, and/or anxiety-like behavior. We tested if three 5-HT4R agonists with varying affinity (e.g., partial or selective agonists) could protect against stress in two mouse strains by utilizing a chronic corticosterone (CORT) administration or a contextual fear conditioning (CFC) paradigm. Mice were administered RS-67,333, prucalopride, or PF-04995274 at varying doses and then 1 week later were subjected to chronic CORT or CFC. Chronic administration of RS-67,333, but not Flx was efficacious as a prophylactic against CORT in C57BL/6NTac mice. A single injection of RS-67,333 attenuated learned fear in male, but not female 129S6/SvEv mice. RS-67,333 was ineffective against stress-induced depressive-like behavior in the forced swim test (FST). A single injection of either prucalopride or PF-04995274 attenuated learned fear and decreased stress-induced depressive-like behavior. These data show that in addition to (R,S)-ketamine, 5-HT4R agonists are also effective prophylactics against stress, suggesting that the 5-HT4R may be a novel target for prophylactic drug development.
ROCK Inhibitor Increases Proacinar Cells in Adult Salivary Gland Organoids
Authors: Koslow Matthew , O’Keefe Kevin J. , Nelson Deirdre A. , Larsen Melinda
Published: July 2019

Salisphere-derived adult epithelial cells enriched for progenitor cells have been used to improve saliva production of irradiated mouse salivary glands. Importantly, optimization of the cellular composition of salispheres could improve their regenerative capabilities. The Rho Kinase (ROCK)1 inhibitor, Y27632, has been used to increase the proliferation and reduce apoptosis of progenitor cells grown in vitro. In this study, we investigated whether Y27632 in different cell media contexts could be used to improve expansion of adult epithelial progenitor cells or to affect their differentiation potential. Application of Y27632 in medium used previously to grow salispheres promoted expansion of Kit+ cells, while in simple serum-containing medium Y27632 increased the number of cells that expressed the K5 basal progenitor marker. When tested in a 3D organoid assay, Y27632 enhanced the contribution of adult salispheres to salivary organoids expressing the secretory proacinar marker Aquaporin 5 (AQP5) in response to FGF2 dependent mesenchymal signals. Optimization of epithelial-mesenchymal interactions organoids can be used to improve application of adult salivary progenitor cells in regenerative medicine strategies.

Highlights

Y27632 promotes Kit+ salisphere cell proliferation in salisphere media.

Y27632 promotes K5 expression in salispheres cultured in serum containing media.

Y27632 treated Kit+ salispheres form salivary organoids expressing AQP5.
Dynamic changes in RNA-chromatin interactome promote endothelial dysfunction
Authors: Calandrelli Riccardo , Xu Lixia , Luo Yingjun , Wu Weixin , Fan Xiaochen , Nguyen Tri , Chen Chienju , Sriram Kiran , Natarajan Rama , Bouman-Chen Zhen , Zhong Sheng
Published: July 2019

Chromatins are pervasively attached by RNAs. Here, we asked whether global RNA-chromatin contacts are altered in a given cell type in a disease context, and whether these alterations impact gene expression and cell function. In endothelial cells (ECs) treated by high-glucose and TNFα, we employed single-cell RNA-sequencing and in situ mapping of RNA-genome interaction (iMARGI) assay to delineate temporal changes in transcriptome and RNA-chromatin interactome. ECs displayed dramatic and heterogeneous changes in single cell transcriptome, accompanied by a dynamic and strong increase in inter-chromosomal RNA-DNA interactions, particularly among super enhancers (SEs). These SEs overlap with genes contributing to inflammatory response and endothelial mesenchymal transition (EndoMT), two key aspects of endothelial dysfunction. Perturbation of a high-glucose and TNFα-activated interaction involving SEs overlapping LINC00607 and SERPINE1 attenuated the pro-inflammatory and pro-EndoMT gene induction and EC dysfunction. Our findings highlight RNA-chromatin contacts as a crucial regulatory feature in biological and disease processes, exemplified by endothelial dysfunction, a major mediator of numerous diseases.
Diverse biological processes coordinate the transcriptional response to nutritional changes in aDrosophila melanogastermultiparent population
Authors: Ng’oma E. , Williams-Simon P.A. , Rahman A. , King E.G.
Published: July 2019

Background

Environmental variation in the amount of resources available to populations challenge individuals to optimize the allocation of those resources to key fitness functions. This coordination of resource allocation relative to resource availability is commonly attributed to key nutrient sensing gene pathways in laboratory model organisms, chiefly the insulin/TOR signaling pathway. However, the genetic basis of diet-induced variation in gene expression is less clear.

Results

To describe the natural genetic variation underlying nutrient-dependent differences, we used an outbred panel derived from a multiparental population, theDrosophilaSynthetic Population Resource. We analyzed RNA sequence data from multiple female tissue samples dissected from flies reared in three nutritional conditions: high sugar (HS), dietary restriction (DR), and control (C) diets. A large proportion of genes in the experiment (19.6% or 2,471 genes) were significantly differentially expressed for the effect of diet, 7.8% (978 genes) for the effect of the interaction between diet and tissue type (LRT,Padj.< 0.05). Interestingly, we observed similar patterns of gene expression relative to the C diet, in the DR and HS treated flies, a response likely reflecting diet component ratios. Hierarchical clustering identified 21 robust gene modules showing intra-modularly similar patterns of expression across diets, all of which were highly significant for diet or diet-tissue interaction effects (false discovery rate, FDRPadj.< 0.05). Gene set enrichment analysis for different diet-tissue combinations revealed a diverse set of pathways and gene ontology (GO) terms (two-sample t-test, FDR < 0.05). GO analysis on individual co-expressed modules likewise showed a large number of terms encompassing a large number of cellular and nuclear processes (Fisher exact test,Padj.< 0.01). Although a handful of genes in the IIS/TOR pathway includingIlp5,Rheb, andSirt2showed significant elevation in expression, known key genes such asInR,chico, insulin peptide genes, and the nutrient-sensing pathways were not observed.

Conclusions

Our results suggest that a more diverse network of pathways and gene networks mediate the diet response in our population. These results have important implications for future studies focusing on diet responses in natural populations.
A few stickleback suffice for the transport of alleles to new lakes
Authors: Galloway Jared , Cresko William A. , Ralph Peter
Published: July 2019

Threespine stickleback populations provide a striking example of local adaptation to divergent habitats in populations that are connected by recurrent gene flow. These small fish occur in marine and freshwater habitats throughout the Northern Hemisphere, and in numerous cases the smaller freshwater populations have been established “de novo” from marine colonists. Independently evolved freshwater populations show similar phenotypes that have been shown to derive largely from the same standing genetic variants. Geographic isolation prevents direct migration between the freshwater populations, strongly suggesting that these shared locally adaptive alleles are transported through the marine population. However it is still largely unknown how gene flow, recombination, and selection jointly impact the standing variation that might fuel this adaptation. Here we use individual-based, spatially explicit simulations to determine the levels of gene flow that best match observed patterns of allele sharing among habitats in stickleback. We aim to better understand how gene flow and local adaptation in large metapopulations determine the speed of adaptation and re-use of standing genetic variation. In our simulations we find that repeated adaptation uses a shared set of alleles that are maintained at low frequency by migration-selection balance in oceanic populations. This process occurs over a realistic range of intermediate levels of gene flow that match previous empirical population genomic studies in stickleback. Examining these simulations more deeply reveals how lower levels of gene flow leads to slow, independent adaptation to different habitats, whereas higher levels gene flow leads to significant mutation load – but an increased probability of successful population genomic scans for locally adapted alleles. Surprisingly, we find that the genealogical origins of most freshwater adapted alleles can be traced back to the original generation of marine individuals that colonized the lakes, as opposed to subsequent migrants. These simulations provide deeper context for existing studies of stickleback evolutionary genomics, and guidance for future empirical studies in this model. More broadly, our results support existing theory of local adaptation but extend it by more completely documenting the genealogical history of adaptive alleles in a metapopulation.
Genomic phylogeography of the White-crowned ManakinPseudopipra pipra(Aves: Pipridae) illuminates a continental-scale radiation out of the Andes
Authors: Berv Jacob S. , Campagna Leonardo , Feo Teresa J. , Castro-Astor Ivandy , Ribas Camila C. , Prum Richard O. , Lovette Irby J.
Published: July 2019

The complex landscape history of the Neotropics has generated opportunities for population isolation and diversification that place this region among the most species-rich in the world. Detailed phylogeographic studies are required to uncover the biogeographic histories of Neotropical taxa, to identify evolutionary correlates of diversity, and to reveal patterns of genetic connectivity, disjunction, and potential differentiation among lineages from different areas of endemism. The White-crowned Manakin (Pseudopipra pipra) is a small suboscine passerine bird that is broadly distributed through the subtropical rainforests of Central America, the lower montane cloud forests of the Andes from Colombia to central Peru, the lowlands of Amazonia and the Guianas, and the Atlantic forest of southeast Brazil.Pseudopiprais currently recognized as a single, polytypic biological species. We studied the effect of the Neotropical landscape on genetic and phenotypic differentiation within this species using genomic data derived from double digest restriction site associated DNA sequencing (ddRAD), and mitochondrial DNA. Most of the genetic breakpoints we identify among populations coincide with physical barriers to gene flow previously associated with avian areas of endemism. The phylogenetic relationships among these populations imply a novel pattern of Andean origination for this group, with subsequent diversification into the Amazonian lowlands. Our analysis of genomic admixture and gene flow reveals a complex history of introgression between some western Amazonian populations. These reticulate processes confound our application of standard concatenated and coalescent phylogenetic methods and raise the question of whether a lineage in the western Napo area of endemism should be considered a hybrid species. Lastly, analysis of variation in vocal and plumage phenotypes in the context of our phylogeny supports the hypothesis thatPseudopiprais a species-complex composed of at least 8, and perhaps up to 17 distinct species which have arisen in the last ∼2.5 Ma.
Identification ofkit-ligand aas the Gene Responsible for the Medaka Pigment Cell Mutantfew melanophore
Authors: Otsuki Yuji , Okuda Yuki , Naruse Kiyoshi , Saya Hideyuki
Published: July 2019

The body coloration of animals is due to pigment cells derived from neural crest cells, which are multipotent and differentiate into diverse cell types. Medaka (Oryzias latipes) possesses four distinct types of pigment cells known as melanophores, xanthophores, iridophores, and leucophores. Thefew melanophore(fm) mutant of medaka is characterized by reduced numbers of melanophores and leucophores. We here identifykit-ligand a(kitlga) as the gene whose mutation gives rise to thefmphenotype. This identification was confirmed by generation ofkitlgaknockout medaka and the findings that these fish also manifest reduced numbers of melanophores and leucophores and fail to rescue thefmmutant phenotype. We also found that expression ofsox5,pax7a,pax3a, andmitfagenes is down-regulated in bothfmandkitlgaknockout medaka, implicating c-Kit signaling in regulation of the expression of these genes as well as the encoded transcription factors in pigment cell specification. Our results may provide insight into the pathogenesis of c-Kit–related pigmentation disorders such as piebaldism in humans, and ourkitlgaknockout medaka may prove useful as a tool for drug screening.
Discovery and Quality Analysis of a Comprehensive Set of Structural Variants and Short Tandem Repeats
Authors: Jakubosky David , Smith Erin N. , D’Antonio Matteo , Bonder Marc Jan , Greenwald William W. Young , D’Antonio-Chronowska Agnieszka , Matsui Hiroko ,   i2QTL Consortium,   HipSci Consortium, Stegle Oliver , Montgomery Stephen B. , DeBoever Christopher , Frazer Kelly A.
Published: July 2019

Structural variants (SVs) and short tandem repeats (STRs) are important sources of genetic diversity but are not routinely analyzed in genetic studies because they are difficult to accurately identify and genotype. Because SVs and STRs range in size and type, it is necessary to apply multiple algorithms that incorporate different types of evidence from sequencing data and employ complex filtering strategies to discover a comprehensive set of high-quality and reproducible variants. Here we assembled a set of 719 deep whole genome sequencing (WGS) samples (mean 42x) from 477 distinct individuals which we used to discover and genotype a wide spectrum of SV and STR variants using five algorithms. We used 177 unique pairs of genetic replicates to identify factors that affect variant call reproducibility and developed a systematic filtering strategy to create of one of the most complete and well characterized maps of SVs and STRs to date.
Sub-sampling of cues in associative learning
Authors: Perez Omar D. , Vogel Edgar H. , Narasiwodeyar Sanjay , Soto Fabian A.
Published: July 2019

Theories of learning distinguish between elemental and configural stimulus processing depending on whether stimuli are processed independently or as whole configurations. Evidence for elemental processing comes from findings of summation in animals where a compound of two dissimilar stimuli is deemed to be more predictive than each stimulus alone, whereas configural processing is supported by experiments employing similar stimuli in which summation is not found. However, in humans the summation effect is robust and impervious to similarity manipulations. In three experiments in human predictive learning, we show that summation can be obliterated when partially reinforced cues are added to the summands in training and test. This lack of summation only holds when the partially reinforced cues are similar to the reinforced cues (Experiment 1) and seems to depend on participants sampling only the most salient cue in each trial (Experiments 2a and 2b) in a sequential visual search process. Instead of attributing our and other’s instances of lack of summation to the customary idea of configural processing, we offer a formal sub-sampling rule that might be applied to situations in which the stimuli are hard to parse from each other.
Passive transfer of fibromyalgia pain from patients to mice
Authors: Goebel Andreas , Gentry Clive , Cuhadar Ulku , Krock Emerson , Vastani Nisha , Sensi Serena , Sandor Katalin , Jurczak Alexandra , Baharpoor Azar , Brieskorn Louisa , Urbina Carlos Morado , Sandstrom Angelika , Tour Janette , Kadetoff Diana , Kosek Eva , Bevan Stuart , Svensson Camilla I. , Andersson David A.
Published: July 2019

Fibromyalgia syndrome (FMS) is a chronic pain condition characterized by widespread pain and tenderness1,2. The etiology and pathophysiology of fibromyalgia are unknown and there are no effective treatments. Here we show that sensory hypersensitivity in FMS is caused by autoantibodies that act by sensitizing nociceptive sensory neurons. Administration of IgG from FMS patients increased mouse pain sensitivities to stimulation with mechanical pressure and cold. In contrast, transfer of IgG depleted samples from FMS patients or IgG from healthy control subjects had no effect on pain sensitivity. Sensory nerve fibres in ex vivo skin-nerve preparations from mice treated with FMS IgG were hypersensitive to mechanical stimulation. Immunohistochemical analysis revealed that IgG from FMS patients specifically labeled satellite glial cells and myelinated fibre tracts, as well as a small number of macrophages and endothelial cells in mouse dorsal root ganglia but not skin, muscle, spinal cord and brain. Our results demonstrate that fibromyalgia pain is caused by IgG autoantibodies that sensitize peripheral nociceptive afferents neurons and suggest that therapies that reduce patient IgG titres may be effective treatments of fibromyalgia pain.
Structural fuzziness of the RNA-organizing protein SERF1a determines a toxic gain-of-interaction
Authors: Meyer N. Helge , Dellago Hanna , Tam-Amersdorfer Carmen , Merle David A. , Parlato Rosanna , Gesslbauer Bernd , Almer Johannes , Gschwandtner Martha , Leon A. , Franzmann Titus M. , Grillari Johannes , Kungl Andreas J. , Zangger Klaus , Falsone S. Fabio
Published: July 2019

Members of the MOAG/SERF protein family have been attributed a neuropathologic significance because of their ability to enhance the proteotoxic polymerization of amyloidogenic proteins such as alpha-Synuclein (aSyn). However, the cellular function remains unknown. Here, we identify SERF1a as an RNA-organizing protein i) with no structural homology to canonical RNA-binding proteins, ii) with an RNA-chaperoning activity which favours the incorporation of RNA into nucleoli and liquid-like artificial RNA-organelles, and iii) with a high degree of conformational disorder in the RNA-bound state. We demonstrate that this type of structural fuzziness determines an undifferentiated interaction of SERF1a with aSyn and RNA. Both molecules bind to one identical, positively charged site of SERF1a by an analogous electrostatic binding mode, with similar binding affinities, and without any observable disorder-to-order transition. The absence of primary or secondary structure discriminants results in SERF1a being unable to distinguish between nucleic acid and amyloidogenic protein, leading the pro-amyloid aSyn:SERF1a interaction to prevail under conditions of cellular stress. We suggest that structural fuzziness of SERF1a accounts for an adverse gain-of-interaction which favours toxic binding to aSyn at the expense of non-toxic RNA binding, therefore promoting a functionally distorted and pathogenic process. Our results provide a direct link between structural disorder, amyloid aggregation and the malfunctioning of RNA-associated cellular processes, three hallmarks of neurodegenerative diseases.
Understanding the interplay of sleep and aging: Methodological challenges
Authors: Muehlroth Beate E. , Werkle-Bergner Markus
Published: July 2019

In quest of new avenues to explain, predict, and treat pathophysiological conditions during aging, research on sleep and aging has flourished. Despite the great scientific potential to pinpoint mechanistic pathways between sleep, aging, and pathology, only little attention has been paid to the suitability of analytic procedures applied to study these interrelations. On the basis of electrophysiological sleep and structural brain data of healthy younger and older adults, we identify, illustrate, and resolve methodological core challenges in the study of sleep and aging. We demonstrate potential biases in common analytic approaches when applied to older populations. We argue that uncovering age-dependent alterations in the physiology of sleep requires the development of adjusted and individualized analytic procedures that filter out age-independent inter-individual differences. Age-adapted methodological approaches are thus required to foster the development of valid and reliable biomarkers of age-associated cognitive pathologies.
Identification of expression quantitative trait loci in the skeletal muscle of Thoroughbreds reveals heritable variation in expression of genes relevant to cofactor metabolism
Authors: Farries G. , Bryan K. , McGivney C. L. , McGettigan P. A. , Gough K. F. , Browne J.A. , MacHugh D. E. , Katz L. M. , Hill E. W.
Published: July 2019

While over ten thousand genetic loci have been associated with phenotypic traits and inherited diseases in genome-wide association studies, in most cases only a relatively small proportion of the trait heritability is explained and biological mechanisms underpinning these traits have not been clearly identified. Expression quantitative trait loci (eQTL) are subsets of genomic loci shown experimentally to influence gene expression. Since gene expression is one of the primary determinants of phenotype, the identification of eQTL may reveal biologically relevant loci and provide functional links between genomic variants, gene expression and ultimately phenotype. Skeletal muscle (gluteus medius) gene expression was quantified by RNA-seq for 111 Thoroughbreds (47 male, 64 female) in race training at a single training establishment sampled at two time-points: at rest (n = 92) and four hours after high-intensity exercise (n = 77); n = 60 were sampled at both time points. Genotypes were generated from the Illumina Equine SNP70 BeadChip. Applying a Benjamini-Hochberg corrected P-value threshold (PFDR < 0.05), association tests identified 3,583 cis-eQTL associated with expression of 1,456 genes at rest; 4,992 cis-eQTL associated with the expression of 1,922 genes post-exercise; 1,703 trans-eQTL associated with 563 genes at rest; and 1,219 trans-eQTL associated with 425 genes post-exercise. The gene with the highest cis-eQTL association at both time-points was the endosome-associated-trafficking regulator 1 gene (ENTR1; Rest: PFDR = 3.81 × 10−27, Post-exercise: PFDR = 1.66 × 10−24), which has a potential role in the transcriptional regulation of the solute carrier family 2 member 1 glucose transporter protein (SLC2A1). Functional analysis of genes with significant eQTL revealed significant enrichment for cofactor metabolic processes. These results suggest heritable variation in genomic elements such as regulatory sequences (e.g. gene promoters, enhancers, silencers), microRNA and transcription factor genes, which are associated with metabolic function and may have roles in determining end-point muscle and athletic performance phenotypes in Thoroughbred horses.
The PASTA domains of Bacillus subtilis PBP2B stabilize the interaction of PBP2B with DivIB
Authors: Angeles Danae Morales , Macia-Valero Alicia , Bohorquez Laura C. , Scheffers Dirk-Jan
Published: July 2019

Bacterial cell division is mediated by a protein complex known as the divisome. Many protein-protein interactions in the divisome have been characterized. In this report, we analyse the role of the PASTA (Penicillin binding protein And Serine Threonine kinase Associated)-domains of Bacillus subtilis PBP2B. PBP2B itself is essential and cannot be deleted, but removing the PBP2B PASTA domains results in impaired cell division and a heat sensitive phenotype. This resembles the deletion of divIB, a known interaction partner of PBP2B. Bacterial two hybrid and co-immunoprecipitation analyses show that the interaction between PBP2B and DivIB is weakened when the PBP2B PASTA domains are removed. Combined, our results show that the PBP2B PASTA domains are required to stabilize the interaction between PBP2B and DivIB.
A computational model tracks whole-lung Mycobacterium tuberculosis infection and predicts factors that inhibit dissemination
Authors: Wessler Timothy , Joslyn Louis R. , Borish H. Jacob , Gideon Hannah P. , Flynn JoAnne L. , Kirschner Denise E. , Linderman Jennifer J.
Published: July 2019

Mycobacterium tuberculosis (Mtb), the causative infectious agent of tuberculosis (TB), kills more individuals per year than any other infectious agent. Granulomas, the hallmark of Mtb infection, are complex structures that form in lungs, composed of immune cells surrounding bacteria, infected cells, and a caseous necrotic core. While granulomas serve to physically contain and immunologically restrain bacteria growth, some granulomas are unable to control Mtb growth, leading to bacteria and infected cells leaving the granuloma and disseminating, either resulting in additional granuloma formation (local or non-local) or spread to airways or lymph nodes. Dissemination is associated with development of active TB. It is challenging to experimentally address specific mechanisms driving dissemination from TB lung granulomas. Herein, we develop a novel hybrid multi-scale computational model, MultiGran, that tracks Mtb infection within multiple granulomas in an entire lung. MultiGran follows cells, cytokines, and bacterial populations within each lung granuloma throughout the course of infection and is calibrated to multiple non-human primate (NHP) cellular, granuloma, and whole-lung datasets. We show that MultiGran can recapitulate patterns of in vivo local and non-local dissemination, predict likelihood of dissemination, and predict a crucial role for multifunctional CD8+ T cells and macrophage dynamics for preventing dissemination.

Author Summary

Tuberculosis (TB) is caused by infection with Mycobacterium tuberculosis (Mtb) and kills 3 people per minute worldwide. Granulomas, spherical structures composed of immune cells surrounding bacteria, are the hallmark of Mtb infection and sometimes fail to contain the bacteria and disseminate, leading to further granuloma growth within the lung environment. To date, the mechanisms that determine granuloma dissemination events have not been characterized. We present a computational multi-scale model of granuloma formation and dissemination within primate lungs. Our computational model is calibrated to multiple experimental datasets across the cellular, granuloma, and whole-lung scales of non-human primates. We match to both individual granuloma and granuloma-population datasets, predict likelihood of dissemination events, and predict a critical role for multifunctional CD8+ T cells and macrophage-bacteria interactions to prevent infection dissemination.
A cell-based evaluation of a non-essential amino acid formulation as a non-bioactive control for activation and stimulation of muscle protein synthesis using ex vivo human serum
Authors: Patel Bijal , Pauk Martina , Amigo-Benavent Miryam , Nongonierma Alice B. , Fitzgerald Richard J. , Jakeman Philip M. , Carson Brian P.
Published: July 2019

Purpose

The purpose of this study was to compare the effect of treating skeletal muscle cells with media conditioned by postprandial ex vivo human serum fed with either isonitrogenous NEAA or a whey protein hydrolysate (WPH) on stimulating MPS in C2C12 skeletal muscle cells.

Methods

Blood was taken from six young healthy males following overnight fast (fasted) and 60 min postprandial (fed) ingestion of either WPH or NEAA (0.33 g.kg-1 Body Mass). C2C12 myotubes were treated with media conditioned by ex vivo human serum (20%) for 4 h. Activation of MPS signalling (phosphorylation of mTOR, P70S6K and 4E-BP1) were determined in vitro by Western Blot and subsequent de novo MPS were determined in vitro by Western Blot and surface sensing of translation technique (SUnSET) techniques, respectively.

Results

Media conditioned by NEAA fed serum had no effect on protein signalling or MPS compared to fasted, whereas media conditioned by WPH fed serum significantly increased mTOR, P70S6K and 4E-BP1 phosphorylation (p<0.01, p<0.05) compared to fasted serum. Furthermore, the effect of media conditioned by WPH fed serum on protein signalling and MPS was significantly increased (p<0.01, p<0.05) compared to NEAA fed serum.

Conclusion

In summary, media conditioned by NEAA fed serum did not result in activation of MPS. Therefore, these in vitro findings suggest the use of isonitrogenous NEAA acts as an effective control for comparing bioactivity of different proteins on activation of MPS. These findings also confirm that activation of MPS in C2C12 myotubes treated with media conditioned by WPH-fed serum is primarily due to circulating EAA.
Evaluation ofTrichoderma harzianummutant lines in the resistance induction against white mold and growth promotion of common bean
Authors: Brandão Renata Silva , Qualhato Thiago Fernandes , Valdisser Paula Arielle Mendes Ribeiro , de C. B. Côrtes Marcio Vinicius , Vieira Pabline Marinho , Silva Roberto Nascimento , Junior Murillo Lobo , Ulhoa Cirano José
Published: July 2019

Genetic engineering has brought improvements ofTrichodermaspecies for biotechnological processes such as agriculture. It has previously been reported thatsm1andaquaglyceroporingenes fromTrichoderma harzianumare strongly expressed during pathogens biocontrol. We have previously shown thatSm1plays a significant role in plant disease resistance and aquaporin (AQP) regulate physiological processes and responses related to water stress. Here, we report the evaluation of mutant lines withsm1(deleated) andaqp(overexpressed) inPhaseoulus vulgarisgrowth promotion and disease resistance induction. It was investigated plants physiological and biochemical responses, plant growth promotion and biocontrol ofSclerotinia sclerotiorum, the causal agent of white mold. Treatments includedTrichoderma harzianumwyld type, mutant line overexpressing aquaporin (Aqua), and deletedΔEpl1mutant line. Our results demonstrated that Aqua mutant line increased leaf area in 25%, in comparison to non-treated plants. It also differed from other treatments, in comparison to plants with treatments ALL-42 andΔEpl1, which increased 28% and 91%, respectively (Isso é abstract, ta confuse e extensor. Specific activity of β-1.3 glucanase was higher in plants treated withT. harzianum ΔEpl1mutant isolate, in leaves and roots respectively with 2.07 Umg−1and 2.57 Umg−1. Plants inoculated or not withS. sclerotiorumpresented with 2.27 Umg-1and 2.34 Umg-1of β-1.3 glucanase on average, respectively, but enzymatic activity was higher on leaves when compared to the roots. TheAquamutant demonstrated excellent growth promoting properties. Both mutants ofT. harzianumdo not induce resistance to white mold on common bean plants.
Cross-species epigenetic regulation of nucleus accumbens KCNN3 transcript variants by excessive ethanol drinking and dependence
Authors: Cervera-Juanes Rita , Padula Audrey E. , Wilhem Larry J. , Park Byung , Grant Kathleen A. , Ferguson Betsy , Mulholland Patrick J.
Published: July 2019

The underlying genetic and epigenetic mechanisms driving functional adaptations in neuronal excitability and excessive alcohol intake are poorly understood. Given that small-conductance Ca2+-activated K+ (KCa2 or SK) channels encoded by the KCNN family of genes have emerged from preclinical studies as a crucial target that contributes to heavy drinking and alcohol-induced functional neuroadaptations, we performed a cross-species analysis of KCNN3 methylation, gene expression, and polymorphisms of alcohol-drinking monkeys and alcohol dependent mice. Because of the alternative promoters in KCNN3, we analyzed expression of the different transcript variants that when translated influence surface trafficking and function of KCa2 channels. In heavy drinking rhesus macaques and alcohol dependent C57BL/6J mice, bisulfite sequencing analysis of the nucleus accumbens revealed a differentially methylated region in exon 1A of KCNN3 that overlaps with a predicted promoter sequence. The hypermethylation of KCNN3 in monkey and mouse accumbens paralleled an increase in expression of alternative transcript variants that encode apamin-insensitive and dominant-negative KCa2 channel isoforms. A polymorphic repeat in macaque KCNN3 encoded by exon 1 did not correlate with alcohol drinking. At the protein level, KCa2.3 channel expression in the accumbens was significantly reduced in very heavy drinking monkeys. Together, our cross-species findings on epigenetic dysregulation of KCNN3 by heavy alcohol drinking and dependence represent a complex mechanism that utilizes alternative promoters to impact firing of accumbens neurons. Thus, these results provide support for hypermethylation of KCNN3 by excessive alcohol drinking as a possible key molecular mechanism underlying harmful alcohol intake and alcohol use disorder.
WiPP: Workflow for improved Peak Picking for Gas Chromatography-Mass Spectrometry (GC-MS) data
Authors: Borgsmüller Nico , Gloaguen Yoann , Opialla Tobias , Blanc Eric , Sicard Emilie , Royer Anne-Lise , Le Bizec Bruno , Durand Stéphanie , Migné Carole , Pétéra Mélanie , Pujos-Guillot Estelle , Giacomoni Franck , Guitton Yann , Beule Dieter , Kirwan Jennifer
Published: July 2019

Lack of reliable peak detection impedes automated analysis of large scale GC-MS metabolomics datasets. Performance and outcome of individual peak-picking algorithms can differ widely depending on both algorithmic approach and parameters as well as data acquisition method. Comparing and contrasting between algorithms is thus difficult. Here we present a workflow for improved peak picking (WiPP), a parameter optimising, multi-algorithm peak detection for GC-MS metabolomics. WiPP evaluates the quality of detected peaks using a machine learning-based classification scheme based on seven peak classes. The quality information returned by the classifier for each individual peak is merged with results from different peak detection algorithms to create one final high quality peak set for immediate down stream analysis. Medium and low quality peaks are kept for further inspection. By applying WiPP to standard compound mixes and a complex biological dataset we demonstrate that peak detection is improved through the novel way to assign peak quality, an automated parameter optimisation, and results integration across different embedded peak picking algorithms. Furthermore, our approach can provide an impartial performance comparison of different peak picking algorithms. WiPP is freely available on GitHub (https://github.com/bihealth/WiPP) under MIT licence.
kASA: Taxonomic Analysis of Metagenomic Data on a Notebook
Authors: Weging Silvio , Gogol-Döring Andreas , Grosse Ivo
Published: July 2019

The taxonomic analysis of sequencing data has become important in many areas of life sciences. However, currently available software tools for that purpose either consume large amounts of RAM or yield an insufficient quality of the results.

Here we present kASA, a k-mer based software capable of identifying and profiling metagenomic DNA sequences with high computational efficiency and a user-definable memory footprint. We ensure both high sensitivity and precision by using an amino acid-like encoding of k-mers with a dynamic length of multiple k’s. Custom algorithms and data structures optimized for external memory storage enable for the first time a full-scale metagenomic analysis without compromise on a standard notebook.
Nesprin-2 accumulates at the front of the nucleus during confined cell migration
Authors: Davidson Patricia M. , Battistella Aude , Déjardin Théophile , Betz Timo , Plastino Julie , Borghi Nicolas , Cadot Bruno , Sykes Cécile
Published: July 2019

The mechanisms by which cells exert forces on their nuclei to migrate through openings smaller than the nuclear diameter remain unclear. In microfluidic devices, the hourglass shape of the nucleus and its strain patterns as it translocates through narrow constrictions suggest pulling forces. We use CRISPR/Cas9 to fluorescently label nesprin-2 giant, a protein that links the cytoskeleton to the interior of the nucleus. We demonstrate that nesprin-2 giant accumulates at the front of the nucleus during nuclear deformation through narrow constrictions, independently of the nuclear lamina. We find that nesprins are more mobile than lamin A/C, at time scales similar to that of the accumulation. Using artificial constructs, we show that the actin-binding domain of nesprin-2 is necessary and sufficient to generate this accumulation, and that microtubules are not necessary. Actin filaments are organized in a barrel structure around the moving nucleus in the direction of movement, suggesting that this structure is responsible for redistribution of nesprins towards the front of the nucleus. Two-photon ablation and the use of drugs inhibiting the cytoskeleton demonstrate a pulling force on the nucleus from the front of the cell that is dependent on formin and actomyosin contractility. This elastic recoil is significantly reduced when nesprins are reduced at the nuclear envelope. We thus show that actin redistributes nesprin-2 giant towards the front of the nucleus and contributes to pulling the nucleus through narrow constrictions, in concert with myosin.
Atlas of transcriptionally active transposable elements in human adult tissues
Authors: Bogu Gireesh K. , Reverter Ferran , Marti-Renom Marc A. , Snyder Michael P. , Guigó Roderic
Published: July 2019

Approximately half of the human genome consists of mobile repetitive DNA sequences known as transposable elements (TEs). They are usually silenced by epigenetic mechanisms, but a few are known to escape silencing at embryonic stages, affecting early human development by regulating nearby protein-coding genes. To investigate transcriptional activity in human adult tissues we systematically investigate the expression landscape of about 4.2 million non-coding TEs in 8,051 RNA-Seq datasets from up to 49 adult tissues and 540 individuals. We show that approximately 79,558 individual TEs (2%). belonging to 856 subfamilies escape epigenetic silencing in adult tissues and become transcriptionally active, often in a very tissue-specific manner. Supporting a role for TEs in the regulation of expression of nearby genes, we found the expression of TEs often correlated with the expression of nearby genes, and significantly stronger when the TEs include eQTLs for the genes. We identified thousands of tissue-elevated, sex-associated TEs in the breast, ethnicity-associated in the skin and age-associated in the tibial artery, where we found a potential implication of two TE subfamilies in atherosclerosis. Our results suggest a functional role of TEs in the regulation of gene expression, support their implication in human phenotypes, and also serve as a comprehensive resource of transcriptionally active TEs in human adult tissues.