bluuebunny/crossref_metadata_2025 huggingface dataset:
Long-lasting salt bridges provide the anchoring mechanism of oncogenic KRas-4B proteins at cell membranes
Authors: Lu Huixia , Martí Jordi
Published: August 2020

Ras is a family of related proteins participating in all animal cell lineages and organs. Ras proteins work as GDP-GTP binary switches and regulate cytoplasmic signalling networks that are able to control several cellular processes, playing an essential role in signal transduction pathways involved in cell growth, differentiation and survival so that overacting Ras signalling can lead to cancer. One of the hardest challenges to face is, with more than hundred different missense mutations found in cancer, the design of mutation-selective therapeutic strategies. In this work, a G12D mutated farnesylated GTP bound KRas-4B protein has been simulated at the interface of a DOPC/DOPS/cholesterol model anionic cell membrane at the all-atom level. A specific long-lasting salt bridge connection between farnesyl and the hypervariable region of the protein has been identified as the main mechanism responsible of the binding of oncogenic farnesylated KRas-4B to the cell membrane, since this particular bond is absent in both wild-type and oncogenic methylated species of KRas-4B. This finding may lead to a deeper understanding of the mechanisms of protein binding and eventual growing and spreading inside cell membranes. From free energy landscapes obtained by well-tempered metadynamics simulations, we have characterised local and global minima of KRas-4B binding to the cell membrane revealing the main pathways between anchored and released states.
Synaptic anchoring of the endoplasmic reticulum depends on myosin V and caldendrin activity
Authors: Konietzny Anja , Grendel Jasper , Hertrich Nathalie , Dekkers Dick H. W. , Demmers Jeroen A. A. , Mikhaylova Marina
Published: August 2020

Excitatory synapses of principal hippocampal neurons are frequently located on dendritic spines. The dynamic strengthening or weakening of individual inputs results in a great structural and molecular diversity of dendritic spines. Active spines with large Ca2+transients are frequently invaded by a single protrusion from the endoplasmic reticulum (ER), which is dynamically transported into and out of spines by the actin-based motor myosin V. An increase in synaptic strength often correlates with stable anchoring of the ER, followed by the formation of the spine apparatus organelle. Here we show that synaptic ER stabilization depends on the interplay of two Ca2+-binding proteins: calmodulin serves as a light chain of myosin V and activates the motor function, whereas caldendrin acts as an inhibitor which transforms myosin into a stationary F-actin tether. Together, they provide a Ca2+-sensing module for fine-tuning myosin V activity and thereby regulate the formation of the spine apparatus in a subset of active dendritic spines.
Mapping the Human Visual Thalamus and its Cytoarchitectonic Subdivisions Using Quantitative MRI
Authors: Müller-Axt Christa , Eichner Cornelius , Rusch Henriette , Kauffmann Louise , Bazin Pierre-Louis , Anwander Alfred , Morawski Markus , von Kriegstein Katharina
Published: August 2020

The human lateral geniculate nucleus (LGN) of the visual thalamus is a key subcortical processing site for visual information analysis. A non-invasive assessment of the LGN and its functionally and microstructurally distinct magnocellular (M) and parvocellular (P) subdivisionsin-vivoin humans is challenging, because of its small size and location deep inside the brain. Here we tested whether recent advances in high-field structural quantitative MRI (qMRI) can enable MR-based mapping of human LGN subdivisions. We employed ultra-high resolution 7 Tesla qMRI of apost-mortemhuman LGN specimen and high-resolution 7 Teslain-vivoqMRI in a large participant sample. We found that a quantitative assessment of the LGN and a differentiation of its subdivisions was possible based on microstructure-informed MR-contrast alone. In both thepost-mortemandin-vivoqMRI data, we identified two components of shorter and longer longitudinal relaxation time (T1) within the LGN that coincided with the known anatomical locations of a dorsal P and a ventral M subdivision, respectively. Through a subsequent ground truth histological examination of the samepost-mortemLGN specimen, we showed that the observed T1contrast pertains to cyto- and myeloarchitectonic differences between LGN subdivisions. These differences were based on cell and myelin density, but not on iron content. Our qMRI-based mapping strategy overcomes shortcomings of previous fMRI-based mapping approaches. It paves the way for an in-depth understanding of the function and microstructure of the LGN in humans. It also enables investigations into the selective contributions of LGN subdivisions to human behavior in health and disease.

Significance Statement

The lateral geniculate nucleus (LGN) is a key processing site for the analysis of visual information. Due to its small size and deep location within the brain, non-invasive mapping of the LGN and its microstructurally distinct subdivisions in humans is challenging. Using quantitative MRI methods that are sensitive to underlying microstructural tissue features, we show that a differentiation of the LGN and its microstructurally distinct subdivisions is feasible in humans non-invasively. These findings are important because they open up novel opportunities to assess the hitherto poorly understood complex role of the LGN in human perception and cognition, as well as the contribution of selective LGN subdivision impairments to various clinical conditions including developmental dyslexia, glaucoma and multiple sclerosis.
Discriminating Lineages ofBatrachochytrium dendrobatidisusing quantitative PCR
Authors: Ghosh Pria N , Verster Ruhan , Sewell Thomas , O’Hanlon Simon , Brookes Lola M , Rieux Adrien , Garner Trenton WJ , Weldon Ché , Fisher Matthew C
Published: August 2020

The ability to detect and monitor infectious disease in a phylogenetically informative manner is critical for their management. Phylogenetically informative diagnostic tests enable patterns of pathogen introduction or changes in the distribution of genotypes to be measured, enabling research into the ecology of the pathogen.Batrachochytrium dendrobatidis(Bd), a causative agent of chytridiomycosis in amphibian populations, emerged worldwide in the 21stcentury and is composed of six lineages which are display varying levels of virulence in their hosts.

Research into the distribution, ecology and pathogenicity of these lineages has been hampered by an inability to type lineage efficiently. Here, we describe a lineage-specific TaqMan qPCR assay that differentiates the two lineages ofBdmost commonly associated with chytridiomycosis:BdGPL andBdCAPE. We demonstrate how this assay can be used for the surveillance of wild populations of amphibians in Southern Africa using skin swabs, tissue samples and cultured isolates.
Capturingin situVirus-Host Range and Interaction Dynamics through Gene Fusion with epicPCR
Authors: Sakowski Eric G. , Arora-Williams Keith , Tian Funing , Zayed Ahmed A , Zablocki Olivier , Sullivan Matthew B. , Preheim Sarah P.
Published: August 2020

Viruses impact microbial diversity, phenotype, and gene flow through virus-host interactions that in turn alter ecology and biogeochemistry. Though metagenomics surveys are rapidly cataloging viral diversity, capturing specific virus-host interactionsin situwould identify hosts for novel viruses and reveal influential ecological or environmental factors. We leveraged metagenomics and a high-throughput, cultivation-independent gene fusion technique (epicPCR) to investigate viral diversity and virus-host interactions over time in a critical estuarine environment, the Chesapeake Bay. EpicPCR capturedin situvirus-host interactions for viral clades with no closely related database representatives. Abundant freshwater Actinobacteria lineages were the most common hosts for these poorly characterized viruses, and observed viral interactions with one abundant Actinobacterial population (Rhodoluna) were correlated with environmental factors. Tracking virus-host interaction dynamics also revealed ecological differences between multi-host (generalist) and single-host (specialist) viruses. Generalist viruses had significantly longer periods with observed virus-host interactions but specialist viruses were observed interacting with hosts at lower minimum abundances, suggesting more efficient interactions. Together, these observations reveal ecological differences between generalist and specialist viruses that provide insight into evolutionary trade-offs. Capturingin situinteractions with epicPCR revealed environmental and ecological factors that shape virus-host interactions, highlighting epicPCR as a scalable new tool in viral ecology.
Probabilistic Thermodynamic Analysis of Metabolic Networks
Authors: Gollub Mattia G. , Kaltenbach Hans-Michael , Stelling Jörg
Published: August 2020

Random sampling of metabolic fluxes can provide an unbiased description of the capabilities of a metabolic network. However, current sampling approaches do not model thermodynamics explicitly, leading to inaccurate predictions of an organism’s potential or actual metabolic operations. We present a probabilistic framework combining thermodynamic quantities with steady-state flux constraints to analyze the properties of a metabolic network. It includes methods for probabilistic metabolic optimization and for joint sampling of thermodynamic and flux spaces. Applied to a model ofE. coli, we use the methods to reveal known and novel mechanisms of substrate channeling, and to accurately predict reaction directions and metabolite concentrations. Interestingly, predicted flux distributions are multimodal, leading to discrete hypotheses onE. coli‘s metabolic capabilities. C++ source code with MATLAB interface available athttps://gitlab.com/csb.ethz/pta.
Adolescent female rats undergo full systems consolidation of an aversive memory, while males of the same age fail to discriminate contexts
Authors: Crestani Ana P. , Lotz Fernanda N. , Casagrande Mirelle A. , Popik Bruno , Guerra Kétlyn T. K. , de Oliveira Alvares Lucas , Quillfeldt Jorge A.
Published: August 2020

Generalization is an adaptive process that allows animals to deal with threatening circumstances similar to prior experiences. Systems consolidation is a time-dependent process in which memory loses it precision concomitantly with reorganizational changes in the brain structures that support memory retrieval. In this, memory becomes progressively independent from the hippocampus and more reliant on cortical structures. Generalization, however, may take place much faster in adult animals depending on the presence of sex hormones. Notwithstanding its relevance, there are few studies on sex differences in memory modulation. Here, a contextual fear discrimination task was used to investigate the onset of memory generalization and hippocampus-independence in adolescent male and female rats (P42-49). Subjects were tested 2, 7, 14, 21 or 28 days after training, with females showing memory generalization from day 21 on, whereas males surprisingly unable to discriminate contexts at any time. Ovariectomized females, however, displayed an early onset of generalization. Consistently, pre-test pharmacological blocking of dorsal hippocampus was able to impair memory retrieval in females, but not in males, which indicate that precise memory is dependent on the hippocampus. To our notice, this is the first report of a memory systems consolidation process – expressed in its two dimensions, neuroanatomical and qualitative – in adolescent female rats, and one that can also be accelerated by the reduction of sex hormones through ovariectomy. It is also unprecedented that despite adolescent male rats being able to remember fear learning, they did not discriminate contexts with any precision.
Common and distinct neural correlates of music and food-induced pleasure: a coordinate-based meta-analysis of neuroimaging studies
Authors: Mas-Herrero Ernest , Maini Larissa , Sescousse Guillaume , Zatorre Robert J.
Published: August 2020

Neuroimaging studies have shown that, despite the abstractness of music, it may mimic biologically rewarding stimuli (e.g. food) in its ability to engage the brain’s reward circuity. However, due to the lack of research comparing music and other types of reward, it is unclear to what extent the recruitment of reward-related structures overlaps among domains. To achieve this goal, we performed a coordinate-based meta-analysis of 38 neuroimaging studies (703 subjects) comparing the brain responses specifically to music and food-induced pleasure. Both engaged a common set of brain regions including the ventromedial prefrontal cortex, ventral striatum, and insula. Yet, comparative analyses indicated a partial dissociation in the engagement of the reward circuitry as a function of the type of reward, as well as additional reward type-specific activations in brain regions related to perception, sensory processing, and learning. These results support the idea that hedonic reactions rely on the engagement of a common reward network, yet through specific routes of access depending on the modality and nature of the reward.
High resolution visual information via a gap junction-mediated spike order code
Authors: Tong Rudi , Trenholm Stuart
Published: August 2020

Gap junctions promote correlated spiking between coupled neurons. Recording from pairs of coupled retinal ganglion cells, we find that differences in firing rate between coupled neurons dictates which cell leads correlated spike-pairs, and that the precise temporal order of spike activity between pairs of cells changes depending on the spatial position of a visual stimulus. We thus demonstrate a spike order spatial code for encoding sensory information.
Low-cost, bottom-up fabrication of large-scale single-molecule nanoarrays by DNA origami placement
Authors: Shetty Rishabh M. , Brady Sarah R. , Rothemund Paul W. K. , Hariadi Rizal F. , Gopinath Ashwin
Published: August 2020

Large-scale nanoarrays of single biomolecules enable high-throughput assays while unmasking the underlying heterogeneity within ensemble populations. Until recently, creating such grids which combine the unique advantages of microarrays and single-molecule experiments (SMEs) has been particularly challenging due to the mismatch between the size of these molecules and the resolution of top-down fabrication techniques. DNA Origami Placement (DOP) combines two powerful techniques to address this issue: (i) DNA origami, which provides a ∼ 100-nm self-assembled template for single-molecule organization with 5 nm resolution, and (ii) top-down lithography, which patterns these DNA nanostructures, transforming them into functional nanodevicesvialarge-scale integration with arbitrary substrates. Presently, this technique relies on state-of-the-art infrastructure and highly-trained personnel, making it prohibitively expensive for researchers. Here, we introduce a bench-top technique to create meso-to-macro-scale DNA origami nanoarrays using self-assembled colloidal nanoparticles, thereby circumventing the need for top-down fabrication. We report a maximum yield of 74%, two-fold higher than the statistical limit of 37% imposed on non-specific molecular loading alternatives. Furthermore, we provide a proof-of-principle for the ability of this nanoarray platform to transform traditionally low-throughput, stochastic, single-molecule assays into high-throughput, deterministic ones, without compromising data quality. Our approach has the potential to democratize single-molecule nanoarrays and demonstrates their utility as a tool for biophysical assays and diagnostics.
Red light imaging for programmed cell death visualization and quantification in plant-pathogen interactions
Authors: Villanueva Sergio Landeo , Malvestiti Michele C. , Joosten Matthieu H.A.J. , van Ieperen Wim , van Kan Jan A.L.
Published: August 2020

Studies on plant-pathogen interactions often involve monitoring disease symptoms or responses of the host plant to pathogen-derived immunogenic patterns, either visually or by staining the plant tissue. Both these methods have limitations with respect to resolution, reproducibility and the ability to quantify the results. In this study we show that red light detection in a multi-purpose fluorescence imaging system that is probably available in many labs can be used to visualize plant tissue undergoing cell death. Red light emission is the result of chlorophyll fluorescence upon thylakoid membrane disassembly during the development of a programmed cell death process. The activation of programmed cell death can occur either during a hypersensitive response to a biotrophic pathogen or an apoptotic cell death triggered by a necrotrophic pathogen. Quantifying the intensity of the red light signal enables to evaluate the magnitude of programmed cell death and provides a non-invasive readout of the plant immune response in a faster and safer manner as compared to chemical staining methodologies previously developed. This application can be implemented to screen for differences in symptom severity in plant-pathogen interactions, and to visualize and quantify in a sensitive and objective manner the intensity of a plant response upon perception of a given immunological pattern. We illustrate the utility and versatility of the method using diverse immunogenic patterns and pathogens.
Involvement of uterine natural killer cells in the natural selection of human embryos at implantation
Authors: Kong Chow-Seng , Ordoñez Alexandra Almansa , Turner Sarah , Tremaine Tina , Muter Joanne , Lucas Emma S. , Salisbury Emma , Vassena Rita , Fouladi-Nashta Ali A. , Tiscornia Gustavo , Hartshorne Geraldine , Brosens Jan J. , Brighton Paul J.
Published: August 2020

Decidualizing endometrial stromal cells (EnSC) critically determine the maternal response to an implanting conceptus, triggering either menstruation-like disposal of low-fitness embryos or creating an environment that promotes further development. However, the mechanism that couples maternal recognition of low-quality embryos to tissue breakdown remains poorly understood. Recently, we demonstrated that successful transition of the cycling endometrium to a pregnancy state requires selective elimination of pro-inflammatory senescent decidual cells by activated uterine natural killer (uNK) cells. Here we report that uNK cells express CD44, the canonical hyaluronan (HA) receptor, and demonstrate that high-molecular weight HA (HMWHA) inhibits uNK cell-mediated killing of senescent decidual cells. By contrast, low-molecular weight HA (LMWHA) did not attenuate uNK cell activity in co-culture experiments. Killing of senescent decidual cells by uNK cells was also inhibited upon exposure to medium conditioned by IVF embryos that failed to implant, but not successful embryos. Embryo-mediated inhibition of uNK cell activity was reversed by recombinant hyaluronidase 2 (HYAL2), which hydrolyses HMWHA. We further report a correlation between the levels of HYAL2 secretion by human blastocysts, morphological scores, and implantation potential. Taken together, the data suggest a pivotal role for uNK cells in embryo biosensing and endometrial fate decisions at implantation.
Rearing water microbiomes in white leg shrimp (Litopenaeus vannamei) larviculture assemble stochastically and are influenced by the microbiomes of live feed products
Authors: Heyse Jasmine , Props Ruben , Kongnuan Pantipa , De Schryver Peter , Rombaut Geert , Defoirdt Tom , Boon Nico
Published: August 2020

The development of effective management strategies to reduce the occurrence of diseases in aquaculture is hampered by the limited knowledge on the microbial ecology of these systems. In this study, the dynamics and dominant community assembly processes in the rearing water ofLitopenaeus vannameilarviculture tanks were determined. Additionally, the contribution of peripheral microbiomes, such as those of live and dry feeds, to the rearing water microbiome were quantified. The community assembly in the hatchery rearing water over time was dominated by stochasticity, which explains the observed heterogeneity between replicate cultivations. The community undergoes two shifts that match with the dynamics of the algal abundances in the rearing water. Source tracking analysis revealed that 37% of all bacteria in the hatchery rearing water were either introduced by the live or dry feeds, or during water exchanges. The contribution of the microbiome from the algae was the largest, followed by that of theArtemia, the exchange water and the dry feeds. Our findings provide fundamental knowledge on the assembly processes and dynamics of rearing water microbiomes and illustrate the crucial role of these peripheral microbiomes in maintaining health-promoting rearing water microbiomes.

Originality-Significance Statement

Most studies on rearing water microbiomes are characterized by sampling resolutions of multiple days and by few replicate cultivations. Through an 18-day sampling campaign in aLitopenaeus vannameihatchery where five replicate cultivations were studied at a sampling resolution of one day, we studied the microbiome dynamics in this system. We show that the community assembly is dominated by stochasticity, which explains the heterogeneity between replicate cultivations. The dynamics of the algal community in the rearing water induced shifts in community composition at two differerent timepoints. Finally, we quantified the contribution of live and dry feed microbiomes to the rearing water community for the first time. We found that the contribution of each source was dependent on its taxonomic composition, the bacterial load caused by the addition of this source and the timing of the introduction. These new insights will aid in the further development of effective microbiome management to reduce the frequency and magnitude of bacterial diseases.
Canonical EEG Microstate Dynamic Properties and Their Associations with fMRI Signals at Resting Brain
Authors: Zoubi Obada Al , Misaki Masaya , Tsuchiyagaito Aki , Mayeli Ahmad , Zotev Vadim ,   Tulsa 1000 Investigators, Refai Hazem , Paulus Martin , Bodurka Jerzy
Published: August 2020

Electroencephalography microstates (EEG-ms) capture and reflect the spatio-temporal neural dynamics of the brain. A growing literature is employing EEG-ms-based analyses to study various mental illnesses and to evaluate brain mechanisms implicated in cognitive and emotional processing. The spatial and functional interpretation of the EEG-ms is still being investigated. Previous works studied the association of EEG-ms time courses with blood-oxygen-level-dependent (BOLD) functional magnetic resonance imaging (fMRI) signal and suggested an association between EEG-ms and resting-state networks (RSNs). However, the distinctive association between EEG-ms temporal dynamics and brain neuronal activities is still not clear, despite the assumption that EEG-ms are an electrophysiological representation of RSNs activity. Recent works suggest a role for brain spontaneous EEG rhythms in contributing to and modulating canonical EEG-ms topographies and determining their classes (coined A through D) and metrics. This work simultaneously utilized EEG and fMRI to understand the EEG-ms and their properties further. We adopted the canonical EEG-ms analysis to extract three types of regressors for EEG-informed fMRI analyses: EEG-ms direct time courses, temporal activity per microstate, and pairwise temporal transitions among microstates (the latter two coined activity regressors). After convolving EEG-ms regressors with a hemodynamic response function, a generalized linear model whole-brain voxel-wise analysis was conducted to associate EEG-ms regressors with fMRI signals. The direct time course regressors replicated prior findings of the association between the fMRI signal and EEG-ms time courses but to a smaller extent. Notably, EEG-ms activity regressors were mostly anticorrelated with fMRI, including brain regions in the somatomotor, visual, dorsal attention, and ventral attention fMRI networks with no significant overlap for default mode, limbic or frontoparietal networks. A similar pattern emerged in using the transition regressors among microstates but not in self-transitions. The relatively short duration of each EEG-ms and the significant association of EEG-ms activity regressors with fMRI signals suggest that EEG-ms manifests successive transition from one brain functional state to another rather than being associated with specific brain functional state or RSN networks.
Methylene Blue has a potent antiviral activity against SARS-CoV-2 in the absence of UV-activation in vitro
Authors: Cagno Valeria , Medaglia Chiara , Cerny Andreas , Cerny Thomas , Tapparel Caroline , Cerny Erich
Published: August 2020

Methylene blue is an FDA and EMA approved drug with an excellent safety profile. It displays broad-spectrum virucidal activity in the presence of UV light and has been shown to be effective in inactivating various viruses in blood products prior to transfusions. In addition, its use has been validated for methemoglobinemia and malaria treatment. Here we show the virucidal activity of methylene blue at low micromolar concentrations and in the absence of UV activation against SARS-CoV2.
Vitamin D status is heritable and under environment-dependent selection in the wild
Authors: Sparks Alexandra M. , Johnston Susan E. , Handel Ian , Pilkington Jill G. , Berry Jacqueline , Pemberton Josephine M. , Nussey Daniel H. , Mellanby Richard J.
Published: August 2020

Vitamin D has a well-established role in skeletal health and is increasingly linked to chronic disease and mortality in humans and companion animals. Despite the clear significance of vitamin D for health and obvious implications for fitness under natural conditions, no longitudinal study has tested whether the circulating concentration of vitamin D is under natural selection in the wild. Here, we show that concentrations of dietary-derived vitamin D2 and endogenously-produced vitamin D3 metabolites are heritable and largely polygenic in a wild population of Soay sheep (Ovis aries). Vitamin D2 status was positively associated with female adult survival, and vitamin D3 status predicted female fecundity in particular, good environment years. Our study provides evidence that vitamin D status has the potential to respond to selection, as well as new insights into how vitamin D metabolism is associated with fitness in the wild.
Life story of Tunisian durum wheat landraces revealed by their genetic and phenotypic diversity
Authors: Ben Krima Safa , Slim Amine , Gélisse Sandrine , Kouki Hajer , Nadaud Isabelle , Sourdille Pierre , Yahyaoui Amor , Ben M’barek Sarrah , Suffert Frédéric , Marcel Thierry C
Published: August 2020

Durum wheat (Triticum turgidumL. subsp.durum) landraces represent a prominent genetic resource for Mediterranean farming systems and breeding programs. Fourteen landraces sampled in Tunisia were genotyped with 9 microsatellite markers and characterized with 15 morphological descriptors, including resistance to the fungal diseaseSeptoria triticiblotch (STB). The genetic diversity, nearly was as important within landraces populations (45%) than between populations (54%). It was structured in seven genetic groups and was only partly explained by the variety name or the locality of origin. Populations were also greatly diversified phenotypically (Shannon-Weaver H’=0.54) with traits related to spike and awn colours being the most diversified. Resistance to STB was either qualitative in two populations or with varying degrees of quantitative resistance in the others. A Pst-Fstcomparison indicate a local adaptation of the populations. Overall, the genetic structure of Tunisian durum wheat landraces revealed a complex selection trajectory and seed exchanges between farmers.
Humanization ofDrosophilaGαo to modelGNAO1paediatric encephalopathies
Authors: Savitsky Mikhail , Solis Gonzalo P. , Katanaev Vladimir L.
Published: August 2020

Several hundred genes have been identified to contribute to epilepsy – the disease affecting 65 million people worldwide. One of these genes isGNAO1encoding Gαo, the major neuronal α-subunit of heterotrimeric G proteins. An avalanche of dominantde novomutations inGNAO1have been recently described in paediatric epileptic patients, suffering in addition to epilepsy from motor dysfunction and developmental delay. Although occurring in amino acids conserved from humans toDrosophila, these mutations and their functional consequences have only poorly been analysed at the biochemical or neuronal levels. Adequate animal models to study molecular aetiology ofGNAO1encephalopathies have also so far been lacking. As the first step towards modelling the disease inDrosophila, we here describe humanization of theGαolocus in the fruit fly. A two-step CRISPR/Cas9-mediated replacement was conducted, first substituting the coding exons 2-3 ofGαowith respective humanGNAO1sequences. At the next step, the remaining exons 4-7 were similarly replaced, keeping intact the geneCyp49a1embedded in-between, as well as the non-coding exon 1 and the surrounding regulatory sequences. The resulting flies, homozygous for the humanizedGNAO1loci, are viable and fertile without any visible phenotypes; their body weight and longevity are also normal. Human Gαo-specific antibodies confirm the endogenous-level expression of the humanized Gαo, which fully replaces theDrosophilafunctions. The genetic model we established will make it easy to incorporate encephalopathicGNAO1mutations and will permit intensive investigations into the molecular aetiology of the human disease through the powerful toolkit ofDrosophilagenetics.
A non-canonical DNA damage checkpoint response in a major fungal pathogen
Authors: Shor Erika , Garcia-Rubio Rocio , DeGregorio Lucius , Perlin David S.
Published: August 2020

To protect genome integrity, eukaryotic cells respond to DNA damage by triggering highly conserved checkpoint mechanisms involving the phosphorylation of Rad53/CHK2 kinase. Budding yeastCandida glabrata, closely related to model eukaryoteSaccharomyces cerevisiae, is an opportunistic pathogen characterized by high genetic diversity and rapid emergence of drug resistant mutants. However, the mechanisms enabling this genetic variability are unclear. Here we show thatC. glabratacells exposed to DNA damage neither induce CgRad53 phosphorylation nor accumulate in S phase, and exhibit higher lethality thanS. cerevisiae. Furthermore, time-lapse microscopy showedC. glabratacells continuing to divide in the presence of DNA damage, resulting in mitotic errors and cell death. Finally, RNAseq analysis revealed transcriptional rewiring of the DNA damage response inC. glabrataand identified several key protectors of genome stability upregulated by DNA damage inS. cerevisiaebut downregulated inC. glabrata, including PCNA. Together, our results reveal a non-canonical fungal DNA damage response, which may contribute to rapidly generating genetic change and drug resistance.
The Multivariate Genome-wide Architecture of Interrelated Literacy, Language and Working Memory Skills Reveals Distinct Etiologies
Authors: Shapland Chin Yang , Verhoef Ellen , Smith George Davey , Fisher Simon E. , Verhulst Brad , Dale Philip S. , Pourcain Beate St
Published: August 2020

There is genetic overlap between many measures of literacy, language and phonological working memory (PWM) though our knowledge of multivariate genetic architectures is incomplete. Here, we directly modeled genetic trait interrelationships in unrelated UK youth (8-13 years, N=6,453), as captured by genome-wide relationship matrices, using novel structural equation modeling techniques. We identified, besides shared genetic factors across different domains (explaining 91-97% genetic variance in literacy-related measures such as passage reading fluency, spelling, phonemic awareness, 44% in oral language and 53% in PWM), evidence for distinct cognitive abilities; trait-specific genetic influences ranged between 47% for PWM to 56% for oral language. Among reading fluency measures (non-word, word and passage reading), single-word reading was genetically most diverse. Multivariate genetic and residual covariance patterns showed concordant effect directionality, except for near-zero residual correlations between oral language and literacy-related abilities. These findings suggest differences in etiological mechanisms and trait modifiability even among genetically highly correlated skills.
The FDA-approved drug Alectinib compromises SARS-CoV-2 nucleocapsid phosphorylation and inhibits viral infection in vitro
Authors: Yaron Tomer M. , Heaton Brook E. , Levy Tyler M. , Johnson Jared L. , Jordan Tristan X. , Cohen Benjamin M. , Kerelsky Alexander , Lin Ting-Yu , Liberatore Katarina M. , Bulaon Danielle K. , Kastenhuber Edward R. , Mercadante Marisa N. , Shobana-Ganesh Kripa , He Long , Schwartz Robert E. , Chen Shuibing , Weinstein Harel , Elemento Olivier , Piskounova Elena , Nilsson-Payant Benjamin E. , Lee Gina , Trimarco Joseph D. , Burke Kaitlyn N. , Hamele Cait E. , Chaparian Ryan R. , Harding Alfred T. , Tata Aleksandra , Zhu Xinyu , Tata Purushothama Rao , Smith Clare M. , Possemato Anthony P. , Tkachev Sasha L. , Hornbeck Peter V. , Beausoleil Sean A. , Anand Shankara K. , Aguet François , Getz Gad , Davidson Andrew D. , Heesom Kate , Kavanagh-Williamson Maia , Matthews David , tenOever Benjamin R. , Cantley Lewis C. , Blenis John , Heaton Nicholas S.
Published: August 2020

While vaccines are vital for preventing COVID-19 infections, it is critical to develop new therapies to treat patients who become infected. Pharmacological targeting of a host factor required for viral replication can suppress viral spread with a low probability of viral mutation leading to resistance. In particular, host kinases are highly druggable targets and a number of conserved coronavirus proteins, notably the nucleoprotein (N), require phosphorylation for full functionality. In order to understand how targeting kinases could be used to compromise viral replication, we used a combination of phosphoproteomics and bioinformatics as well as genetic and pharmacological kinase inhibition to define the enzymes important for SARS-CoV-2 N protein phosphorylation and viral replication. From these data, we propose a model whereby SRPK1/2 initiates phosphorylation of the N protein, which primes for further phosphorylation by GSK-3α/β and CK1 to achieve extensive phosphorylation of the N protein SR-rich domain. Importantly, we were able to leverage our data to identify an FDA-approved kinase inhibitor, Alectinib, that suppresses N phosphorylation by SRPK1/2 and limits SARS-CoV-2 replication. Together, these data suggest that repurposing or developing novel host-kinase directed therapies may be an efficacious strategy to prevent or treat COVID-19 and other coronavirus-mediated diseases.
The Protein Hourglass: Exact First Passage Time Distributions for Protein Thresholds
Authors: Rijal Krishna , Prasad Ashok , Das Dibyendu
Published: August 2020

Protein thresholds have been shown to act as an ancient timekeeping device, such as in the time to lysis ofE. coliinfected with bacteriophagelambda. The time taken for protein levels to reach a particular threshold for the first time is defined as the first passage time of the protein synthesis system, which is a stochastic quantity. The first few moments of the distribution of first passage times were known earlier, but an analytical expression for the full distribution was not available. In this work, we derive an analytical expression for the first passage times for a long-lived protein. This expression allows us to calculate the full distribution not only for cases of no self-regulation, but also for both positive and negative self-regulation of the threshold protein. We show that the shape of the distribution matches previous experimental data on lambda-phage lysis time distributions. We also provide analytical expressions for the FPT distribution with non-zero degradation in Laplace space. Furthermore, we study the noise in the precision of the first passage times described by coefficient of variation (CV) of the distribution as a function of the protein threshold value. We show that under conditions of positive self-regulation, the CV declines monotonically with increasing protein threshold, while under conditions of linear negative self-regulation, there is an optimal protein threshold that minimizes the noise in the first passage times.
Belowground competition favors character convergence but not character displacement in root traits
Authors: Colom Sara M , Baucom Regina S
Published: August 2020

Character displacement can play a major role in species ecology and evolution, however, research testing whether character displacement can influence the evolution of root traits in plant systems remains scarce in the literature. Here we investigated the potential that character displacement may influence the evolution of root traits using two closely related morning glory species,Ipomoea purpureaandI. hederacea.

We performed a field experiment where we grew the common morning glory,I. purpurea, in the presence and absence of competition fromI. hederaceaand examined the potential that the process of character displacement could influence the evolution of root traits.

We found maternal line variation in root phenotypes and evidence that belowground competition acts as an agent of selection on these traits. Our test of character displacement, however, showed evidence of characterconvergence on our measure of root architecture rather thandisplacement. These results suggest that plants may be constrained by their local environments to express a phenotype that enhances fitness. Therefore, the conditions of the competitive environment experienced by a plant may influence the potential for character convergence or displacement to influence the evolution of root traits.
Rab7 mediates Wnt-dependent mesoderm patterning and gastrulation inXenopus
Authors: Kreis Jennifer , Wielath Fee M. , Vick Philipp
Published: August 2020

Early embryogenesis requires tightly controlled temporal and spatial activity of growth factors. Modulation is achieved at multiple levels, from cellular transcription to tissue-scale propagation, concerning activation, maintenance, or termination of signaling. Intracellularly, the endolysosomal system emerged as important factor for such regulations. Endocytosis and early endosomes mainly orchestrate activation and transduction of signals, while late endosomes are thought to mediate lysosome-based termination. Only few reports attributed activating roles to late endosomes, mainly for Wnt pathway activation. InXenopus, few is known about developmental roles of endosomal regulators, or their function for signaling, especially true for late endosomes. Therefore, we chose to analyze a hypothesized positive role of Rab7 in this context, a small GTPase mainly known for its role as a late endosomal regulator. We analyzed if Rab7 participates in early activation of growth factor signaling, focusing on Wnt signaling. We foundrab7expressed very localized throughout development, and inhibition resulted in strong gastrulation defects. Concomitantly, Rab7 was strictly required for both, exogenous activation and endogenous Wnt-mediated patterning of the mesoderm.

Summary Statement

The late endosomal regulator Rab7 is required for gastrulation movements and axis elongation inXenopusby positively regulating Wnt signaling-mediated mesoderm patterning.
MethylSPWNet and MethylCapsNet: Biologically Motivated Organization of DNAm Neural Network, Inspired by Capsule Networks
Authors: Levy Joshua J. , Chen Youdinghuan , Azizgolshani Nasim , Petersen Curtis L. , Titus Alexander J. , Moen Erika L. , Vaickus Louis J. , Salas Lucas A. , Christensen Brock C.
Published: August 2020

DNA methylation (DNAm) alterations have been heavily implicated in carcinogenesis and the pathophysiology of diseases through upstream regulation of gene expression. DNAm deep-learning approaches are able to capture features associated with aging, cell type, and disease progression, but lack incorporation of prior biological knowledge. Here, we present modular, user-friendly deep learning methodology and software,MethylCapsNetandMethylSPWNet, that group CpGs into biologically relevant capsules – such as gene promoter context, CpG island relationship, or user-defined groupings – and relate them to diagnostic and prognostic outcomes. We demonstrate these models’ utility on 3,897 individuals in the classification of central nervous system (CNS) tumors.MethylCapsNetandMethylSPWNetprovide an opportunity to increase DNAm deep learning analyses’ interpretability by enabling a flexible organization of DNAm data into biologically relevant capsules.
Learned adaptive multiphoton illumination microscopy
Authors: Pinkard Henry , Baghdassarian Hratch , Mujal Adriana , Roberts Ed , Hu Kenneth H. , Friedman Daniel Haim , Malenica Ivana , Shagam Taylor , Fries Adam , Corbin Kaitlin , Krummel Matthew F. , Waller Laura
Published: August 2020

Multiphoton microscopy is a powerful technique for deepin vivoimaging in scattering samples. However, it requires precise, sample-dependent increases in excitation power with depth in order to maintain signal while minimizing photodamage. We show that cells with identical fluorescent labels imagedin situcan be used to train a physics-based machine learning model that solves this problem. After this training has been performed, the correct illumination power can be predicted and adaptively adjusted at each point in a 3D volume on subsequent samples as a function of the sample’s shape, without the need for specialized fluorescent labelling. We use this technique forin vivoimaging of immune responses in mouse lymph nodes following vaccination, with imaging volumes 2-3 orders of magnitude larger than previously reported. We achieve visualization of physiologically realistic numbers of antigen-specific T cells for the first time, and demonstrate changes in the global organization and motility of dendritic cell networks during the early stages of the immune response.
A chlorzoxazone-baclofen combination improves cerebellar impairment in spinocerebellar ataxia type 1
Authors: Bushart David D. , Huang Haoran , Man Luke J. , Morrison Logan M. , Shakkottai Vikram G.
Published: August 2020

Background

A combination of central muscle relaxants, chlorzoxazone and baclofen (chlorzoxazone-baclofen), has been proposed for treatment of cerebellar symptoms in human spinocerebellar ataxia (SCA). However, central muscle relaxants can worsen balance. The optimal dose for target engagement without toxicity remains unknown.

Objectives

Using the genetically preciseAtxn1154Q/2Qmodel of SCA1, we determine the role of cerebellar dysfunction in motor impairment. We also aim to identify appropriate concentrations of chlorzoxazone-baclofen needed for target engagement without toxicity to plan for human clinical trials.

Methods

We use patch clamp electrophysiology in acute cerebellar slices and immunostaining to identify the specific ion channels targeted by chlorzoxazone-baclofen. Behavioral assays for coordination and grip strength are used to determine specificity of chlorzoxazone-baclofen for improving cerebellar dysfunction without off-target effects inAtxn1154Q/2Qmice.

Results

We identify irregular Purkinje neuron firing in association with reduced expression of the ion channelsKcnma1andCacna1ginAtxn1154Q/2Qmice. Usingin vitroelectrophysiology in brain slices, we identify concentrations of chlorzoxazone-baclofen that improve Purkinje neuron spike regularity without reducing firing frequency. At a disease stage inAtxn1154Q/2Qmice when motor impairment is due to cerebellar dysfunction, orally administered chlorzoxazone-baclofen improves motor performance without affecting muscle strength.

Conclusion

We identify a tight relationship between baclofen-chlorzoxazone concentrations needed to engage target, and levels above which cerebellar function will be compromised. We propose to use this information for a novel clinical trial design, using sequential dose escalation within each subject, to identify dose levels that are likely to improve ataxia symptoms while minimizing toxicity.
CopR, a global regulator of transcription to maintain copper homeostasis inPyrococcus furiosus
Authors: Grünberger Felix , Reichelt Robert , Waege Ingrid , Ned Verena , Bronner Korbinian , Kaljanac Marcell , Weber Nina , El Ahmad Zubeir , Knauss Lena , Madej M. Gregor , Ziegler Christine , Grohmann Dina , Hausner Winfried
Published: August 2020

Although copper is in many cases an essential micronutrient for cellular life, higher concentrations are toxic. Therefore, all living cells have developed strategies to maintain copper homeostasis. In this manuscript, we have analysed the transcriptome-wide response ofPyrococcus furiosusto increased copper concentrations and described the essential role of the putative copper-sensing metalloregulator CopR in the detoxification process.

To this end, we employed biochemical and biophysical methods to characterise the role of CopR. Additionally, acopRknockout strain revealed an amplified sensitivity in comparison to the parental strain towards increased copper levels, which designates an essential role of CopR for copper homeostasis. To learn more about the CopR-regulated gene network, we performed differential gene expression and ChIP-seq analysis under normal and 20 μM copper-shock conditions. By integrating the transcriptome and genome-wide binding data, we found that CopR binds to the upstream regions of many copper-induced genes. Negative-stain transmission electron microscopy and 2D class averaging revealed an octameric assembly formed from a tetramer of dimers for CopR, similar to published crystal structures from the Lrp family. In conclusion, we propose a model for CopR-regulated transcription and highlight the complex regulatory network that enablesPyrococcusto respond to increased copper concentrations.
Methylation of RNA Cap in SARS-CoV-2 captured by serial crystallography
Authors: Wilamowski M. , Sherrell D.A. , Minasov G. , Kim Y. , Shuvalova L. , Lavens A. , Chard R. , Maltseva N. , Jedrzejczak R. , Rosas-Lemus M. , Saint N. , Foster I.T. , Michalska K. , Satchell K.J.F. , Joachimiak A
Published: August 2020

The genome of the SARS-CoV-2 coronavirus contains 29 proteins, of which 15 are nonstructural. Nsp10 and Nsp16 form a complex responsible for the capping of mRNA at the 5′ terminus. In the methylation reaction the S-adenosyl-L-methionine serves as the donor of the methyl group that is transferred to Cap-0 at the first transcribed nucleotide to create Cap-1. The presence of Cap-1 makes viral RNAs mimic the host transcripts and prevents their degradation. To investigate the 2′-O methyltransferase activity of SARS-CoV-2 Nsp10/16, we applied fixed-target serial synchrotron crystallography (SSX) which allows for physiological temperature data collection from thousands of crystals, significantly reducing the x-ray dose while maintaining a biologically relevant temperature. We determined crystal structures of Nsp10/16 that revealed the states before and after the methylation reaction, for the first time illustrating coronavirus Nsp10/16 complexes with them7GpppAm2′-OCap-1, where 2′OH of ribose is methylated. We compare these structures with structures of Nsp10/16 at 297 K and 100 K collected from a single crystal. This data provide important mechanistic insight and can be used to design small molecules that inhibit viral RNA maturation making SARS-CoV-2 sensitive to host innate response.