bluuebunny/crossref_metadata_2025 huggingface dataset:
Spatio-temporal mRNA dynamics in the early zebrafish embryo
Authors: Holler Karoline , Neuschulz Anika , Drewe-Boß Philipp , Mintcheva Janita , Spanjaard Bastiaan , Arsiè Roberto , Ohler Uwe , Landthaler Markus , Junker Jan Philipp
Published: November 2020

Early stages of embryogenesis depend heavily on subcellular localization and transport of maternally deposited mRNA. However, systematic analysis of these processes is currently hindered by a lack of spatio-temporal information in single-cell RNA sequencing. Here, we combined spatially-resolved transcriptomics and single-cell RNA labeling to study the spatio-temporal dynamics of the transcriptome during the first few hours of zebrafish development. We measured spatial localization of mRNA molecules with sub-single-cell resolution at the one-cell stage, which allowed us to identify a class of mRNAs that are specifically localized at an extraembryonic position, the vegetal pole. Furthermore, we established a method for high-throughput single-cell RNA labeling in early zebrafish embryos, which enabled us to follow the fate of individual maternal transcripts until gastrulation. This approach revealed that many localized transcripts are specifically transported to the primordial germ cells. Finally, we acquired spatial transcriptomes of two xenopus species, and we compared evolutionary conservation of localized genes as well as enriched sequence motifs. In summary, we established sub-single-cell spatial transcriptomics and single-cell RNA labeling to reveal principles of mRNA localization in early vertebrate development.
Fast Vesicle Fusion Mediated by Hydrophobic Dipeptides
Authors: Wei C. , Pohorille A.
Published: November 2020

To understand the transition from inanimate matter to life, we studied a process that directly couples simple metabolism to evolution via natural selection, demonstrated experimentally by Adamala and Szostak (Nat. Chem. 2013, 5, 495–501). In this process, dipeptides synthesized inside precursors of cells promote absorption of fatty acid micelles to vesicles inducing their preferential growth and division at the expense of other vesicles. The process is explained on the basis of coarse-grained molecular dynamics simulations, each extending for tens of microseconds, carried out to model fusion between a micelle and a membrane, both made of fatty acids in the absence and presence of hydrophobic dipeptides. In all systems with dipeptides, but not in their absence, fusion events were observed. They involve the formation of a stalk made by hydrophobic chains from the micelle and the membrane, similar to that postulated for vesicle-vesicle fusion. The emergence of a stalk is facilitated by transient clusters of dipeptides, side chains of which formed hydrophobic patches at the membrane surface. Committor probability calculations indicate that the size of a patch is a suitable reaction coordinate and allow for identifying the transition state for fusion. Free energy barrier to fusion is greatly reduced in the presence of dipeptides to only 4-5 kcal/mol, depending of the hydrophobicity of side chains. The mechanism of mediated fusion, which is expected to apply to other small peptides and hydrophobic molecules, provides a robust means by which a nascent metabolism can confer evolutionary advantage to precursors of cells.

Significance

We study fusion of micelles and vesicles in the presence and absence of hydrophobic dipeptides by way of molecular dynamics simulations and demonstrate the spontaneous formation of a hydrophobic stalk that has been also long postulated as the key process in vesicle-vesicle fusion but shown only in forced simulations. We show that fusion is facilitated by clusters of dipeptides that form hydrophobic patches at the membrane surface. In order to understand the transition from inanimate matter to life, we explain and generalize experimental findings that hydrophobic dipeptides synthesized inside precursors of cells promote absorption of fatty acid micelles to vesicles inducing their preferential growth and division, thus providing cells endowed with such metabolism with evolutionary advantage.
AI-Driven Multiscale Simulations Illuminate Mechanisms of SARS-CoV-2 Spike Dynamics
Authors: Casalino Lorenzo , Dommer Abigail , Gaieb Zied , Barros Emilia P. , Sztain Terra , Ahn Surl-Hee , Trifan Anda , Brace Alexander , Bogetti Anthony , Ma Heng , Lee Hyungro , Turilli Matteo , Khalid Syma , Chong Lillian , Simmerling Carlos , Hardy David J. , Maia Julio D. C. , Phillips James C. , Kurth Thorsten , Stern Abraham , Huang Lei , McCalpin John , Tatineni Mahidhar , Gibbs Tom , Stone John E. , Jha Shantenu , Ramanathan Arvind , Amaro Rommie E.
Published: November 2020

We develop a generalizable AI-driven workflow that leverages heterogeneous HPC resources to explore the time-dependent dynamics of molecular systems. We use this workflow to investigate the mechanisms of infectivity of the SARS-CoV-2 spike protein, the main viral infection machinery. Our workflow enables more efficient investigation of spike dynamics in a variety of complex environments, including within a complete SARS-CoV-2 viral envelope simulation, which contains 305 million atoms and shows strong scaling on ORNL Summit using NAMD. We present several novel scientific discoveries, including the elucidation of the spike’s full glycan shield, the role of spike glycans in modulating the infectivity of the virus, and the characterization of the flexible interactions between the spike and the human ACE2 receptor. We also demonstrate how AI can accelerate conformational sampling across different systems and pave the way for the future application of such methods to additional studies in SARS-CoV-2 and other molecular systems.

ACM Reference Format

Lorenzo Casalino1†, Abigail Dommer1†, Zied Gaieb1†, Emilia P. Barros1, Terra Sztain1, Surl-Hee Ahn1, Anda Trifan2,3, Alexander Brace2, Anthony Bogetti4, Heng Ma2, Hyungro Lee5, Matteo Turilli5, Syma Khalid6, Lillian Chong4, Carlos Simmerling7, David J. Hardy3, Julio D. C. Maia3, James C. Phillips3, Thorsten Kurth8, Abraham Stern8, Lei Huang9, John McCalpin9, Mahidhar Tatineni10, Tom Gibbs8, John E. Stone3, Shantenu Jha5, Arvind Ramanathan2∗, Rommie E. Amaro1∗. 2020. AI-Driven Multiscale Simulations Illuminate Mechanisms of SARS-CoV-2 Spike Dynamics. In Supercomputing ’20: International Conference for High Performance Computing, Networking, Storage, and Analysis. ACM, New York, NY, USA, 14 pages. https://doi.org/finalDOI
Null effects of levodopa on reward- and error-based motor adaptation, savings, and anterograde interference
Authors: Palidis Dimitrios J. , McGregor Heather R. , Vo Andrew , MacDonald Penny A. , Gribble Paul L.
Published: November 2020

Dopamine signaling is thought to mediate reward-based learning. We tested for a role of dopamine in motor adaptation by administering the dopamine precursor levodopa to healthy participants in two experiments involving reaching movements. Levodopa has been shown to impair reward-based learning in cognitive tasks. Thus, we hypothesized that levodopa would selectively impair aspects of motor adaptation that depend on reinforcement of rewarding actions.

In the first experiment, participants performed two separate tasks in which adaptation was driven either by visual error-based feedback of the hand position or binary reward feedback. We used EEG to measure event-related potentials evoked by task feedback. We hypothesized that levodopa would specifically diminish adaptation and the neural responses to feedback in the reward learning task. However, levodopa did not affect motor adaptation in either task nor did it diminish event-related potentials elicited by reward outcomes.

In the second experiment, participants learned to compensate for mechanical force field perturbations applied to the hand during reaching. Previous exposure to a particular force field can result in savings during subsequent adaptation to the same force field or interference during adaptation to an opposite force field. We hypothesized that levodopa would diminish savings and anterograde interference, as previous work suggests that these phenomena result from a reinforcement learning process. However, we found no reliable effects of levodopa.

These results suggest that reward-based motor adaptation, savings, and interference may not depend on the same dopaminergic mechanisms that have been shown to be disrupted by levodopa during various cognitive tasks.

New and Noteworthy

Motor adaptation relies on multiple processes including reinforcement of successful actions. Cognitive reinforcement learning is impaired by levodopa-induced disruption of dopamine function. We administered levodopa to healthy adults who participated in multiple motor adaptation tasks. We found no effects of levodopa on any component of motor adaptation. This suggests that motor adaptation may not depend on the same dopaminergic mechanisms as cognitive forms or reinforcement learning that have been shown to be impaired by levodopa.
SeRenDIP-CE: Sequence-based Interface Prediction for Conformational Epitopes
Authors: Hou Qingzhen , Stringer Bas , Waury Katharina , Capel Henriette , Haydarlou Reza , Abeln Sanne , Heringa Jaap , Feenstra K. Anton
Published: November 2020

Motivation

Antibodies play an important role in clinical research and biotechnology, with their specificity determined by the interaction with the antigen’s epitope region, as a special type of protein-protein interaction (PPI) interface. The ubiquitous availability of sequence data, allows us to predicting epitopes from sequence in order to focus time-consuming wet-lab experiments onto the most promising epitope regions. Here, we extend our previously developed sequence-based predictors for homodimer and heterodimer PPI interfaces to predict epitope residues that have the potential to bind an antibody.

Results

We collected and curated a high quality epitope dataset from the SAbDaB database. Our generic PPI heterodimer predictor obtained an AUC-ROC of 0.666 when evaluated on the epitope test set. We then trained a random forest model specifically on the epitope dataset, reaching AUC 0.694. Further training on the combined heterodimer and epitope datasets, improves our final predictor to AUC 0.703 on the epitope test set. This is better than the best state-of-the-art sequence-based epitope predictor BepiPred-2.0. On one solved antibody-antigen structure of the COVID19 virus spike RNA binding domain, our predictor reaches AUC 0.778. We added the SeRenDIP-CE Conformational Epitope predictors to our webserver, which is simple to use and only requires a single antigen sequence as input, which will help make the method immediately applicable in a wide range of biomedical and biomolecular research.

Availability

Webserver, source code and datasets are available atwww.ibi.vu.nl/programs/serendipwww/

Contact

k.a.feenstra@vu.nl
Low intrinsic efficacy alone cannot explain the improved side effect profiles of new opioid agonists
Authors: Stahl Edward L. , Bohn Laura M.
Published: November 2020

In a recent report in Science Signaling (DOI: 10.1126/scisignal.aaz3140), it was suggested that low intrinsic agonism, and not biased agonism, leads to an improvement in the separation of potency in opioid-induced respiratory suppression versus antinociception. Although many of the compounds that were tested have been shown to display G protein signaling bias in prior publications, the authors conclude that since they cannot detect biased agonism in their cellular signaling studies the compounds are therefore not biased agonists. Rather, they conclude that it is low intrinsic efficacy that leads to the therapeutic window improvement. Intrinsic efficacy is the extent to which an agonist can stimulate a G protein-coupled receptor (GPCR) response in a system, while biased agonism takes into consideration not only intrinsic efficacy, but also potency of an agonist in an assay. Herein, we have re-analyzed the data presented in the published work (DOI: 10.1126/scisignal.aaz3140) (including the recent Erratum: DOI: 10.1126/scisignal.abf9803) to derive intrinsic efficacy and bias factors as ΔΔlog(τ/KA) and ΔΔlog(Emax/EC50). Based on this reanalysis, the data support the conclusion that biased agonism, favoring G protein signaling, was observed. Moreover, a conservation of rank order intrinsic efficacy was not observed upon comparing responses in each assay, further suggesting that multiple active receptor states were present. These observations agree with prior studies wherein oliceridine, PZM21 and SR-17018 were first described as biased agonists with improvement in antinociception over respiratory suppression in mice. Therefore, the data in the Science Signaling manuscript does provide strong corroborating evidence that G protein signaling bias may be a means to improve opioid analgesia while avoiding certain undesirable side effects.
Discovery of a Promising Susceptibility Factor for Fusarium Head Blight in Wheat
Authors: Chhabra Bhavit , Tiwari Vijay , Gill Bikram S , Dong Yanhong , Rawat Nidhi
Published: November 2020

Fusarium head blight (FHB) disease of wheat caused byFusariumspp. deteriorates both quantity and quality of the crop. Manipulation of susceptibility factors, the genes facilitating disease development in plants, offers a novel and alternative strategy for enhancing FHB resistance in plants. In this study, a major effect susceptibility gene for FHB was identified on the short arm of chromosome 7A (7AS). Nullisomic-tetrasomic lines for homoeologous group-7 of wheat revealed dosage effect of the gene, with tetrasomic 7A being more susceptible than control Chinese Spring wheat, qualifying it as a bonafide susceptibility factor. The gene locus was conserved in six chromosome 7A inter-varietal wheat substitution lines of diverse origin and a tetraploidTriticum dicoccoidesgenotype. The susceptibility gene was named asSF7ASFHBand mapped on chromosome 7AS to 48.5-50.5 Mb peri-centromeric region between del7AS-3 and del7AS-8. Our results showed that deletion ofSF7ASFHBimparts ~ 50-60% type 2 FHB resistance (against the spread of the fungal pathogen) and its manipulation may lead to enhanced resistance against FHB in wheat.

Highlight

Discovery and mapping of a conserved susceptibility factor located on the short arm of wheat chromosome 7A whose deletion makes plants resistant to Fusarium Head Blight.
PCQC: Selecting optimal principal components for identifying clusters with highly imbalanced class sizes in single-cell RNA-seq data
Authors: Burstein David , Fullard John F. , Roussos Panos
Published: November 2020

Prior to identifying clusters in single cell gene expression experiments, selecting the top principal components is a critical step for filtering out noise in the data set. Identifying these top principal components typically focuses on the total variance explained, and principal components that explain small clusters from rare populations will not necessarily capture a large percentage of variance in the data. We present a computationally efficient alternative for identifying the optimal principal components based on the tails of the distribution of variance explained for each observation. We then evaluate the efficacy of our approach in three different single cell RNA-sequencing data sets and find that our method matches, or outperforms, other selection criteria that are typically employed in the literature.

Availability and implementation

pcqc is written in Python and available atgithub.com/RoussosLab/pcqc
Impacts of water quality onAcroporacoral settlement: The relative importance of substrate quality and light
Authors: Ricardo Gerard F. , Harper Charlotte E. , Negri Andrew P. , Luter Heidi M. , Wahab Muhammad Azmi Abdul , Jones Ross J.
Published: November 2020

Coral larval settlement patterns are influenced by a vast array of factors; however, the relative roles of individual factors are rarely tested in isolation, leading to confusion about which are most crucial for settlement. For example, direct effects of light environment are often cited as a major determinate of settlement patterns, yet this has not been demonstrated under environmentally realistic lighting regimes in the absence of confounding factors. Here we apply programmable multispectral lights to create realistic light spectra, while removing correlating (but not obvious) factors that are common in laboratory settlement experiments. Using two common species ofAcropora– key framework builders of the Great Barrier Reef – we find little evidence that light intensity or changes in the spectral profile play a substantial role in larval settlement under most environmentally realistic settings but can under more extreme or artificial settings. We alternatively hypothesise and provide evidence that chronic conditions of light and recent sediment exposures that impact benthic substrates (e.g., crustose coralline algae) preceding settlement have a greater impact, with up to 74% decrease in settlement observed on substrates with prior exposure and poor water quality conditions. Management of water quality conditions that impact the quality of benthic-settlement substrates therefore should present a priority area of focus for improving coral recruitment.
Clipper: p-value-free FDR control on high-throughput data from two conditions
Authors: Ge Xinzhou , Chen Yiling Elaine , Song Dongyuan , McDermott MeiLu , Woyshner Kyla , Manousopoulou Antigoni , Wang Ning , Li Wei , Wang Leo D. , Li Jingyi Jessica
Published: November 2020

High-throughput biological data analysis commonly involves identifying features such as genes, genomic regions, and proteins, whose values differ between two conditions, from numerous features measured simultaneously. The most widely-used criterion to ensure the analysis reliability is the false discovery rate (FDR), which is primarily controlled based on p-values. However, obtaining valid p-values relies on either reasonable assumptions of data distribution or large numbers of replicates under both conditions. Clipper is a general statistical framework for FDR control without relying on p-values or specific data distributions. Clipper outperforms existing methods for a broad range of applications in high-throughput data analysis.
A newly identified prophage-encoded gene,ymfM, causes SOS-inducible filamentation inEscherichia coli
Authors: Ansari Shirin , Walsh James C. , Bottomley Amy L. , Duggin Iain G. , Burke Catherine , Harry Elizabeth J.
Published: November 2020

Rod-shaped bacteria such asEscherichia colican regulate cell division in response to stress, leading to filamentation, a process where cell growth and DNA replication continues in the absence of division, resulting in elongated cells. The classic example of stress is DNA damage which results in the activation of the SOS response. While the inhibition of cell division during SOS has traditionally been attributed to SulA inE. coli, a previous report suggests that the e14 prophage may also encode an SOS-inducible cell division inhibitor, previously named SfiC. However, the exact gene responsible for this division inhibition has remained unknown for over 35 years. A recent high-throughput over-expression screen inE. coliidentified the e14 prophage gene,ymfM, as a potential cell division inhibitor. In this study, we show that the inducible expression ofymfMfrom a plasmid causes filamentation. We show that this expression ofymfMresults in the inhibition of Z ring formation and is independent of the well characterised inhibitors of FtsZ ring assembly inE. coli, SulA, SlmA and MinC. We confirm thatymfMis the gene responsible for the SfiC+phenotype as it contributes to the filamentation observed during the SOS response. This function is independent of SulA, highlighting that multiple division inhibition pathways exist during the stress-induced SOS response. Our data also highlight that our current understanding of cell division regulation during the SOS response is incomplete and raises many questions regarding how many inhibitors there actually are and their purpose for the survival of the organism.

Importance

Filamentation is an important biological mechanism which aids in the survival, pathogenesis and antibiotic resistance of bacteria within different environments, including pathogenic bacteria such as uropathogenicEscherichia coli. Here we have identified a bacteriophage-encoded cell division inhibitor which contributes to the filamentation that occurs during the SOS response. Our work highlights that there are multiple pathways that inhibit cell division during stress. Identifying and characterising these pathways is a critical step in understanding survival tactics of bacteria which become important when combating the development of bacterial resistance to antibiotics and their pathogenicity.
Comparative genomics of Exiguobacterium reveals what makes a cosmopolitan bacterium
Authors: Zhang De-Chao , Zhu Zhaolu , Li Xudong , Guan Ziyu , Zheng Jinshui
Published: November 2020

Although the adaptation strategies of bacteria to specific environmental conditions are widely reported, fewer studies have addressed how microbe with cosmopolitan distribution adapted to diverse habitats. Exiguobacterium is a versatile genus whose members have been commonly found in great variety of habitats. To understand the mechanism behind the universally of Exiguobacterium, we isolated 103 strains from diverse environments, and performed large-scale metabolic and adaptive ability tests. We found that the capacities of survival in a wide range of temperature, salinity and pH are common for most Exiguobacterium members. According to the core genome based phylogeny and ANI analysis, 26 putative species including 13 putative new ones were identified and two genetic groups were classified as Group I and II. Comparative genomic analysis revealed that Exiguobacterium members can not only utilize a variety of complex polysaccharides and proteins that are ubiquitous in both terrestrial and marine environments, but also have a number of chaperonins and transporters which could support them to survive in different extreme environments. In addition, we found that the species from Group I can be found in more diverse environments with larger genome size compared to those of Group II. Twenty-five transporter families involved in transport of organic or inorganic substrates and environments stresses resistance were predicted to be enriched in Group I strains. This study provided the comprehensive insight into general genetic basis of the cosmopolitan distribution of a bacteria genus and deciphered putative determinants behind the ecological difference of different groups belonging to the same genus.

IMPORTANCE

The wide distribution characteristics make Exiguobacterium a valuable model for studying adaptive strategy of bacteria adapted to multiple habitats. In this study, we found that comprehensive capacity of diverse polysaccharides utilization and environmental stress resistance is the important basis for survival, and selective expansion of transporters is an evolution and adaptation strategy for extensive distribution. Our findings are significant for understanding the adaptation and evolution mechanisms of cosmopolitan bacteria and explaining the vital genomic traits that facilitate niches adaptation.
Spatio-temporal Control of ERK Pulse Frequency Coordinates Fate Decisions during Mammary Acinar Morphogenesis
Authors: Ender Pascal , Gagliardi Paolo Armando , Dobrzyński Maciej , Frismantiene Agne , Dessauges Coralie , Höhener Thomas , Jacques Marc-Antoine , Cohen Andrew R. , Pertz Olivier
Published: November 2020

The signaling events controlling proliferation, survival, and apoptosis during mammary epithelial acinar morphogenesis remain poorly characterized. By imaging single-cell ERK activity dynamics in MCF10A acini, we find that these fates depend on the average frequency of non-periodic ERK pulses. High pulse frequency is observed during initial acinus growth, correlating with rapid cell motility. Subsequent decrease in motility correlates with lower ERK pulse frequency and quiescence. Later, during lumen formation, coordinated ERK waves emerge across multiple cells of an acinus, correlating with high and low ERK pulse frequency in outer surviving and inner dying cells respectively. Optogenetic entrainment of ERK pulses causally connects high ERK pulse frequency with inner cell survival. Acini harboring the PIK3CA H1047R mutation, commonly observed in breast cancer, display increased ERK pulse frequency, inner cell survival and loss of lumen formation. Thus, fate decisions during acinar morphogenesis are fine-tuned by different spatio-temporal coordination modalities of ERK pulse frequency.
How a Hemicarcerand Incarcerates Guests at Room Temperature Decoded with Modular Simulations
Authors: McFerrin Katherine G. , Pang Yuan-Ping
Published: November 2020

Hemicarcerands are host molecules created to study constrictive binding with guest molecules for insights into the rules of molecular complexation. However, the molecular dynamics simulations that facilitate such studies have been limited because three-dimensional models of hemicarcerands are tedious to build and their atomic charges are complicated to derive. There have been no molecular dynamics simulations of the reported water-soluble hemicarcerand (Octacid4) that explain how it uniquely encapsulates its guests at 298 K and keeps them encapsulated at 298 K in NMR experiments. Herein we report a modular approach to hemicarcerand simulations that simplifies the model building and charge derivation in a manner reminiscent of the approach to protein simulations with truncated amino acids as building blocks. We also report thatapoOctacid4 in water adopts two clusters of conformations, one of which has an equatorial portal open thus allowing guests to enter the cavity of Octacid4, in microsecond molecular dynamics simulations performed using the modular approach at 298 K. Under the same simulation conditions, the guest-bound Octacid4 adopts one cluster of conformations with all equatorial portals closed thus keeping the guests incarcerated. These results explain the unique constrictive binding of Octacid4 and suggest that the guest-induced host conformational change that impedes decomplexation is a previously unrecognized conformational characteristic that promotes strong molecular complexation.
IBEX – A versatile multi-plex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues
Authors: Radtke Andrea J. , Kandov Evelyn , Lowekamp Bradley , Speranza Emily , Chu Colin J. , Gola Anita , Thakur Nishant , Shih Rochelle , Yao Li , Yaniv Ziv Rafael , Beuschel Rebecca T. , Kabat Juraj , Croteau Joshua , Davis Jeremy , Hernandez Jonathan M. , Germain Ronald N.
Published: November 2020

The diverse composition of mammalian tissues poses challenges for understanding the cell-cell interactions required for organ homeostasis and how spatial relationships are perturbed during disease. Existing methods such as single-cell genomics, lacking a spatial context, and traditional immunofluorescence, capturing only 2-6 molecular features, cannot resolve these issues. Imaging technologies have been developed to address these problems, but each possesses limitations that constrain widespread use. Here we report a new method that overcomes major impediments to highly multi-plex tissue imaging.IterativeBleachingExtends multi-pleXity (IBEX) uses an iterative staining and chemical bleaching method to enable high resolution imaging of >65 parameters in the same tissue section without physical degradation. IBEX can be employed with various types of conventional microscopes and permits use of both commercially available and user-generated antibodies in an ‘open’ system to allow easy adjustment of staining panels based on ongoing marker discovery efforts. We show how IBEX can also be used with amplified staining methods for imaging strongly fixed tissues with limited epitope retention and with oligonucleotide-based staining, allowing potential cross-referencing between flow cytometry, Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-Seq), and IBEX analysis of the same tissue. To facilitate data processing, we provide an open source platform for automated registration of iterative images. IBEX thus represents a technology that can be rapidly integrated into most current laboratory workflows to achieve high content imaging to reveal the complex cellular landscape of diverse organs and tissues.

Significance Statement

Single cell flow cytometry and genomic methods are rapidly increasing our knowledge of the diversity of cell types in metazoan tissues. However, suitably robust methods for placing these cells in a spatial context that reveal how their localization and putative interactions contribute to tissue physiology and pathology are still lacking. Here we provide a readily accessible pipeline (IBEX) for highly multi-plex immunofluorescent imaging that enables a fine-grained analysis of cells in their tissue context. Additionally, we describe extensions of the IBEX workflow to handle hard to image tissue preparations and a method to facilitate direct integration of the imaging data with flow cytometry and sequencing technologies.
Genome-wide screen of genomic imprinting in endosperm and population-level analysis reveal allelic variation for imprinting in flax
Authors: Jiang Haixia , Guo Dongliang , Ye Jiali , Gao Yanfang , Liu Huiqing , Wang Yue , Xue Min , Yan Qingcheng , Chen Jiaxun , Duan Lepeng , Li Gongzhe , Li Xiao , Xie Liqiong
Published: November 2020

Genomic imprinting is an epigenetic phenomenon caused by the biased expression of maternally and paternally inherited alleles. In flowering plants, genomic imprinting predominantly occurs in triploid endosperm and plays a vital role in seed development. In this study, we identified 241 candidate imprinted genes including 143 maternally expressed imprinted genes (MEGs) and 98 paternally expressed imprinted genes (PEGs) in flax (Linum usitatissimumL.) endosperm using deep RNA sequencing. The conservation of imprinting in plants is very limited and imprinting clustering is not a general feature. MEGs tends to be endosperm expression specific, while PEGs are non-tissue specific. Imprinted SNPs differentiated 200 flax cultivars into oil flax, oil-fiber dual purpose flax (OF) and fiber flax subgroups, suggesting that genomic imprinting contributes to intraspecific variation in flax. The nucleotide diversity (π) of imprinted genes in oil flax subgroup is significantly higher than that in fiber flax subgroup, indicating that some imprinted genes undergo positive selection during flax domestication from oil flax to fiber flax. Imprinted genes undergo positive selection is related to the functions. Eleven imprinted genes related to seed size and weight were identified using the candidate gene-based association study. Our study provides information for further exploring the function and genomic variation of imprinted genes in flax population.
Bioinformatic Analysis Linking Genomic Defects to Chemosensitivity and Mechanism of Action
Authors: Covell David G.
Published: November 2020

A joint analysis of NCI60 small molecule screening data, their genetically defective genes and mechanisms of action (MOA) of FDA approved cancer drugs screened in the NCI60 is proposed for identifying links between chemosensitivity, genomic defects and MOA. Self-organizing-maps (SOMs) are used to organize the chemosensitivity data. Student’s t-tests are used to identify SOM clusters with chemosensitivity for tumor cells harboring genetically defective genes. Fisher’s exact tests are used to reveal instances where defective gene to chemosensitivity associations have enriched MOAs. The results of this analysis find a relatively small set of defective genes, inclusive of ABL1, AXL, BRAF, CDC25A, CDKN2A, IGF1R, KRAS, MECOM, MMP1, MYC, NOTCH1, NRAS, PIK3CG, PTK2, RPTOR, SPTBN1, STAT2, TNKS and ZHX2, as possible candidates for roles in chemosensitivity for compound MOAs that target primarily, but not exclusively, kinases, nucleic acid synthesis, protein synthesis, apoptosis and tubulin. This analysis may contribute towards the goals of cancer drug discovery, development decision making, and explanation of mechanisms.
Evaluation of Genomic and Proteomic Expression of Surfactant Protein D in the Mouse Retina
Authors: Vieira Frederico , Kung Johannes W. , Bhatti Faizah
Published: November 2020

Surfactant Protein D (SP-D), an essential protein related to innate immunity, is expressed in multiple tissue types throughout the body. A closely-related protein, Surfactant Protein A (SP-A), is present in the mouse retina and is associated with neovascularization (NV) in the Oxygen-Induced Retinopathy (OIR) mouse model, mimicking retinopathy of prematurity (ROP). We hypothesized that SP-D would be present in the retina and is also associated with OIR and ROP, which is one of the leading causes of pediatric blindness due to increasing survival rates of extremely preterm newborns. In our study, we did not detect SP-D in the mouse retina through proteomic and genomic investigation at baseline and in pathways known to up-regulate SP-D in other mammal tissues. It is therefore unlikely that SP-D participates in neovascularization in the mouse retina.
Structural dynamics of the functional nonameric Type III translocase export gate
Authors: Yuan Biao , Portaliou Athina G. , Parakra Rinky , Smit Jochem H. , Wald Jiri , Li Yichen , Srinivasu Bindu , Loos Maria S. , Dhupar Harveer Singh , Fahrenkamp Dirk , Kalodimos Charalampos G. , Duong van Hoa Franck , Cordes Thorben , Karamanou Spyridoula , Marlovits Thomas C. , Economou Anastassios
Published: November 2020

Type III protein secretion is widespread in Gram-negative pathogens. It comprises the injectisome with a surface-exposed needle and an inner membrane translocase. The translocase contains the SctRSTU export channel enveloped by the export gate subunit SctV that binds chaperone/exported clients and forms a putative ante- chamber. We probed the assembly, function, structure and dynamics of SctV from enteropathogenicE.coli(EPEC). In both EPEC andE.colilab strains, SctV forms peripheral oligomeric clusters that are detergent-extracted as homo-nonamers. Membrane-embedded SctV9is necessary and sufficient to act as a receptor for different chaperone/exported protein pairs with distinct C-domain binding sites that are essential for secretion. Negative staining electron microscopy revealed that peptidisc-reconstituted His-SctV9forms a tripartite particle of ∼22 nm with a N- terminal domain connected by a short linker to a C-domain ring structure with a ∼5 nm-wide inner opening. The isolated C-domain ring was resolved with cryo-EM at 3.1 Å and structurally compared to other SctV homologues. Its four sub-domains undergo a three-stage “pinching” motion. Hydrogen-deuterium exchange mass spectrometry revealed this to involve dynamic and rigid hinges and a hyper-flexible sub-domain that flips out of the ring periphery and binds chaperones on and between adjacent protomers. These motions are coincident with pore surface and ring entry mouth local conformational changes that are also modulated by the ATPase inner stalk. We propose a model that the intrinsic dynamics of the SctV protomer are modulated by chaperones and the ATPase and could affect allosterically the other subunits of the nonameric ring during secretion.
Molecular signatures of resource competition: clonal interference favors ecological diversification and can lead to incipient speciation
Authors: Amicone Massimo , Gordo Isabel
Published: November 2020

Microbial ecosystems harbor an astonishing diversity that can persist for long times. To understand how such diversity is structured and maintained, ecological and evolutionary processes need to be integrated at similar timescales. Here, we study a model of resource competition that allows for evolutionvia de novomutation, and focus on rapidly adapting asexual populations with large mutational inputs, as typical of many bacteria species. We characterize the adaptation and diversification of an initially maladapted population and show how the eco-evolutionary dynamics are shaped by the interaction between simultaneously emerging lineages – clonal interference. We find that in large populations, more intense clonal interference fosters diversification under sympatry, increasing the probability that phenotypically and genetically distinct clusters stably coexist. In smaller populations, the accumulation of deleterious and compensatory mutations can push further the diversification process and kick-start speciation. Our findings have implications beyond microbial populations, providing novel insights about the interplay between ecology and evolution in clonal populations.
Improved prediction of femoral fracture toughness in mice by combining standard medical imaging with Raman spectroscopy
Authors: Massie Christine , Knapp Emma , Chen Keren , Berger Andrew J , Awad Hani A
Published: November 2020

Bone fragility and fracture risk are assessed by measuring the areal bone mineral density (aBMD) using dual-energy X-ray absorptiometry (DXA). While aBMD correlates with bone strength, it is a poor predictor of fragility fracture risk. Alternatively, fracture toughness assesses the bone’s resistance to crack propagation and fracture, making it a suitable bone quality metric. Here, we explored how femoral midshaft measurements from DXA, micro-computed tomography (μCT), and Raman spectroscopy could predict fracture toughness. We hypothesized that ovariectomy (OVX) decreases aBMD and fracture toughness compared to controls and we can optimize a multivariate assessment of bone quality by combining results from X-ray and Raman spectroscopy. Female mice underwent an OVX (n=5) or sham (n=5) surgery at 3 months of age. Femurs were excised 3 months after ovariectomy and assessed with Raman spectroscopy, μCT, and DXA. Subsequently, a notch was created on the anterior side of the mid-diaphysis of the femurs. Three-point bending induced a controlled fracture that initiated at the notch. The OVX mice had a significantly lower aBMD, cortical thickness, and fracture toughness when compared to controls (p<0.05). A leave one out cross-validated (LOOCV) partial least squares regression (PLSR) model based only on the combination of aBMD and cortical thickness showed no significant predictive correlations with fracture toughness, whereas a PLSR model based on principal components derived from the full Raman spectra yielded significant prediction (r2=0.71, p<0.05). Further, the PLSR model was improved by incorporating aBMD, cortical thickness, and principal components from Raman spectra (r2=0.92, p<0.001). This exploratory study demonstrates combining X-ray with Raman spectroscopy leads to a more accurate assessment of bone fracture toughness and could be a useful diagnostic tool for the assessment of fragility fracture risk.
A causal analysis of the effect of age and sex differences on brain atrophy in the elderly brain
Authors: Gómez-Ramírez Jaime , Fernández-Blázquez Miguel A. , González-Rosa Javier
Published: November 2020

We study how brain volume loss at old age is affected by factors such as age, APOE gene, sex, and school level. The quantitative characterization of brain volume loss at old age relative to young age requires at least in principle two MRI scans performed at both young and old age. There is, however, a way to address the problem by having only one MRI scan at old age. We compute the total brain loss of elderly subjects as the ratio between the estimated brain volume and the estimated total intracranial volume. Magnetic resonance imaging (MRI) scans of 890 healthy subjects aged 70 to 85 were assessed. The causal analysis of factors affecting brain atrophy was performed using Probabilistic Bayesian Modeling and the Mathematics of Causal Inference. We find that healthy subjects get into their seventies with an average brain volume loss of 30% from their maximum brain volume at a young age. Both age and the sexes are causally related to brain atrophy, with women getting to elderly age with 1% larger brain volume relative to intracranial volume than men. How the brain ages and what are the reasons for sex differences in adult lifespan are causal questions that need to be addressed with causal inference and empirical data. The graphical causal modeling presented here can be instrumental in understanding a puzzling scientific inquiry -the biological age of the brain.
Mesenchymal Stromal Cell Delivery via an Ex Vivo Bioreactor Preclinical Test System Attenuates Clot Formation for Intravascular Application
Authors: O’Rourke Brian , Nguyen Sunny , Tilles Arno W. , Bynum James A. , Cap Andrew P , Parekkadan Biju , Barcia Rita N.
Published: November 2020

While mesenchymal stromal cells (MSCs) are an appealing therapeutic option for a range of clinical applications, their potential to induce clotting when used systemically remains a safety concern, particularly in hypercoagulable conditions, such as in patients with severe COVID-19, trauma, or cancers. Here, we tested a novelex vivoapproach aimed at improving the safety of MSC systemic administration by use of a bioreactor. In this device, MSCs are seeded on the outside of a hollow-fiber filter, sequestering them behind a hemocompatible membrane, while still maintaining cross talk with blood cells and circulating signaling molecules. The potential for these bioreactor MSCs to induce clots in coagulable plasma was compared against “free” MSCs, as a model of systemic administration, which were directly injected into the circuit. Our results showed that physical isolation of the MSCs via a bioreactor extends the time necessary for clot formation to occur when compared to “free” MSCs. Measurement of cell surface data indicates the presence of known clot inducing factors, namely tissue factor and phosphatidylserine. Results also showed that recovering cells and flushing the bioreactor prior to use further prolonged clot formation time. Further, application of this technology in twoin vivomodels did not require additional heparin to maintain target ACT levels relative to the acellular device. Taken together, the use of hollow fiber filters to house MSCs, if adopted clinically, could offer a novel method to control systemic MSC exposure and prolong clot formation time.
Clinical positioning of the IAP antagonist tolinapant (ASTX660) in colorectal cancer
Authors: Crawford Nyree , Stott Katie , Sessler Tamas , McCann Christopher , McDaid William , Latimer Cheryl , Fox Jennifer , Munck Joanne M , Smyth Tomoko , Shah Alpesh , Martins Vanessa , Lawler Mark , Dunne Philip , Kerr Emma , McDade Simon S , Coyle Vicky , Longley Daniel B
Published: November 2020

Cancer cells frequently express elevated levels of Inhibitor of Apoptosis Proteins (IAPs): cIAPI, cIAP2 and XIAP. Elevated expression of cIAP1 and cIAP2 (but not XIAP) significantly correlated with poor prognosis in microsatellite stable (MSS) stage-III colorectal cancer (CRC) patients treated with adjuvant chemotherapy, suggesting their involvement in promoting resistance. Preclinical analysis of the IAP inhibitor tolinapant in CRC cell lines demonstrated robust on-target effects and caspase-8-dependent apoptosis that was inhibited by the caspase-8 paralogs FLIP and, unexpectedly, caspase-10. Importantly, tolinipant-induced apoptosis was augmented by standard-of-care chemotherapy (FOLFOX) in CRC disease models, due (at least in part) to FOLFOX-induced downregulation of Class-I histone deacetylases, leading to acetylation of the FLIP-binding partner Ku70 and downregulation of FLIP. Moreover, this effect could be phenocopied using a Class-I HDAC inhibitor. Further analyses revealed that caspase-8-knockout RIPK3-positive CRC models were sensitive to tolinostat-induced necroptosis, an effect that could be exploited with the FDA-approved caspase inhibitor emricasan. Our study provides evidence for immediate clinical exploration of tolinapant in combination with FOLFOX chemotherapy in poor prognosis MSS CRC with elevated cIAP1/2 expression.
Three-dimensional interrelationship between osteocyte network and forming mineral during human bone remodeling
Authors: Ayoubi Mahdi , van Tol Alexander F. , Weinkamer Richard , Roschger Paul , Brugger Peter C. , Berzlanovich Andrea , Bertinetti Luca , Roschger Andreas , Fratzl Peter
Published: November 2020

During bone remodeling, osteoblasts are known to deposit unmineralized collagenous tissue (osteoid), which mineralizes after some time lag. Some of the osteoblasts differentiate into osteocytes, forming a cell network within the lacunocanalicular network (LCN) of bone. To get more insight into the potential role of osteocytes in the mineralization process of osteoid, sites of bone formation were three-dimensionally imaged in nine forming human osteons using focused ion beam-scanning electron microscopy (FIB-SEM). In agreement with previous observations, the mineral concentration was found to gradually increase from the central Haversian canal towards preexisting mineralized bone. Most interestingly, a similar feature was discovered on a length scale more than 100-times smaller, whereby mineral concentration increased from the LCN, leaving around the canaliculi a zone virtually free of mineral, the size of which decreases with progressing mineralization. This suggests that the LCN controls mineral formation but not just by diffusion of mineralization precursors, which would lead to a continuous decrease of mineral concentration from the LCN. Our observation is, however, compatible with the codiffusion and reaction of precursors and inhibitors from the LCN into the bone matrix.
Highly multiplexed spatially resolved gene expression profiling of mouse organogenesis
Authors: Lohoff T. , Ghazanfar S. , Missarova A. , Koulena N. , Pierson N. , Griffiths J.A. , Bardot E.S. , Eng C.-H.L. , Tyser R.C.V. , Argelaguet R. , Guibentif C. , Srinivas S. , Briscoe J. , Simons B.D. , Hadjantonakis A.-K. , Göttgens B. , Reik W. , Nichols J. , Cai L. , Marioni J.C.
Published: November 2020

Transcriptional and epigenetic profiling of single-cells has advanced our knowledge of the molecular bases of gastrulation and early organogenesis. However, current approaches rely on dissociating cells from tissues, thereby losing the crucial spatial context that is necessary for understanding cell and tissue interactions during development. Here, we apply an image-based single-cell transcriptomics method, seqFISH, to simultaneously and precisely detect mRNA molecules for 387 selected target genes in 8-12 somite stage mouse embryo tissue sections. By integrating spatial context and highly multiplexed transcriptional measurements with two single-cell transcriptome atlases we accurately characterize cell types across the embryo and demonstrate how spatially-resolved expression of genes not profiled by seqFISH can be imputed. We use this high-resolution spatial map to characterize fundamental steps in the patterning of the midbrain-hindbrain boundary and the developing gut tube. Our spatial atlas uncovers axes of resolution that are not apparent from single-cell RNA sequencing data – for example, in the gut tube we observe early dorsal-ventral separation of esophageal and tracheal progenitor populations. In sum, by computationally integrating high-resolution spatially-resolved gene expression maps with single-cell genomics data, we provide a powerful new approach for studying how and when cell fate decisions are made during early mammalian development.