bluuebunny/crossref_metadata_2025 huggingface dataset:
Neural circuits in the mouse retina support color vision in the upper visual field
Authors: Szatko Klaudia P. , Korympidou Maria M. , Ran Yanli , Berens Philipp , Dalkara Deniz , Schubert Timm , Euler Thomas , Franke Katrin
Published: August 2019

Color vision is essential to the survival of most animals. Its neural basis lies in the retina, where chromatic signals from different photoreceptor types sensitive to distinct wavelengths are locally compared by neural circuits. Mice, like most mammals, are generally dichromatic and have two cone photoreceptor types. However, in the ventral retina most cones display the same spectral preference, impairing spectral comparisons necessary for color vision. This conflicts with behavioral evidence showing that mice can discriminate colors only in the corresponding upper visual field. Here, we systematically investigated the neural circuits underlying mouse color vision across three processing stages of the retina by recording the output of cones, bipolar and ganglion cells using two-photon imaging. Surprisingly, we found that across all retinal layers most color-opponent cells were located in the ventral retina. This started at the level of the cone output, where color-opponency was mediated by horizontal cells and likely involving rod photoreceptors. Next, bipolar cells relayed the chromatic information to ganglion cells in the inner retina, where type-specific, non-linear center-surround interactions resulted in specific color-opponent output channels to the brain. This suggests that neural circuits in the mouse retina are specifically tuned to extract color information from the upper visual field, aiding robust detection of aerial predators and ensuring the animal’s survival.
Can natural selection favour indiscriminate spite?
Authors: Patel Matishalin , West Stuart A. , Biernaskie Jay M.
Published: August 2019

Spiteful behaviours occur when an actor harms its own fitness to inflict harm on the fitness of others. Several papers have predicted that spite can be favoured in sufficiently small populations, even when the harming behaviour is directed indiscriminately at others. However, it is not clear that truly spiteful behaviour could be favoured without the harm being directed at a subset of social partners with relatively low genetic similarity to the actor (kin discrimination). Using mathematical models, we show that: (1) the evolution of spite requires kin discrimination; (2) previous models suggesting indiscriminate spite involve scenarios where the actor gains a direct feedback benefit from harming others, and so the harming is selfish rather than spiteful; (3) extreme selfishness can be favoured in small populations (and in some cases small groups) because this is where the feedback benefit of harming is greatest.
Cross frequency coupling in next generation inhibitory neural mass models
Authors: Ceni Andrea , Olmi Simona , Torcini Alessandro , Angulo-Garcia David
Published: August 2019

Coupling among neural rhythms is one of the most important mechanisms at the basis of cognitive processes in the brain. In this study we consider a neural mass model, rigorously obtained from the microscopic dynamics of an inhibitory spiking network with exponential synapses, able to autonomously generate collective oscillations (COs). These oscillations emerge via a super-critical Hopf bifurcation, and their frequencies are controlled by the synaptic time scale, the synaptic coupling and the excitability of the neural population. Furthermore, we show that two inhibitory populations in a master-slave configuration with different synaptic time scales can display various collective dynamical regimes: namely, damped oscillations towards a stable focus, periodic and quasi-periodic oscillations, and chaos. Finally, when bidirectionally coupled the two inhibitory populations can exhibit different types of θ-γ cross-frequency couplings (CFCs): namely, phase-phase and phase-amplitude CFC. The coupling between θ and γ COs is enhanced in presence of a external θ forcing, reminiscent of the type of modulation induced in Hippocampal and Cortex circuits via optogenetic drive.

In healthy conditions, the brain’s activity reveals a series of intermingled oscillations, generated by large ensembles of neurons, which provide a functional substrate for information processing. How single neuron properties influence neuronal population dynamics is an unsolved question, whose solution could help in the understanding of the emergent collective behaviors arising during cognitive processes. Here we consider a neural mass model, which reproduces exactly the macroscopic activity of a network of spiking neurons. This mean-field model is employed to shade some light on an important and ubiquitous neural mechanism underlying information processing in the brain: the θ-γ cross-frequency coupling. In particular, we will explore in detail the conditions under which two coupled inhibitory neural populations can generate these functionally relevant coupled rhythms.
An impressive capacity for cold tolerance plasticity protects against ionoregulatory collapse in the disease vector,Aedes aegypti
Authors: Jass Amanda , Yerushalmi Gil Y. , Davis Hannah E. , Donini Andrew , MacMillan Heath A.
Published: August 2019

The mosquitoAedes aegyptiis largely confined to tropical and subtropical regions but its range has recently been spreading to colder climates. As insect biogeography is closely tied to environmental temperature, understanding the limits ofAe. aegyptithermal tolerance and their capacity for phenotypic plasticity is important in predicting the spread of this species.

In this study we report on the chill coma onset and recovery, as well as low temperature survival phenotypes of larvae and adults ofAedes aegyptithat developed or were acclimated to 15°C (cold) or 25°C (warm).

Developmental cold acclimation did not affect chill coma onset of larvae but substantially reduced chill coma onset temperatures in adults. Chill coma recovery time was affected by both temperature and the duration of exposure, and developmental and adult acclimation both strongly mitigated these effects and increased rates of survival following prolonged chilling.

Female adults were far less likely to take a blood meal when cold acclimated and simply exposing females to blood (without feeding) attenuated some of the beneficial effects of cold acclimation on chill coma recovery time.

Lastly, larvae suffered from hemolymph hyperkalemia when chilled, but development in the cold attenuated the imbalance, which suggests that acclimation can prevent cold-induced ionoregulatory collapse in this species.

Our results demonstrate thatAedes aegyptilarvae and adults have the capacity to acclimate to cold temperatures and do so at least in part by better maintaining ion balance in the cold. This ability for cold acclimation may facilitate the spread of this species to higher latitudes, particularly in an era of climate change.
A Latent Variable Model for Evaluation of Disparate Ratings of Stem Cell Colonies by Two Experts
Authors: Halter Michael , Lund Steven , Peskin Adele , Li-Baboud Ya-Shian , Bajcsy Peter , Aulov Oleg , Hoeppner Daniel J. , Chenoweth Joshua G. , Kim Suel-Kee , McKay Ronald D. , Plant Anne L.
Published: August 2019

The visual inspection of pluripotent stem cell colonies by microscopy is widely used as a primary method to assess the quality of the preparations and degree of pluripotency. The lack of ground truth and the possible inconsistency of evaluations from multiple experts within and between stem cell laboratories are sources of uncertainty about the state of the cells, the reproducibility of preparations, and the efficiency of expansion protocols. To examine how to evaluate the level of confidence one has in disparate rating from experts, we explored a statistical method for assessing the differences in ratings of pluripotent stem cells by two different experts. Two experts rated phase contrast microscope images of human embryonic stem cell (hESC) colonies on a scale of 1 (poor) to 5 (maximum pluripotency character) but agreed with one another only 48% of the time. To assess whether experts used similar criteria to rate colonies, we developed custom image feature algorithms based on the stated visual criteria provided by the experts for selection of colonies. These features, plus others, were then used to develop pluripotency scoring algorithms trained to reflect ratings of both experts. We treated expert ratings as inexact indicators of a continuous pluripotency score and considered the inconsistency between expert ratings in developing our models. The model suggests that the two experts use somewhat different scales for discriminating between colony quality. Covariance analysis indicated that both experts use features that are not included in the model. Two image features, colony perimeter and a feature based on texture, were the most important for both experts for predicting the ratings. Interestingly, colony perimeter was not one of the expert-provided criteria for rating colonies, showing that this modeling approach allowed identification of features that the experts were not aware they were using. A linear model based on both experts identified each expert’s top-rated colonies as well as, or better than, the ratings of the other expert, as indicated by receiver operator characteristic curve analysis. By providing an understanding of the differences and similarities in disparate sets of expert ratings, this analysis helps to establish confidence in the ratings and the criteria for ratings, even when the experts disagree.
Evolution of resistance in vitro reveals a novel mechanism of artemisinin activity in Toxoplasma gondii
Authors: Rozenberg Alex , Luth Madeline R. , Winzeler Elizabeth A. , Behnke Michael , Sibley L. David
Published: August 2019

Artemisinins are effective against a variety of parasites and provide the first line of treatment for malaria. Laboratory studies have identified several mechanisms for artemisinin resistance in Plasmodium falciparum, including mutations in Kelch13 that are associated with delayed clearance in some clinical isolates, although other mechanisms are likely involved. To explore other potential mechanisms of resistance in parasites, we took advantage of the genetic tractability of T. gondii, a related apicomplexan parasite that shows moderate sensitivity to artemisinin. Resistant populations of T. gondii were selected by culture in increasing drug concentrations and whole genome sequencing identified several non-conservative point mutations that emerged in the population and were fixed over time. Genome editing using CRISPR/Cas9 was used to introduce point mutations conferring amino acids changes in a serine protease homologous to DegP and a serine/threonine protein kinase of unknown function. Single and double mutations conferred a competitive advantage over wild type parasites in the presence of drug, despite not changing EC50 values. Additionally, the evolved resistant lines showed dramatic amplification of the mitochondrial genome, including genes encoding cytochrome b and cytochrome oxidase I. Consistent with prior studies in yeast and mammalian tumor cells that implicate the mitochondrion as a target of artemisinins, treatment of wild type parasites with artemisinin decreased mitochondrial membrane potential, and resistant parasites showed altered morphology and decreased membrane potential. These findings extend the repertoire of mutations associated with artemisinin resistance and suggest that the mitochondrion may be an important target of inhibition in T. gondii.

Significance

Artemisinins provide important therapeutic agents for treatment of malaria and have potential for use in other infections and in cancer. Their use is threatened by the potential for resistance development, so understanding their mechanism of action and identifying genetic changes that alter sensitivity are important for improving clinical outcomes. Our findings suggest that mutations in novel targets can contribute to the emergence of parasites with increased tolerance to artemisinin treatment and that such mutations can confer a fitness advantage even in the absence of a notable shift in EC50. Our findings also support the idea that inhibition of mitochondrial function may be an important target in T. gondii, as previously suggested by studies in yeast and human cancer cells.
Phylogenomics investigation of sparids (Teleostei: Spariformes) using high-quality proteomes highlights the importance of taxon sampling
Authors: Natsidis P , Tsakogiannis A , Pavlidis P , Tsigenopoulos CS , Manousaki T
Published: August 2019

Sparidae (Teleostei: Spariformes) are a family of fish constituted by approximately 150 species with high popularity and commercial value, such as porgies and seabreams. Although the phylogeny of this family has been investigated multiple times, its position among other teleost groups remains ambiguous. Most studies have used a single or few genes to decipher the phylogenetic relationships of sparids. Here, we conducted a phylogenomic attempt to resolve the position of the family using five recently available Sparidae gene-sets and 26 available fish proteomes from species with a sequenced genome, to ensure higher quality of the predicted genes. A thorough phylogenomic analysis suggested that Tetraodontiformes (puffer fish, sunfish) are the closest relatives to sparids than all other groups used, a finding that contradicts our previous phylogenomic analysis that proposed the yellow croaker and the european seabass as closest taxa of sparids. By analytically comparing the methodologies applied in both cases, we show that this discordance is not due to different orthology assignment algorithms; on the contrary, we prove that it is caused by the increased taxon sampling of the present study, outlining the great importance of this aspect in phylogenomic analyses in general.
Stochastic model of BKPy Virus replication and assembly
Authors: Stiegelmeyer Suzy M. , Jeffers-Francis Liesl K. , Giddings Morgan C. , Webster-Cyriaque Jennifer
Published: August 2019

BK Polyomavirus (BKPyV), belongs to the same family as SV40 and JC Virus and has recently been associated with both Sjögrens Syndrome and HIV associated Salivary Gland Disease. BKPyV was previously only known for causing the rejection of kidney transplants. As such, BKPyV infection of salivary gland cells implicates oral transmission of the virus. BKPyV replicates slowly in salivary gland cells, producing infectious virus after 72-96 hours. However, it remains unclear how this virus infects or replicates within salivary gland cells, blocking the development of therapeutic strategies to inhibit the virus. Thus, an intracellular, computational model using agent-based modeling was developed to model BKPyV replication within a salivary gland cell. In addition to viral proteins, we modeled host cell machinery that aids transcription, translation and replication of BKPyV. The model has separate cytosolic and nuclear compartments, and represents all large molecules such as proteins, RNAs, and DNA as individual computer “agents” that move and interact within the simulated salivary gland cell environment. An application of the Boids algorithm was implemented to simulate molecular binding and formation of BKPyV virions and BKPyV virus-like particles (VLPs). This approach enables the direct study of spatially complex processes such as BKPyV virus self-assembly, transcription, and translation. This model reinforces experimental results implicating the processes that result in the slow accumulation of viral proteins. It revealed that the slow BKPyV replication rate in salivary gland cells might be explained by capsid subunit accumulation rates. BKPyV particles may only form after large concentrations of capsid subunits have accumulated. In addition, salivary gland specific transcription factors may enable early region transcription of BKPyV.
Enzyme-free synthesis of natural phospholipids in water
Authors: Liu Luping , Zou Yike , Bhattacharya Ahanjit , Zhang Dongyang , Lang Susan Q. , Houk K. N. , Devaraj Neal K.
Published: August 2019

All living organisms synthesize phospholipids as the primary constituent of their cell membranes. While phospholipids can spontaneously self-assemble in water to form membrane-bound vesicles, their aqueous synthesis requires pre-existing membrane-embedded enzymes. This limitation has led to models in which the first cells used simpler types of membrane building blocks and has hampered integration of phospholipid synthesis into artificial cells. Here we demonstrate that a combination of ion pairing and self-assembly of reactants allows high-yielding synthesis of cellular phospholipids in water. Acylation of 2-lysophospholipids using cationic thioesters occurs in mildly alkaline solutions resulting in the formation of cell-like membranes. A variety of membrane-forming natural phospholipids can be synthesized. Membrane formation takes place in water from natural alkaline sources, such as soda lakes and hydrothermal oceanic vents. When formed vesicles are transferred to more acidic solutions, electrochemical proton gradients are spontaneously established and maintained.
Brain wiring and supragranular-enriched genes linked to protracted human frontal cortex development
Authors: Hendy Jasmine P. , Takahashi Emi , van der Kouwe Andre J. , Charvet Christine J.
Published: August 2019

The human frontal cortex is unusually large compared with many other species. The expansion of the human frontal cortex is accompanied by both connectivity and transcriptional changes. Yet, the developmental origins generating variation in frontal cortex circuitry across species remain unresolved. Nineteen genes, which encode filaments, synapse, and voltage-gated channels (e.g., NEFH, SYT2, VAMP1) are especially enriched in the supragranular layers of the cerebral cortex in humans relative to mice. The increased expression of these genes suggests enhanced cortico-cortical projections emerging from layer III in humans. We confirm that the expression of these supragranular-enriched genes is preferentially expressed in frontal cortex layer III in humans relative to mice. We demonstrate a concomitant expansion in cortico-cortical pathways projecting within the frontal cortex white matter in humans with diffusion MR tractography. To identify developmental sources of such variation, we compare frontal cortical white matter growth and developmental trajectories of transcriptional profiles of supragranular-enriched genes in humans and mice. We also use temporal changes in gene expression during postnatal development to control for variation in developmental schedules across species. The growth of the frontal cortex white matter and transcriptional profiles of supragranular genes are both protracted in humans relative to the timing of other transformations. These findings demonstrate that an expansion of projections emerging from the human frontal cortex is achieved by extending the duration of cortical circuitry development. Integrating RNA sequencing with neuroimaging level phenotypes is an effective strategy to assess deviations in developmental programs leading to variation in connections across species.
Practical Aspects of Phylogenetic Network Analysis Using PhyloNet
Authors: Cao Zhen , Liu Xinhao , Ogilvie Huw A. , Yan Zhi , Nakhleh Luay
Published: August 2019

Phylogenetic networks extend trees to enable simultaneous modeling of both vertical and horizontal evolutionary processes. PhyloNet is a software package that has been under constant development for over 10 years and includes a wide array of functionalities for inferring and analyzing phylogenetic networks. These functionalities differ in terms of the input data they require, the criteria and models they employ, and the types of information they allow to infer about the networks beyond their topologies. Furthermore, PhyloNet includes functionalities for simulating synthetic data on phylogenetic networks, quantifying the topological differences between phylogenetic networks, and evaluating evolutionary hypotheses given in the form of phylogenetic networks.

In this paper, we use a simulated data set to illustrate the use of several of PhyloNet’s functionalities and make recommendations on how to analyze data sets and interpret the results when using these functionalities. All inference methods that we illustrate are incomplete lineage sorting (ILS) aware; that is, they account for the potential of ILS in the data while inferring the phylogenetic network. While the models do not include gene duplication and loss, we discuss how the methods can be used to analyze data in the presence of polyploidy.

The concept of species is irrelevant for the computational analyses enabled by PhyloNet in that species-individuals mappings are user-defined. Consequently, none of the functionalities in PhyloNet deals with the task of species delimitation. In this sense, the data being analyzed could come from different individuals within a single species, in which case population structure along with potential gene flow is inferred (assuming the data has sufficient signal), or from different individuals sampled from different species, in which case the species phylogeny is being inferred.
ER reorganization and intracellular retention of CD58 are functionally independent properties of the human cytomegalovirus ER resident glycoprotein UL148
Authors: Nguyen Christopher C. , Domma Anthony J. , Zhang Hongbo , Kamil Jeremy P.
Published: August 2019

The human cytomegalovirus (HCMV) endoplasmic reticulum (ER)-resident glycoprotein UL148 is posited to play roles in immune evasion and regulation of viral cell tropism. UL148 prevents cell surface presentation of the immune cell costimulatory ligand CD58 while promoting maturation and virion incorporation of glycoprotein O, a receptor binding subunit for an envelope glycoprotein complex involved in entry. Meanwhile, UL148 activates the unfolded protein response (UPR) and causes large-scale reorganization of the ER. In an effort to determine whether the seemingly disparate effects of UL148 are related or discrete, we generated charged-cluster-to-alanine (CCTA) mutants of six charged clusters within the UL148 ectodomain, and compared them against wildtype UL148, in the context of recombinant viruses and in ectopic expression, assaying for effects on ER remodeling and CD58 surface presentation. Two mutants, targeting charged clusters spanning residues 79-83 (CC3) and 133-136 (CC4), respectively, retained the potential to impede CD58 presentation, and did so to an extent comparable to wildtype. Of the six mutants, only CC3 retained the capacity to reorganize the ER, showing a partial phenotype. Wildtype UL148 accumulates in a detergent-insoluble form during infection. However, all six CCTA mutants were fully soluble, which may imply a relationship between insolubility and organelle remodeling. Additionally, we found that the chimpanzee cytomegalovirus UL148 homolog suppresses CD58 presentation but fails to reorganize the ER, while the homolog from rhesus cytomegalovirus shows neither activity. Collectively, our findings illustrate varying degrees of functional divergence between homologous primate cytomegalovirus immunevasins and suggest that ER reorganization is unique to HCMV UL148.

IMPORTANCE

In myriad examples, viral gene products cause striking effects on cells, such as activation of stress responses. It can be challenging to decipher how such effects contribute to the biological roles of the proteins. The HCMV glycoprotein UL148 retains CD58 within the ER, thereby preventing it from reaching the cell surface where it functions to stimulate cell-mediated antiviral responses. Intriguingly, UL148 also triggers the formation of large, ER-derived membranous structures, and activates the UPR, a set of signaling pathways involved in adaptation to ER stress. We demonstrate that the potential of UL148 to reorganize the ER and to retain CD58 are separable by mutagenesis and possibly, by evolution, since chimpanzee cytomegalovirus UL148 retains CD58 but does not remodel the ER. Our findings imply that ER reorganization contributes to other roles of UL148, such as modulation of alternative viral glycoprotein complexes that govern the virus’ ability to infect different cell types.
Evolution of linkage and genome expansion in protocells
Authors: Szilágyi András , Péter Kovács Viktor , Szathmáry Eörs , Santos Mauro
Published: August 2019

Chromosomes are likely to have followed unlinked genes in early evolution. Genetic linkage reduces the assortment load and intragenomic conflict in reproducing protocell models to the extent that chromosomes can go to fixation even if chromosomes suffer from a replicative disadvantage, relative to unlinked genes, proportional to their length. Here we show that chromosomes spread within protocells even if recurrent deleterious mutations affecting replicating genes (as ribozymes) are taken into account. Dosage effect selects for optimal genomic composition within protocells that carries over to the genic composition of emerging chromosomes. Lacking an accurate segregation mechanism protocells continue to benefit from the stochastic corrector principle (group selection of early replicators), but now at the chromosome level. A remarkable feature of this process is the appearance of multigene families (in optimal genic proportions) on chromosomes. An added benefit of chromosome formation is an increase in the selectively maintainable genome size (number of different genes), primarily due to the marked reduction of the assortment load. This result complements the established benefit conferred by chromosomes on protocells allowing for the fixation of highly specific and efficient enzymes.
A systematic survey of PRMT interactomes reveals key roles of arginine methylation in the global control of RNA splicing and translation
Authors: Wei Huan-Huan , Fan Xiao-Juan , Hu Yue , Tian Xiao-Xu , Guo Meng , Fang Zhao-Yuan , Wu Ping , Gao Shuai-Xin , Peng Chao , Yang Yun , Wang Zefeng
Published: August 2019

Thousands of proteins undergo arginine methylation, a widespread post-translational modification catalyzed by various protein arginine methyltransferases (PRMTs). However, a full picture of the catalytic network for each PRMT is lacking and the global understanding of their biological roles remains limited. Here we systematically identified interacting proteins for all human PRMTs and demonstrated that they are functionally important for mRNA splicing and translation. We showed that the interactomes of human PRMTs are significantly overlapped with the known methylarginine containing proteins, and different PRMTs are functionally complementary with a high degree of overlap in their substrates and high similarities between their putative methylation motifs. Importantly, arginine methylation is significantly enriched in RNA binding proteins involved in regulating RNA splicing and translation, and inhibition of PRMTs leads to global alteration of alternative splicing and suppression of translation. In particular, ribosomal proteins are pervasively modified with methylarginine, and mutations on their methylation sites suppress ribosome assembly, translation, and eventually cell growth. Collectively, our study provides a novel global view of different PRMT networks and uncovers critical functions of arginine methylation in the regulation of mRNA splicing and translation.
Galactose-modified duocarmycin prodrugs as senolytics
Authors: Guerrero Ana , Guiho Romain , Herranz Nicolás , Uren Anthony , Withers Dominic J. , Martínez-Barbera Juan Pedro , Tietze Lutz F. , Gil Jesús
Published: August 2019

Senescence is a stable growth arrest that impairs the replication of damaged, old or preneoplastic cells, therefore contributing to tissue homeostasis. Senescent cells accumulate during ageing and are associated with diseases, such as cancer, fibrosis and many age-related pathologies. Recent evidence suggests that the selective elimination of senescent cells can be effective on the treatment of many of these senescence-associated diseases. A universal characteristic of senescent cells is that they display elevated activity of the lysosomal β-galactosidase this has been exploited as a marker for senescence (senescence-associated β-galactosidase activity). Consequently, we hypothesised that galactose-modified cytotoxic prodrugs will be preferentially processed by senescent cells, resulting in their selective killing. Here, we show that different galactose-modified duocarmycin (GMD) derivatives preferentially kill senescent cells. GMD prodrugs induce selective apoptosis of senescent cells in a lysosomal β-galactosidase (GLB1)-dependent manner. GMD prodrugs can eliminate a broad range of senescent cells in culture, and treatment with a GMD prodrug enhances the elimination of bystander senescent cells that accumulate upon whole body irradiation or doxorubicin treatment of mice. Moreover, taking advantage of a mouse model of human adamantinomatous craniopharyngioma (ACP), we show that treatment with a GMD pro-drug result selectively reduced the number of β-catenin-positive preneoplastic senescent cells, what could have therapeutic implications. In summary, the above results show that galactose-modified duocarmycin prodrugs behave as senolytics, suggesting that they could be used to treat a wide range of senescence-related pathologies.
Molecular Basis for G-protein-Coupled Receptor (GPCR) Activation and Biased Signalling at the Platelet Thrombin Receptor Proteinase Activated Receptor-4 (PAR4)
Authors: Thibeault Pierre E. , LeSarge Jordan C. , Arends D’Arcy , Fernandes Michaela , Chidiac Peter , Stathopulos Peter B. , Luyt Leonard G. , Ramachandran Rithwik
Published: August 2019

Proteinase Activated Receptor-4 (PAR4) is a member of the proteolytically-activated PAR family of G-Protein-coupled Receptors (GPCRs). PARs are activated following proteolytic cleavage of the receptor N-terminus by enzymes such as thrombin, trypsin, and cathepsin-G to reveal the receptor-activating motif termed the tethered ligand. The tethered ligand binds intramolecularly to the receptor and triggers receptor signalling and cellular responses. In spite of this unusual mechanism of activation, PARs are fundamentally peptide receptors and can also be activated by exogenous application of short synthetic peptides derived from the tethered ligand sequence. In order to gain a better understanding of the molecular basis for PAR4-dependent signalling, we examined signalling responses to a library of peptides derived from the canonical PAR4 activating peptide (PAR4-AP), AYPGKF-NH2. We examined peptide residues involved in activation of the Gαq/11-coupled calcium signalling pathway, β-arrestin recruitment, and mitogen-activated protein kinase pathway activation. The peptideN-methyl-alanine-YPGKF-NH2was identified as a compound that is a poor activator of PAR4-dependent calcium signalling but was fully competent in recruiting β-arrestin-1 and -2. In order to gain a better understanding of the ligand-binding pocket, we usedin silicodocking to identify key residues involved in PAR4 interaction with AYPGKF-NH2. The predicted interactions were verified by site-directed mutagenesis and analysis of calcium signalling and β-arrestin-1/-2 recruitment following proteolytic activation (with thrombin) or activation with the synthetic agonist peptide (AYPGKF-NH2). We determined that a key extracellular loop-2 aspartic acid residue (Asp230) is critical for signalling following both proteolytic and peptide activation of PAR4. Finally, we investigated platelet aggregation in response to AyPGKF-NH2(a peptide with D-tyrosine in position two) which is unable to activate calcium signalling, and AYPGRF-NH2a peptide that is equipotent to the parental peptide AYPGKF-NH2for calcium signalling but is more potent at recruiting β-arrestins. We found that AyPGKF-NH2fails to activate platelets while AYPGRF-NH2causes a platelet aggregation response that is greater than that seen with the parental peptide and is comparable to that seen with thrombin stimulation. Overall, these studies uncover molecular determinants for agonist binding and signalling through a non-canonically activated GPCR and provide a template for development of small molecule modulators of PAR4.
Ongoing repair of migration-coupled DNA damage allows stem cells to reach wound sites
Authors: Sahu Sounak , Sridhar Divya , Abnave Prasad , Kosaka Nobuyoshi , Dattani Anish , Thompson James M. , Hill Mark A. , Aboobaker A. Aziz
Published: August 2019

The impact of mechanical stress during cell migration may be a previously unappreciated source of genome instability [1–3], but to what extent this happens in vivo remains unknown. Here we consider an in vivo system where the adult stem cells of planarian flatworms are required to migrate to a distal wound site [4]. We observe a relationship between adult stem cell migration and ongoing DNA damage and repair during tissue regeneration. Migrating planarian stem cells undergo changes in nuclear shape and increased levels of DNA damage. Increased DNA damage levels resolve once stem cells reach the wound site and stop migrating. Stem cells in which DNA damage is induced prior to wounding take longer to initiate migration suggesting migration activity is sensitive to DNA damage. Migrating stem cells populations are more sensitive to further DNA damage than stationary stem cells, providing evidence that levels of migration-coupled-DNA-damage (MCDD) are significant. RNAi mediated knockdown of DNA repair pathway components blocks normal stem cell migration, confirming that DNA repair pathways are required to allow successful migration to a distal wound site. Together these lines of evidence demonstrate that migration leans to DNA damage in vivo and requires DNA repair mechanisms. Our findings reveal that migration of stem cells represents an unappreciated source of damage, that could be a significant source of mutations in animals during development or during long term tissue homeostasis.
In vitrohepatitis C virus infection and hepatic choline metabolism
Authors: Odai Kaelan Gobeil , O’Dwyer Conor , Steenbergen Rineke , Shaw Tyler A. , Renner Tyler M. , Ghorbani Peyman , Rezaaifar Mojgan , Han Shauna , Langlois Marc-André , Crawley Angela M. , Russell Rodney S. , Pezacki John P. , Tyrrell D. Lorne , Fullerton Morgan D.
Published: August 2019

Choline is an essential nutrient required for normal neuronal and muscular development, as well as homeostatic regulation of hepatic metabolism. In the liver, choline is incorporated into the main eukaryotic phospholipid, phosphatidylcholine (PC) and can enter one carbon metabolism via mitochondrial oxidation. Hepatitis C virus (HCV) is a hepatotropic positive-strand RNA virus that similar to other positive-strand RNA viruses can impact phospholipid metabolism. In the current study we sought to interrogate the link between choline transport and early HCV infection. Namely, we aimed to investigate how HCV modulates markers of choline metabolism followingin vitroinfection, while subsequently assessing how the inhibition of choline uptake and metabolism upon concurrent HCV infection may alter early viral replication. Finally, we assessed whether these parameters were consistent between cells cultured in fetal bovine serum (FBS) or human serum (HS), conditions known to differentially affectin vitroHCV infection. We observed that choline transport in FBS-cultured Huh7.5 cells is facilitated by the intermediate affinity transporter choline transporter-like family (CTL), and that CTL1 expression and the incorporation of choline into PC is diminished in 24 h infected FBS-cultured cells. Reciprocally, limiting the availability of choline for PC synthesis resulted in increased HCV replication at this early stage. In chronically HS-cultured Huh7.5 cells, there were no differences in the expression of choline transporters upon HCV infection or alterations to viral replication when choline transport was inhibited compared to control treatments. However, inhibiting choline uptake and metabolism in this system significantly impaired the production of infectious virions in HS-cultured cells. These results suggest that in addition to a known role of choline kinase, the transport of choline, potentially via CTL1, might also represent an important and regulated process during HCV infection.

Abstract Figure
Interpretable Machine Learning for Perturbation Biology
Authors: Yuan Bo , Shen Ciyue , Luna Augustin , Korkut Anil , Marks Debora S. , Ingraham John , Sander Chris
Published: August 2019

Systematic perturbation of cells followed by comprehensive measurements of molecular and phenotypic responses provides an informative data resource for constructing computational models of cell biology. Models that generalize well beyond training data can be used to identify combinatorial perturbations of potential therapeutic interest. Major challenges for machine learning on large biological datasets are to find global optima in an enormously complex multi-dimensional solution space and to mechanistically interpret the solutions. To address these challenges, we introduce a hybrid approach that combines explicit mathematical models of dynamic cell biological processes with a machine learning framework, implemented in Tensorflow. We tested the modeling framework on a perturbation-response dataset for a melanoma cell line after drug treatments. The models can be efficiently trained to accurately describe cellular behavior, as tested by cross-validation. Even though completely data-driven and independent of prior knowledge, the resultingde novonetwork models recapitulate some known interactions. The main predictive application is the identification of combinatorial candidates for cancer therapy. The approach is readily applicable to a wide range of kinetic models of cell biology.
Reducing module size bias of participation coefficient
Authors: Pedersen Mangor , Omidvarnia Amir , Shine James M. , Jackson Graeme D. , Zalesky Andrew
Published: August 2019

Both natural and engineered networks are often modular. Whether a network node interacts with only nodes from its own module or nodes from multiple modules provides insight into its functional role. The participation coeffcient (PC) is typically used to measure this attribute although its value also depends on the size of the module it belongs to, often leading to non-intuitive identification of highly connected nodes. Here, we develop a normalized PC that overcomes the module size bias associated with the conventional PC. Using brain, C.elegans, airport and simulated networks, we show that our measure of participation alleviates the module size bias, while preserving conceptual and mathematical properties, of the classic formulation of PC. Unlike the conventional PC, we identify London and New York as high participators in the air traffic network and demonstrate stronger associations with working memory in human brain networks, yielding new insights into nodal participation across network modules.
Culture under normoxic conditions and enhanced virulence of phase IICoxiella burnetiitransformed with a RSF1010-based shuttle vector
Authors: Luo Shengdong , He Zemin , Sun Zhihui , Yu Yonghui , Jiang Yongqiang , Tong Yigang , Song Lihua
Published: August 2019

Coxiella burnetiiis a Gram-negative, facultative intracellular microorganism that can cause acute or chronic Q fever in human. It was recognized as an obligate intracellular organism until the revolutionary design of an axenic cystine culture medium (ACCM). Present axenic culture ofC. burnetiistrictly requires a hypoxic condition (<10% oxygen). Here we investigated the normoxic growth ofC. burnetiistrains in ACCM-2 with or without tryptophan supplementation. ThreeC. burnetiistrains - Henzerling phase I, Nine Mile phase II and a Nine Mile phase II transformant, were included. The transformant contains a pMMGK plasmid that is composed of a RSF1010 ori, a repABC operon, an eGFP gene and a kanamycin resistance cassette. We found that, under normoxia if staring from an appropriate concentration of fresh age inocula, Nine Mile phase II can grow significantly in ACCM-2 with tryptophan, while the transformant can grow robustly in ACCM-2 with or without tryptophan. In contrast, long-term frozen stocks of phase II and its transformant, and Henzerling phase I of different ages had no growth capability under normoxia under any circumstances. Furthermore, frozen stocks of the transformant consistently caused large splenomegaly in SCID mice, while wild type Nine Mile phase II induced a lesser extent of splenomegaly. Taken together, our data show that normoxic cultivation of phase IIC. burnetiican be achieved under certain conditions. Our data suggests that tryptophan and an unknown temperature sensitive signal are involved in the expression of genes for normoxic growth regulated by quorum sensing inC. burnetii.
Free-ranging dogs are capable of comprehending complex human pointing cues
Authors: Bhattacharjee Debottam , Mandal Sarab , Shit Piuli , Varghese Mebin George , Vishnoi Aayushi , Bhadra Anindita
Published: August 2019

Dogs are the most common species to be found as pets and have been subjects of human curiosity leading to extensive research on their socialization with humans. One of the dominant themes in dog cognition pertains to their capacity of understanding and responding to human referential gestures. The remarkable socio-cognitive skills of pet dogs, while interacting with humans, is quite well established. However, studies regarding the free-ranging subpopulations are greatly lacking. Free-ranging dogs represent an ideal system to investigate interspecific communication with unfamiliar humans, nullifying any contribution of indirect conditioning. The interactions of these dogs with humans are quite complex and multidimensional. For the first time, we tested free-ranging dogs’ ability to understand relatively complex human referential gestures using dynamic and momentary distal pointing cues. We found that these dogs are capable of apprehending distal pointing cues from humans. However, approximately half of the population tested showed a lack of tendency to participate even after successful familiarization with the experimental set-up. A closer inspection revealed anxious behavioural states of the individuals were responsible for such an outcome. We assume that life experiences with humans probably shape personalities of free-ranging dogs, which in turn influence their responsiveness to human communicative gestures.
Bacteria- and temperature-regulated peptides modulate beta-catenin signaling inHydra
Authors: Taubenheim J , Willoweit-Ohl D , Knop M , Franzenburg S , He J , Bosch TCG , Fraune S
Published: August 2019

Animal development has traditionally been viewed as an autonomous process directed by the host genome. But in many animals biotic and abiotic cues, like temperature and bacterial colonizers, provide signals for multiple developmental steps.Hydraoffers unique features to encode these complex interactions of developmental processes with biotic and abiotic factors. Here, we used the model animalHydrato investigate the impact of bacterial colonizers and temperature on the pattern formation process. InHydra, formation of the head organizer involves the canonical Wnt pathway. Treatment with alsterpaullone (ALP) results in acquiring characteristics of the head organizer in the body column. Intriguingly, germ-freeHydrapolyps are significantly more sensitive to ALP compared to control polyps. In addition to microbes, β-catenin dependent pattern formation is also affected by temperature. Gene expression analyses led to the identification of two small secreted peptides, named Eco1 and Eco2, being upregulated in the response to both,Curvibactersp, the main bacterial colonizer ofHydra, and low temperatures. Loss-of function experiments revealed that Eco peptides are involved in the regulation of pattern formation and have an antagonistic function to Wnt signaling inHydra.
Extracellular electron uptake inMethanosarcinalesis independent of multihemec-type cytochromes
Authors: Yee Mon Oo , Rotaru Amelia-Elena
Published: August 2019

The co-occurrence ofGeobacterandMethanosarcinalesis often used as a proxy for the manifestation of direct interspecies electron transfer (DIET) in man-made and natural aquatic environments. We previously reported that not allGeobacterare capable of DIET withMethanosarcina. Here we tested 15 new artificial co-culture combinations with methanogens and electrogenic bacteria, including an electrogen outside of theGeobacterclade –Rhodoferax ferrireducens.

Consistently, highly effective electrogenic bacteria (G. metallireducens, G. hydrogenophilusandR. ferrireducens) formed successful associations withMethanosarcinales. Highly effective electrogens could not sustain the growth of H2-utilizing methanogens of the generaMethanococcus,Methanobacterium, Methanospirillum, MethanolaciniaorMethanoculleus.

Methanosarcinales, including strict non-hydrogenotrophic methanogens of the genusMethanothrix (Mtx. harundinaceaandMtx. shoeghenii) andMethanosarcina horonobensis, conserved their ability to interact with electrogens.Methanosarcinaleswere classified as the only methanogens containingc-type cytochromes, unlike strict hydrogenotrophic methanogens. It was then hypothesized that multiheme c-type cytochromes giveMethanosarcinalestheir ability to retrieve extracellular electrons. However, multiheme c-type cytochromes are neither unique to this group of methanogens nor universal. Only two of the sevenMethanosarcinalestested had multiheme c-type cytochromes (MCH). In one of these two species -M. mazeia deletion mutant for its MCH was readily available. Here we tested if the absence of this MHC impacts extracellular electron uptake. Deletion of the MHC inM. mazeidid not impact the ability of this methanogens to retrieve extracellular electrons fromG. metallireducensor a poised cathode. SinceMethanosarcinadid not require multiheme c-type cytochromes for direct electron uptake we proposed an alternative strategy for extracellular electron uptake.
Dormancy-to-death transition in yeast spores occurs due to gradual loss of gene-expressing ability
Authors: Maire Théo , Allertz Tim , Betjes Max A. , Youk Hyun
Published: August 2019

Dormancy is colloquially considered as extending lifespan by being still. Starved yeasts form dormant spores that wake-up (germinate) when nutrients reappear but cannot germinate (die) after some time. What sets their lifespans and how they age are open questions because what processes occur - and by how much - within each dormant spore remains unclear. With single-cell-level measurements, we discovered how dormant yeast spores age and die: spores have a quantifiable gene-expressing ability during dormancy that decreases over days to months until it vanishes, causing death. Specifically, each spore has a different probability of germinating that decreases because its ability to - without nutrients - express genes decreases, as revealed by a synthetic circuit that forces GFP expression during dormancy. Decreasing amounts of molecules required for gene expression - including RNA polymerases - decreases gene-expressing ability which then decreases chances of germinating. Spores gradually lose these molecules because they are produced too slowly compared to their degradations, causing gene-expressing ability to eventually vanish and, thus, death. Our work provides a systems-level view of dormancy-to-death transition.

Short summary

This study identifies systems-level quantities that decay during dormancy in Saccharomyces cerevisiae spores and thereby reveals the meaning of ageing for dormant yeast spores and shows that they die when their gene-expressing ability is irreversibly lost.

Highlights

For a given glucose concentration, a dormant yeast spore has a well-defined probability of germinating (“germination ability”).

A spore’s germination ability positively correlates with its RNAP I-III levels and the gene-expression (GFP) level it can realize when the expression is forced without nutrients.

Ageing during dormancy means gradual decreases in germination ability, RNAP levels, and GFP-level realizable when expression is forced.

Spores die after sufficiently losing gene-expressing ability and drugs that inhibit gene expression during dormancy shorten spores’ lifespans (e.g., from months to a day).