title: A new sugar for an old phage: A c-di-GMP dependent polysaccharide pathway sensitizes E. coli for bacteriophage infection

abstract:
Bacteriophages are ubiquitous parasites of bacteria and major drivers of bacterial ecology and evolution. Despite an ever-growing interest in their biotechnological and therapeutic applications, detailed knowledge of the molecular mechanisms underlying phage-host interactions remains scarce. Here, we show that bacteriophage N4 exploits a novel surface glycan, NGR, as a receptor to infect its host Escherichia coli. We demonstrate that this process is regulated by the second messenger c-di-GMP and that N4 infection is specifically stimulated by the diguanylate cyclase DgcJ while the phosphodiesterase PdeL effectively protects E. coli from N4-mediated killing. PdeL-mediated protection requires its catalytic activity to reduce c-di-GMP and includes a secondary role as a transcriptional repressor. We demonstrate that PdeL binds to and represses the promoter of the wec operon, which encodes components of the ECA exopolysaccharide pathway. However, only the acetylglucosamine epimerase WecB but none of the other ECA components is required for N4 infection. Based on this, we postulate that NGR is an N-acetylmannosamine-based carbohydrate polymer that is produced and exported to the cell surface of E. coli in a c-di-GMP dependent manner where it serves as a receptor for N4. This novel carbohydrate pathway is conserved in E. coli and other bacterial pathogens, serves as the primary receptor for a range of N4-like bacteriophages, and is induced at elevated temperature and by specific amino acid-based nutrients. These studies provide an entry point into understanding how bacteria use specific regulatory mechanisms to balance costs and benefits of highly conserved surface structures.

ImportanceBecause bacterial surface glycans are in direct contact with the environment they can provide essential protective functions during infections or against competing bacteria. But such structures are also  Achilles heels as they can serve as primary receptors for bacteriophages. Bacteria thus need to carefully control the exposure of conserved surface glycans to balance costs and benefits. Here, we identify a novel exopolysaccharide that is widely conserved in E. coli and is used by N4 and related bacteriophages as primary receptor. We demonstrate that the synthesis of NGR (N4 glycan receptor) is tightly controlled by the second messenger c-di-GMP in a highly specific manner and by a single diguanylate cyclase. These studies provide an example of how bacteria can alleviate the strong selective pressure imposed on them by bacteriophages entering through conserved surface structures by carefully regulating their synthesis and secretion.

category: microbiology


title: Evolutionary dynamics of Begomoviruses causing Papaya leaf curl disease in India

abstract: The genus begomovirus represents a group of multipartite viruses that significantly damages many agricultural crops, including papaya and drastically influence the overall production. Understanding the genetic variations, mutations and recombination of the begomovirus population infecting papaya has several important implications in alleviating substantial losses, mainly in developing countries, including India. In an attempt for a bioinformatics driven study of diversity and variability of papaya leaf curl disease in India, a total of thirty-two (32) DNA-A and sixteen (16) betasatellite sequences were retrieved from GenBank. An uneven distribution of evolutionary divergence has been observed across the branch length, which triggered the estimated recombinational event. Interestingly, a maximum of the begomoviruses were found to be intra-species recombinants. Further genetic variability, selection pressure, and substitution rate acting on the population were estimated and found to be high enough to support the evolution of geminiviruses. Genetic divergence composition in all begomovirus datasets revealed predominance of nucleotide diversity driven by mutation. The analysis indicates that even though a significant fraction of the genetic variations might be due to recombination but, it was constantly lower than the mutation rate. Thus, the diversification of the begomovirus population is principally impelled by mutational dynamics.

category: pathology


title: Impact of maternal intermittent fasting during pregnancy on cardiovascular, metabolic and renal function in adult rat offspring

abstract: Pregnant Muslim women are exempt from fasting during Ramadan; however a majority are reported to fast. The impact of this form of maternal intermittent fasting (IF) on fetal development and offspring health is not well defined. Using a rat model, we have shown previously that maternal IF results in fetal growth restriction accompanied by changes in placental nutrient transport function. The aim of this study was to assess cardiovascular, metabolic and renal function in adult offspring of IF-exposed dams. Food was withheld from Wistar rats from 17:00 to 09:00 daily throughout pregnancy; controls had ad libitum access to food. Birth weight was unaffected; however male IF pups grew more slowly up to 10 weeks of age (P < 0.01) whereas IF females matched their control counterparts. Systolic blood pressure (SBP), glucose tolerance and basal renal function at 14 weeks were not affected by IF exposure. When offered saline solutions (0.9-2.1%) to drink, females showed a greater salt preference than males (P < 0.01); however there were no differences between dietary groups. A separate group of pups was weaned onto a 4% NaCl diet. SBP increased in IF pups sooner, at 7 weeks (P < 0.01), than controls which became hypertensive from 10 weeks. Renal function did not appear to differ; however markers of renal injury were elevated in IF males (P < 0.05). Maternal IF does not affect resting cardiovascular, metabolic and renal function; but when challenged by dietary salt load male IF offspring are more prone to renal injury.

category: physiology


title: Multiple metals influence distinct properties of the Arabidopsis circadian clock

abstract: Circadian rhythms coordinate endogenous events with external signals, and are essential to biological function. When environmental contaminants affect these rhythms, the organism may experience fitness consequences such as reduced growth or increased susceptibility to pathogens. In their natural environment plants may be exposed to a wide range of industrial and agricultural pollutants. Here, we investigate how the addition of various metal salts to the environment can impact plant-circadian rhythms, via the promoter:luciferase system. The consequences of these environmental changes were found to be varied and complex. Therefore, in addition to Fourier-based analyses, we apply novel wavelet-based spectral hypothesis testing and clustering methodologies to organize and understand the data. We are able to classify broad sets of responses to environmental contaminants, including pollutants which increase, or decrease, the period, or which induce a lack of precision or disrupt any meaningful periodicity. The methods are general, and may be applied to discover common responses and hidden structures within a wide range of biological time series data.

category: plant biology


title: The exocyst complex and intracellular vesicles mediate soluble protein trafficking to the primary cilium

abstract: The efficient transport of proteins into the primary cilium is a crucial step for many signaling pathways. Dysfunction of this process can lead to the disruption of signaling cascades or cilium assembly, resulting in developmental disorders and cancer. Previous studies on the protein delivery to the cilium were mostly focused on the membrane-embedded receptors. In contrast, how soluble proteins are delivered into the cilium is poorly understood. In our work, we identify the exocyst complex as a key player in the ciliary trafficking of soluble Gli transcription factors. In line with the known function of the exocyst in intracellular vesicle transport, we demonstrate that soluble proteins, including Gli2/3 and Lkb1, can use the endosome recycling machinery for their delivery to the primary cilium. Finally, we identify GTPases: Rab14, Rab18, Rab23, and Arf4 that are involved in vesicle-mediated Gli protein ciliary trafficking. Our data pave the way for a better understanding of ciliary transport and uncover novel transport mechanisms inside the cell.

category: cell biology


title: Intraspecific variation stabilizes classic predator-prey dynamics

abstract: In 1920, Alfred J. Lotka found that, to his "considerable surprise", the dynamics of a simple predatorprey model he had devised led "to undamped, and hence indefinitely continued, oscillations" 1,2-- which he thought epitomized the "rhythm of Nature" dear to the Victorians. In 1926, the same model was proposed independently by mathematician Vito Volterra 3,4, who was inspired by the work of his son-in-law, fish biologist Umberto DAncona 5. For over a century, the equations that now bear their names have served as a template for the development of sophisticated models for population dynamics 6-10. Coexistence in this classic predator-prey model is fragile--stochasticity or temporal variability in parameter values result in extinctions. The dynamics can be stabilized by intraspecific competition or other forms of self-regulation, but the prevalence of these processes in large food webs has been questioned 11,12. Here we show that when we consider populations characterized by intraspecific variability, dynamics are stable--despite the absence of any direct self-regulation. Our results can be generalized further, defining a new class of consumer-resource models 8,13. By accounting for intraspecific variation, which is manifest in all biological populations, we obtain dynamics that differ qualitatively and quantitatively from those found for homogeneous populations--challenging a central assumption of many ecological models.

category: ecology


title: Community assessment of crustose calcifying red algae as coral recruitment substrates

abstract: Successful recruitment of invertebrate larvae to reef substrates is essential to the health of tropical coral reef ecosystems and their capacity to recover from disturbances. Crustose calcifying red algae (CCRA) have been identified as important recruitment substrates for scleractinian corals. As such, CCRA as a whole or subgroups (e.g., crustose coralline algae, CCA) are often used at the functional group level in experimental, ecological, and monitoring studies. Species of CCRA, however, differ in their ecological roles and their value as coral recruitment substrates. Here, we (1) investigate the species richness and community composition of CCRA on experimental coral recruitment tiles, and (2) assess if there is a recruitment preference of the coral Acropora surculosa for any of these CCRA species. 27 species of two orders of CCRA (Corallinales and Peyssonneliales) were identified from the recruit tiles. None of the DNA sequences of these species matched released sequences in GenBank or sequences of CCRA collected from natural reef systems in Guam. The similarity in CCRA communities between the recruitment tiles was high. Two species of CCRA were significantly preferred as recruitment substrates over the other CCRA species. Both of these species belonged to the subfamily of the Lithophylloideae. These two species are closely related to Pacific species that have been referred to as Titanoderma -but probably have to be assigned to another genus- and many of the latter have been attributed to be preferred coral recruitment substrates. Of all CCRA, Lithophylloideae sp. 1 had the highest benthic cover on the recruitment tiles and was the most preferred recruitment substrate. These findings highlight the high taxonomic diversity of CCRA communities and provide insight into species-specific ecological roles of CCRA that are often overlooked.


title: Generative Adversarial Networks (GAN) for the simulation of central-place foraging trajectories

abstract:
AO_SCPLOWBSTRACTC_SCPLOWO_LIMiniature electronic device such as GPS have enabled ecologists to document relatively large amount of animal trajectories. Modeling such trajectories may attempt (1) to explain mechanisms underlying observed behaviors and (2) to elucidate ecological processes at the population scale by simulating multiple trajectories. Existing approaches to animal movement modeling mainly addressed the first objective and they are yet soon limited when used for simulation. Individual-based models based on ad-hoc formulation and empirical parametrization lack of generability, while state-space models and stochastic differential equations models, based on rigorous statistical inference, consist in 1st order Markovian models calibrated at the local scale which can lead to overly simplistic description of trajectories.
C_LIO_LIWe introduce a  state-of-the-art tool from artificial intelligence - Generative Adversarial Networks (GAN) - for the simulation of animal trajectories. GAN consist in a pair of deep neural networks that aim at capturing the data distribution of some experimental dataset, and that enable the generation of new instances of data that share statistical similarity. In this study, we aim on one hand to identify relevant deep networks architecture for simulating central-place foraging trajectories and on the second hand to evaluate GAN benefits over classical methods such as state-switching Hidden Markov Models (HMM).
C_LIO_LIWe demonstrate the outstanding ability of GAN to simulate  realistic seabirds foraging trajectories. In particular, we show that deep convolutional networks are more efficient than LSTM networks and that GAN-derived synthetic trajectories reproduce better the Fourier spectral density of observed trajectories than those simulated using HMM. Therefore, unlike HMM, GAN capture the variability of large-scale descriptive statistics such as foraging trips distance, duration and tortuosity.
C_LIO_LIGAN offer a relevant alternative to existing approaches to modeling animal movement since it is calibrated to reproduce multiple scales at the same time, thus freeing ecologists from the assumption of first-order markovianity. GAN also provide an ultra-flexible and robust framework that could further take environmental conditions, social interactions or even bio-energetics model into account and tackle a wide range of key challenges in movement ecology.
C_LI


title: The Clock:cycle complex is a major transcriptional regulator of Drosophila photoreceptors that protects the eye from retinal degeneration and oxidative stress

abstract: The aging eye experiences physiological changes that include decreased visual function and increased risk of retinal degeneration. Although there are transcriptomic signatures in the aging retina that correlate with these physiological changes, the gene regulatory mechanisms that contribute to cellular homeostasis during aging remain to be determined. Here, we integrated ATAC-seq and RNA-seq data to identify 61 transcription factors that showed differential activity in aging Drosophila photoreceptors. These 61 age-regulated transcription factors include two circadian regulators, Clock and cycle, that showed sustained increases in activity during aging. When we disrupted Clock activity in adult photoreceptors, we observed changes in expression of 15 - 20% of genes including key components of the phototransduction machinery and many eye-specific transcription factors. Using ATAC-seq, we showed that loss of Clock activity leads to changes in activity of 31 transcription factors and causes a progressive decrease in global levels of chromatin accessibility in photoreceptors. Supporting a key role for Clock-dependent transcription in the eye, disruption of Clock activity in photoreceptors also induced light-dependent retinal degeneration and increased oxidative stress, independent of light exposure. Together, our data suggests that the circadian regulators Clock and cycle act as neuroprotective factors in the aging eye by directing gene regulatory networks that maintain expression of the phototransduction machinery and counteract oxidative stress.

category: genomics


title: Polygenic enrichment distinguishes disease associations of individual cells in single-cell RNA-seq data

abstract: Gene expression at the individual cell-level resolution, as quantified by single-cell RNA-sequencing (scRNA-seq), can provide unique insights into the pathology and cellular origin of diseases and complex traits. Here, we introduce single-cell Disease Relevance Score (scDRS), an approach that links scRNA-seq with polygenic risk of disease at individual cell resolution without the need for annotation of individual cells to cell types; scDRS identifies individual cells that show excess expression levels for genes in a disease-specific gene set constructed from GWAS data. We determined via simulations that scDRS is well-calibrated and powerful in identifying individual cells associated to disease. We applied scDRS to GWAS data from 74 diseases and complex traits (average N =346K) in conjunction with 16 scRNA-seq data sets spanning 1.3 million cells from 31 tissues and organs. At the cell type level, scDRS broadly recapitulated known links between classical cell types and disease, and also produced novel biologically plausible findings. At the individual cell level, scDRS identified subpopulations of disease-associated cells that are not captured by existing cell type labels, including subpopulations of CD4+ T cells associated with inflammatory bowel disease, partially characterized by their effector-like states; subpopulations of hippocampal CA1 pyramidal neurons associated with schizophrenia, partially characterized by their spatial location at the proximal part of the hippocampal CA1 region; and subpopulations of hepatocytes associated with triglyceride levels, partially characterized by their higher ploidy levels. At the gene level, we determined that genes whose expression across individual cells was correlated with the scDRS score (thus reflecting co-expression with GWAS disease genes) were strongly enriched for gold-standard drug target and Mendelian disease genes.

category: bioinformatics


title: Gene Expression Analysis of Glioma Neural Stem Cells Shows Disturbed Amino Acids Metabolism and Axonal Growth Cone Dynamics in Glioblastoma Multiforme.

abstract: Glioblastoma multiforme(GBM) is a group of fatal and aggressive tumors of the central nervous system. Despite advancements in the treatment of GBM, patients diagnosed with these tumors typically have a poor prognosis and poor quality of life as the disease develops. The single-cell RNA high-throughput sequencing processed data for Glioma cancer stem cells were taken from GEO and analyzed to find out the underlying expression differences at the gene level between glioma neural stem cells(GSCs) and Normal neural stem cells(NSCs). In the current study, we have performed an RNA-sequencing analysis between GSCs and NSCs to better understand the origin of GBM. We have performed bioinformatics analysis on the transcriptional profile of 134 samples which consisted of 75 GSCs and 59 NSCs obtained from the NCBI bio project(PRJNA546254). First, an exploratory analysis was performed which showed significant variation patterns between GSCs and NSCs. Subsequently, Deseq2 differential gene expression analysis identified 1436 differentially expressed genes between GSCs and NSCs[(padj. value <0.05, log2 fold change (>=+/-1.5)]. This study reveals genes like MAOA, MAOB, GATM, GLDC, AMT, and SHMT1 as the key features contributing to the disturbed processes of Glycine, threonine, and serine amino acid metabolism, axonal cone growth curve, and cell migration in Glioma. Conclusively, our study also depicts gene expression changes in amyloid beta-binding protein in between GSCs and NSCs which plays an important role in tumor microenvironment formation. Besides, the results presented here reveal new insight into the progression of GBM and the identification of novel genes involved in gliomagenesis.

category: cancer biology


title: Modeling tissue-specific breakpoint proximity of structural variations from 2,382 whole-genomes to identify cancer drivers

abstract: Structural variations (SVs) in cancer cells often impact large genomic regions with functional consequences. However, little is known about the genomic features related to the breakpoint distribution of SVs in different cancers, a prerequisite to distinguish loci under positive selection from those with neutral evolution. We developed a method that uses a generalized additive model to investigate the breakpoint proximity curves from 2,382 whole-genomes of 32 cancer types. We find that a multivariate model, which includes linear and nonlinear partial contributions of various tissue-specific features and their interaction terms, can explain up to 57% of the observed deviance of breakpoint proximity. In particular, three-dimensional genomic features such as topologically associating domains (TADs), TAD-boundaries and their interaction with other features show significant contributions. The model is validated by identification of known cancer genes and revealed putative drivers in novel cancers that have previous evidence of therapeutic relevance in other cancers.


title: Spatiotemporal co-dependency between macrophages and exhausted CD8+ T cells in cancer

abstract:
T cell exhaustion is a major impediment to anti-tumor immunity. However, it remains elusive how other immune cells in the tumor microenvironment (TME) contribute to this dysfunctional state. Here we show that the biology of tumor-associated macrophages (TAM) and exhausted T cells (Tex) in the TME is extensively linked. We demonstrate that in vivo depletion of TAM reduces exhaustion programs in tumor-infiltrating CD8+ T cells and reinvigorates their effector potential. Reciprocally, transcriptional and epigenetic profiling reveals that Tex express factors that actively recruit monocytes to the TME and shape their differentiation. Using lattice light sheet microscopy, we show that TAM and CD8+ T cells engage in unique long-lasting antigen-specific synaptic interactions that fail to activate T cells, but prime them for exhaustion, which is then accelerated in hypoxic conditions. Spatially resolved sequencing supports a spatiotemporal self-enforcing positive feedback circuit that is aligned to protect rather than destroy a tumor.

Graphical Abstract

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title: Nanopore sequencing methods detect cell-free DNA associated with minimal residual disease and central nervous system infiltration in pediatric Acute Lymphoblastic Leukemia

abstract:
Acute Lymphoblastic Leukemia (ALL) patients that are positive for minimal residual disease (MRD) after therapy or have leukemic infiltration into the central nervous system (CNS) are considered high risk and receive intensive chemotherapy regimens. Current methods to diagnose MRD and CNS infiltration rely on detecting leukemic cells in patient samples using pathology, flow cytometry, or next-generation sequencing. However, leukemic blasts may persist in the patient but not be physically present in bone marrow biopsy or biofluid sample, leading to inaccurate or delayed patient diagnosis. We have developed a nanopore sequencing workflow to detect B-ALL-associated cell-free DNA (cfDNA) in blood and cerebrospinal fluid (CSF) samples. Quantitation of B-cell specific VDJ recombination events in cfDNA samples defined B-ALL clonal heterogeneity. This workflow allowed us to track the response of individual B-ALL clones throughout treatment. Detection of cfDNA also predicted the clinical diagnosis of MRD and CNS disease. Importantly, we identified patients diagnosed as CNS negative who had low B-cell derived cell-free DNA levels in their CSF sample that correlated with B-cell clones present in the bone marrow. These data suggest that cfDNA assays may be useful in detecting the presence of ALL in the patient even when blasts are not in the biofluid sample. Nanopore analysis of cell-free DNA is a simple, rapid, and inexpensive assay that can serve as a valuable complement to traditional clinical diagnostic approaches for ALL.

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title: Midkine noncanonically suppresses AMPK activation through disrupting the LKB1-STRAD-Mo25 complex

abstract: Midkine (MDK), an extracellular growth factor, regulates signal transduction and cancer progression by interacting with receptors, and it can be internalized into the cytoplasm by endocytosis. However, its intracellular function and signaling regulation remain unclear. Here, we show that intracellular MDK interacts with LKB1 and STRAD to disrupt the LKB1-STRAD-Mo25 complex. Consequently, MDK decreases the activity of LKB1 to dampen both the basal and stress-induced activation of AMPK by glucose starvation or treatment of 2-DG. We also found that MDK accelerates cancer cell proliferation by inhibiting the activation of the LKB1-AMPK axis. In human cancers, compared to other well-known growth factors, MDK expression is most significantly upregulated in cancers, especially in liver, kidney and breast cancers, correlating with clinical outcomes and inversely correlating with PRKAA1 (encoding AMPK1) expression and phosphorylated AMPK levels. Our study elucidates an inhibitory mechanism for AMPK activation, which is mediated by the intracellular MDK through disrupting the LKB1-STRAD-Mo25 complex.


title: Reconstructing Sample-Specific Networks using LIONESS

abstract: We recently developed LIONESS, a method to estimate sample-specific networks based on the output of an aggregate network reconstruction approach. In this manuscript, we describe how to apply LIONESS to different network reconstruction algorithms and data types. We highlight how decisions related to data preprocessing may affect the output networks, discuss expected outcomes, and give examples of how to analyze and compare single sample networks.

category: systems biology


title: Overcoming the effect of ASH1L haploinsufficiency on stem cells amenability to genome editing and differentiation into the neuronal lineage - a technical report

abstract: Genome editing and neuronal differentiation protocols have proliferated in the last decade. Mutations in genes that control pluripotency could lead to a potential obstacle with regards to the survival and differentiation potential of the genome-edited cell lines. Here we describe a protocol for the generation, and differentiation, of cell lines containing CRISPR/Cas9 induced mutations in the histone methyl transferase ASH1L. This chromatin modifier was previously implicated in hematopoietic stem cell pluripotency and is a major genetic risk factor for autism spectrum disorders (ASD). We find that haploinsufficiency of ASH1L leads to decreased NANOG gene expression leading to reduce cell survival and increased spontaneous differentiation. We report a method that provides improved single-cell survival with higher colony formation efficiency in ASH1L mutant stem cells. Additionally, we describe a modified dual-SMAD inhibition neuronal induction methodology that permits the successful generation of human neurons with mutations in ASH1L, in a smaller scale than previously reported methods. With our modified CRISPR-genome editing and neuronal differentiation protocols, it is possible to generate genome-edited stem cells containing mutations in genes that impact pluripotency and could affect subsequent cell lineage specific differentiation. Our detailed technical report presents cost-effective strategies that will benefit researchers focusing on both translational and basic science using stem cell experimental systems.

category: neuroscience


title: Towards optimization of oscillatory stimulation during sleep

abstract:
BackgroundOscillatory rhythms during sleep such as slow oscillations (SO) and spindles, and most importantly their coupling, are thought to underlie processes of memory consolidation. External slow oscillatory transcranial direct current stimulation (so-tDCS) with a frequency of 0.75 Hz has been shown to improve this coupling and memory consolidation, however, effects varied quite markedly between individuals, studies, and species. Here, we aimed to determine how precisely the frequency of stimulation has to match the naturally occurring SO frequency in individuals to optimally improve SO-spindle coupling. Moreover, we systematically tested stimulation durations necessary to induce changes.

MethodsWe addressed these questions by comparing so-tDCS with individualized frequency to standardized frequency of 0.75 Hz in a within-subject design with 28 older participants during napping while systematically varying stimulation train durations between 30 s, 2 min, and 5 min.

ResultsStimulation trains as short as 30 s were sufficient to modulate the coupling between SOs and spindle activity. Contrary to our expectations, so-tDCS with standardized frequency indicated stronger aftereffects with regard to SO-spindle coupling compared to individualized frequency. Angle and variance of spindle maxima occurrence during the SO cycle were similarly modulated.

ConclusionIn sum, short stimulation trains were sufficient to induce significant changes in sleep physiology allowing for more trains of stimulation, which provides methodological advantages and possibly even larger behavioral effects in future studies. With regard to individualized stimulation frequency, further options of optimization need to be investigated, such as closed-loop stimulation to calibrate stimulation frequency to the SO frequency at time of stimulation onset.

Significance statementApplication of slow oscillatory transcranial direct current stimulation during sleep has been shown to enhance specific memory-relevant sleep parameters and memory performance after sleep, albeit with a high degree of variability. Here, we systematically explored two major stimulation parameters possibly accounting for this variability in humans: frequency and duration of stimulation. We found, contrary to our expectations, standardized frequency stimulation with 0.75 Hz being superior to individualized frequency stimulation in enhancing specific sleep parameters. Moreover short stimulation trains of 30 seconds were as effective as 5 min in modulating aftereffects. These are encouraging findings, implying methodological advantages as larger quantity of aftereffects data can be obtained within the same time window, which may also lead to enhanced behavioral stimulation effects.


title: The activities of LRRK2 and GCase are positively correlated in clinical biospecimens and experimental models of Parkinson's disease

abstract: LRRK2 is a kinase involved in different cellular functions, including autophagy, endolysosomal pathways and vesicle trafficking. Mutations in LRRK2 cause autosomal dominant forms of Parkinsons disease (PD). Heterozygous mutations in GBA1, the gene encoding the lysosomal enzyme glucocerebrosidase (GCase), are the most common genetic risk factors for PD. Moreover, GCase function is altered in idiopathic PD and in other genetic forms of the disease. Recent work suggests that LRRK2 kinase activity can regulate GCase function. However, both a positive and a negative correlation have been described. To gain insights into the impact of LRRK2 on GCase, we investigated GCase levels and activity in LRRK2 G2019S knockin mice, in clinical biospecimens from PD patients carrying this mutation and in patient-derived cellular models. In these models we found a positive correlation between the activities of LRRK2 and GCase, which was further confirmed in cell lines with genetic and pharmacological manipulation of LRRK2 kinase activity. Overall, our study indicates that LRRK2 kinase activity affects both the levels and the catalytic activity of GCase.


title: Interactions of cytosolic termini of the Jen1 monocarboxylate transporter are critical for trafficking, transport activity and endocytosis

abstract: Plasma membrane (PM) transporters of the major facilitator superfamily (MFS) are essential for cell metabolism and growth, as well as for survival in response to stress or cytotoxic drugs, in both prokaryotes and eukaryotes. In the yeast Saccharomyces cerevisiae, Jen1 is a monocarboxylate/H+ symporter that has been used to dissect the molecular details underlying control of cellular expression, transport mechanism and turnover of MFS transporters. Here, we present evidence supporting previously non-described roles of the cytosolic N- and C- termini in Jen1 biogenesis, PM stability and activity, through functional analyses of rationally designed truncations and chimeric constructs with UapA, a S. cerevisiae endocytosis-insensitive purine transporter from Aspergillus nidulans. Our results reveal a cryptic role of the N-terminal region and thus show that both cytosolic N- and C-termini are critical for Jen1 trafficking to the PM, transport activity and endocytosis. In particular, we provide evidence that the N- and the C-cytosolic termini of Jen1 undergo transport-dependent dynamic intra-molecular interactions, which critically affect the mechanism of transport and turnover of Jen1. Our results support an emerging concept where the cytosolic tails of PM transporters control transporter expression and function, through flexible intra-molecular interactions with each other and the transmembrane core of the protein. This idea may be extended to other MFS members providing a deeper understanding of conserved, but also evolving, mechanisms underlying MFS transporter structure-function relationships.


title: Large-scale mapping of positional changes of hypoxia-responsive genes upon activation

abstract: Chromosome structure and nuclear organization are important factors in the regulation of gene expression. Transcription of a gene is influenced by local and global chromosome features such as condensation status and histone modifications. The relationship between the position of a gene in the cell nucleus and its activity is less clear. Here, we used high-throughput imaging to perform a large-scale analysis of the spatial location of a set of nearly 100 hypoxia-inducible genes to determine whether their location within the nucleus is correlated with their activity state upon stimulation. Radial distance analysis demonstrated that the majority of HIF- and CREB-inducible hypoxia responsive genes are located in the intermediate region of the nucleus. Radial position of numerous responsive genes changed upon hypoxic treatment. Analysis of the relative distances amongst a subset of HIF target gene groups revealed that some gene pairs also altered their relative location to each other upon hypoxic treatment, suggesting higher order chromatin rearrangements. While these changes in location occurred in response to hypoxic activation of the target genes, they did not correlate with the extent of their activation. These results suggest that induction of the hypoxia-responsive gene expression program is accompanied by spatial alterations of the genome, but that radial and relative gene positions are not directly related to gene activity.


title: A serotonergic axon-cilium synapse drives nuclear signaling to maintain chromatin accessibility

abstract: Chemical synapses between axons and dendrites mediate much of the brains intercellular communication. Here we describe a new kind of synapse - the axo-ciliary synapse - between axons and primary cilia. By employing enhanced focused ion beam - scanning electron microscopy on samples with optimally preserved ultrastructure, we discovered synapses between the serotonergic axons arising from the brainstem, and the primary cilia of hippocampal CA1 pyramidal neurons. Functionally, these cilia are enriched in a ciliary-restricted serotonin receptor, 5-hydroxytryptamine receptor 6 (HTR6), whose mutation is associated with learning and memory defects. Using a newly developed cilia-targeted serotonin sensor, we show that optogenetic stimulation of serotonergic axons results in serotonin release onto cilia. Ciliary HTR6 stimulation activates a non-canonical Gq/11-RhoA pathway. Ablation of this pathway results in nuclear actin and chromatin accessibility changes in CA1 pyramidal neurons. Axo-ciliary synapses serve as a distinct mechanism for neuromodulators to program neuron transcription through privileged access to the nuclear compartment.


title: Small extracellular vesicles from iPSC-MSC lose their regenerative potential upon UV-C irradiation

abstract: Mesenchymal Stem Cells derived from induced Pluripotent Stem cells (iPSC-MSC) have become a promising alternative to classical Mesenchymal Stem Cells in regenerative medicine. Their properties -as immunomodulatory and regenerative capacities-are in part due to the secretion of Extracellular Vesicles (EVs). Small EVs (sEVs) with sizes that range from 50 to 120 nm contain proteins, lipids, and nucleic acids that exert a role in cellular communication. Their content will depend on the cell of origin and its physiological state, thus the message they convey might change in response to changes in cellular conditions. In particular, the DNA damage response (DDR) has been reported to modulate sEVs secretion. In this work, we analyze how UV-C radiation upon iPSC-MSC alter sEVs secretion, cargo and bystander effect. Here, we confirm that UV-C radiation causes DDR in a dose dependent manner. In addition, we found that UV-C induced stress did not modulate the expression of genes that participate in sEVs biogenesis pathway. Consequently, we found that the amount of sEVs secreted by radiated and non-irradiated cells remained stable. However, sEVs from radiated cells were unable to promote cell migration in their target cells. Moreover, a label-free proteomic analysis revealed that UV-C induced DDR produces sEVs with an altered cargo, rich in migration-inhibiting proteins, and resulting in a less stromal-oriented repertoire.


title: PCSK9 affects expression of key surface proteins in human pancreatic beta cells through intra- and extracellular regulatory circuits

abstract:
Aims/hypothesisProprotein convertase subtilisin/kexin 9 (PCSK9) is involved in the degradation of LDLR. However, PCSK9 can target other proteins in a cell-type specific manner. While PCSK9 has been detected in pancreatic islets, its expression in insulin-producing pancreatic beta cells is debated. Herein, we studied PCSK9 expression, regulation and function in the human pancreatic beta cell line EndoC-{beta}H1.

MethodsWe assessed PCSK9 expression in mouse and human pancreatic islets, and in the pancreatic beta cell line EndoC-{beta}H1. We also studied PCSK9 regulation by cholesterol, lipoproteins, Mevastatin, and by SREBPs transcription factors. To evaluate PCSK9 function in pancreatic beta cells, we performed PCSK9 gain-and loss-of-function experiments in EndoC-{beta}H1 using siPCSK9 or recombinant PCSK9 treatments, respectively.

ResultsWe demonstrate that PCSK9 is expressed and secreted by pancreatic beta cells. In EndoC-{beta}H1 cells, PCSK9 expression is regulated by cholesterol and by SREBPs transcription factors. Importantly, PCSK9 knockdown results in multiple transcriptome, proteome and secretome deregulations and impaired insulin secretion. By gain- and loss-of-function experiments, we observed that PCSK9 regulates the expression levels of LDLR and VLDLR through an extracellular mechanism while CD36, PD-L1 and HLA-ABC are regulated through an intracellular mechanism.

Conclusions/interpretationCollectively, these results highlight PCSK9 as an important regulator of CD36, PD-L1 and HLA-ABC cell surface expression in pancreatic beta cells.

Data availabilityRNA-seq data have been deposited to GEO database with accession number GSE182016. Mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the following identifiers: PXD027921, PXD027911 and PXD027913.


title: Reovirus infection is regulated by NPC1 and endosomal cholesterol homeostasis

abstract:
Cholesterol homeostasis is required for the replication of many viruses, including Ebola virus, hepatitis C virus, and human immunodeficiency virus-1. Niemann-Pick C1 (NPC1) is an endosomal-lysosomal membrane protein involved in cholesterol trafficking from late endosomes and lysosomes to the endoplasmic reticulum. We identified NPC1 in CRISPR and RNA interference screens as a putative host factor for infection by mammalian orthoreovirus (reovirus). Following internalization via clathrin-mediated endocytosis, the reovirus outer capsid is proteolytically removed, the endosomal membrane is disrupted, and the viral core is released into the cytoplasm where viral transcription, genome replication, and assembly take place. We found that reovirus infection is significantly impaired in cells lacking NPC1, but infection is restored by treatment of cells with hydroxypropyl-{beta}-cyclodextrin, which binds and solubilizes cholesterol. Absence of NPC1 did not dampen infection by infectious subvirion particles, which are reovirus disassembly intermediates that bypass the endocytic pathway for infection of target cells. NPC1 is not required for reovirus attachment to the plasma membrane, internalization into cells, or uncoating within endosomes. Instead, NPC1 is required for delivery of transcriptionally active reovirus core particles into the cytoplasm. These findings suggest that cholesterol homeostasis, ensured by NPC1 transport activity, is required for reovirus penetration into the cytoplasm, pointing to a new function for NPC1 and cholesterol homeostasis in viral infection.

Author summaryGenetic screens are useful strategies to identify host factors required for viral infection. NPC1 was identified in independent CRISPR and RNA interference screens as a putative host factor required for reovirus replication. We discovered that NPC1-mediated cholesterol transport is dispensable for reovirus attachment, internalization, and disassembly but required for penetration of the viral disassembly intermediate from late endosomes into the cytoplasm. These findings pinpoint an essential function for cholesterol in the entry of reovirus and raise the possibility that cholesterol homeostasis regulates the entry of other viruses that penetrate late endosomes to initiate replication.


title: Omics profiling identifies MAPK/ERK pathway as a gatekeeper of nephron progenitor metabolism

abstract: Nephron endowment is defined by fetal kidney growth and critically dictates renal health in adults. Despite the advances in understanding the molecular regulation of nephron progenitors, the causes for low congenital nephron count and contribution of basic metabolism to nephron progenitor biology remain poorly understood. Here we characterized the metabolic consequences of MAPK/ERK-deficiency in nephron progenitors, whose maintenance and propagation in developing kidney critically depends on ERK activation. Our LC/MS-based metabolomics profiling identified 42 reduced metabolites, of which 26 were further supported by in vivo transcriptional characterization of MAPK/ERK-deficient nephron progenitors. This revealed a severe shortage of energy and nucleotide biosynthesis precursors, blockage in glycolysis and diminished pyruvate and proline metabolism. Utilization of in vitro kidney cultures demonstrated a dosage-specific function for glycolytic pyruvate as an energy source that controls the shape of the ureteric bud tip kwon to serve as a niche for nephron progenitor regulation. Analysis of the proline biosynthesis effects in developing kidney in vivo revealed premature loss of nephron progenitor maintenance in the absence of Pycr1/2 functions. Our results suggest that MAPK/ERK-dependent nephron progenitor metabolism functionally contributes to progenitor preservation by controlling pyruvate availability and proline metabolism in developing kidneys.

category: developmental biology


title: Mechano-Biochemical Regulation of the C. elegans HMP1-HMP2 protein complex

abstract: The HMP1-HMP2 protein complex, a counterpart of -catenin-{beta}-catenin complex in C. elegans, mediates the tension transmission between HMR1 (cadherin) and actin cytoskeleton and serves as a critical mechanosensor at the cell-cell adherens junction. The complex has been shown to play critical roles in embryonic development and tissue integrity in C. elegans. The complex is subject to tension due to internal actomyosin contractility and external mechanical micro-environmental perturbations. However, how tension regulates the stability and interaction of HMP1-HMP2 complex has yet to be investigated. Here, we directly quantify the mechanical stability of the full-length HMP1 and its force-bearing modulation domains (M1-M3), and show that they unfold within physiological level of tension (pico-newton scale). The inter-domain interactions within the modulation domain leads to strong mechanical stabilization of M1 in HMP1, resulting in a significantly stronger force threshold to expose the buried vinculin binding site compared to the M1 domain in -catenins. Moreover, we also quantify the mechanical stability of the inter-molecular HMP1-HMP2 interface and show that it is mechanically stable enough to support the tension-transmission and tension-sensing of the HMP1 modulation domains. Further, we show that single-residue phosphomimetic mutation (Y69E) on HMP2 weakens the mechanical stability of the HMP1-HMP2 interface and thus weakens the force-transmission molecular linkage and the associated mechanosensing functions. Together, these results provide a mechano-biochemical understanding of C. elegans HMP1-HMP2 protein complexs roles in mechanotransduction.

category: biophysics


title: Generation of iPSC lines with high cytogenetic stability from peripheral blood mononuclear cells (PBMCs)

abstract: The utility of human induced pluripotent stem cells (hiPSCs) is contingent upon genomic integrity and stability. Recurrent genomic aberrations have been observed in human iPSC lines upon long-term culture, [~]10-25% demonstrate karyotype abnormalities. We describe a new and reliable non-integrating episomal plasmid reprogramming method for fresh (unexpanded) peripheral blood mononuclear cells (PBMC) into iPSCs (PBMC-iPSCs). PBMC-iPSCs produced using this method have a superior chromosome-level karyotype stability rate ([~]5% abnormality rate for all chromosomes; 2.8% for autosomes). After extended culture PBMC-iPSCs maintain a low rate of abnormalities (2% for autosomes). Deep coverage whole genome sequencing in a subset of PBMC-iPSC lines showed no shared single nucleotide polymorphisms (SNPs) or structural variants are introduced during reprogramming and maintenance of PBMC-iPSCs. iPSCs reprogrammed from unexpanded PBMCs have consistently high cytogenetic stability and minimal genomic aberrations, suggesting this method is highly suited for iPSCs in research and therapeutic clinical applications.


title: Dynein acts to cluster glutamate receptors and traffic the PIP5 kinase, Skittles, to regulate postsynaptic membrane organization at the neuromuscular junction

abstract: Cytoplasmic dynein is essential in motoneurons for retrograde cargo transport that sustains neuronal connectivity. Little, however, is known about dyneins function on the postsynaptic side of the circuit. Here we report distinct postsynaptic roles for dynein at neuromuscular junctions (NMJs). Intriguingly, we show that dynein punctae accumulate postsynaptically at glutamatergic synaptic terminals. Moreover, Skittles, a phosphatidylinositol 4-phosphate 5-kinase that produces PI(4,5)P2 to organize the spectrin cytoskeleton, also localizes specifically to glutamatergic synaptic terminals. Depletion of postsynaptic dynein disrupts the accumulation of Skittles, PI(4,5)P2 phospholipid, and organization of the spectrin cytoskeleton at the postsynaptic membrane. Coincidental with dynein depletion, we observe an increase in the clusters size of ionotropic glutamate receptor (iGluR), and an increase in the amplitude and frequency of mEJPs. However, PI(4,5)P2 levels do not affect iGluR clustering and dynein does not affect the protein levels of iGluR subunits at the NMJ, suggesting a separate, transport independent function for dynein in iGluR cluster organization. As dynein punctae closely associate with iGluR clusters, we propose that dynein physically tethers iGluR clusters at the postsynaptic membrane to ensure proper synaptic transmission.


title: Optogenetic control of NOTCH1 signalling

abstract: The Notch signalling pathway is a crucial regulator of cell differentiation as well as tissue organisation. Dysregulation of Notch signalling has been linked to the pathogenesis of different diseases. Notch plays a key role in breast cancer progression by controlling the interaction between the tumour cells and the microenvironment as well as by increasing cell motility and invasion. NOTCH1 is a mechanosensitive receptor, where mechanical force is required to activate the proteolytic cleavage and release of the Notch intracellular domain (NICD). Here, we circumvent this step by regulating Notch activity by light. To achieve this, we have engineered a membrane-bound optogenetic NOTCH1 receptor (optoNotch) to control the activation of NOTCH1 intracellular domain (N1ICD) and its downstream transcriptional activities. Using optoNotch we confirm that NOTCH1 activation increases cell proliferation in MCF7 and MDA-MB-468 breast cancer cells in 2D and spheroid 3D cultures. OptoNotch allows fine-tuning ligand-independent regulation of N1ICD to understand the spatiotemporal complexity of Notch signalling.


title: Biomimetic Vasculatures by 3D-Printed Porous Molds

abstract: Anatomically and biologically relevant vascular models are critical to progress our understanding of cardiovascular diseases (CVDs) that can lead to effective therapies. Despite advances in 3D bioprinting, recapitulating complex architectures (i.e., freestanding, branching, multilayered, perfusable) of a cell-laden vascular construct remains technically challenging, and the development of new techniques that can recapitulate both anatomical and biological features of blood vessels is of paramount importance. In this work, we introduce a unique, microfluidics-enabled molding technique that allows us to fabricate anatomically-relevant, cell-laden hydrogel vascular models. Our approach employed 3D-printed porous molds of poly(ethylene glycol) diacrylate (PEGDA) as templates to cast alginate-containing bioinks. Due to the porous and aqueous nature of the PEGDA mold, the calcium ion (Ca2+) was diffusively released to crosslink the bioinks to create hollow structures. Applying this technique, multiscale, multilayered vascular constructs that were freestanding and perfusable were readily fabricated using cell-compatible bioinks (i.e., alginate and gelatin methacryloyl (GelMA)). The bioinks were also readily customizable to either improve the compatibility with specific vascular cells or tune the mechanical modulus to mimic native blood vessels. Importantly, we successfully integrated smooth muscle cells and endothelial cells in a biomimetic organization within our vessel constructs and demonstrated a significant increase in monocyte adhesion upon stimulation with an inflammatory cytokine, tumor necrosis factor-alpha (TNF-). We also demonstrated that the fabricated vessels were amenable for testing percutaneous coronary interventions (i.e., drug-eluting balloons and stents) under physiologically-relevant mechanical states, such as vessel stretching and bending. Overall, we introduce a versatile fabrication technique with multi-faceted possibilities of generating biomimetic vascular models that can benefit future research in mechanistic understanding of CVD progression and the development of therapeutic interventions.

category: bioengineering


title: Practices for Measuring 3D Organelle Morphology and Generating Surfaces with Amira

abstract:
Analysis of 3D structures is of paramount importance in cellular biology. Although light microscopy and transmission electron microscopy (TEM) have remained staples for imaging cellular structures, they lack the ability to image in 3D. However, recent technological advances, such as serial block-face scanning electron microscopy (SBF-SEM) and focused ion beam scanning electron microscopy (FIB-SEM), have allowed researchers to observe cellular ultrastructure in 3D. Here, we propose a standardized protocol using the visualization software Amira to quantify organelle morphologies in 3D; this method allows researchers to produce accurate and reproducible measurements of cellular structure characteristics. We demonstrate this applicability by utilizing SBF-SEM and Amira to quantify mitochondria and endoplasmic reticulum (ER) structures.

GRAPHICAL ABSTRACT

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title: Protein-protein interaction-Gaussian accelerated molecular dynamics (PPI-GaMD): Characterization of protein binding thermodynamics and kinetics

abstract: Protein-protein interactions (PPIs) play key roles in many fundamental biological processes such as cellular signaling and immune responses. However, it has proven challenging to simulate repetitive protein association and dissociation in order to calculate binding free energies and kinetics of PPIs, due to long biological timescales and complex protein dynamics. To address this challenge, we have developed a new computational approach to all-atom simulations of PPIs based on a robust Gaussian accelerated molecular dynamics (GaMD) technique. The method, termed "PPI-GaMD", selectively boosts interaction potential energy between protein partners to facilitate their slow dissociation. Meanwhile, another boost potential is applied to the remaining potential energy of the entire system to effectively model the proteins flexibility and rebinding. PPI-GaMD has been demonstrated on a model system of the ribonuclease barnase interactions with its inhibitor barstar. Six independent 2 s PPI-GaMD simulations have captured repetitive barstar dissociation and rebinding events, which enable calculations of the protein binding thermodynamics and kinetics simultaneously. The calculated binding free energies and kinetic rate constants agree well with the experimental data. Furthermore, PPI-GaMD simulations have provided mechanistic insights into barstar binding to barnase, which involve long-range electrostatic interactions and multiple binding pathways, being consistent with previous experimental and computational findings of this model system. In summary, PPI-GaMD provides a highly efficient and easy-to-use approach for binding free energy and kinetics calculations of PPIs.


title: Global Metabolome Analysis of Dunaliella tertiolecta, Phaeobacter italicus R11 Co-cultures using Thermal Desoprtion - Comprehensive Two-dimensional Gas Chromatography - Time-of-Flight Mass Spectrometry

abstract:
Dunaliella tertiolecta is a marine microalgae that has been studied extensively as a potential carbon-neutral biofuel source [1]. Microalgae oil contains high quantities of energy-rich fatty acids and lipids, but is not yet commercially viable as an alternative fuel. Carefully optimised growth conditions, and more recently, algal-bacterial co-cultures have been explored as a way of improving the yield of Dunaliella tertiolecta microalgae oils. The relationship between the host microalgae and bacterial co-cultures is currently poorly understood. Here, a complete workflow is proposed to analyse the global metabolomic profile of co-cultured Dunaliella tertiolectra and Phaeobacter italicus R11, which will enable researchers to explore the chemical nature of this relationship in more detail.

Graphical Abstract

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C_LIO_LIAdvanced chemometric tools can extract, useful and discriminating metabolomic features even from very noisy complex datasets.
C_LI


title: Pseudolaguvia moinensis, a new benthic miniature Asian catfish (Teleostei: Sisoridae) from Arunachal Pradesh, north-eastern India

abstract: A new species of miniature sisorid catfish is described, from the upper Brahmaputra river drainage in northeastern India. The new species is distinguished from all its congeners except P. ferruginea and P. focusa by having an elongate light brown to cream marks either side on ventro-lateral margin just above anus. Further, distinguished from its congeners by the following combination of characters: presence of a W-shaped dark brown band on caudal fin, thoracic adhesive apparatus extending closer to pelvic-fin base, smooth anterior margin of dorsal spine, a narrow V-shaped light brown to cream bands on side of the body, dorsal-spine length (10.7-14.7% SL), dorsal-fin base length (10.7-14.2% SL), pectoral-fin spine length (14.0-21.1% SL), pelvic-fin length (13.5-16.6% SL), caudal peduncle length (14.4-18.2% SL) and depth (7.8-9.9% SL), total vertebrae (30-31), caudal fin with a complete medial hyaline bands towards its anterior end, reaching outer margin of each lobe. Other combination of characters differentiating the new species from its congeners are provided in the respective diagnoses.

category: zoology


title: Fragile X syndrome patient-derived neurons developing in the mouse brain show FMR1 -dependent phenotypes

abstract: Abnormal neuronal development in Fragile X syndrome (FXS) is poorly understood. Data on FXS patients remain scarce and FXS animal models have failed to yield successful therapies. In vitro models do not fully recapitulate the morphology and function of human neurons. Here, we co-injected neural precursor cells (NPCs) from FXS patient-derived and corrected isogenic control induced pluripotent stem cells into the brain of neonatal immune-deprived mice. The transplanted cells populated the brain and a proportion differentiated into neurons and glial cells. Single-cell RNA sequencing of transplanted cells revealed upregulated excitatory synaptic transmission and neuronal differentiation pathways in FXS neurons. Immunofluorescence analyses showed accelerated maturation of FXS neurons after an initial delay. Additionally, increased percentages of Arc- and Egr1-positive FXS neurons and wider dendritic protrusions of mature FXS striatal medium spiny neurons pointed to an increase in synaptic activity and synaptic strength as compared to control. This transplantation approach provides new insights into the alterations of neuronal development in FXS by facilitating physiological development of cells in a 3D context, and could be used to test new therapeutic compounds correcting neuronal development defects in FXS.


title: Compartment-specific tuning of hippocampal dendritic feature selectivity by intracellular Ca2+ release

abstract: Dendritic Ca2+ signaling is central to neural plasticity mechanisms allowing animals to adapt to the environment. Intracellular Ca2+ release (ICR) from endoplasmic reticulum has long been thought to shape these mechanisms. However, ICR has not been investigated in mammalian neurons in vivo. We combined electroporation of single CA1 pyramidal neurons, simultaneous imaging of dendritic and somatic activity during spatial navigation, optogenetic place field induction, and acute genetic augmentation of ICR cytosolic impact to reveal that ICR supports the establishment of dendritic feature selectivity and shapes integrative properties determining output-level receptive fields. This role for ICR was more prominent in apical than in basal dendrites. Thus, ICR cooperates with circuit-level architecture in vivo to promote the emergence of behaviorally-relevant plasticity in a compartment-specific manner.


title: Distinct neural representations for prosocial and self-benefitting effort

abstract: Prosocial behaviours - actions that benefit others - are central to individual and societal well-being. Most prosocial acts are effortful. Yet, how the brain encodes effort costs when actions benefit others is unknown. Here, using a combination of multivariate representational similarity analysis and model-based univariate analysis during fMRI, we reveal how the costs of prosocial efforts are processed. Strikingly, we identified a unique neural signature of effort in the anterior cingulate gyrus for prosocial acts both when choosing to help others and when exerting force for their benefit. This pattern was absent for similar self-benefitting behaviour and correlated with individual levels of empathy. In contrast, the ventral tegmental area and the ventral insula signalled subjective value preferentially when choosing whether to exert effort to benefit oneself. These findings demonstrate partially distinct brain areas guide the evaluation and exertion of effort costs when acts are prosocial or self-benefitting.


title: mSOD1-NSG mice: a new in vivo model to test human T cells in ALS

abstract:
Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease with unclear etiology and few treatment options. Engineering of human cells for therapy has great potential to provide a means to create long-lived therapeutics that can respond to local signals within tissues. One challenge for development of human cell therapeutics is to have disease models that do not reject transplanted human cells. G93A mutant superoxide dismutase-1 (mSOD1) transgenic mouse model is a robust disease model for ALS, with development of local inflammation, activation of microglia and astrocytes, motor neuron death, and development of paralysis. To create a mouse model for ALS that permits the transplantation of human T cells without immune-mediate rejection, we bred the G93A transgene onto the NOD-SCID-IL-2R{gamma}-deficient (NSG) mouse model to create mSOD1-NSG mice. We report that mSOD1-NSG mice develop a progressive ALS-like disease with microgliosis, astrogliosis, and paralysis development with an average onset at 11 weeks and end-stage at 14 weeks. Transplanted human T regulatory cells survive in these mice for at least 60 days. This mSOD1-NSG mouse model for ALS can be used to test human T cell-based therapeutics, and it may be helpful to test any human cell-based therapy for ALS.

Significance StatementG93A SOD1 transgenic NSG mice develop a progressive ALS-like disease and are an in vivo model for testing human T cell-based therapies for ALS.

category: immunology


title: Genetic barriers to gene flow separate divergent substitution rates across a butterfly hybrid zone

abstract: Hybridization is a major evolutionary force that can erode genetic differentiation between species, whereas reproductive isolation maintains such differentiation. In studying a hybrid zone between the swallowtail butterflies Papilio syfanius and Papilio maackii, we made the unexpected discovery that genomic substitution rates are unequal between the parental species. This phenomenon creates a novel process in hybridization, where genomic regions most affected by gene flow evolve at similar rates, while genomic regions with greater reproductive isolation evolve at divergent rates. Thus, hybridization mixes evolutionary rates in a way similar to its effect on ancestry. Using coalescent theory, we show that the rate-mixing process provides distinct information about levels of gene flow across different parts of genomes, and that maintenance of divergent substitution rates can be predicted quantitatively from relative sequence divergence (FST) between the hybridizing species at equilibrium. A corollary is that divergent rates will be maintained in regions linked to barrier loci. Overall, we demonstrate that reproductive isolation maintains not only the final outcome of genomic differentiation, but also the rate at which differentiation accumulates. This new information also suggests that the separation of evolutionary rates co-localizes with the separation of gene pools between genomes of incipient species.

category: evolutionary biology


title: Admixture of evolutionary rates across a hybrid zone


title: Single cell Iso-Sequencing enables rapid genome annotation for scRNAseq analysis

abstract: Single cell RNA sequencing (scRNAseq) is a powerful technique that continues to expand across various biological applications. However, incomplete 3' UTR annotations in less developed or non-model systems can impede single cell analysis resulting in genes that are partially or completely uncounted. Performing scRNAseq with incomplete 3' UTR annotations can impede the identification of cell identities and gene expression patterns and lead to erroneous biological inferences. We demonstrate that performing single cell isoform sequencing (ScISOr-Seq) in tandem with scRNAseq can rapidly improve 3' UTR annotations. Using threespine stickleback fish (Gasterosteus aculeatus), we show that gene models resulting from a minimal embryonic ScISOr-Seq dataset retained 26.1% greater scRNAseq reads than gene models from Ensembl alone. Furthermore, pooling our ScISOr-Seq isoforms with a previously published adult bulk Iso-Seq dataset from stickleback, and merging the annotation with the Ensembl gene models, resulted in a marginal improvement (+0.8%) over the ScISOr-Seq only dataset. In addition, isoforms identified by ScISOr-Seq included thousands of new splicing variants. The improved gene models obtained using ScISOr-Seq lead to successful identification of cell types and increased the reads identified of many genes in our scRNAseq stickleback dataset. Our work illuminates ScISOr-Seq as a cost-effective and efficient mechanism to rapidly annotate genomes for scRNAseq.

category: genetics


title: Prime-seq, efficient and powerful bulk RNA-sequencing

abstract: With the advent of Next Generation Sequencing, RNA-sequencing (RNA-seq) has become the major method for quantitative gene expression analysis. Reducing library costs by early barcoding has propelled single-cell RNA-seq, but has not yet caught on for bulk RNA-seq. Here, we optimized and validated a bulk RNA-seq method we call prime-seq. We show that with respect to library complexity, measurement accuracy, and statistical power it performs equivalent to TruSeq, a standard bulk RNA-seq method, but is four-fold more cost-efficient due to almost 50-fold cheaper library costs. We also validate a direct RNA isolation step that further improves cost and time-efficiency, show that intronic reads are derived from RNA, validate that prime-seq performs optimal with only 1,000 cells as input, and calculate that prime-seq is the most cost-efficient bulk RNA-seq method currently available. We discuss why many labs would profit from a cost-efficient early barcoding RNA-seq protocol and argue that prime-seq is well suited for setting up such a protocol as it is well validated, well documented, and requires no specialized equipment.


title: OnePetri: accelerating common bacteriophage Petri dish assays with computer vision

abstract:
IntroductionBacteriophage plaque enumeration is a critical step in a wide array of protocols. The current gold standard for plaque enumeration on Petri dishes is through manual counting. This approach is time-intensive, has low-throughput, is limited to Petri dishes which have a countable number of plaques, and can have variable results upon recount due to human error.

MethodsWe present OnePetri, a collection of trained machine learning models and open-source mobile application for the rapid enumeration of bacteriophage plaques on circular Petri dishes.

ResultsWhen compared against the current gold standard of manual counting, OnePetri was significantly faster, with minimal error. Compared against two other similar tools, Plaque Size Tool and CFU.AI, OnePetri had higher plaque recall and reduced detection times on most test images.

ConclusionsThe OnePetri application can rapidly enumerate phage plaques on circular Petri dishes with high precision and recall.


title: Expert-integrated automated machine learning uncovers hemodynamic predictors in spinal cord injury

abstract: Automated machine learning (AutoML) is positioned to democratize artificial intelligence (AI) by reducing the amount of human input and ML expertise needed to create prediction models. However, successful translation of ML in biomedicine requires moving beyond optimizing only for prediction accuracy and towards discovering reproducible clinical and biological inferences. Here, we present a model-agnostic framework to reinforce AutoML using strategies and tools of explainable and reproducible AI, including novel metrics for performance precision and feature instability. The framework enables clinicians to interpret AutoML-generated models for clinical and biological verifiability and consequently integrate domain expertise during model development. We applied the framework towards spinal cord injury prognostication and identified a detrimental relationship between intraoperative hypertension and patient outcome. Furthermore, our analysis captured evolving clinical practices such as faster time-to-surgery and blood pressure management that affected clinical model validation. Altogether, we illustrate how augmenting AutoML for inferential reproducibility empowers biomedical discovery and builds trust in AI processes towards effective clinical integration.


title: PINFORMED1 polarity in the shoot is insensitive to the polarity of neighbouring cells

abstract: In plants, epidermal cells are planar-polarized along an axis marked by the asymmetric localization patterns of several proteins including PIN-FORMED1 (PIN1), which facilitates the directional efflux of the plant hormone auxin to pattern phyllotaxis (Heisler et al., 2010; Mansfield et al., 2018; Reinhardt et al., 2003). While PIN1 polarity is known to be regulated non-cell autonomously via the MONOPTEROS (MP) transcription factor, how this occurs has not been determined (Bhatia et al., 2016). Here we use mosaic expression of the serine threonine kinase PINOID (PID) to test whether PIN1 polarizes according to the polarity of neighbouring cells. Our findings reveal that PIN1 is insensitive to the polarity of PIN1 in neighbouring cells arguing against auxin flux or extracellular auxin concentrations acting as a polarity cue, in contrast to previous model proposals (Abley et al., 2016; Stoma et al., 2008).


title: PIN-FORMED1 polarity in the shoot is insensitive to the polarity of neighbouring cells


title: A merger between compatible but divergent genomes supports allopolyploidization in the Brassicaceae family

abstract: Hybridization and polyploidization are pivotal to plant evolution. Genetic crosses between distantly related species rarely occur in nature mainly due to reproductive barriers but how such hurdles can be overcome is largely unknown. xBrassicoraphanus is a fertile intergeneric allopolyploid synthesized between Brassica rapa and Raphanus sativus in the Brassicaceae family. Genomes of B. rapa and R. sativus are diverged enough to suppress synapsis formation between non-homologous progenitor chromosomes during meiosis, and we found that both genomes reside in the single nucleus of xBrassicoraphanus without genome loss or rearrangement. Expressions of syntenic orthologs identified in B. rapa and R. sativus were adjusted to a hybrid nuclear environment of xBrassicoraphanus, which necessitates reconfiguration of transcription network by rewiring cis-trans interactions. B. rapa coding sequences have a higher level of gene-body methylation than R. sativus, and such methylation asymmetry is maintained in xBrassicoraphanus. B. rapa-originated transposable elements were transcriptionally silenced in xBrassicoraphanus, rendered by gain of CHG methylation in trans via small RNAs derived from the same sequences of R. sativus subgenome. Our work proposes that not only transcription compatibility but also a certain extent of genome divergence supports hybrid genome stabilization, which may explain great diversification and expansion of angiosperms during evolution.


title: The APC/CCCS52A2 E3-Ligase Complex targeted PKN1 Protein is a Plant Lineage-Specific Driver of Cell Division

abstract:
The anaphase-promoting complex/cyclosome (APC/C) marks key cell cycle proteins for proteasomal breakdown, thereby ensuring unidirectional progression through the cell cycle. Its target recognition is temporally regulated by activating subunits, one of which is called CELL CYCLE SWITCH 52 A2 (CCS52A2). We sought to expand the knowledge of identified APC/C targets by using the severe growth phenotypes of CCS52A2-deficient Arabidopsis thaliana plants as a readout in a suppressor mutagenesis screen, resulting in the identification of the previously undescribed gene called PIKMIN1 (PKN1). PKN1 deficiency rescues the disorganized root stem cell phenotype of the ccs52a2-1 mutant, whereas an excess of PKN1 inhibits growth of ccs52a2-1 plants, indicating the importance of PKN1 abundance for proper development. Accordingly, the lack of PKN1 in a wild-type background negatively impacts cell division, while its ectopic expression promotes proliferation. PKN1 shows a cell cycle phase-dependent accumulation pattern, localizing to microtubular structures, including the preprophase band, the mitotic spindle, and phragmoplast. PKN1 is conserved throughout the plant kingdom, with its function in cell division being evolutionary conserved in the liverwort Marchantia polymorpha. Our data thus demonstrate that PKN1 represents a novel, plant-specific gene with a rate-limiting role in cell division, which is proteolytically controlled by the CCS52A2-activated APC/C.

One-Sentence SummaryPKN1 is a conserved plant-specific protein that is rate-limiting for cell division, likely through its interaction with microtubuli, and is proteolytically controlled by APC/CCCS52A2.


title: Tasselyzer, a machine learning method to quantify anther extrusion in maize, based on PlantCV

abstract:
Male fertility in maize is controlled by development and genetic programming and is directly impacted by environmental factors such as light, temperature, water, and nutrient availability; the control of this trait has substantial agronomic utility. Maize anthers emerge from male florets, which are clustered to form the tassel at the top of the plant separated from the female ear. Quantification of anther extrusion is one important aspect in the determination of male fertility. To address the lack of an automated method to measure anther extrusion on a large scale, we developed  Tasselyzer, a quantitative, image-based color trait analysis pipeline for tassel image segmentation, based on the existing PlantCV platform, and we applied it to determine the proportion of anther extrusion. We evaluated Tasselyzer in maize during the seven-day period of pollen shedding as well as in the temperature-sensitive male sterile mutant dcl5. With tassel images obtained with a smart phone camera, we show that the anther scores positively correlate with anther extrusion, and such methods can be used to measure environmental impacts on the dcl5 mutant. Altogether, this work establishes an automated and inexpensive method to quantify anther extrusion in maize, which would be useful for research and breeding.

Significance StatementTasselyzer is a novel image-based segmentation tool, to address the lack of an automated method to measure anther extrusion, and the impact of genetic and environment variation on male fertility in maize.


title: The Open Field test as a tool for behavior analysis in pigs - is a standardization of setup necessary? A systematic review.

abstract: The Open Field test is a common tool to measure anxiety and behavioral changes in rodents. However, scientific findings of rodent experiments may not translate adequately to humans and it has been shown that larger animal models might perform better in that regard. As a result, the number of published studies involving the Open Field test in domestic pig models is increasing. The aim of our review was to investigate the Open Field set-ups in published studies as well as similarity between performance and parameters published. Following the PRISMA guidelines for reviews we selected 69 studies for data extraction in this systematic review. We were able to determine specific set-up conditions such as size, duration and daytime for most of the included studies and found a high variability within these test specifiers. Results indicate a non-uniform performance of set-up including size, timing, parameters and additional combined tests such as the novel object test. We would like to point out the need for standardization of Open Field test for pigs in order to improve result, comparability and reduce inconsistencies.

category: scientific communication and education


title: Fast and cost-effective age estimation in Bengal tiger and Asiatic lion: applicability of cementum analysis method

abstract: Age estimation methods, through cementum analysis, for wild animals are rarely developed in Southeast Asian Countries. In the present study, we describe the applicability of the cementum analysis technique for developing a fast ({+/-}19 hours) and cost-effective age estimation method for Bengal tiger (Panthera tigris tigris) and Asiatic lion (Panthera leo persica) using incisor tooth. The I2 and I3 incisor teeth from the right mandible of a tiger and I2 and I3 from the left maxilla of a lion were used in the study. The longitudinal sections of the tooth were made using a low cost hand grinding technique on sand papers followed by decalcification and staining with hematoxylin. The cementum layers were counted under the microscope at 100X or 200X magnifications. Two cementum layers were observed in each of the I2 and I3 incisor tooth of tiger and six cementum layers were observed in each of the I2 and I3 incisor teeth of lion. The permanent incisors in tiger and lion erupt between 12-14 months; hence, we added 1 year to the counted number of cementum layers to estimate the final age of tiger and lion incisors. The absolute age of tiger and lion incisors was estimated to be of 2+1 years and 6+1 years, respectively. The same number of cementum layers in both incisors respective to the tiger and lion were observed. Therefore, we suggest (i) undertake the blind test and (ii) collect incisor teeth from naturally died or killed individuals for strengthening the database on the age of the wild population. This optimized method may be suitable for many carnivore species, applicable in wildlife forensic studies and can be used by researchers with minimum expertise, time, and funds requirements throughout the world.


title: Horn trophy seized in illegal wildlife trade assigned to swamp buffalo using a combined morphometric and DNA based approach in wildlife forensics in India

abstract: The illegal wildlife trade has threatened the existence of many extant wild animal species throughout the world. While dealing with the illegal wildlife trade of horns, we face problems of not having a proper protocol and lack of reference database to assign the species for proper implementation of wildlife laws. In one such condition, a horn trophy suspected to be of a wild buffalo was seized by authorities and sent to us for species identification. We used a combined approach of morphological and DNA analysis to ascertain the seized horns species. The two measurements, circumference at the base (CAB) and length on the front curve (LOFC) were measured for the seized and other horns of different bovid species, showing morphological resemblance with the seized horn. The 3-D scatter plot, generated by the values of CAB, LOFC and CAB/ LOFC, differentiated the different bovid species into distinct clusters and placed the seized horn in the proximity of domestic buffaloes. The Bayesian evolutionary analysis of the partial D-loop gene (521bp) placed the seized horn in a clade with swamp buffaloes. Since swamp buffaloes are domestic buffaloes, both these approaches concluded the same results. Hence, the current protocol developed may also be used to differentiate among wild buffalo, domestic buffalo, Cattle, Wild yak, Gaur and Takin using a combined approach of morphometric and DNA-based analysis, which may be used to deal with illegal wildlife trade of different bovid species at the world level.


title: Rapid Parameter Estimation for Selective Inversion Recovery Myelin Imaging using an Open-Source Julia Toolkit

abstract:
BackgroundMagnetic resonance imaging (MRI) is used extensively to quantify myelin content, however computational bottlenecks remain challenging for advanced imaging techniques in clinical settings. We present a fast, open-source toolkit for processing quantitative magnetization transfer derived from selective inversion recovery (SIR) acquisitions that allows parameter map estimation, including the myelin-sensitive macromolecular pool size ratio (PSR). Significant progress has been made in reducing SIR acquisition times to improve clinically feasibility. However, parameter map estimation from the resulting data remains computationally expensive. To overcome this computational limitation, we developed a computationally efficient, open-source toolkit implemented in the Julia language.

MethodsTo test the accuracy of this toolkit, we simulated SIR images with varying PSR and spin-lattice relaxation time of the free water pool (R1f) over a physiologically meaningful scale from 5 to 20% and 0.5 to 1.5 s-1, respectively. Rician noise was then added, and the parameter maps were estimated using our Julia toolkit. Probability density histogram plots and Lins concordance correlation coefficients (LCCC) were used to assess accuracy and precision of the fits to our known simulation data. To further mimic biological tissue, we generated five cross-linked bovine serum albumin (BSA) phantoms with concentrations that ranged from 1.25 to 20%. The phantoms were imaged at 3T using SIR, and data were fit to estimate PSR and R1f. Similarly, a healthy volunteer was imaged at 3T, and SIR parameter maps were estimated to demonstrate the reduced computational time for a real-world clinical example.

ResultsEstimated SIR parameter maps from our Julia toolkit agreed with simulated values (LCCC> 0.98). This toolkit was further validated using BSA phantoms and a whole brain scan at 3T. In both cases, SIR parameter estimates were consistent with published values using MATLAB. However, compared to earlier work using MATLAB, our Julia toolkit provided an approximate 20-fold reduction in computational time.

ConclusionsPresented here, we developed a fast, open-source, toolkit for rapid and accurate SIR MRI using Julia. The reduction in computational cost should allow SIR parameters to be accessible in clinical settings.


title: Feature-reweighted RSA: A method for improving the fit between computational models, brains, and behavior

abstract: Representational Similarity Analysis (RSA) has emerged as a popular method for relating representational spaces from human brain activity, behavioral data, and computational models. RSA is based on the comparison of representational (dis-)similarity matrices (RDM or RSM), which characterize the pairwise (dis-)similarities of all conditions across all features (e.g. fMRI voxels or units of a model). However, classical RSA treats each feature as equally important. This  equal weights assumption contrasts with the flexibility of multivariate decoding, which reweights individual features for predicting a target variable. As a consequence, classical RSA may lead researchers to underestimate the correspondence between a model and a brain region and, in case of model comparison, may lead them to select an inferior model. The aim of this work is twofold: First, we sought to broadly test feature-reweighted RSA (FR-RSA) applied to computational models and reveal the extent to which reweighting model features improves RSM correspondence and affects model selection. Previous work suggested that reweighting can improve model selection in RSA but it has remained unclear to what extent these results generalize across datasets and data modalities. To draw more general conclusions, we utilized a range of publicly available datasets and three popular deep neural networks (DNNs). Second, we propose voxel-reweighted RSA, a novel use case of FR-RSA that reweights fMRI voxels, mirroring the rationale of multivariate decoding of optimally combining voxel activity patterns. We found that reweighting individual model units markedly improved the fit between model RSMs and target RSMs derived from several fMRI and behavioral datasets and affected model selection, highlighting the importance of considering FR-RSA. For voxel-reweighted RSA, improvements in RSM correspondence were even more pronounced, demonstrating the utility of this novel approach. We additionally show that classical noise ceilings can be exceeded when FR-RSA is applied and propose an updated approach for their computation. Taken together, our results broadly validate the use of FR-RSA for improving the fit between computational models, brain, and behavioral data, possibly allowing us to better adjudicate between competing computational models. Further, our results suggest that FR-RSA applied to brain measurement channels could become an important new method to assess the correspondence between representational spaces.


title: Feature-reweighted representational similarity analysis: A method for improving the fit between computational models, brains, and behavior


title: The cytokine receptor DR3 identifies and activates thymic NKT17 cells

abstract: Invariant natural killer T (iNKT) cells are thymus-generated T cells with innate-like characteristics and effector function. Several functionally distinct iNKT subsets have been identified, but NKT17 is the only iNKT subset that produces the proinflammatory cytokine IL-17. NKT17 cells are generated in the thymus and then exported into the periphery to populate lymphoid organs and barrier tissues, such as the lung, to provide critical support in host defense. However, the molecular mechanisms that drive the thymic development and subset-specific activation of NKT17 cells remain mostly unknown. Here, we identify the cytokine receptor DR3, a member of the TNF receptor superfamily, being selectively expressed on NKT17 cells but absent on all other thymic iNKT subsets. We further demonstrate that DR3 ligation leads to the in vivo activation of thymic NKT17 cells and provides in vitro costimulatory effects upon -GalCer-stimulation. Thus, our study reports the identification of a specific surface marker for thymic NKT17 cells that selectively triggers their activation both in vivo and in vitro. These findings provide new insights for deciphering the role and function of IL-17-producing NKT17 cells and for understanding the development and activation mechanisms of iNKT cells in general.


title: TEMPERATURE AND NUTRIENT CONDITIONS MODIFY THE EFFECTS OF PHENOLOGICAL SHIFTS IN PREDATOR-PREY COMMUNITIES

abstract: While there is mounting evidence indicating that the relative timing of predator and prey phenologies shapes the outcome of trophic interactions, we still lack a comprehensive understanding of how important the environmental context (e.g. abiotic conditions) is for shaping this relationship. Environmental conditions not only frequently drive shifts in phenologies, but they can also affect the very same processes that mediate the effects of phenological shifts on species interactions. Thus, identifying how environmental conditions shape the effects of phenological shifts is key to predict community dynamics across a heterogenous landscape and how they will change with ongoing climate change in the future. Here I tested how environmental conditions shape effects of phenological shifts by experimentally manipulating temperature, nutrient availability, and relative phenologies in two predator-prey freshwater systems (mole salamander-bronze frog vs dragonfly larvae-leopard frog). This allowed me to (1) isolate the effect of phenological shifts and different environmental conditions, (2) determine how they interact, and (3) how consistent these patterns are across different species and environments. I found that delaying prey arrival dramatically increased predation rates, but these effects were contingent on environmental conditions and predator system. While both nutrient addition and warming significantly enhanced the effect of arrival time, their effect was qualitatively different: Nutrient addition enhanced the positive effect of early arrival while warming enhanced the negative effect of arriving late. Predator responses varied qualitatively across predator-prey systems. Only in the system with strong gape-limitation were predators (salamanders) significantly affected by prey arrival time and this effect varied with environmental context. Correlations between predator and prey demographic rates suggest that this was driven by shifts in initial predator-prey size ratios and a positive feedback between size-specific predation rates and predator growth rates. These results highlight the importance of accounting for temporal and spatial correlation of local environmental conditions and gape-limitation in predator-prey systems when predicting the effects of phenological shifts and climate change on predator-prey systems.


title: Involvement of non-LTR retrotransposons in the cancer incidence and lifespan of mammals

abstract: The presence in nature of closely related species showing drastic differences in lifespan and cancer incidence has recently increased the interest of the scientific community on these topics. In particular, the adaptations and genomic characteristics underlying the evolution of cancer-resistant and long-lived species have recently focused on the presence of alterations in the number of non-coding RNAs, on epigenetic regulation and, finally, on the activity of transposable elements (TEs). In this study, we compared the content and dynamics of TE activity in the genomes of four rodent and six bat species exhibiting different lifespans and cancer susceptibility. Mouse, rat and guinea pig (short-lived and cancer-prone organisms) were compared with the naked mole rat (Heterocephalus glaber) which is the rodent with the longest lifespan. The long-lived and cancer-resistant bats of the genera Myotis, Rhinolophus, Pteropus and Rousettus were instead compared with the Molossus, which is instead a short-lived and cancer-resistant organism. Analyzing the patterns of recent accumulations of TEs in the genome in these species, we found a strong suppression or negative selection to accumulation, of non-LTR retrotransposons in long-lived and cancer-resistant organisms. On the other hand, all short-lived and cancer-prone species have shown recent accumulation of this class of TEs. Among bats, the Molossus molossus turned out to be a very particular species and, at the same time, an important model because, despite being susceptible to rapid ageing, it is resistant to cancer. In particular, we found that its genome has the highest density of SINE (non-LTR retrotransposons), but, on the other hand, a total lack of active LINE retrotransposons. Our hypothesis is that the lack of LINEs presumably makes the Molossus cancer resistant due to lack of retrotransposition but, at the same time, the high presence of SINE, may be related to their short life span due to "sterile inflammation" and high mutation load. We suggest that research on ageing and cancer evolution should put particular attention to the involvement of non-LTR retrotransposons in these phenomena.


title: Integration of a radiofrequency coil and commercial field camera for ultra-high-field MRI

abstract:
PurposeTo develop an RF coil with an integrated commercial field camera for ultra-high field (7 T) neuroimaging. The RF coil will operate within a head-only gradient coil and be subject to the corresponding design constraints. The RF coil can thereafter be used for subject-specific correction of k-space trajectories--notably in gradient-sensitive sequences such as single-shot spiral imaging.

MethodsThe transmit and receive performance was evaluated before and after the integration of field probes, while field probes were evaluated when in an optimal configuration external to the coil and after their integration. Diffusion-weighted EPI and single-shot spiral acquisitions were employed to evaluate the efficacy of correcting higher order field perturbations and the consequent effect on image quality.

ResultsField probes had a negligible effect on RF-coil performance, including the transmit efficiency, transmit uniformity, and mean SNR over the brain. Modest reductions in field-probe signal lifetimes were observed, caused primarily by non-idealities in the gradient and shim fields of the head-only gradient coil at the probe positions. The field monitoring system could correct up to second-order field perturbations in single-shot spiral imaging.

ConclusionThe integrated RF coil and field camera was capable of concurrent field monitoring within a 7T head-only scanner and facilitated the subsequent correction of k-space trajectories during spiral imaging.


title: LORE homomerization is required for 3-OH-C10:0 induced immune signaling

abstract: Perception and processing of various internal and external signals is essential for all living organisms. Plants have an expanded and diversified repertoire of cell surface-localized receptor-like kinases (RLKs) that transduce signals across the plasma membrane. RLKs often assemble into higher-order receptor complexes with co-receptors, regulators and scaffolds to convert extracellular stimuli into cellular responses. To date, the only S-domain-RLK from Arabidopsis thaliana with a known ligand and function is AtLORE, a pattern recognition receptor that senses bacterial 3-hydroxy fatty acids of medium chain length, such as 3-hydroxy decanoic acid (3-OH-C10:0), to activate pattern-triggered immunity. Here we show that AtLORE forms receptor homomers, which is essential for 3-OH-C10:0-induced immune signaling. AtLORE homomerization is mediated by the transmembrane and extracellular domain. We show natural variation in the perception of 3-OH-C10:0 within the Brassicaceae family. Arabidopsis lyrata and Arabidopsis halleri do not respond to 3-OH-C10:0, although they possess a putative LORE orthologue. We found that LORE orthologues of these 3-OH-C10:0 nonresponsive species have defective extracellular domains that can bind the 3-OH-C10:0 ligand but lack the ability to homomerize. Our findings shed light on the activation mechanisms of AtLORE and explain natural variation of 3-OH-C10:0 perception within the Brassicaceae family.


title: Phosphorus uptake and toxicity is delimited by mycorrhizal symbiosis in P-sensitive Eucalyptus marginata but not in P-tolerant Acacia celastrifolia

abstract: Many plant species from regions with ancient, highly-weathered nutrient-depleted soils have specialised adaptations for acquiring P and are sensitive to excess P-supply. Mycorrhizal associations may regulate P-uptake at high external P-concentrations, potentially reducing P-toxicity. We predicted that excess P-application will negatively impact species from the nutrient-depleted jarrah forest of Western Australia and that mycorrhizal inoculation will reduce P-toxicity by regulating P-uptake. For seedlings of the N2-fixing legume Acacia celastrifolia and the tree species Eucalyptus marginata, we measured growth at P-concentrations of 0 to 90 mg kg-1 soil and in relation to inoculation with the arbuscular mycorrhizal fungus (AMF) Rhizophagus irregularis. Non-inoculated A. celastrifolia maintained leaf P-concentrations at <2 mg g-1 dry mass (DM) across the range of external P-concentrations. However, for non-inoculated E. marginata, as external P-concentrations increased leaf P also increased, reaching >9 mg g-1 DM at 30 mg P kg-1 soil. A. celastrifolia DM increased with increasing external P-concentrations, while E. marginata DM was maximal at 15 mg P kg-1 soil, declining at higher external P concentrations. Neither DM nor leaf P of A. celastrifolia were affected by inoculation with AMF. For E. marginata, even at 90 mg P kg-1 soil, inoculation with AMF resulted in leaf P remaining <1 mg g-1 DM, and DM being maintained. These data strengthen the evidence base that AMF may not only facilitate P-uptake at low external P-concentrations, but are also important for moderating P-uptake at elevated external P-concentrations and maintaining plant P concentrations within a relatively narrow concentration range.


title: Inhibition of chloroplast translation as a new target for herbicides

abstract: The rise in herbicide resistance over recent decades threatens global agriculture and food security and so discovery of new modes of action is increasingly important. Here we reveal linezolid, an oxazolidinone antibiotic that inhibits microbial translation, is also herbicidal. To validate the herbicidal mode of action of linezolid we confirmed its micromolar inhibition is specific to chloroplast translation and did not affect photosynthesis directly. To assess the herbicide potential of linezolid, testing against a range of weed and crop species found it effective pre- and post-emergence. Using structure-activity analysis we identified the critical elements for herbicidal activity, but importantly also show, using antimicrobial susceptibility assays, that separation of antibacterial and herbicidal activities was possible. Overall these results validate chloroplast translation as a viable herbicidal target.