Bipolar outflows out to 10 kpc for massive galaxies at redshift z ≈ 1.

Galactic outflows are believed to play a critical role in the evolution of galaxies by regulating their mass build-up and star formation1. Theoretical models assume bipolar shapes for the outflows that extend well into the circumgalactic medium (CGM), up to tens of kiloparsecs (kpc) perpendicular to the galaxies. They have been directly observed in the local Universe in several individual galaxies, for example, around the Milky Way and M82 (refs. 2,3). At higher redshifts, cosmological simulations of galaxy formation predict an increase in the frequency and efficiency of galactic outflows owing to the increasing star-formation activity4. Galactic outflows are usually of low gas density and low surface brightness and therefore difficult to observe in emission towards high redshifts. Here we present an ultra-deep Multi-Unit Spectroscopic Explorer (MUSE) image of the mean Mg II emission surrounding a sample of galaxies at z ≈ 1 that strongly suggests the presence of outflowing gas on physical scales of more than 10 kpc. We find a strong dependence of the detected signal on the inclination of the central galaxy, with edge-on galaxies clearly showing enhanced Mg II emission along the minor axis, whereas face-on galaxies show much weaker and more isotropic emission. We interpret these findings as supporting the idea that outflows typically have a bipolar cone geometry perpendicular to the galactic disk. We demonstrate that this CGM-scale outflow is prevalent among galaxies with stellar mass M* ≳ 109.5M⊙.

In vivo transcriptomic profiling using cell encapsulation identifies effector pathways of systemic aging.

Sustained exposure to a young systemic environment rejuvenates aged organisms and promotes cellular function. However, due to the intrinsic complexity of tissues it remains challenging to pinpoint niche-independent effects of circulating factors on specific cell populations. Here, we describe a method for the encapsulation of human and mouse skeletal muscle progenitors in diffusible polyethersulfone hollow fiber capsules that can be used to profile systemic aging in vivo independent of heterogeneous short-range tissue interactions. We observed that circulating long-range signaling factors in the old systemic environment lead to an activation of Myc and E2F transcription factors, induce senescence, and suppress myogenic differentiation. Importantly, in vitro profiling using young and old serum in 2D culture does not capture all pathways deregulated in encapsulated cells in aged mice. Thus, in vivo transcriptomic profiling using cell encapsulation allows for the characterization of effector pathways of systemic aging with unparalleled accuracy.

AXR1 affects DNA methylation independently of its role in regulating meiotic crossover localization.

Meiotic crossovers (COs) are important for reshuffling genetic information between homologous chromosomes and they are essential for their correct segregation. COs are unevenly distributed along chromosomes and the underlying mechanisms controlling CO localization are not well understood. We previously showed that meiotic COs are mis-localized in the absence of AXR1, an enzyme involved in the neddylation/rubylation protein modification pathway in Arabidopsis thaliana. Here, we report that in axr1-/-, male meiocytes show a strong defect in chromosome pairing whereas the formation of the telomere bouquet is not affected. COs are also redistributed towards subtelomeric chromosomal ends where they frequently form clusters, in contrast to large central regions depleted in recombination. The CO suppressed regions correlate with DNA hypermethylation of transposable elements (TEs) in the CHH context in axr1-/- meiocytes. Through examining somatic methylomes, we found axr1-/- affects DNA methylation in a plant, causing hypermethylation in all sequence contexts (CG, CHG and CHH) in TEs. Impairment of the main pathways involved in DNA methylation is epistatic over axr1-/- for DNA methylation in somatic cells but does not restore regular chromosome segregation during meiosis. Collectively, our findings reveal that the neddylation pathway not only regulates hormonal perception and CO distribution but is also, directly or indirectly, a major limiting pathway of TE DNA methylation in somatic cells.

Melatonin Levels and Low-Frequency Magnetic Fields in Humans and Rats: New Insights From a Bayesian Logistic Regression.

The present analysis revisits the impact of extremely low-frequency magnetic fields (ELF-MF) on melatonin (MLT) levels in human and rat subjects using both a parametric and non-parametric approach. In this analysis, we use 62 studies from review articles. The parametric approach consists of a Bayesian logistic regression (LR) analysis and the non-parametric approach consists of a Support Vector analysis, both of which are robust against spurious/false results. Both approaches reveal a unique well-ordered pattern, and show that human and rat studies are consistent with each other once the MF strength is restricted to cover the same range (with B ≲ 50 µT). In addition, the data reveal that chronic exposure (longer than ∼22 days) to ELF-MF appears to decrease MLT levels only when the MF strength is below a threshold of ~30 µT ( log B thr [ µ T ] = 1 . 4 - 0 . 4 + 0 . 7 ), i.e., when the man-made ELF-MF intensity is below that of the static geomagnetic field. Studies reporting an association between ELF-MF and changes to MLT levels and the opposite (no association with ELF-MF) can be reconciled under a single framework. Bioelectromagnetics. 2019;40:539-552. © 2019 Bioelectromagnetics Society.

Post-transcriptional gene silencing triggers dispensable DNA methylation in gene body in Arabidopsis.

Spontaneous post-transcriptional silencing of sense transgenes (S-PTGS) is established in each generation and is accompanied by DNA methylation, but the pathway of PTGS-dependent DNA methylation is unknown and so is its role. Here we show that CHH and CHG methylation coincides spatially and temporally with RDR6-dependent products derived from the central and 3' regions of the coding sequence, and requires the components of the RNA-directed DNA methylation (RdDM) pathway NRPE1, DRD1 and DRM2, but not CLSY1, NRPD1, RDR2 or DCL3, suggesting that RDR6-dependent products, namely long dsRNAs and/or siRNAs, trigger PTGS-dependent DNA methylation. Nevertheless, none of these RdDM components are required to establish S-PTGS or produce a systemic silencing signal. Moreover, preventing de novo DNA methylation in non-silenced transgenic tissues grafted onto homologous silenced tissues does not inhibit the triggering of PTGS. Overall, these data indicate that gene body DNA methylation is a consequence, not a cause, of PTGS, and rule out the hypothesis that a PTGS-associated DNA methylation signal is transmitted independent of a PTGS signal.

Cell Mass Increase Associated with Formation of Glucose-Controlling ß-Cell Mass in Device-Encapsulated Implants of hiPS-Derived Pancreatic Endoderm.

Device-encapsulated human stem cell-derived pancreatic endoderm (PE) can generate functional ß-cell implants in the subcutis of mice, which has led to the start of clinical studies in type 1 diabetes. Assessment of the formed functional ß-cell mass (FBM) and its correlation with in vivo metabolic markers can guide clinical translation. We recently reported ex vivo characteristics of device-encapsulated human embryonic stem cell-derived (hES)-PE implants in mice that had established a metabolically adequate FBM during 50-week follow-up. Cell suspensions from retrieved implants indicated a correlation with the number of formed ß cells and their maturation to a functional state comparable to human pancreatic ß cells. Variability in metabolic outcome was attributed to differences in number of PE-generated ß cells. This variability hinders studies on processes involved in FBM-formation. This study reports modifications that reduce variability. It is undertaken with device-encapsulated human induced pluripotent stem cell-derived-PE subcutaneously implanted in mice. Cell mass of each cell type was determined on intact tissue inside the device to obtain more precise data than following isolation and dispersion. Implants in a preformed pouch generated a glucose-controlling ß-cell mass within 20 weeks in over 60% of recipients versus less than 20% in the absence of a pouch, whether the same or threefold higher cell dose had been inserted. In situ analysis of implants indicated a role for pancreatic progenitor cell expansion and endocrine differentiation in achieving the size of ß- and α-cell mass that correlated with in vivo markers of metabolic control. Stem Cells Translational Medicine 2019;8:1296&1305.

Macroencapsulated Human iPSC-Derived Pancreatic Progenitors Protect against STZ-Induced Hyperglycemia in Mice.

In type 1 diabetes, a renewable source of human pancreatic ß cells, in particular from human induced pluripotent stem cell (hiPSC) origin, would greatly benefit cell therapy. Earlier work showed that pancreatic progenitors differentiated from human embryonic stem cells in vitro can further mature to become glucose responsive following macroencapsulation and transplantation in mice. Here we took a similar approach optimizing the generation of pancreatic progenitors from hiPSCs. This work demonstrates that hiPSCs differentiated to pancreatic endoderm in vitro can be efficiently and robustly generated under large-scale conditions. The hiPSC-derived pancreatic endoderm cells (HiPECs) can further differentiate into glucose-responsive islet-like cells following macroencapsulation and in vivo implantation. The HiPECs can protect mice from streptozotocin-induced hyperglycemia and maintain normal glucose homeostasis and equilibrated plasma glucose concentrations at levels similar to the human set point. These results further validate the potential use of hiPSC-derived islet cells for application in clinical settings.

Antagonistic Actions of FPA and IBM2 Regulate Transcript Processing from Genes Containing Heterochromatin.

Repressive epigenetic marks, such as DNA and histone methylation, are sometimes located within introns. In Arabidopsis (Arabidopsis thaliana), INCREASE IN BONSAI METHYLATION2 (IBM2), an RNA-binding protein containing a bromo-adjacent homology domain, is required to process functional transcript isoforms of genes carrying intronic heterochromatin. In a genetic screen for suppressors of the ibm2 mutation, we identified FPA, an RNA-binding protein that promotes use of proximal polyadenylation sites in genes targeted by IBM2, including IBM1 encoding an essential H3K9 histone demethylase and the disease resistance gene RECOGNITION OF PERONOSPORA PARASITICA7 Both IBM2 and FPA are involved in the processing of their common mRNA targets: Transcription of IBM2 target genes is restored when FPA is mutated in ibm2 and impaired in transgenic plants overexpressing FPA By contrast, transposons targeted by IBM2 and localized outside introns are not under this antagonistic control. The DNA methylation patterns of some genes and transposons are modified in fpa plants, including the large intron of IBM1, but these changes are rather limited and reversed when the mutant is complemented, indicating that FPA has a restricted role in mediating silencing. These data reveal a complex regulation by IBM2 and FPA pathways in processing mRNAs of genes bearing heterochromatic marks.

Redistribution of CHH Methylation and Small Interfering RNAs across the Genome of Tomato ddm1 Mutants.

In plants, cytosine methylation, an epigenetic mark critical for transposon silencing, is maintained over generations by key enzymes that directly methylate DNA and is facilitated by chromatin remodelers, like DECREASE IN DNA METHYLATION1 (DDM1). Short-interfering RNAs (siRNAs) also mediate transposon DNA methylation through a process called RNA-directed DNA methylation (RdDM). In tomato (Solanum lycopersicum), siRNAs are primarily mapped to gene-rich chromosome arms, and not to pericentromeric regions as in Arabidopsis thaliana Tomato encodes two DDM1 genes. To better understand their functions and interaction with the RdDM pathway, we targeted the corresponding genes via the CRISPR/Cas9 technology, resulting in the isolation of Slddm1a and Slddm1b knockout mutants. Unlike the single mutants, Slddm1a Slddm1b double mutant plants display pleiotropic vegetative and reproductive phenotypes, associated with severe hypomethylation of the heterochromatic transposons in both the CG and CHG methylation contexts. The methylation in the CHH context increased for some heterochromatic transposons and conversely decreased for others localized in euchromatin. We found that the number of heterochromatin-associated siRNAs, including RdDM-specific small RNAs, increased significantly, likely limiting the transcriptional reactivation of transposons in Slddm1a Slddm1b Taken together, we propose that the global production of siRNAs and the CHH methylation mediated by the RdDM pathway are restricted to chromosome arms in tomato. Our data suggest that both pathways are greatly enhanced in heterochromatin when DDM1 functions are lost, at the expense of silencing mechanisms normally occurring in euchromatin.

Encapsulated Optically Responsive Cell Systems: Toward Smart Implants in Biomedicine.

Managing increasingly prevalent chronic diseases will require close continuous monitoring of patients. Cell-based biosensors may be used for implantable diagnostic systems to monitor health status. Cells are indeed natural sensors in the body. Functional cellular systems can be maintained in the body for long-term implantation using cell encapsulation technology. By taking advantage of recent progress in miniaturized optoelectronic systems, the genetic engineering of optically responsive cells may be combined with cell encapsulation to generate smart implantable cell-based sensing systems. In biomedical research, cell-based biosensors may be used to study cell signaling, therapeutic effects, and dosing of bioactive molecules in preclinical models. Today, a wide variety of genetically encoded fluorescent sensors have been developed for real-time imaging of living cells. Here, recent developments in genetically encoded sensors, cell encapsulation, and ultrasmall optical systems are highlighted. The integration of these components in a new generation of biosensors is creating innovative smart in vivo cell-based systems, bringing novel perspectives for biomedical research and ultimately allowing unique health monitoring applications.

An Arabidopsis Natural Epiallele Maintained by a Feed-Forward Silencing Loop between Histone and DNA.

The extent of epigenetic variation is currently well documented, but the number of natural epialleles described so far remains very limited. Determining the relevance of epigenetic changes for natural variation is an important question of research that we investigate by isolating natural epialleles segregating in Arabidopsis recombinant populations. We previously described a genetic incompatibility among Arabidopsis strains based on the silencing of a gene involved in fitness. Here, we isolated a new epiallele resulting from the silencing of a transfer-RNA editing gene in an Arabidopsis accession from the Netherlands (Nok-1). Crosses with the reference accession Col-0 show a complete incompatibility between this epiallele and another locus localized on a different chromosome. We demonstrate that conversion of an unmethylated version of this allele occurs in hybrids, associated with modifications of small RNA populations. These epialleles can also spontaneously revert within the population. Furthermore, we bring evidence that neither METHYLTRANSFERASE 1, maintaining methylation at CGs, nor components of RNA-directed DNA methylation, are key factors for the transmission of the epiallele over generations. This depends only on the self-reinforcing loop between CHROMOMETHYLASE 3 and KRYPTONITE, involving DNA methylated in the CHG context and histone H3 lysine 9 methylation. Our findings reveal a predominant role of this loop in maintaining a natural epiallele.

Encapsulation of Insulin-Secreting Cells Expressing a Genetically Encoded Fluorescent Calcium Indicator for Cell-Based Sensing In Vivo.

The development of cell-based biosensors that give insight into cell and tissue function in vivo is an attractive technology for biomedical research. Here, the development of a cell line expressing a fluorescent calcium sensor for the study of beta-cell function in vivo is reported. The bioresponsive cell model is based on INS-1E pancreatic beta-cells, stably expressing the genetically encoded cameleon-based fluorescent sensor YC3.6cyto . Following single-cell selection and expansion, functional testing and in vitro encapsulation experiments are used to identify a suitable clone of INS-1E cells expressing the calcium sensor. This clone is transplanted subcutaneous in mouse using a cell macroencapsulation system based on flat sheet porous membranes. Cells in the implanted capsules are able to respond to glucose in vivo by secreting insulin and thereby contributing to the regulation of glycaemia in the mice. Furthermore, fluorescence imaging of explanted devices shows that encapsulated cells maintain high level expression of YC3.6cyto in vivo. In conclusion, these data show that encapsulated INS-1E cells stably expressing a genetically encoded calcium sensor can be successfully implanted in vivo, and therefore serve as biosensing element or in vivo model to longitudinally monitor the function of pancreatic beta-cells.

The Scientific Challenge of Expanding the Frontiers of Nutrition.

Nutritional research is entering a paradigm shift which necessitates the modeling of complex interactions between diet, genetics, lifestyle, and environmental factors. This requires the development of analytical and processing capabilities for multiple data and information sources to be able to improve targeted and personalized nutritional approaches for the maintenance of health. Ideally, such knowledge will be employed to underpin the development of concepts that combine consumer and medical nutrition with diagnostic targeting for early intervention designed to maintain proper metabolic homeostasis and delay the onset of chronic diseases. Nutritional status is fundamental to any description of health, and when combined with other data on lifestyle, environment, and genetics, it can be used to drive stratified or even personalized nutritional strategies for health maintenance and preventive medicine. In this work, we will discuss the importance of developing new nutrient assessment methods and diagnostic capabilities for nutritional status to generate scientific hypotheses and actionable concepts from which to develop targeted and eventually personalized nutritional solutions for health protection. We describe efforts to develop algorithms for dietary nutrient intake and a holistic nutritional profiling platform as the basis of understanding the complex nutrition and health interactome.

Post-transcriptional gene silencing triggered by sense transgenes involves uncapped antisense RNA and differs from silencing intentionally triggered by antisense transgenes.

Although post-transcriptional gene silencing (PTGS) has been studied for more than a decade, there is still a gap in our understanding of how de novo silencing is initiated against genetic elements that are not supposed to produce double-stranded (ds)RNA. Given the pervasive transcription occurring throughout eukaryote genomes, we tested the hypothesis that unintended transcription could produce antisense (as)RNA molecules that participate to the initiation of PTGS triggered by sense transgenes (S-PTGS). Our results reveal a higher level of asRNA in Arabidopsis thaliana lines that spontaneously trigger S-PTGS than in lines that do not. However, PTGS triggered by antisense transgenes (AS-PTGS) differs from S-PTGS. In particular, a hypomorphic ago1 mutation that suppresses S-PTGS prevents the degradation of asRNA but not sense RNA during AS-PTGS, suggesting a different treatment of coding and non-coding RNA by AGO1, likely because of AGO1 association to polysomes. Moreover, the intended asRNA produced during AS-PTGS is capped whereas the asRNA produced during S-PTGS derives from 3' maturation of a read-through transcript and is uncapped. Thus, we propose that uncapped asRNA corresponds to the aberrant RNA molecule that is converted to dsRNA by RNA-DEPENDENT RNA POLYMERASE 6 in siRNA-bodies to initiate S-PTGS, whereas capped asRNA must anneal with sense RNA to produce dsRNA that initiate AS-PTGS.

Interplay between chromatin and RNA processing.

The processing of pre-mRNAs, including the selection of polyadenylation sites, is influenced by the surrounding chromatin context. We review here recent studies in Arabidopsis thaliana highlighting the intricate and reciprocal interplay between chromatin state and RNA processing. The studies have revealed that transcription can be influenced by the presence, in gene introns, of combination of epigenetic marks typical of heterochromatin. New factors binding to these marks have been identified and shown to play key roles in controlling the use of polyadenylation sites and processing of functional mRNAs. Concomitantly, several proteins of both the splicing and the polyadenylation machineries are also emerging as regulators of DNA methylation patterns and chromatin silencing.

A non-canonical plant microRNA target site.

Plant microRNAs (miRNAs) typically form near-perfect duplexes with their targets and mediate mRNA cleavage. Here, we describe an unconventional miRNA target of miR398 in Arabidopsis, an mRNA encoding the blue copper-binding protein (BCBP). BCBP mRNA carries an miR398 complementary site in its 5'-untranslated region (UTR) with a bulge of six nucleotides opposite to the 5' region of the miRNA. Despite the disruption of a target site region thought to be especially critical for function, BCBP mRNAs are cleaved by ARGONAUTE1 between nucleotides 10th and 11th, opposite to the miRNA, like conventional plant target sites. Levels of BCBP mRNAs are inversely correlated to levels of miR398 in mutants lacking the miRNA, or transgenic plants overexpressing it. Introducing two mutations that disrupt the miRNA complementarity around the cleavage site renders the target cleavage-resistant. The BCBP site functions outside of the context of the BCBP mRNA and does not depend on 5'-UTR location. Reducing the bulge does not interfere with miR398-mediated regulation and completely removing it increases the efficiency of the slicing. Analysis of degradome data and target predictions revealed that the miR398-BCBP interaction seems to be rather unique. Nevertheless, our results imply that functional target sites with non-perfect pairings in the 5' region of an ancient conserved miRNA exist in plants.

SHOOT GROWTH1 maintains Arabidopsis epigenomes by regulating IBM1.

Maintaining correct DNA and histone methylation patterns is essential for the development of all eukaryotes. In Arabidopsis, we identified SHOOT GROWTH1 (SG1), a novel protein involved in the control of gene methylation. SG1 contains both a Bromo-Adjacent Homology (BAH) domain found in several chromatin regulators and an RNA-Recognition Motif (RRM). The sg1 mutations are associated with drastic pleiotropic phenotypes. The mutants degenerate after few generations and are similar to mutants of the histone demethylase INCREASE IN BONSAI METHYLATION1 (IBM1). A methylome analysis of sg1 mutants revealed a large number of gene bodies hypermethylated in the cytosine CHG context, associated with an increase in di-methylation of lysine 9 on histone H3 tail (H3K9me2), an epigenetic mark normally found in silenced transposons. The sg1 phenotype is suppressed by mutations in genes encoding the DNA methyltransferase CHROMOMETHYLASE3 (CMT3) or the histone methyltransferase KRYPTONITE (KYP), indicating that SG1 functions antagonistically to CMT3 or KYP. We further show that the IBM1 transcript is not correctly processed in sg1, and that the functional IBM1 transcript complements sg1. Altogether, our results suggest a function for SG1 in the maintenance of genome integrity by regulating IBM1.

Rapid establishment of genetic incompatibility through natural epigenetic variation.

Epigenetic variation is currently being investigated with the aim of deciphering its importance in both adaptation and evolution [1]. In plants, epimutations can underlie heritable phenotypic diversity [2-4], and epigenetic mechanisms might contribute to reproductive barriers between [5] or within species [6]. The extent of epigenetic variation begins to be appreciated in Arabidopsis [7], but the origin of natural epialleles and their impact in the wild remain largely unknown. Here we show that a genetic incompatibility among Arabidopsis thaliana strains is related to the epigenetic control of a pair of duplicate genes involved in fitness: a transposition event results in a rearranged paralogous structure that causes DNA methylation and transcriptional silencing of the other copy. We further show that this natural, strain-specific epiallele is stable over numerous generations even after removal of the duplicated, rearranged gene copy through crosses. Finally, we provide evidence that the rearranged gene copy triggers de novo DNA methylation and silencing of the unlinked native gene by RNA-directed DNA methylation. Our findings suggest an important role of naturally occurring epialleles originating from structural variation in rapidly establishing genetic incompatibilities following gene duplication events.

Chronic delivery of antibody fragments using immunoisolated cell implants as a passive vaccination tool.

BACKGROUND: Monoclonal antibodies and antibody fragments are powerful biotherapeutics for various debilitating diseases. However, high production costs, functional limitations such as inadequate pharmacokinetics and tissue accessibility are the current principal disadvantages for broadening their use in clinic. METHODOLOGY AND PRINCIPAL FINDINGS: We report a novel method for the long-term delivery of antibody fragments. We designed an allogenous immunoisolated implant consisting of polymer encapsulated myoblasts engineered to chronically release scFv antibodies targeted against the N-terminus of the Aß peptide. Following a 6-month intracerebral therapy we observed a significant reduction of the production and aggregation of the Aß peptide in the APP23 transgenic mouse model of Alzheimer's disease. In addition, functional assessment showed prevention of behavioral deficits related to anxiety and memory traits. CONCLUSIONS AND SIGNIFICANCE: The chronic local release of antibodies using immunoisolated polymer cell implants represents an alternative passive vaccination strategy in Alzheimer's disease. This novel technique could potentially benefit other diseases presently treated by local and systemic antibody administration.

New insights into miR398 functions in Arabidopsis.

We recently identified a new target of microRNA398 (miR398), a conserved miRNA in plants. In Arabidopsis, miR398 targets the mRNAs of two copper/zinc superoxide dismutases (Cu/Zn SODs) by triggering their cleavage or repressing their translation. We analysed the transcriptomes of mutants impaired in miR398 production, revealing that the mRNAs encoding the chaperone (CCS1), essential for copper delivering to the Cu/Zn SODs of Arabidopsis and to generate the mature proteins, were undiscovered targets of miR398. It is likely that CCS1 was not identified by previous bioinformatic predictions because of the number of mismatches between the mRNA and its target. Since CCS1 has four mismatches and one GU wobble, it would have been excluded by the majority of prediction algorithms. miR398 directs the post-transcriptional regulation of CCS1 mRNAs by cleavage and ARGONAUTE10 (AGO10)-mediated translational repression. Indeed, CCS1 protein accumulate in zwille (ago10) mutants while both miR398 and CCS1 mRNAs levels remain identical to the Landsberg erecta WT plants. Moreover, since AGO10 is a negative regulator of AGO1, the CCS1 protein is more abundant in a double ago1-27 ago10-3 Col mutant compared to the single hypomorphic ago1-27 mutant, as previously shown for CSD2.