Impaired DNA demethylation of C/EBP sites causes premature aging.

Changes in DNA methylation are among the best-documented epigenetic alterations accompanying organismal aging. However, whether and how altered DNA methylation is causally involved in aging have remained elusive. GADD45α (growth arrest and DNA damage protein 45A) and ING1 (inhibitor of growth family member 1) are adapter proteins for site-specific demethylation by TET (ten-eleven translocation) methylcytosine dioxygenases. Here we show that Gadd45a/Ing1 double-knockout mice display segmental progeria and phenocopy impaired energy homeostasis and lipodystrophy characteristic of Cebp (CCAAT/enhancer-binding protein) mutants. Correspondingly, GADD45α occupies C/EBPß/δ-dependent superenhancers and, cooperatively with ING1, promotes local DNA demethylation via long-range chromatin loops to permit C/EBPß recruitment. The results indicate that enhancer methylation can affect aging and imply that C/EBP proteins play an unexpected role in this process. Our study suggests a causal nexus between DNA demethylation, metabolism, and organismal aging.

Flagellin hypervariable region determines symbiotic properties of commensal Escherichia coli strains.

Escherichia coli represents a classical intestinal gram-negative commensal. Despite this commensalism, different E. coli strains can mediate disparate immunogenic properties in a given host. Symbiotic E. coli strains such as E. coli Nissle 1917 (EcN) are attributed beneficial properties, e.g., promotion of intestinal homeostasis. Therefore, we aimed to identify molecular features derived from symbiotic bacteria that might help to develop innovative therapeutic alternatives for the treatment of intestinal immune disorders. This study was performed using the dextran sodium sulphate (DSS)-induced colitis mouse model, which is routinely used to evaluate potential therapeutics for the treatment of Inflammatory Bowel Diseases (IBDs). We focused on the analysis of flagellin structures of different E. coli strains. EcN flagellin was found to harbor a substantially longer hypervariable region (HVR) compared to other commensal E. coli strains, and this longer HVR mediated symbiotic properties through stronger activation of Toll-like receptor (TLR)5, thereby resulting in interleukin (IL)-22-mediated protection of mice against DSS-induced colitis. Furthermore, using bone-marrow-chimeric mice (BMCM), CD11c+ cells of the colonic lamina propria (LP) were identified as the main mediators of these flagellin-induced symbiotic effects. We propose flagellin from symbiotic E. coli strains as a potential therapeutic to restore intestinal immune homeostasis, e.g., for the treatment of IBD patients.

Energy Barriers for Steroid Hormone Transport in Nanofiltration.

Nanofiltration (NF) membranes can retain micropollutants (MPs) to a large extent, even though adsorption into the membrane and gradual permeation result in breakthrough and incomplete removal. The permeation of MPs is investigated by examining the energy barriers (determined using the Arrhenius concept) for adsorption, intrapore diffusion, and permeation encountered by four different steroid hormones in tight and loose NF membranes. Results show that the energy barriers for steroid hormone transport in tight membrane are entropically dominated and underestimated because of the high steric exclusion at the pore entrance. In contrast, the loose NF membrane enables steroid hormones partitioning at the pore entrance, with a permeation energy barrier (from feed toward the permeate side) ranging between 96 and 116 kJ/mol. The contribution of adsorption and intrapore diffusion to the energy barrier for steroid hormone permeation reveals a significant role of intrapore diffusive transport on the obtained permeation energy barrier. Overall, the breakthrough phenomenon observed during the NF of MPs is facilitated by the low energy barrier for adsorption. Experimental evidence of such principles is relevant for understanding mechanisms and ultimately improving the selectivity of NF.

Long noncoding RNA TARID directs demethylation and activation of the tumor suppressor TCF21 via GADD45A.

DNA methylation is a dynamic and reversible process that governs gene expression during development and disease. Several examples of active DNA demethylation have been documented, involving genome-wide and gene-specific DNA demethylation. How demethylating enzymes are targeted to specific genomic loci remains largely unknown. We show that an antisense lncRNA, termed TARID (for TCF21 antisense RNA inducing demethylation), activates TCF21 expression by inducing promoter demethylation. TARID interacts with both the TCF21 promoter and GADD45A (growth arrest and DNA-damage-inducible, alpha), a regulator of DNA demethylation. GADD45A in turn recruits thymine-DNA glycosylase for base excision repair-mediated demethylation involving oxidation of 5-methylcytosine to 5-hydroxymethylcytosine in the TCF21 promoter by ten-eleven translocation methylcytosine dioxygenase proteins. The results reveal a function of lncRNAs, serving as a genomic address label for GADD45A-mediated demethylation of specific target genes.

Gadd45α modulates aversive learning through post-transcriptional regulation of memory-related mRNAs.

Learning is essential for survival and is controlled by complex molecular mechanisms including regulation of newly synthesized mRNAs that are required to modify synaptic functions. Despite the well-known role of RNA-binding proteins (RBPs) in mRNA functionality, their detailed regulation during memory consolidation is poorly understood. This study focuses on the brain function of the RBP Gadd45α (growth arrest and DNA damage-inducible protein 45 alpha, encoded by the Gadd45a gene). Here, we find that hippocampal memory and long-term potentiation are strongly impaired in Gadd45a-deficient mice, a phenotype accompanied by reduced levels of memory-related mRNAs. The majority of the Gadd45α-regulated transcripts show unusually long 3' untranslated regions (3'UTRs) that are destabilized in Gadd45a-deficient mice via a transcription-independent mechanism, leading to reduced levels of the corresponding proteins in synaptosomes. Moreover, Gadd45α can bind specifically to these memory-related mRNAs. Our study reveals a new function for extended 3'UTRs in memory consolidation and identifies Gadd45α as a novel regulator of mRNA stability.

Ing1 functions in DNA demethylation by directing Gadd45a to H3K4me3.

Active DNA demethylation regulates epigenetic gene activation in numerous processes, but how the target site specificity of DNA demethylation is determined and what factors are involved are still poorly understood. Here we show that the tumor suppressor inhibitor of growth protein 1 (Ing1) is required for targeting active DNA demethylation. Ing1 functions by recruiting the regulator of DNA demethylation growth arrest and DNA damage protein 45a (Gadd45a) to histone H3 trimethylated at Lys 4 (H3K4me3). We show that reduced H3K4 methylation impairs recruitment of Gadd45a/Ing1 and gene-specific DNA demethylation. Our results indicate that histone methylation directs DNA demethylation.

Weak Agonistic LPS Restores Intestinal Immune Homeostasis.

Generated by gram-negative bacteria, lipopolysaccharides (LPSs) are one of the most abundant and potent immunomodulatory substances present in the intestinal lumen. Interaction of agonistic LPS with the host myeloid-differentiation-2/Toll-like receptor 4 (MD-2/TLR4) receptor complex results in nuclear factor κB (NF-κB) activation, followed by the robust induction of pro-inflammatory immune responses. Here we have isolated LPS from a common gut commensal, Bacteroides vulgatus mpk (BVMPK), which provides only weak agonistic activity. This weak agonistic activity leads to the amelioration of inflammatory immune responses in a mouse model for experimental colitis, and it was in sharp contrast to strong agonists and antagonists. In this context, the administration of BVMPK LPS into mice with severe intestinal inflammation re-established intestinal immune homeostasis within only 2 weeks, resulting in the clearance of all symptoms of inflammation. These inflammation-reducing properties of weak agonistic LPS are grounded in the induction of a special type of endotoxin tolerance via the MD-2/TLR4 receptor complex axis in intestinal lamina propria CD11c+ cells. Thus, weak agonistic LPS represents a promising agent to treat diseases involving pathological overactivation of the intestinal immune system, e.g., in inflammatory bowel diseases.

Recycled and desalinated water: Consumers' associations, and the influence of affect and disgust on willingness to use.

Traditional water sources are increasingly coming under pressure from urbanization, population and industrial growth as well as climate variability and alternatives such as desalinated and recycled water will need to be considered in the future. However, available data indicate that consumers are apprehensive about these alternative water sources, with many reactions centered on disgust. It is not clear though, whether trait or state disgust is responsible for these disgust reactions. Secondly, the associations the public has with desalinated and recycled water are not well described, even though knowledge of these associations would provide the holistic understanding required for addressing perceptions. Using respondents from Australia and South Africa, therefore, this study investigated the associations people have with recycled and desalinated water, as well as the influence of affect, institutional trust, cultural/religious beliefs, water scarcity, and demographic variables, on willingness to use (WTU) these waters. The results show that whereas recycled water is associated with negative affect from notions of disgust, desalinated water is often thought of in terms of minerals, or as "simply water". We also found that affect was the most significant predictor, and that state disgust as well as trait disgust influenced WTU. Together these data suggest that disgust-related rejection of is likely due to the disgust elicited by the waters rather than disgust-proneness of individuals. It is plausible, therefore, that efforts to induce positive affect through making salient positive associations e.g. NEWater in Singapore, may increase WTU.

TAF12 recruits Gadd45a and the nucleotide excision repair complex to the promoter of rRNA genes leading to active DNA demethylation.

Many studies have detailed the repressive effects of DNA methylation on gene expression. However, the mechanisms that promote active demethylation are just beginning to emerge. Here, we show that methylation of the rDNA promoter is a dynamic and reversible process. Demethylation of rDNA is initiated by recruitment of Gadd45a (growth arrest and DNA damage inducible protein 45 alpha) to the rDNA promoter by TAF12, a TBP-associated factor that is contained in Pol I- and Pol II-specific TBP-TAF complexes. Once targeted to rDNA, Gadd45a triggers demethylation of promoter-proximal DNA by recruiting the nucleotide excision repair (NER) machinery to remove methylated cytosines. Knockdown of Gadd45a, XPA, XPG, XPF, or TAF12 or treatment with drugs that inhibit NER causes hypermethylation of rDNA, establishes heterochromatic histone marks, and impairs transcription. The results reveal a mechanism that recruits the DNA repair machinery to the promoter of active genes, keeping them in a hypomethylated state.

Symbiotic gut commensal bacteria act as host cathepsin S activity regulators.

Cathepsin S (CTSS) is a lysosomal protease whose activity regulation is important for MHC-II signaling and subsequent activation of CD4+ T cell mediated immune responses. Dysregulation of its enzymatic activity or enhanced secretion into extracellular environments is associated with the induction or progression of several autoimmune diseases. Here we demonstrate that commensal intestinal bacteria influence secretion rates and intracellular activity of host CTSS and that symbiotic bacteria, i.e. Bacteroides vulgatus mpk, may actively regulate this process and help to maintain physiological levels of CTSS activities in order to prevent from induction of pathological inflammation. The symbiont-controlled regulation of CTSS activity is mediated by anticipating reactive oxygen species induction in dendritic cells which, in turn, maintains cystatin C (CysC) monomer binding to CTSS. CysC monomers are potent endogenous CTSS inhibitors. This Bacteroides vulgatus caused and CysC dependent CTSS activity regulation is involved in the generation of tolerant intestinal dendritic cells contributing to prevention of T-cell mediated induction of colonic inflammation. Taken together, we demonstrate that symbionts of the intestinal microbiota regulate host CTSS activity and secretion and might therefore be an attractive approach to deal with CTSS associated autoimmune diseases.

GADD45a physically and functionally interacts with TET1.

DNA demethylation plays a central role during development and in adult physiology. Different mechanisms of active DNA demethylation have been established. For example, Growth Arrest and DNA Damage 45-(GADD45) and Ten-Eleven-Translocation (TET) proteins act in active DNA demethylation but their functional relationship is unresolved. Here we show that GADD45a physically interacts--and functionally cooperates with TET1 in methylcytosine (mC) processing. In reporter demethylation GADD45a requires endogenous TET1 and conversely TET1 requires GADD45a. On GADD45a target genes TET1 hyperinduces 5-hydroxymethylcytosine (hmC) in the presence of GADD45a, while 5-formyl-(fC) and 5-carboxylcytosine (caC) are reduced. Likewise, in global analysis GADD45a positively regulates TET1 mediated mC oxidation and enhances fC/caC removal. Our data suggest a dual function of GADD45a in oxidative DNA demethylation, to promote directly or indirectly TET1 activity and to enhance subsequent fC/caC removal.

Removal of glyphosate (GLY) and aminomethylphosphonic acid (AMPA) by ultrafiltration with permeate-side polymer-based spherical activated carbon (UF-PBSAC).

Glyphosate (GLY) is the most commonly used herbicide worldwide, and aminomethylphosphonic acid (AMPA) is one of its main metabolites. GLY and AMPA are toxic to humans, and their complex physicochemical properties present challenges in their removal from water. Several technologies have been applied to remove GLY and AMPA such as adsorption, filtration, and degradation with varied efficiencies. In previous works, an ultrafiltration membrane with permeate-side polymer-based spherical activated carbon (UF-PBSAC) showed the feasibility of removing uncharged micropollutants via adsorption in a flow-through configuration. The same UF-PBSAC was investigated for GLY and AMPA adsorption to assess the removal of charged and lower molecular weight micropollutants. The results indicated that both surface area and hydraulic residence time were limiting factors in GLY/AMPA adsorption by UF-PBSAC. The higher external surface of PBSAC with strong affinity for GLY and AMPA showed higher removal in a dynamic process where the hydraulic residence time was short (tens of seconds). Extending hydraulic residence times (hundreds of seconds) resulted in higher GLY/AMPA removal by allowing GLY/AMPA to diffuse into the PBSAC pores and reach more surfaces. Enhancement was achieved by minimising both limiting factors (external surface and hydraulic residence time) with a low flux of 25 L/m2.h, increased PBSAC layer of 6 mm, and small PBSAC particle size of 78 µm. With this configuration, UF-PBSAC could remove 98 % of GLY and 95 % of AMPA from an initial concentration of 1000 ng/L at pH 8.2 ± 0.2 and meet European Union (EU) regulation for herbicides (100 ng/L for individuals and 500 ng/L for total herbicides). The results implied that UF-PBSAC was able to remove charged micropollutants to the required levels and had potential for application in wastewater treatment and water reuse.

Tutorial review of error evaluation in experimental water research at the example of membrane filtration.

Experimental water research lacks clear methodology to estimate experimental error. Especially when natural waters are involved, the characterization tools bear method-specific artifacts while the varying environmental conditions prevent regular repeats. This tutorial review identifies common mistakes, and proposes a practical procedure to determine experimental errors at the example of membrane filtration. Statistical analysis is often applied to an insufficient number of repeated measurements, while not all error sources and contributions are considered. This results in an underestimation of the experimental error. Variations in relevant experimental parameters need to be investigated systematically, and the related errors are quantified as a half of the variation between the max and min values when standard deviation is not applicable. Error of calculated parameters (e.g. flux, pollutant removal and mass loss) is estimated by applying error propagation, where weighing contributions of the experimental parameters are considered. Appropriate judgment and five-time repetition of a selected experiment under identical conditions are proposed to validate the propagated experimental error. For validation, the five repeated data points should lie within the estimated error range of the error bar. The proposed error evaluation procedure is adaptable in experimental water research and intended for researchers to identify the contributing factors of an experimental error and carry out appropriate error quantification and validation. The most important aim is to raise awareness of the necessity to question error methodology and reproducibility of experimental data, to produce and publish high quality research.

Heterostructure g-C3N4/Bi2MoO6 PVDF nanofiber composite membrane for the photodegradation of steroid hormone micropollutants.

Photocatalytic membrane reactors (PMRs) are a promising technology for micropollutant removal. Sunlight utilization and catalyst surface sites limit photodegradation. A poly(vinylidene fluoride) (PVDF) nanofiber composite membrane (NCM) with immobilized visible-light-responsive g-C3N4/Bi2MoO6 (BMCN) were developed. Photodegradation of steroid hormones with the PVDF-BMCN NCM was investigated with varying catalyst properties, operating conditions, and relevant solution chemistry under solar irradiation. Increasing CN ratio (0-65 %) enhanced estradiol (E2) degradation from 20 ± 10 to 75 ± 7 % due to improved sunlight utilization and photon lifetime. PVDF nanofibers reduced self-aggregation of catalysts. Hydraulic residence time and light intensity enhanced the photodegradation. With the increasing pH value, the E2 removal decreased from 84 ± 4 to 67 ± 7 % owing to electrical repulsion and thus reduced adsorption between catalysts and E2. A removal of 96 % can be attained at environmentally relevant feed concentration (100 ng.L-1) with a flux of 60 L.m-2.h-1, irradiance of 100 mW.cm-2, and 1 mg.cm-2 BMCN65 loading. This confirmed that heterojunction photocatalysts can enhance micropollutants degradation in PMRs.

Removal of strontium by nanofiltration: Role of complexation and speciation of strontium with organic matter.

Strontium (Sr) removal from water is required because excessive naturally occurring Sr exposure is hazardous to human health. Climate and seasonal changes cause water quality variations, in particular quality and quantity of organic matter (OM) and pH, and such variations affect Sr removal by nanofiltration (NF). The mechanisms for such variations are not clear and thus OM complexation and speciation require attention. Sr removal by NF was investigated with emphasis on the role of OM (type and concentration) and pH (2-12) on possible removal mechanisms, specifically size and/or charge exclusion as well as solute-solute interactions. The filtration results show that the addition of various OM (10 types) and an increase of OM concentration (2-100 mgC.L-1) increased Sr removal by 10-15%. The Sr-OM interaction was enhanced with increasing OM concentration, implying enhanced size exclusion via Sr-OM interaction as the main mechanism. Such interactions were quantified by asymmetric flow field-flow fractionation (FFFF) coupled with an inductively coupled plasma mass spectrometer (ICP-MS). Both extremely low and high pH increased Sr removal due to the enhanced charge exclusion and Sr-OM interactions. This work elucidated and verified the mechanism of OM and pH on Sr removal by NF membranes.

Adsorption of uranium (VI) complexes with polymer-based spherical activated carbon.

Adsorption processes with carbon-based adsorbents have received substantial attention as a solution to remove uranium from drinking water. This study investigated uranium adsorption by a polymer-based spherical activated carbon (PBSAC) characterised by a uniformly smooth exterior and an extended surface of internal cavities accessible via mesopores. The static adsorption of uranium was investigated applying varying PBSAC properties and relevant solution chemistry. Spatial time-of-flight secondary ion mass spectrometry (ToF-SIMS) was employed to visualise the distribution of the different uranium species in the PBSAC. The isotherms and thermodynamics calculations revealed monolayer adsorption capacities of 28-667 mg/g and physical adsorption energies of 13-21 kJ/mol. Increasing the surface oxygen content of the PBSAC to 10 % enhanced the adsorption and reduced the equilibrium time to 2 h, while the WHO drinking water guideline of 30 µgU/L could be achieved for an initial concentration of 250 µgU/L. Uranium adsorption with PBSAC was favourable at the pH 6-8. At this pH range, uranyl carbonate complexes (UO2CO3(aq), UO2(CO3)22-, (UO2)2CO3(OH)3-) predominated in the solution, and the ToF-SIMS analysis revealed that the adsorption of these complexes occurred on the surface and inside the PBSAC due to intra-particle diffusion. For the uranyl cations (UO22+, UO2OH+) at pH 2-4, only shallow adsorption in the outermost PBSAC layers was observed. The work demonstrated the effective removal of uranium from contaminated natural water (67 µgU/L) and meeting both German (10 µgU/L) and WHO guideline concentrations. These findings also open opportunities to consider PBSAC in hybrid treatment technologies for uranium removal, for instance, from high-level radioactive waste.

Nitrogen compounds removal from brackish water by electrodialysis at fixed electric potential and dynamic current density operations.

Nitrogen (N) compounds can occur in water resources from natural and anthropogenic activities. It is ideal that these contaminants be removed before water consumption. As water quality has been affected by increased salinity and pH variation, more advanced and robust technologies such as electrodialysis (ED) can be considered for simultaneous desalination and pollutant removal. In this context, the removal of N-species (NO3-, NO2-, NH4+, and CH4N2O) from brackish water by ED was investigated for different feed water quality, considering increased salinity (0 - 10g/L NaCl) and pH variation (3 - 11), under limit current density (LCD) at fixed electric potential condition. The applied electric potential (5 - 25V) under, at, and over the LCD at fixed electric potential and dynamic current density (DCD), as a percentage of LCD (0.4 - 1.2), were analyzed to improve the process. In addition, energy efficiency in the form of specific energy consumption (SEC) and current efficiency (CE) were assessed for ED at fixed electric potential and DCD. The results showed that, at extreme pH of the feed water, the removal of NO2- and NH4+ can be affected, while NO3-was the most stable compound with pH variation. An increase in feed water salinity just slightly impacted the removal of N-compounds, due to the similar characteristics of the ions in the water. The increase in electric potential at fixed electric potential or DCD increased the removal and molar flux of N-compounds. However, operating over the LCD increased the SEC of the ED process while changes in removal were not significant. DCD procedures resulted in higher CE and shorter run time of the experiments. Therefore, ED proved to be a suitable treatment technique to produce fresh water due to the selective removal of the studied ions, especially at 15V (fixed electrical potential) and 0.8 LCD (DCD) related to removal, molar flux, and run time to achieve guidelines.

Interplay of the forces governing steroid hormone micropollutant adsorption in vertically-aligned carbon nanotube membrane nanopores.

Vertically-aligned carbon nanotube (VaCNT) membranes allow water to conduct rapidly at low pressures and open up the possibility for water purification and desalination, although the ultralow viscous stress in hydrophobic and low-tortuosity nanopores prevents surface interactions with contaminants. In this experimental investigation, steroid hormone micropollutant adsorption by VaCNT membranes is quantified and explained via the interplay of the hydrodynamic drag and friction forces acting on the hormone, and the adhesive and repulsive forces between the hormone and the inner carbon nanotube wall. It is concluded that a drag force above 2.2 × 10-3 pN overcomes the friction force resulting in insignificant adsorption, whereas lowering the drag force from 2.2 × 10-3 to 4.3 × 10-4 pN increases the adsorbed mass of hormones from zero to 0.4 ng cm-2. At a low drag force of 1.6 × 10-3 pN, the adsorbed mass of four hormones is correlated with the hormone-wall adhesive (van der Waals) force. These findings explain micropollutant adsorption in nanopores via the forces acting on the micropollutant along and perpendicular to the flow, which can be exploited for selectivity.

Experimental energy barriers to anions transporting through nanofiltration membranes.

Environmentally relevant contaminants fluoride, chloride, nitrate, and nitrite face Arrhenius energy barriers during transport through nanofiltration (NF) membranes. The energy barriers were quantified using crossflow filtration experiments and were in the range of 7-17 kcal·mol(-1), according to ion type and membrane type (Filmtec NF90 and NF270). Fluoride faced a comparatively high energy barrier for both membranes. This can be explained by the strong hydration energy of fluoride rather than other ion properties such as bare ion radius, fully hydrated radius, Stokes radius, diffusion coefficient, or ion charge. The energy barrier for fluoride decreased with pressure, indicating an impact of directional force on energy barriers. The influence of temperature-induced pore radius variability and viscosity on energy barriers was considered. The novel link between energy barriers and ion properties emphasizes the importance of ion hydration and/or partial dehydration mechanisms in determining transport in NF.

Gadd45a promotes epigenetic gene activation by repair-mediated DNA demethylation.

DNA methylation is an epigenetic modification that is essential for gene silencing and genome stability in many organisms. Although methyltransferases that promote DNA methylation are well characterized, the molecular mechanism underlying active DNA demethylation is poorly understood and controversial. Here we show that Gadd45a (growth arrest and DNA-damage-inducible protein 45 alpha), a nuclear protein involved in maintenance of genomic stability, DNA repair and suppression of cell growth, has a key role in active DNA demethylation. Gadd45a overexpression activates methylation-silenced reporter plasmids and promotes global DNA demethylation. Gadd45a knockdown silences gene expression and leads to DNA hypermethylation. During active demethylation of oct4 in Xenopus laevis oocytes, Gadd45a is specifically recruited to the site of demethylation. Active demethylation occurs by DNA repair and Gadd45a interacts with and requires the DNA repair endonuclease XPG. We conclude that Gadd45a relieves epigenetic gene silencing by promoting DNA repair, which erases methylation marks.