DNMT3B PWWP mutations cause hypermethylation of heterochromatin.

The correct establishment of DNA methylation patterns is vital for mammalian development and is achieved by the de novo DNA methyltransferases DNMT3A and DNMT3B. DNMT3B localises to H3K36me3 at actively transcribing gene bodies via its PWWP domain. It also functions at heterochromatin through an unknown recruitment mechanism. Here, we find that knockout of DNMT3B causes loss of methylation predominantly at H3K9me3-marked heterochromatin and that DNMT3B PWWP domain mutations or deletion result in striking increases of methylation in H3K9me3-marked heterochromatin. Removal of the N-terminal region of DNMT3B affects its ability to methylate H3K9me3-marked regions. This region of DNMT3B directly interacts with HP1α and facilitates the bridging of DNMT3B with H3K9me3-marked nucleosomes in vitro. Our results suggest that DNMT3B is recruited to H3K9me3-marked heterochromatin in a PWWP-independent manner that is facilitated by the protein's N-terminal region through an interaction with a key heterochromatin protein. More generally, we suggest that DNMT3B plays a role in DNA methylation homeostasis at heterochromatin, a process which is disrupted in cancer, aging and Immunodeficiency, Centromeric Instability and Facial Anomalies (ICF) syndrome.

BRCA1-BARD1 combines multiple chromatin recognition modules to bridge nascent nucleosomes.

Chromatin association of the BRCA1-BARD1 heterodimer is critical to promote homologous recombination repair of DNA double-strand breaks (DSBs) in S/G2. How the BRCA1-BARD1 complex interacts with chromatin that contains both damage induced histone H2A ubiquitin and inhibitory H4K20 methylation is not fully understood. We characterised BRCA1-BARD1 binding and enzymatic activity to an array of mono- and di-nucleosome substrates using biochemical, structural and single molecule imaging approaches. We found that the BRCA1-BARD1 complex preferentially interacts and modifies di-nucleosomes over mono-nucleosomes, allowing integration of H2A Lys-15 ubiquitylation signals with other chromatin modifications and features. Using high speed- atomic force microscopy (HS-AFM) to monitor how the BRCA1-BARD1 complex recognises chromatin in real time, we saw a highly dynamic complex that bridges two nucleosomes and associates with the DNA linker region. Bridging is aided by multivalent cross-nucleosome interactions that enhance BRCA1-BARD1 E3 ubiquitin ligase catalytic activity. Multivalent interactions across nucleosomes explain how BRCA1-BARD1 can recognise chromatin that retains partial di-methylation at H4 Lys-20 (H4K20me2), a parental histone mark that blocks BRCA1-BARD1 interaction with nucleosomes, to promote its enzymatic and DNA repair activities.

An Emerging Trend in the Synthesis of Iron Titanate Photocatalyst Toward Water Splitting.

Hydrogen gas is a prominent focus in pursuing renewable and clean alternative energy sources. The quest for maximizing hydrogen production yield involves the exploration of an ideal photocatalyst and the development of a simple, cost-effective technique for its generation. Iron titanate has garnered attention in this context due to its photocatalytic properties, affordability, and non-toxic nature. Over the years, different synthesis routes, different morphologies, and some modifications of iron titanate have been carried out to improve its photocatalytic performance by enhancing light absorption in the visible region, boosting charge carrier transfer, and decreasing recombination of electrons and holes. The use of iron titanate photocatalyst for hydrogen evolution reaction has seen an upward trend in recent times, and based on available findings, more can be done to improve the performance. This review paper provides a comprehensive overview of the fundamental principles of photocatalysis for hydrogen generation, encompassing the synthesis, morphology, and application of iron titanate-based photocatalysts. The discussion delves into the limitations of current methodologies and present and future perspectives for advancing iron titanate photocatalysts. By addressing these limitations and contemplating future directions, the aim is to enhance the properties of materials fabricated for photocatalytic water splitting.

Spatially Resolved Metabolites in Stable and Unstable Human Atherosclerotic Plaques Identified by Mass Spectrometry Imaging.

BACKGROUND: Impairments in carbohydrate, lipid, and amino acid metabolism drive features of plaque instability. However, where these impairments occur within the atheroma remains largely unknown. Therefore, we sought to characterize the spatial distribution of metabolites within stable and unstable atherosclerosis in both the fibrous cap and necrotic core. METHODS: Atherosclerotic tissue specimens from 9 unmatched individuals were scored based on the Stary classification scale and subdivided into stable and unstable atheromas. After performing mass spectrometry imaging on these samples, we identified over 850 metabolite-related peaks. Using MetaboScape, METASPACE, and Human Metabolome Database, we confidently annotated 170 of these metabolites and found over 60 of these were different between stable and unstable atheromas. We then integrated these results with an RNA-sequencing data set comparing stable and unstable human atherosclerosis. RESULTS: Upon integrating our mass spectrometry imaging results with the RNA-sequencing data set, we discovered that pathways related to lipid metabolism and long-chain fatty acids were enriched in stable plaques, whereas reactive oxygen species, aromatic amino acid, and tryptophan metabolism were increased in unstable plaques. In addition, acylcarnitines and acylglycines were increased in stable plaques whereas tryptophan metabolites were enriched in unstable plaques. Evaluating spatial differences in stable plaques revealed lactic acid in the necrotic core, whereas pyruvic acid was elevated in the fibrous cap. In unstable plaques, 5-hydroxyindoleacetic acid was enriched in the fibrous cap. CONCLUSIONS: Our work here represents the first step to defining an atlas of metabolic pathways involved in plaque destabilization in human atherosclerosis. We anticipate this will be a valuable resource and open new avenues of research in cardiovascular disease.

Genetic manipulation of photosynthesis to enhance crop productivity under changing environmental conditions.

Current global agricultural production needs to be increased to feed the unconstrained growing population. The changing climatic condition due to anthropogenic activities also makes the conditions more challenging to meet the required crop productivity in the future. The increase in crop productivity in the post green revolution era most likely became stagnant, or no major enhancement in crop productivity observed. In this review article, we discuss the emerging approaches for the enhancement of crop production along with dealing to the future climate changes like rise in temperature, increase in precipitation and decrease in snow and ice level, etc. At first, we discuss the efforts made for the genetic manipulation of chlorophyll metabolism, antenna engineering, electron transport chain, carbon fixation, and photorespiratory processes to enhance the photosynthesis of plants and to develop tolerance in plants to cope with changing environmental conditions. The application of CRISPR to enhance the crop productivity and develop abiotic stress-tolerant plants to face the current changing climatic conditions is also discussed.

Development of loop-mediated isothermal method and comparison with conventional PCR assay for rapid on spot identification of tissue of cattle origin.

Loop-mediated isothermal amplification (LAMP) is a diagnostic method for meat speciation with rapid and minimal equipment requirements. In this study, we developed cattle-specific tube-based LAMP assays targeting mitochondrial Cyt b gene sequence, compared with conventional PCR assay for specificity, sensitivity, and validation of the assay was made. The LAMP reaction was carried at 64 °C for 45 min, and results were confirmed by SYBR Green I dye and agarose gel-electrophoresis. The specificity of the assays was cross-tested with DNA of buffalo, goat, sheep, and pork. The amplification was observed with samples from cattle only without cross-reactivity with other meat species. The analytical sensitivity of LAMP and PCR method for cattle DNA detection was 0.0001 ng and 1 ng, respectively. Repeatability of the assay was achieved on samples from known/blind and admixture meat with other than cattle at the relative percentage of 20%, 10%, 5%, and 1%. The study concluded that the developed assay can be easily employed for the rapid identification of tissue of cattle origin in meat and meat products in low resource areas.

On point identification of species origin of food animals by recombinase polymerase amplification-lateral flow (RPA-LF) assay targeting mitochondrial gene sequences.

The present study was aimed to develop and standardize Recombinase polymerase amplification-lateral flow (RPA-LF) assays for on point identification of species origin of food animals viz: cattle, buffalo and pig. Species specific RPA primers sets for cattle, buffalo and pig were designed by homology comparisons of the sequences of mitochondrial cytochrome b gene and d-loop region from common food species viz: cattle, buffalo, sheep, goat, pig and chicken. The RPA assays for designed primers sets were optimized using the reaction components from Twist Amp basic kit and instructions in its manual. Endpoint detection of species specific amplified RPA products were made by gel electrophoresis and designed species specific RPA-LFA strips. The developed assays were evaluated for their specificity, diagnostic sensitivity, and validated on coded samples and binary meat admixtures with relative percentage of 20, 10, 5 & 1% target species. The developed RPA assays resulted in amplification of DNA template exclusively of cattle, buffalo and pig origin to product sizes of 294, 405 and 283 bp respectively. The diagnostic sensitivities of developed assays were up to 10 pg of genomic DNA and highly correlated with species specific PCR assays taken as gold standard. Developed species specific RPA assays also identified the target species in coded samples and binary meat admixture up to 1%.

Early cirrhosis and a preserved bone marrow niche favour regenerative response to growth factors in decompensated cirrhosis.

BACKGROUND: Exogenous growth factor-mobilized bone marrow (BM) stem cells have shown a differential response in the management of decompensated cirrhosis (DC). This study was designed to evaluate potential clinical benefit of adding Erythropoietin (EPO) in granulocyte-colony stimulating factor (G-CSF)-mobilized stem cell therapy, possible mechanisms of regeneration and predictive factors of regenerative response. METHODS: Sixty consecutive DC patients received either G-CSF with EPO (Group A; n = 30) or G-CSF and placebo (Group B; n = 30) for 2 months and were carefully followed up for 1 year. Baseline and post-treatment liver biopsy, BM biopsy and BM aspirate were analysed for fibro-inflammatory and regenerative response and BM hematopoietic reservoir. RESULTS: Addition of EPO to G-CSF showed a significant improvement in Child-Pugh score (P = 0.03) and MELD score (P = 0.003) as compared to G-CSF alone, with reduction in mortality (16.6% vs 36.7%, P = 0.09). The combination arm also demonstrated a decreased incidence of acute kidney injury (P < 0.001), encephalopathy (P = 0.005) and refilling of ascites (P = 0.03). Compared to monotherapy, it increased CD163+ macrophages (P = 0.013), Ki67+ index (P < 0.001) with decrease in α-SMA levels (P < 0.001) in liver tissue. The response was better with grade 1 and 2 than with grade 3 ascites; Child B cirrhosis and MELD < 16. Non-responders had lower hematopoietic stem cells (HSCs) at baseline. On multivariate analysis, the liver disease severity (MELD < 16) and a relatively preserved BM (BM-HSCs > 0.4) predicted therapeutic response (AUROC = 0.82). CONCLUSIONS: Early DC (MELD < 16) patients with mild-moderate ascites and those with a healthy cellular baseline BM respond better to growth factor therapy. Addition of EPO to G-CSF provides better regenerative response than G-CSF monotherapy.

Identification and molecular characterization of a trans-acting small interfering RNA producing locus regulating leaf rust responsive gene expression in wheat (Triticum aestivum L.).

MAIN CONCLUSION: A novel leaf rust responsive ta-siRNA-producing locus was identified in wheat showing similarity to 28S rRNA and generated four differentially expressing ta-siRNAs by phasing which targeted stress responsive genes. Trans-acting-small interfering RNAs (Ta-siRNAs) are plant specific molecules generally involved in development and are also stress responsive. Ta-siRNAs identified in wheat till date are all responsive to abiotic stress only. Wheat cultivation is severely affected by rusts and leaf rust particularly affects grain filling. This study reports a novel ta-siRNA producing locus (TAS) in wheat which is a segment of 28S ribosomal RNA but shows differential expression during leaf rust infestation. Four small RNA libraries prepared from wheat Near Isogenic Lines were treated with leaf rust pathogen and compared with untreated controls. A TAS with the ability to generate four ta-siRNAs by phasing events was identified along with the microRNA TamiR16 as the phase initiator. The targets of the ta-siRNAs included α-gliadin, leucine rich repeat, trans-membrane proteins, glutathione-S-transferase, and fatty acid desaturase among others, which are either stress responsive genes or are essential for normal growth and development of plants. Expression of the TAS, its generated ta-siRNAs, and their target genes were profiled at five different time points after pathogen inoculation of susceptible and resistant wheat isolines and compared with mock-inoculated controls. Comparative analysis of expression unveiled differential and reciprocal relationship as well as discrete patterns between susceptible and resistant isolines. The expression profiles of the target genes of the identified ta-siRNAs advocate more towards effector triggered susceptibility favouring pathogenesis. The study helps in discerning the functions of wheat genes regulated by ta-siRNAs in response to leaf rust.

Uncovering leaf rust responsive miRNAs in wheat (Triticum aestivum L.) using high-throughput sequencing and prediction of their targets through degradome analysis.

MAIN CONCLUSION: Deep sequencing identified 497 conserved and 559 novel miRNAs in wheat, while degradome analysis revealed 701 targets genes. QRT-PCR demonstrated differential expression of miRNAs during stages of leaf rust progression. Bread wheat (Triticum aestivum L.) is an important cereal food crop feeding 30 % of the world population. Major threat to wheat production is the rust epidemics. This study was targeted towards identification and functional characterizations of micro(mi)RNAs and their target genes in wheat in response to leaf rust ingression. High-throughput sequencing was used for transcriptome-wide identification of miRNAs and their expression profiling in retort to leaf rust using mock and pathogen-inoculated resistant and susceptible near-isogenic wheat plants. A total of 1056 mature miRNAs were identified, of which 497 miRNAs were conserved and 559 miRNAs were novel. The pathogen-inoculated resistant plants manifested more miRNAs compared with the pathogen infected susceptible plants. The miRNA counts increased in susceptible isoline due to leaf rust, conversely, the counts decreased in the resistant isoline in response to pathogenesis illustrating precise spatial tuning of miRNAs during compatible and incompatible interaction. Stem-loop quantitative real-time PCR was used to profile 10 highly differentially expressed miRNAs obtained from high-throughput sequencing data. The spatio-temporal profiling validated the differential expression of miRNAs between the isolines as well as in retort to pathogen infection. Degradome analysis provided 701 predicted target genes associated with defense response, signal transduction, development, metabolism, and transcriptional regulation. The obtained results indicate that wheat isolines employ diverse arrays of miRNAs that modulate their target genes during compatible and incompatible interaction. Our findings contribute to increase knowledge on roles of microRNA in wheat-leaf rust interactions and could help in rust resistance breeding programs.

Bone marrow stem cells and their niche components are adversely affected in advanced cirrhosis of the liver.

UNLABELLED: Bone marrow (BM) is a reservoir for immune and hematopoietic cells and critical for tissue repair and regeneration. All of these functions are severely altered in cirrhosis. We investigated the cellular and functional state of BM in cirrhosis patients. We studied the histological, cellular, and molecular changes in BM of cirrhosis patients (n = 168) and controls (n = 44). Hematopoietic stem cells (HSCs) and associated niche cells, mesenchymal stem cells, Schwann cells, neural fibers, and endothelial cells were evaluated by immunohistochemistry. Cytokines and growth factors were analyzed in peripheral blood and BM plasma. Cirrhotic BM showed an inverse correlation between cluster of differentiation 34+HSCs and Model of End-Stage Liver Disease (ρ = -0.582, P < 0.001) and Child's scores (P < 0.038). BMs of cirrhosis patients with higher Model of End-Stage Liver Disease (>15) showed significantly decreased HSCs, mesenchymal stem cells, Schwann cells, and neural fibers; increased interleukin-1ß (P = 0.004), tumor necrosis factor-α (P = 0.040), and interferon-γ (P = 0.03); and decreased oncostatin M (P = 0.04), stem cell factor (P = 0.05), and stromal cell-derived factor 1 (P = 0.03) compared to those with lower Model of End-Stage Liver Disease scores (≤15). The cluster of differentiation 34+ cell population was a predictor for the development of sepsis (P < 0.001), and per unit loss increased the probability of sepsis by 16%. Cirrhosis patients with fewer HSCs had lower hemoglobin (P = 0.05) and platelet counts (P = 0.05) and showed early graft dysfunction. CONCLUSIONS: Increasing severity of cirrhosis causes derangement of the hematopoietic niche and loss of HSCs, contributing to the hematological and immunological dysfunctions and reduced potential for regeneration; restoring BM functions could provide new therapeutic options in cirrhosis. (Hepatology 2016;64:1273-1288).

Potential of cyanobacterial biofilms in phosphate removal and biomass production.

Four cyanobacterial biofilms, raised from cyanobacterial mats and dominated by Phormidium and Oscillatoria spp., were successfully grown attached to polyester mesh discs, and were tested for their probable application in [Formula: see text] -P removal from domestic sewage and other nutrient enriched wastewaters. Biofilm # 2, dominated by Phormidium fragile, best removed [Formula: see text] -P; nevertheless, some of it also grew outside the substrate making harvesting difficult. Other biofilms also efficiently removed [Formula: see text] -P from the medium in the following order: Biofilm # 1 > Biofilm # 3 > Biofilm # 4. Their growths were restricted to discs and are therefore better candidates as they can be efficiently harvested after [Formula: see text] -P removal. [Formula: see text] -P removal was primarily due to its uptake during growth of the biofilm rather than because of precipitation as pH of the medium remained <8.5. [Formula: see text] -N concentration in the medium determined [Formula: see text] -P removal efficiency of the test biofilms and therefore optimum N:P ratio is necessary for optimizing [Formula: see text] -P removal. The test biofilms could also efficiently remove [Formula: see text] -N from the medium.

Transcriptome-wide analysis of WRKY transcription factors in wheat and their leaf rust responsive expression profiling.

WRKY, a plant-specific transcription factor family, has important roles in pathogen defense, abiotic cues and phytohormone signaling, yet little is known about their roles and molecular mechanism of function in response to rust diseases in wheat. We identified 100 TaWRKY sequences using wheat Expressed Sequence Tag database of which 22 WRKY sequences were novel. Identified proteins were characterized based on their zinc finger motifs and phylogenetic analysis clustered them into six clades consisting of class IIc and class III WRKY proteins. Functional annotation revealed major functions in metabolic and cellular processes in control plants; whereas response to stimuli, signaling and defense in pathogen inoculated plants, their major molecular function being binding to DNA. Tag-based expression analysis of the identified genes revealed differential expression between mock and Puccinia triticina inoculated wheat near isogenic lines. Gene expression was also performed with six rust-related microarray experiments at Gene Expression Omnibus database. TaWRKY10, 15, 17 and 56 were common in both tag-based and microarray-based differential expression analysis and could be representing rust specific WRKY genes. The obtained results will bestow insight into the functional characterization of WRKY transcription factors responsive to leaf rust pathogenesis that can be used as candidate genes in molecular breeding programs to improve biotic stress tolerance in wheat.

Synthesis of 2-N/S/C-substituted benzothiazoles via intramolecular cyclative cleavage of benzotriazole ring.

The synthesis of numerous 2-N/S/C-substituted benzothiazoles was achieved from substituted thiocarbonylbenzotriazoles via free-radical intramolecular cyclative cleavage of the benzotriazole ring in the presence of (TMS)3SiH and AIBN under mild conditions. The developed methodology demonstrates significant compatibility under microwave conditions and is important as it avoids the use of toxic metals for radical cyclization.

Cultivation of algal biofilm and mat communities from the Garhwal Himalayas for possible use in diverse biotechnological applications.

The current study aimed to screen biofilm-/mat-forming and fast-growing algal communities from the Garhwal Himalayas, India. A total of 15 biofilm/mat-forming algal samples were collected, 8 biofilms out of these could be cultured and analyzed for their growth and development with time. Light microscopy was used to identify different types of cyanobacteria and algae present in the different collected biofilms/mats. Four biofilm and mat communities, namely biofilms #E, #F, #G, and #H, were found to have fast growth and were quick to colonize the substratum. Nylon net was identified as the most cost-effective and best-supporting material for biofilm development and biomass production. The study also found that increasing the harvesting frequency from the nylon net-enmeshed biofilms at least once a week would enhance the final biomass yield compared to harvesting the community once after a longer growth duration. Nevertheless, the findings reported here will be useful for researchers in developing phototrophic biofilm-based technology using nylon net, as it will be mechanically strong, supportive, and easy to handle.

Dental Metal Matrix Composites: The Effects of the Addition of Titanium Nanoparticle Particles on Dental Amalgam.

Dental amalgams have been used by dentists for the restoration of posterior human teeth. However, there have been concerns about the release of mercury from amalgams into the oral cavity. The objective of the present research is to study the effect of titanium (Ti) nanoparticles on the microstructural mechanism of the release of mercury vapor in two commonly used brands of dental amalgam (the Dispersalloy: 11.8% Cu; the Sybralloy: 33% Cu). Ti powder was added to both the Dispersalloy and the Sybralloy in increments of 10 mg up to 80 mg. The addition of Ti powder to both brands of dental amalgam has been found to result in a considerable decrease in Hg vapor release. The decrease in the Hg vapor release due to Ti addition has been explained by the formation of strong Hg-Ti covalent bonds, which reduce the availability of Hg atoms for evaporation. The Ti atoms in excess of the solubility limit of Ti in Hg reside in the grain boundaries, which also reduces the evaporation of Hg from the amalgam. The binding of Hg with Ti via a strong covalent bond also results in a significant improvement in mechanical properties such as Vickers hardness.

Strain imaging as an early predictor in acute myocardial infarction - An augmented cross-sectional study.

INTRODUCTION: Cardiac fibres are affected invariably in myocardial infarction, with longitudinal strain being the earliest to be detected in the ischaemic cascade. The present study aims to assess strain imaging in acute myocardial infarction (AMI) patients admitted to the cardiology department at our institute and correlate GLS and DESL findings with other markers for myocardial function. METHODS AND MATERIAL: This augmented cross-sectional study was conducted amongst the patients admitted with diagnosis of AMI. During the study period, 157 subjects were sampled through convenience sampling, and examined as well as tested with routine investigations at baseline. The subjects were then followed through at first, third and six months, and findings noted. Chi-square was used to assess the crude association between sample characteristics. Pearson correlation and student t-test were used to find association between continuous variables. RESULTS: After screening 564 patients, 157 patients were included in the study after fulfilment of inclusion and exclusion criteria. A significant difference was found in baseline GLS scores and NTproBNP levels at 6 months in alive patients with STEMI, t (21.728) = -5.717, p < .001. Out of the 50 NSTEMI patients, 35 (70 %) were positive for ESL, similarly out of 43 STEMI patients without any RWMA, ESL was positive in 39 (90.02 %) patients. CONCLUSIONS: GLS by STE has good correlation with LVEF, WMSI and NT pro-BNP and it is an independent predictor of mortality and heart failure among patients with AMI.

Synthesis of glycoconjugate benzothiazoles via cleavage of benzotriazole ring.

A concise and efficacious benzotriazole-mediated novel two-step protocol has been developed for easy access to glycoconjugate benzothiazoles from protected carbohydrates. The benzotriazolemethanethione 3, prepared by the reaction of free alcohol with bis(1H-benzo[1,2,3]triazol-1-yl)methanethione, on treatment with silanes or stannane under heating or microwave irradiation undergoes free radical ß-scission of N-N bond and affords diverse range of 2-O-substituted benzothiazoles 4 via cyclative elimination of molecular nitrogen. The structures of all of the compounds have been elucidated using IR, NMR, MS, and elemental analysis, and five of them have been characterized by single-crystal X-ray analysis.

Therapeutic potential of growth factors in pulmonary emphysematous condition.

Pulmonary emphysema is a major manifestation of chronic obstructive pulmonary disease (COPD), which is characterized by progressive destruction of alveolar parenchyma with persistent inflammation of the small airways. Such destruction in the distal respiratory tract is irreversible and irreparable. All-trans-retinoic acid was suggested as a novel therapy for regeneration of lost alveoli in emphysema. However, profound discrepancies were evident between studies. At present, no effective therapeutic options are available that allow for the regeneration of lost alveoli in emphysematous human lungs. Recently, some reports on rodent's models have suggested the beneficial effects of various growth factors toward alveolar maintenance and repair processes.