Modular Oxidation of Cytosine Modifications and Their Application in Direct and Quantitative Sequencing of 5-Hydroxymethylcytosine.

Selective, efficient, and controllable oxidation of cytosine modifications is valuable for epigenetic analyses, yet only limited progress has been made. Here, we present two modular chemical oxidation reactions: conversion of 5-hydroxymethylcytosine (5hmC) into 5-formylcytosine (5fC) using 4-acetamido-2,2,6,6-tetramethylpiperidine-1-oxoammonium tetrafluoroborate (ACT+BF4-) and further transformation of 5fC into 5-carboxycytosine (5caC) through Pinnick oxidation. Both reactions are mild and efficient on double-stranded DNA. We integrated these two oxidations with borane reduction to develop chemical-assisted pyridine borane sequencing plus (CAPS+), for direct and quantitative mapping of 5hmC. Compared with CAPS, CAPS+ improved the conversion rate and false-positive rate. We applied CAPS+ to mouse embryonic stem cells, human normal brain, and glioblastoma DNA samples and demonstrated its superior sensitivity in analyzing the hydroxymethylome.

Identification of natural xanthine oxidase inhibitors: Virtual screening, anti-xanthine oxidase activity, and interaction mechanism.

Xanthine oxidase (XO) is a crucial target for hyperuricemia treatment(s). Naturally occurred XO inhibitors with minimal toxicity and high efficacy have attracted researchers' attention. With the goal of quickly identifying natural XO inhibitors, an integrated computational screening strategy was constructed by molecular docking and calculating the free energy of binding. Twenty-seven hits were achieved from a database containing 19,377 natural molecules. This includes fourteen known XO inhibitors and four firstly-reported inhibitors (isolicoflavonol, 5,7-dihydroxycoumarin, parvifolol D and clauszoline M, IC50 < 40 µM). Iolicoflavonol (hit 8, IC50 = 8.45 ± 0.68 µM) and 5,7-dihydroxycoumarin (hit 25, IC50 = 10.91 ± 0.71 µM) displayed the great potency as mixed-type inhibitors. Docking study and molecular dynamics simulation revealed that both hits could interact with XO's primarily active site residues ARG880, MOS1328, and ASN768 of XO. Fluorescence spectroscopy studies showed that hit 8 bound to the active cavity region of XO, causing changes in XO's conformation and hydrophobicity. Hits 8 and 25 exhibit favorable Absorption, Distribution, Metabolism, and Excretion (ADME) properties. Additionally, no cytotoxicity against human liver cells was observed at their median inhibition concentrations against XO. Therefore, the present study offers isolicoflavonol and 5,7-dihydroxycoumarin with the potential to be disease-modifying agents for hyperuricemia.

Mapping epigenetic modifications by sequencing technologies.

The "epigenetics" concept was first described in 1942. Thus far, chemical modifications on histones, DNA, and RNA have emerged as three important building blocks of epigenetic modifications. Many epigenetic modifications have been intensively studied and found to be involved in most essential biological processes as well as human diseases, including cancer. Precisely and quantitatively mapping over 100 [1], 17 [2], and 160 [3] different known types of epigenetic modifications in histone, DNA, and RNA is the key to understanding the role of epigenetic modifications in gene regulation in diverse biological processes. With the rapid development of sequencing technologies, scientists are able to detect specific epigenetic modifications with various quantitative, high-resolution, whole-genome/transcriptome approaches. Here, we summarize recent advances in epigenetic modification sequencing technologies, focusing on major histone, DNA, and RNA modifications in mammalian cells.

A grammar for interpreting geo-analytical questions as concept transformations.

Geographic Question Answering (GeoQA) systems can automatically answer questions phrased in natural language. Potentially this may enable data analysts to make use of geographic information without requiring any GIS skills. However, going beyond the retrieval of existing geographic facts on particular places remains a challenge. Current systems usually cannot handle geo-analytical questions that require GIS analysis procedures to arrive at answers. To enable geo-analytical QA, GeoQA systems need to interpret questions in terms of a transformation that can be implemented in a GIS workflow. To this end, we propose a novel approach to question parsing that interprets questions in terms of core concepts of spatial information and their functional roles in context-free grammar. The core concepts help model spatial information in questions independently from implementation formats, and their functional roles indicate how concepts are transformed and used in a workflow. Using our parser, geo-analytical questions can be converted into expressions of concept transformations corresponding to abstract GIS workflows. We developed our approach on a corpus of 309 GIS-related questions and tested it on an independent source of 134 test questions including workflows. The evaluation results show high precision and recall on a gold standard of concept transformations.

Unveiling the inhibitory mechanism of aureusidin targeting xanthine oxidase by multi-spectroscopic methods and molecular simulations.

Xanthine oxidase (XO) is a key target for gout treatment. Great efforts have been made towards the discovery and development of new XO inhibitors. Aureusidin (AUR), a natural compound, emerges as the second reported XO inhibitor with an aurone skeleton with an IC50 value of 7.617 ± 0.401 µM in vitro. The inhibitory mechanism of AUR against XO was explored through enzyme kinetic studies, multi-spectroscopic methods, computer simulation techniques, and ADME prediction. The results showed that AUR acts as a rapid reversible and mixed-type XO inhibitor and its binding to XO was driven by hydrogen bonding and hydrophobic interaction. Moreover, AUR presented a strong fluorescence quenching effect through a static quenching process and induced a conformation change of XO. Its binding pattern with XO was revealed through molecular docking, and its affinity toward XO was enhanced through interactions with key amino acid residues in the active pocket of XO. Further, AUR demonstrated good stability and pharmacokinetic behavior properties in molecular dynamics simulation and ADME prediction. In short, the current work clarified in depth the inhibitory mechanism of AUR on XO firstly and then provided fresh insights into its further development as a natural potent XO inhibitor with aurone skeleton.

Detection of low-frequency mutations in clinical samples by increasing mutation abundance via the excision of wild-type sequences.

The efficiency of DNA-enrichment techniques is often insufficient to detect mutations that occur at low frequencies. Here we report a DNA-excision method for the detection of low-frequency mutations in genomic DNA and in circulating cell-free DNA at single-nucleotide resolution. The method is based on a competitive DNA-binding-and-digestion mechanism, effected by deoxyribonuclease I (DNase) guided by single-stranded phosphorothioated DNA (sgDNase), for the removal of wild-type DNA strands. The sgDNase can be designed against any wild-type DNA sequences, allowing for the uniform enrichment of all the mutations within the target-binding region of single-stranded phosphorothioated DNA at mild-temperature conditions. Pretreatment with sgDNase enriches all mutant strands with initial frequencies down to 0.01% and leads to high discrimination factors for all types of single-nucleotide mismatch in multiple sequence contexts, as we show for the identification of low-abundance mutations in samples of blood or tissue from patients with cancer. The method can be coupled with next-generation sequencing, droplet digital polymerase chain reaction, Sanger sequencing, fluorescent-probe-based assays and other mutation-detection methods.

DMRT1 regulates human germline commitment.

Germline commitment following primordial germ cell (PGC) specification during early human development establishes an epigenetic programme and competence for gametogenesis. Here we follow the progression of nascent PGC-like cells derived from human embryonic stem cells in vitro. We show that switching from BMP signalling for PGC specification to Activin A and retinoic acid resulted in DMRT1 and CDH5 expression, the indicators of migratory PGCs in vivo. Moreover, the induction of DMRT1 and SOX17 in PGC-like cells promoted epigenetic resetting with striking global enrichment of 5-hydroxymethylcytosine and locus-specific loss of 5-methylcytosine at DMRT1 binding sites and the expression of DAZL representing DNA methylation-sensitive genes, a hallmark of the germline commitment programme. We provide insight into the unique role of DMRT1 in germline development for advances in human germ cell biology and in vitro gametogenesis.

The regulatory roles of the E3 ubiquitin ligase NEDD4 family in DNA damage response.

E3 ubiquitin ligases, an important part of ubiquitin proteasome system, catalyze the covalent binding of ubiquitin to target substrates, which plays a role in protein ubiquitination and regulates different biological process. DNA damage response (DDR) is induced in response to DNA damage to maintain genome integrity and stability, and this process has crucial significance to a series of cell activities such as differentiation, apoptosis, cell cycle. The NEDD4 family, belonging to HECT E3 ubiquitin ligases, is reported as regulators that participate in the DDR process by recognizing different substrates. In this review, we summarize recent researches on NEDD4 family members in the DDR and discuss the roles of NEDD4 family members in the cascade reactions induced by DNA damage. This review may contribute to the further study of pathophysiology for certain diseases and pharmacology for targeted drugs.

The role of E3 ubiquitin ligase WWP2 and the regulation of PARP1 by ubiquitinated degradation in acute lymphoblastic leukemia.

Acute lymphoblastic leukemia (ALL) has been a huge threat for people's health and finding effective target therapy is urgent and important. WWP2, as one of E3 ubiquitin ligase, is involved in many biological processes by specifically binding to substrates. PARP1 plays a role in cell apoptosis and is considered as a therapeutic target of certain cancers. In this study, we firstly found that WWP2 expressed higher in newly diagnosed ALL patients comparing with complete remission (CR) ALL patients and normal control people, and WWP2 in relapse ALL patients expressed higher than normal control people. WWP2 expression was related with the FAB subtype of ALL and the proportion of blast cells in bone marrow blood tested by flow cytometry. We demonstrated knockout WWP2 inhibited the ALL growth and enhanced apoptosis induced by Dox in vitro and vivo for the first time. WWP2 negatively regulated and interacted with PARP1 and WWP2 mechanically degraded PARP1 through polyubiquitin-proteasome pathway in ALL. These findings suggested WWP2 played a role in ALL development as well as growth and apoptosis, and also displayed a regulatory pathway of PARP1, which provided a new potential therapeutic target for the treatment of ALL.

Lactose azocalixarene drug delivery system for the treatment of multidrug-resistant pseudomonas aeruginosa infected diabetic ulcer.

Diabetic wound is one of the most intractable chronic wounds that is prone to bacterial infection. Hypoxia is an important feature in its microenvironment. However, it is challenging for antimicrobial therapy to directly apply the existing hypoxia-responsive drug delivery systems due to the active targeting deficiency and the biofilm obstacle. Herein, we customizes a hypoxia-responsive carrier, lactose-modified azocalix[4]arene (LacAC4A) with the ability to actively target and inhibit biofilm. By loading ciprofloxacin (Cip), the resultant supramolecular nanoformulation Cip@LacAC4A demonstrates enhanced antibacterial efficacy resulting from both the increased drug accumulation and the controlled release at the site of infection. When applied on diabetic wounds together with multidrug-resistant Pseudomonas aeruginosa infection in vivo, Cip@LacAC4A induces definitely less inflammatory infiltration than free Cip, which translates into high wound healing performance. Importantly, such design principle provides a direction for developing antimicrobial drug delivery systems.

Single-Molecule Fluorescence Techniques for Membrane Protein Dynamics Analysis.

Fluorescence-based single-molecule techniques, mainly including fluorescence correlation spectroscopy (FCS) and single-molecule fluorescence resonance energy transfer (smFRET), are able to analyze the conformational dynamics and diversity of biological macromolecules. They have been applied to analysis of the dynamics of membrane proteins, such as membrane receptors and membrane transport proteins, due to their superior ability in resolving spatio-temporal heterogeneity and the demand of trace amounts of analytes. In this review, we first introduced the basic principle involved in FCS and smFRET. Then we summarized the labeling and immobilization strategies of membrane protein molecules, the confocal-based and TIRF-based instrumental configuration, and the data processing methods. The applications to membrane protein dynamics analysis are described in detail with the focus on how to select suitable fluorophores, labeling sites, experimental setup, and analysis methods. In the last part, the remaining challenges to be addressed and further development in this field are also briefly discussed.

Why geographic data science is not a science.

"Data Science" has taken many disciplines by storm. And for a good reason: New forms and unseen quantities of data enter nearly every scientific field, substantially changing the ways how scientists do science, and potentially allowing them to answer old questions or to pose them in novel ways. The recent success of Data Science is also reflected in corresponding study programs and curricula and the emergence of specialized branches, such as Geographic Data Science (GDS). Some researchers, therefore, claim that Data Science and GDS should be treated as autonomous scientific disciplines, while others fear that it sells nothing but old wine in new bottles. In an attempt to sober the discussion, we investigate GDS and Data Science from the perspective of meta-science. We provide arguments why today's GDS and Data Science should be seen as an interdisciplinary community of practice of data-driven scientists, rather than a scientific discipline. We also discuss what is missing for GDS and Data Science to become genuine scientific disciplines.

A novel CpG-based signature for survival prediction of lung adenocarcinoma patients.

Lung adenocarcinoma (LACA) is the leading cause of cancer-associated death worldwide. The present study intended to identify DNA methylation patterns that may serve as diagnostic and prognostic biomarkers for LACA. Data on DNA methylation and the survival data of the patients of LACA were obtained from The Cancer Genome Atlas. Kaplan-Meier curves and receiver operating characteristic curve analysis were utilized to build diagnostic and prognostic models. A total of 13 CpG sites were identified and validated as the optimal diagnostic and prognostic signature for overall survival. It was concluded that the CpG-based signature is a reliable predictor for the diagnosis and prognosis of patients with LACA.

m6A Reader: Epitranscriptome Target Prediction and Functional Characterization of N 6-Methyladenosine (m6A) Readers.

N 6-methyladenosine (m6A) is the most abundant post-transcriptional modification in mRNA, and regulates critical biological functions via m6A reader proteins that bind to m6A-containing transcripts. There exist multiple m6A reader proteins in the human genome, but their respective binding specificity and functional relevance under different biological contexts are not yet fully understood due to the limitation of experimental approaches. An in silico study was devised to unveil the target specificity and regulatory functions of different m6A readers. We established a support vector machine-based computational framework to predict the epitranscriptome-wide targets of six m6A reader proteins (YTHDF1-3, YTHDC1-2, and EIF3A) based on 58 genomic features as well as the conventional sequence-derived features. Our model achieved an average AUC of 0.981 and 0.893 under the full-transcript and mature mRNA model, respectively, marking a substantial improvement in accuracy compared to the sequence encoding schemes tested. Additionally, the distinct biological characteristics of each individual m6A reader were explored via the distribution, conservation, Gene Ontology enrichment, cellular components and molecular functions of their target m6A sites. A web server was constructed for predicting the putative binding readers of m6A sites to serve the research community, and is freely accessible at: http://m6areader.rnamd.com.

Celecoxib Exerts a Therapeutic Effect Against Demyelination by Improving the Immune and Inflammatory Microenvironments.

BACKGROUND: The myelin sheath can be damaged by genetic and/or environmental factors, leading to demyelinating diseases, for which effective treatments are lacking. Recently, cyclooxygenase-2 (COX-2) overexpression was detected in demyelinating lesions both in patients and animal models, opening an avenue for promoting endogenous remyelination. The aim of this study was to investigate the therapeutic effect of celecoxib, a selective COX-2 inhibitor, against demyelination in a zebrafish model. METHODS: The biotoxicity of celecoxib was evaluated on zebrafish embryos. Metronidazole was used to deplete the oligodendrocytes in Tg (mbp:nfsB-egfp) transgenic fish. Celecoxib was then administered both in larvae and adults. The regeneration of the myelin sheath and the underlying mechanisms were explored by immunohistochemistry, flow cytometry, Western blot analysis, quantitative real-time polymerase chain reaction, and behavioral test. RESULTS: Celecoxib had low in vivo toxicity. A stable and practical demyelination model was established by metronidazole induction. Following celecoxib treatment, the number of oligodendrocytes was increased significantly and the concentric structure of the myelin sheath reappeared. The locomotor ability was notably improved and was close to its physiological levels. The expression of arg1, mrc1, il-10, and il-4 was upregulated, while that of il-1ß, il-12, tnf-α, il-6, caspase-3 and caspase-7 was downregulated. CONCLUSION: Inhibition of COX-2 contributed to the transformation of microglia/macrophages from the M1 to the M2 phenotype, improved the inflammatory microenvironment, and suppressed caspase-dependent apoptosis, thus exerting a therapeutic effect against demyelination.

Clinicopathological Significance and Diagnostic Value of DLEC1 Hypermethylation in Lung Cancer: A Meta-analysis.

BACKGROUND: DLEC1 is a tumor-suppressor gene which plays a role in carcinogenesis. The purpose of the current study was to help establish the diagnostic performance of DLEC1 methylation in lung cancer. METHODS: PubMed, Embase, CNKI, and Wanfang databases were searched to obtain eligible studies. The pooled odds ratios (ORs) and 95% confidence intervals (CIs) were calculated to estimate the strength of the associations. The diagnostic value was assessed by the summary receiver operating characteristics test. RESULTS: A total of 7 articles, with 8 studies that included 673 lung cancer and 581 control samples, were collected in this meta-analysis. Our results showed a significant association of DLEC1 hypermethylation with lung cancer (P < 0.00001, OR = 13.93, 95% CI = 9.44-20.55). The frequency of DLEC1 methylation was significantly higher in squamous cell carcinoma (SCC) than adenocarcinoma (AC). Moreover, DLEC1 was more frequently methylated in patients with lung cancer aged 60 years or over, patients with lymphatic metastasis, or patients with stage III/IV lung cancer. In addition, there was a sensitivity value of 0.90 (95% CI = 0.86-0.93) and a specificity value of 0.60 (95% CI = 0.56-0.63), a pooled positive-likelihood ratio (PLR) of 2.27 (95% CI = 2.08-2.48), a pooled negative-likelihood ratio (NLR) of 0.17 (95% CI = 0.12-0.23), a diagnostic odds ratio (DOR) of 14.72 (10.09-21) and an area under the curve (AUC) of 0.8146 using DLEC1 methylation in the prediction of lung cancer risk. CONCLUSION: This meta-analysis confirms that DLEC1 methylation is a promising biomarker for lung cancer.

Nomograms to Predict Survival in Patients with Lung Squamous Cell Cancer: A Population-Based Study.

BACKGROUND: This study aimed to identify risk factors affecting cancer-specific survival (CSS) and overall survival (OS) in patients with lung squamous cell carcinoma (LSCC) and to develop nomograms for prognostic prediction in these patients. METHODS: Patients who received an LSCC diagnosis between 2007 and 2013 were selected from the Surveillance, Epidemiology, and End Results (SEER) database. The prognostic effect of each variable on survival was evaluated with Cox regression and Kaplan-Meier analysis, and nomograms were developed to predict 3-, 5-, and 7-year CSS and OS rates. RESULTS: Data from 23,004 patients with LSCC were analyzed. Nomograms were first developed by using variables that were significantly associated with CSS and OS and then validated by using an internal bootstrap resampling approach, which showed that they had a sufficient level of discrimination, according to the C-index. CONCLUSIONS: The nomograms satisfactorily predicted 3-, 5-, and 7-year CSS and OS rates for patients with LSCC.