Circular RNA circSMAD4 regulates cardiac fibrosis by targeting miR-671-5p and FGFR2 in cardiac fibroblasts.

Heart failure is a leading cause of death and is often accompanied by activation of quiescent cardiac myofibroblasts, which results in cardiac fibrosis. In this study, we aimed to identify novel circular RNAs that regulate cardiac fibrosis. We applied transverse aortic constriction (TAC) for 1, 4, and 8 weeks in mice. RNA sequencing datasets were obtained from cardiac fibroblasts isolated by use of a Langendorff apparatus and then further processed by use of selection criteria such as differential expression and conservation in species. CircSMAD4 was upregulated by TAC in mice or by transforming growth factor (TGF)-ß1 in primarily cultured human cardiac fibroblasts. Delivery of si-circSMAD4 attenuated myofibroblast activation and cardiac fibrosis in mice treated with isoproterenol (ISP). si-circSmad4 significantly reduced cardiac fibrosis and remodeling at 8 weeks. Mechanistically, circSMAD4 acted as a sponge against the microRNA miR-671-5p in a sequence-specific manner. miR-671-5p was downregulated during myofibroblast activation and its mimic form attenuated cardiac fibrosis. miR-671-5p mimic destabilized fibroblast growth factor receptor 2 (FGFR2) mRNA in a sequence-specific manner and interfered with the fibrotic action of FGFR2. The circSMAD4-miR-671-5p-FGFR2 pathway is involved in the differentiation of cardiac myofibroblasts and thereby the development of cardiac fibrosis.

Photocontrolled RAFT polymerization: past, present, and future.

In this review, we provide a brief history, progress, and applications, and discuss the remaining challenges of photocontrolled reversible addition-fragmentation chain transfer (RAFT) polymerization (i.e., photoinduced electron/energy transfer-RAFT (PET-RAFT), photoiniferter, and photomediated cationic RAFT polymerization). Among these, visible-light-driven RAFT polymerization has attracted particular attention in recent years due to its benefits, including low energy consumption and the safe reaction procedure. Moreover, the incorporation of visible-light photocatalysis in the polymerization has conferred attractive features, such as spatiotemporal control and oxygen tolerance; however, a clear understanding of the reaction mechanism has not been completely provided. We also present recent research efforts to elucidate the polymerization mechanisms with the aid of quantum chemical calculations combined with experimental evidence. This review offers an insight into the better design of polymerization systems for desired applications and helps realize the full potential of photocontrolled RAFT polymerization in both academic- and industrial-scale applications.

Formation and degradation of strongly reducing cyanoarene-based radical anions towards efficient radical anion-mediated photoredox catalysis.

Cyanoarene-based photocatalysts (PCs) have attracted significant interest owing to their superior catalytic performance for radical anion mediated photoredox catalysis. However, the factors affecting the formation and degradation of cyanoarene-based PC radical anion (PC•‒) are still insufficiently understood. Herein, we therefore investigate the formation and degradation of cyanoarene-based PC•‒ under widely-used photoredox-mediated reaction conditions. By screening various cyanoarene-based PCs, we elucidate strategies to efficiently generate PC•‒ with adequate excited-state reduction potentials (Ered*) via supra-efficient generation of long-lived triplet excited states (T1). To thoroughly investigate the behavior of PC•‒ in actual photoredox-mediated reactions, a reductive dehalogenation is carried out as a model reaction and identified the dominant photodegradation pathways of the PC•‒. Dehalogenation and photodegradation of PC•‒ are coexistent depending on the rate of electron transfer (ET) to the substrate and the photodegradation strongly depends on the electronic and steric properties of the PCs. Based on the understanding of both the formation and photodegradation of PC•‒, we demonstrate that the efficient generation of highly reducing PC•‒ allows for the highly efficient photoredox catalyzed dehalogenation of aryl/alkyl halides at a PC loading as low as 0.001 mol% with a high oxygen tolerance. The present work provides new insights into the reactions of cyanoarene-based PC•‒ in photoredox-mediated reactions.

A Water-Soluble Organic Photocatalyst Discovered for Highly Efficient Additive-Free Visible-Light-Driven Grafting of Polymers from Proteins at Ambient and Aqueous Environments.

Since the pioneering discovery of a protein bound to poly(ethylene glycol), the utility of protein-polymer conjugates (PPCs) is rapidly expanding to currently emerging applications. Photoinduced energy/electron-transfer reversible addition-fragmentation chain-transfer (PET-RAFT) polymerization is a very promising method to prepare structurally well-defined PPCs, as it eliminates high-cost and time-consuming deoxygenation processes due to its oxygen tolerance. However, the oxygen-tolerance behavior of PET-RAFT polymerization is not well-investigated in aqueous environments, and thereby the preparation of PPCs using PET-RAFT polymerization needs a substantial amount of sacrificial reducing agents or inert-gas purging processes. Herein a novel water-soluble and biocompatible organic photocatalyst (PC) is reported, which enables visible-light-driven additive-free "grafting-from" polymerizations of a protein in ambient and aqueous environments. Interestingly, the developed PC shows unconventional "oxygen-acceleration" behavior for a variety of acrylic and acrylamide monomers in aqueous conditions without any additives, which are apparently distinct from previously reported systems. With such a PC, "grafting-from" polymerizations are successfully performed from protein in ambient buffer conditions under green light-emitting diode (LED) irradiation, which result in various PPCs that have neutral, anionic, cationic, and zwitterionic polyacrylates, and polyacrylamides. It is believed that this PC will be widely employed for a variety of photocatalysis processes in aqueous environments, including the living cell system.

SRF is a nonhistone methylation target of KDM2B and SET7 in the regulation of skeletal muscle differentiation.

The demethylation of histone lysine residues, one of the most important modifications in transcriptional regulation, is associated with various physiological states. KDM2B is a demethylase of histones H3K4, H3K36, and H3K79 and is associated with the repression of transcription. Here, we present a novel mechanism by which KDM2B demethylates serum response factor (SRF) K165 to negatively regulate muscle differentiation, which is counteracted by the histone methyltransferase SET7. We show that KDM2B inhibited skeletal muscle differentiation by inhibiting the transcription of SRF-dependent genes. Both KDM2B and SET7 regulated the balance of SRF K165 methylation. SRF K165 methylation was required for the transcriptional activation of SRF and for the promoter occupancy of SRF-dependent genes. SET7 inhibitors blocked muscle cell differentiation. Taken together, these data indicate that SRF is a nonhistone target of KDM2B and that the methylation balance of SRF as maintained by KDM2B and SET7 plays an important role in muscle cell differentiation.

Sumoylation of histone deacetylase 1 regulates MyoD signaling during myogenesis.

Sumoylation, the conjugation of a small ubiquitin-like modifier (SUMO) protein to a target, has diverse cellular effects. However, the functional roles of the SUMO modification during myogenesis have not been fully elucidated. Here, we report that basal sumoylation of histone deacetylase 1 (HDAC1) enhances the deacetylation of MyoD in undifferentiated myoblasts, whereas further sumoylation of HDAC1 contributes to switching its binding partners from MyoD to Rb to induce myocyte differentiation. Differentiation in C2C12 skeletal myoblasts induced new immunoblot bands above HDAC1 that were gradually enhanced during differentiation. Using SUMO inhibitors and sumoylation assays, we showed that the upper band was caused by sumoylation of HDAC1 during differentiation. Basal deacetylase activity was not altered in the SUMO modification-resistant mutant HDAC1 K444/476R (HDAC1 2R). Either differentiation or transfection of SUMO1 increased HDAC1 activity that was attenuated in HDAC1 2R. Furthermore, HDAC1 2R failed to deacetylate MyoD. Binding of HDAC1 to MyoD was attenuated by K444/476R. Binding of HDAC1 to MyoD was gradually reduced after 2 days of differentiation. Transfection of SUMO1 induced dissociation of HDAC1 from MyoD but potentiated its binding to Rb. SUMO1 transfection further attenuated HDAC1-induced inhibition of muscle creatine kinase luciferase activity that was reversed in HDAC1 2R. HDAC1 2R failed to inhibit myogenesis and muscle gene expression. In conclusion, HDAC1 sumoylation plays a dual role in MyoD signaling: enhancement of HDAC1 deacetylation of MyoD in the basally sumoylated state of undifferentiated myoblasts and dissociation of HDAC1 from MyoD during myogenesis.

Fructus Amomi Cardamomi Extract Inhibit Coxsackievirus-B3 Induced Myocarditis in Murine Myocarditis Model.

Coxsackievirus B3 (CVB3) is the main cause of acute myocarditis and dilated cardiomyopathy. Plant extracts are considered as useful materials to develop new antiviral drugs. We had previously selected candidate plant extracts, which showed anti-inflammatory effects. We examined the antiviral effects by using a HeLa cell survival assay. Among these extracts, we chose the Amomi Cardamomi (Amomi) extract, which showed strong antiviral effect and preserved cell survival in CVB3 infection. We investigated the mechanisms underlying the ability of Amomi extract to inhibit CVB3 infection and replication. HeLa cells were infected by CVB3 with or without Amomi extract. Erk and Akt activities, and their correlation with virus replication were observed. Live virus titers in cell supernatants and viral positive- and negative-strand RNA amplification were measured. Amomi extract significantly increased HeLa cell survival in different concentrations (100-10 µg/ml). CVB3 capsid protein VP1 expression (76%) and viral protease 2A-induced eIF4G1 cleavage (70%) were significantly decreased in Amomi extract (100 µg/ml) treated cells. The levels of positive- (20%) and negative-strand (80%) RNA were dramatically decreased compared with the control, as revealed by reverse transcription-PCR. In addition, Amomi extract improved mice survival (51% vs 26%) and dramatically reduced heart inflammation in a CVB3-induced myocarditis mouse model. These results suggested that Amomi extract significantly inhibited Enterovirus replication and myocarditis damage. Amomi may be developed as a therapeutic drug for Enterovirus.

Transcriptomic Features of Bovine Blastocysts Derived by Somatic Cell Nuclear Transfer.

Reprogramming incompletely occurs in most somatic cell nuclear transfer (SCNT) embryos, which results in misregulation of developmentally important genes and subsequent embryonic malfunction and lethality. Here we examined transcriptome profiles in single bovine blastocysts derived by in vitro fertilization (IVF) and SCNT. Different types of donor cells, cumulus cell and ear-skin fibroblast, were used to derive cSCNT and fSCNT blastocysts, respectively. SCNT blastocysts expressed 13,606 genes on average, similar to IVF (13,542). Correlation analysis found that both cSCNT and fSCNT blastocyst groups had transcriptomic features distinctive from the IVF group, with the cSCNT transcriptomes closer to the IVF ones than the fSCNT. Gene expression analysis identified 56 underrepresented and 78 overrepresented differentially expressed genes in both SCNT groups. A 400-kb locus harboring zinc-finger protein family genes in chromosome 18 were found coordinately down-regulated in fSCNT blastocysts, showing a feature of reprogramming-resistant regions. Probing into different categories of genes important for blastocyst development revealed that genes involved in trophectoderm development frequently were underrepresented, and those encoding epigenetic modifiers tended to be overrepresented in SCNT blastocysts. Our effort to identify reprogramming-resistant, differentially expressed genes can help map reprogramming error-prone loci onto the genome and elucidate how to handle the stochastic events of reprogramming to improve cloning efficiency.

Whole-genome resequencing analyses of five pig breeds, including Korean wild and native, and three European origin breeds.

Pigs have been one of the most important sources of meat for humans, and their productivity has been substantially improved by recent strong selection. Here, we present whole-genome resequencing analyses of 55 pigs of five breeds representing Korean native pigs, wild boar and three European origin breeds. 1,673.1 Gb of sequence reads were mapped to the Swine reference assembly, covering ∼99.2% of the reference genome, at an average of ∼11.7-fold coverage. We detected 20,123,573 single-nucleotide polymorphisms (SNPs), of which 25.5% were novel. We extracted 35,458 of non-synonymous SNPs in 9,904 genes, which may contribute to traits of interest. The whole SNP sets were further used to access the population structures of the breeds, using multiple methodologies, including phylogenetic, similarity matrix, and population structure analysis. They showed clear population clusters with respect to each breed. Furthermore, we scanned the whole genomes to identify signatures of selection throughout the genome. The result revealed several promising loci that might underlie economically important traits in pigs, such as the CLDN1 and TWIST1 genes. These discoveries provide useful genomic information for further study of the discrete genetic mechanisms associated with economically important traits in pigs.

Development, validation and genetic analysis of a large soybean SNP genotyping array.

Cultivated soybean (Glycine max) suffers from a narrow germplasm relative to other crop species, probably because of under-use of wild soybean (Glycine soja) as a breeding resource. Use of a single nucleotide polymorphism (SNP) genotyping array is a promising method for dissecting cultivated and wild germplasms to identify important adaptive genes through high-density genetic mapping and genome-wide association studies. Here we describe a large soybean SNP array for use in diversity analyses, linkage mapping and genome-wide association analyses. More than four million high-quality SNPs identified from high-depth genome re-sequencing of 16 soybean accessions and low-depth genome re-sequencing of 31 soybean accessions were used to select 180,961 SNPs for creation of the Axiom(®) SoyaSNP array. Validation analysis for a set of 222 diverse soybean lines showed that 170,223 markers were of good quality for genotyping. Phylogenetic and allele frequency analyses of the validation set data indicated that accessions showing an intermediate morphology between cultivated and wild soybeans collected in Korea were natural hybrids. More than 90 unanchored scaffolds in the current soybean reference sequence were assigned to chromosomes using this array. Finally, dense average spacing and preferential distribution of the SNPs in gene-rich chromosomal regions suggest that this array may be suitable for genome-wide association studies of soybean germplasm. Taken together, these results suggest that use of this array may be a powerful method for soybean genetic analyses relating to many aspects of soybean breeding.

Population structure and domestication revealed by high-depth resequencing of Korean cultivated and wild soybean genomes.

Despite the importance of soybean as a major crop, genome-wide variation and evolution of cultivated soybeans are largely unknown. Here, we catalogued genome variation in an annual soybean population by high-depth resequencing of 10 cultivated and 6 wild accessions and obtained 3.87 million high-quality single-nucleotide polymorphisms (SNPs) after excluding the sites with missing data in any accession. Nuclear genome phylogeny supported a single origin for the cultivated soybeans. We identified 10-fold longer linkage disequilibrium (LD) in the wild soybean relative to wild maize and rice. Despite the small population size, the long LD and large SNP data allowed us to identify 206 candidate domestication regions with significantly lower diversity in the cultivated, but not in the wild, soybeans. Some of the genes in these candidate regions were associated with soybean homologues of canonical domestication genes. However, several examples, which are likely specific to soybean or eudicot crop plants, were also observed. Consequently, the variation data identified in this study should be valuable for breeding and for identifying agronomically important genes in soybeans. However, the long LD of wild soybeans may hinder pinpointing causal gene(s) in the candidate regions.

Bioinformatics interpretation of exome sequencing: blood cancer.

We had analyzed 10 exome sequencing data and single nucleotide polymorphism chips for blood cancer provided by the PGM21 (The National Project for Personalized Genomic Medicine) Award program. We had removed sample G06 because the pair is not correct and G10 because of possible contamination. In-house software somatic copy-number and heterozygosity alteration estimation (SCHALE) was used to detect one loss of heterozygosity region in G05. We had discovered 27 functionally important mutations. Network and pathway analyses gave us clues that NPM1, GATA2, and CEBPA were major driver genes. By comparing with previous somatic mutation profiles, we had concluded that the provided data originated from acute myeloid leukemia. Protein structure modeling showed that somatic mutations in IDH2, RASGEF1B, and MSH4 can affect protein structures.