Rapid and accurate alignment of nucleotide conversion sequencing reads with HISAT-3N.

Sequencing technologies using nucleotide conversion techniques such as cytosine to thymine in bisulfite-seq and thymine to cytosine in SLAM seq are powerful tools to explore the chemical intricacies of cellular processes. To date, no one has developed a unified methodology for aligning converted sequences and consolidating alignment of these technologies in one package. In this paper, we describe hierarchical indexing for spliced alignment of transcripts-3 nucleotides (HISAT-3N), which can rapidly and accurately align sequences consisting of any nucleotide conversion by leveraging the powerful hierarchical index and repeat index algorithms originally developed for the HISAT software. Tests on real and simulated data sets show that HISAT-3N is faster than other modern systems, with greater alignment accuracy, higher scalability, and smaller memory requirements. HISAT-3N therefore becomes an ideal aligner when used with converted sequence technologies.

SeqWho: reliable, rapid determination of sequence file identity using k-mer frequencies in Random Forest classifiers.

MOTIVATION: With the vast improvements in sequencing technologies and increased number of protocols, sequencing is being used to answer complex biological problems. Subsequently, analysis pipelines have become more time consuming and complicated, usually requiring highly extensive prevalidation steps. Here, we present SeqWho, a program designed to assess heuristically the quality of sequencing files and reliably classify the organism and protocol type by using Random Forest classifiers trained on biases native in k-mer frequencies and repeat sequence identities. RESULTS: Using one of our primary models, we show that our method accurately and rapidly classifies human and mouse sequences from nine different sequencing libraries by species, library and both together, 98.32%, 97.86% and 96.38% of the time, respectively. Ultimately, we demonstrate that SeqWho is a powerful method for reliably validating the quality and identity of the sequencing files used in any pipeline. AVAILABILITY AND IMPLEMENTATION: https://github.com/DaehwanKimLab/seqwho. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Automated classification of cytogenetic abnormalities in hematolymphoid neoplasms.

MOTIVATION: Algorithms for classifying chromosomes, like convolutional deep neural networks (CNNs), show promise to augment cytogeneticists' workflows; however, a critical limitation is their inability to accurately classify various structural chromosomal abnormalities. In hematopathology, recurrent structural cytogenetic abnormalities herald diagnostic, prognostic and therapeutic implications, but are laborious for expert cytogeneticists to identify. Non-recurrent cytogenetic abnormalities also occur frequently cancerous cells. Here, we demonstrate the feasibility of using CNNs to accurately classify many recurrent cytogenetic abnormalities while being able to reliably detect non-recurrent, spurious abnormal chromosomes, as well as provide insights into dataset assembly, model selection and training methodology that improve overall generalizability and performance for chromosome classification. RESULTS: Our top-performing model achieved a mean weighted F1 score of 96.86% on the validation set and 94.03% on the test set. Gradient class activation maps indicated that our model learned biologically meaningful feature maps, reinforcing the clinical utility of our proposed approach. Altogether, this work: proposes a new dataset framework for training chromosome classifiers for use in a clinical environment, reveals that residual CNNs and cyclical learning rates confer superior performance, and demonstrates the feasibility of using this approach to automatically screen for many recurrent cytogenetic abnormalities while adeptly classifying non-recurrent abnormal chromosomes. AVAILABILITY AND IMPLEMENTATION: Software is freely available at https://github.com/DaehwanKimLab/Chromosome-ReAd. The data underlying this article cannot be shared publicly due to it being protected patient information. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Knockdown of Maged1 inhibits cell cycle progression and causes cell death in mouse embryonic stem cells.

Mouse embryonic stem cells (mESCs) are characterized by self-renewal and pluripotency and can undergo differentiation into the three germ layers (ectoderm, mesoderm, and endoderm). Melanoma-associated antigen D1 (Maged1), which is expressed in all developing and adult tissues, modulates tissue regeneration and development. In the present study, we examined the expression and function of Maged1 in mESCs. Maged1 protein and mRNA expression increased during mESC differentiation. The pluripotency of mESCs was significantly reduced through extracellular signal-regulated kinase 1/2 phosphorylation upon knockdown of Maged1, and through G1 cell cycle arrest during cell division, resulting in significantly reduced mESC proliferation. Moreover, the diameter of the embryoid bodies was significantly reduced, accompanied by increased levels of ectodermal differentiation markers and decreased levels of mesodermal and endodermal differentiation markers. Maged1-knockdown mESC lines showed significantly reduced teratoma volumes and inhibition of teratoma formation in nude mice. Additionally, we observed increased ectodermal markers but decreased mesodermal and endodermal markers in teratoma tissues. These findings show that Maged1 affects mESC pluripotency, proliferation, cell cycle, and differentiation, thereby contributing to our understanding of the basic molecular biological mechanisms and potential roles of Maged1 as a regulator of various mESC properties.

Suppression of transient receptor potential melastatin 7 regulates pluripotency, proliferation, and differentiation of mouse embryonic stem cells via mechanistic target of rapamycin-extracellular signal-regulated kinase activation.

Mouse embryonic stem cells (mESCs) are a widely used model for their diverse availability in studying early embryonic development and their application in regenerative treatment of various intractable diseases. Transient receptor potential melastatin 7 (Trpm7) regulates Ca2+ as a nonselective ion channel and is essential for early embryonic development; however, the precise role of Trpm7 in mESCs has not been clearly elucidated. In this study, we showed that the inhibition of Trpm7 affects the pluripotency and self-renewal of mESCs. We found that short hairpin RNA (shRNA)-mediated suppression of Trpm7 resulted in decreased expression of transcriptional regulators, Oct4 and Sox2, which maintain stemness in mESCs. In addition, Trpm7 knockdown led to alterations in the basic properties of mESCs, such as decreased proliferation, cell cycle arrest at the G0/G1 phase, and increased apoptosis. Furthermore, embryoid body (EB) formation and teratoma formation assays revealed abnormal regulation of differentiation due to Trpm7 knockdown, including the smaller size of EBs, elevated ectodermal differentiation, and diminished endodermal and mesodermal differentiation. We found that EB Day 7 samples displayed decreased intracellular Ca2+ levels compared to those of the scrambled group. Finally, we identified that these alterations induced by Trpm7 knockdown occurred due to decreased phosphorylation of mechanistic target of rapamycin (mTOR) and subsequent activation of extracellular signal-regulated kinase (ERK) in mESCs. Our findings suggest that Trpm7 could be a novel regulator for maintaining stemness and modulating the differentiation of mESCs.

Bleeding of the Superior Vena Cava Due to an Iatrogenic Injury to It during the Ultrasound-Guided Central Venous Cannulation.

Central venous cannulation (CVC) is a procedure that is frequently performed to facilitate resuscitation, nutritional support and long-term vascular access. It may often cause mechanical complications during placement of a cannula in association with the anatomical relationship with central veins. A 68-year-old man visited our medical institution with a chief complaint of foreign-body-induced esophageal perforation. This patient presented with bleeding of the superior vena cava due to an iatrogenic injury to it during the CVC in the right internal jugular vein. Our case indicates that it would be mandatory to insert a cannula at an optimal depth considering the anatomical relationship between the central veins during the CVC.

LSD1 activates a lethal prostate cancer gene network independently of its demethylase function.

Medical castration that interferes with androgen receptor (AR) function is the principal treatment for advanced prostate cancer. However, clinical progression is universal, and tumors with AR-independent resistance mechanisms appear to be increasing in frequency. Consequently, there is an urgent need to develop new treatments targeting molecular pathways enriched in lethal prostate cancer. Lysine-specific demethylase 1 (LSD1) is a histone demethylase and an important regulator of gene expression. Here, we show that LSD1 promotes the survival of prostate cancer cells, including those that are castration-resistant, independently of its demethylase function and of the AR. Importantly, this effect is explained in part by activation of a lethal prostate cancer gene network in collaboration with LSD1's binding protein, ZNF217. Finally, that a small-molecule LSD1 inhibitor-SP-2509-blocks important demethylase-independent functions and suppresses castration-resistant prostate cancer cell viability demonstrates the potential of LSD1 inhibition in this disease.

Design of Grating Al2O3 Passivation Structure Optimized for High-Efficiency Cu(In,Ga)Se2 Solar Cells.

In this paper, we propose an optimized structure of thin Cu(In,Ga)Se2 (CIGS) solar cells with a grating aluminum oxide (Al2O3) passivation layer (GAPL) providing nano-sized contact openings in order to improve power conversion efficiency using optoelectrical simulations. Al2O3 is used as a rear surface passivation material to reduce carrier recombination and improve reflectivity at a rear surface for high efficiency in thin CIGS solar cells. To realize high efficiency for thin CIGS solar cells, the optimized structure was designed by manipulating two structural factors: the contact opening width (COW) and the pitch of the GAPL. Compared with an unpassivated thin CIGS solar cell, the efficiency was improved up to 20.38% when the pitch of the GAPL was 7.5-12.5 µm. Furthermore, the efficiency was improved as the COW of the GAPL was decreased. The maximum efficiency value occurred when the COW was 100 nm because of the effective carrier recombination inhibition and high reflectivity of the Al2O3 insulator passivation with local contacts. These results indicate that the designed structure has optimized structural points for high-efficiency thin CIGS solar cells. Therefore, the photovoltaic (PV) generator and sensor designers can achieve the higher performance of photosensitive thin CIGS solar cells by considering these results.

Strategy to Establish Embryo-Derived Pluripotent Stem Cells in Cattle.

Stem cell research is essential not only for the research and treatment of human diseases, but also for the genetic preservation and improvement of animals. Since embryonic stem cells (ESCs) were established in mice, substantial efforts have been made to establish true ESCs in many species. Although various culture conditions were used to establish ESCs in cattle, the capturing of true bovine ESCs (bESCs) has not been achieved. In this review, the difficulty of establishing bESCs with various culture conditions is described, and the characteristics of proprietary induced pluripotent stem cells and extended pluripotent stem cells are introduced. We conclude with a suggestion of a strategy for establishing true bESCs.

A Rho Kinase (ROCK) Inhibitor, Y-27632, Inhibits the Dissociation-Induced Cell Death of Salivary Gland Stem Cells.

Salivary gland stem cells (SGSCs) are potential cell sources for the treatment of salivary gland diseases. The control of cell survival is an essential factor for applying stem cells to regenerative medicine or stem cell-based research. The purpose of this study was to investigate the effects of the ROCK inhibitor Y-27632 on the survival of SGSCs and its underlying mechanisms. SGSCs were isolated from mouse submandibular glands and cultured in suspension. Treatment with Y-27632 restored the viability of SGSCs that was significantly decreased during isolation and the subsequent culture. Y-27632 upregulated the expression of anti-apoptotic protein BCL-2 in SGSCs and, in the apoptosis assay, significantly reduced apoptotic and necrotic cell populations. Matrigel was used to mimic the extracellular environment of an intact salivary gland. The expression of genes regulating apoptosis and the ROCK signaling pathway was significantly reduced when SGSCs were embedded in Matrigel. SGSCs cultured in Matrigel and treated with Y-27632 showed no difference in the total numbers of spheroids and expression levels of apoptosis-regulating genes. Matrigel-embedded SGSCs treated with Y-27632 increased the number of spheroids with budding structures and the expression of acinar cell-specific marker AQP5. We demonstrate the protective effects of Y-27632 against dissociation-induced apoptosis of SGSCs during their culture in vitro.

Fibroblast growth factor receptor 4 increases epidermal growth factor receptor (EGFR) signaling by inducing amphiregulin expression and attenuates response to EGFR inhibitors in colon cancer.

Fibroblast growth factor receptor 4 (FGFR4) is known to induce cancer cell proliferation, invasion, and antiapoptosis through activation of RAS/RAF/ERK and PI3K/AKT pathways, which are also known as major molecular bases of colon cancer carcinogenesis related with epidermal growth factor receptor (EGFR) signaling. However, the interaction between FGFR4 and EGFR signaling in regard to colon cancer progression is unclear. Here, we investigated a potential cross-talk between FGFR4 and EGFR, and the effect of anti-EGFR therapy in colon cancer treatment. To explore the biological roles of FGFR4 in cancer progression, RNA sequencing was carried out using FGFR4 transfected colon cell lines. Gene ontology data showed the upregulation of genes related to EGFR signaling, and we identified that FGFR4 overexpression secretes EGFR ligands such as amphiregulin (AREG) with consequent activation of EGFR and ErbB3. This result was also shown in in vivo study and the cooperative interaction between EGFR and FGFR4 promoted tumor growth. In addition, FGFR4 overexpression reduced cetuximab-induced cytotoxicity and the combination of FGFR4 inhibitor (BLU9931) and cetuximab showed profound antitumor effect compared to cetuximab alone. Clinically, we found the positive correlation between FGFR4 and AREG expression in tumor tissue, but not in normal tissue, from colon cancer patients and these expressions were significantly correlated with poor overall survival in patients treated with cetuximab. Therefore, our results provide the novel mechanism of FGFR4 in connection with EGFR activation and the combination of FGFR4 inhibitor and cetuximab could be a promising therapeutic option to achieve the optimal response to anti-EGFR therapy in colon cancer.

UBE3A Suppresses Overnutrition-Induced Expression of the Steatosis Target Genes of MLL4 by Degrading MLL4.

Regulation of the protein stability of epigenetic regulators remains ill-defined despite its potential applicability in epigenetic therapies. The histone H3-lysine 4-methyltransferase MLL4 is an epigenetic transcriptional coactivator that directs overnutrition-induced obesity and fatty liver formation, and Mll4+/- mice are resistant to both. Here we show that the E3 ubiquitin ligase UBE3A targets MLL4 for degradation, thereby suppressing high-fat diet (HFD)-induced expression of the hepatic steatosis target genes of MLL4. In contrast to Mll4+/- mice, Ube3a+/- mice are hypersensitive to HFD-induced obesity and fatty liver development. Ube3a+/-;Mll4+/- mice lose this hypersensitivity, supporting roles of increased MLL4 levels in both phenotypes of Ube3a+/- mice. Correspondingly, our comparative studies with wild-type, Ube3a+/- and Ube3a-/- and UBE3A-overexpressing transgenic mouse livers demonstrate an inverse correlation of UBE3A protein levels with MLL4 protein levels, expression of the steatosis target genes of MLL4, and their decoration by H3-lysine 4-monomethylation, a surrogate marker for the epigenetic action of MLL4. Conclusion: UBE3A indirectly exerts an epigenetic regulation of obesity and steatosis by degrading MLL4. This UBE3A-MLL4 regulatory axis provides a potential therapeutic venue for treating various MLL4-directed pathogeneses, including obesity and hepatic steatosis.

Biodegradation of Polystyrene by Pseudomonas sp. Isolated from the Gut of Superworms (Larvae of Zophobas atratus).

Recently, various attempts have been made to solve plastic waste problems, such as development of biodegradation without producing pollution. Polystyrene (PS) is the fifth most used plastic in many industries; therefore, degrading PS becomes a critical global issue. Here, we reported Pseudomonas aeruginosa strain DSM 50071, initially isolated from the gut of the superworms, Zophobas atratus, and the PS degradation by Pseudomonas sp. DSM 50071. We examined PS degradation using electronic microscopy and measured changes in atomic composition and contact angles with water droplets on the PS surface that represents a chemical change from hydrophobicity to hydrophilicity. We have further examined chemical structural changes using X-ray photoelectron spectroscopy, Fourier-transform-infrared spectroscopy, and nuclear magnetic resonance (NMR) to confirm the formation of carbonyl groups (C═O) in the oxidation pathway during PS biodegradation. In reverse transcription quantitative polymerase chain reaction analysis, the gene expression level of serine hydrolase (SH) in Pseudomonas sp. DSM 50071 was highly increased during PS degradation, and the enzyme-mediated biodegradation of PS was further confirmed by the SH inhibitor treatment test. Thus, the significance of these findings goes beyond the discovery of a novel function of Pseudomonas sp. DSM 50071 in the gut of superworms, highlighting a potential solution for PS biodegradation.

Enhanced Power Conversion Efficiency of Dye-Sensitized Solar Cells by Band Edge Shift of TiO2 Photoanode.

By simple soaking titanium dioxide (TiO2) films in an aqueous Na2S solution, we could prepare surface-modified photoanodes for application to dye-sensitized solar cells (DSSCs). An improvement in both the open-circuit voltage (Voc) and the fill factor (FF) was observed in the DSSC with the 5 min-soaked photoanode, compared with those of the control cell without any modification. The UV-visible absorbance spectra, UPS valence band spectra, and dark current measurements revealed that the Na2S modification led to the formation of anions on the TiO2 surface, and thereby shifted the conduction band edge of TiO2 in the negative (upward) direction, inducing an increase of 29 mV in the Voc. It was also found that the increased FF value in the surface-treated device was attributed to an elevation in the shunt resistance.

Effect of Lignin Content on Cellulolytic Saccharification of Liquid Hot Water Pretreated Sugarcane Bagasse.

Lignin contributes to the rigid structure of the plant cell wall and is partially responsible for the recalcitrance of lignocellulosic materials to enzymatic digestion. Overcoming this recalcitrance is one the most critical issues in a sugar-flat form process. This study addresses the effect of low lignin sugarcane bagasse on enzymatic hydrolysis after liquid hot water pretreatment at 190 °C and 20 min (severity factor: 3.95). The hydrolysis of bagasse from a sugarcane line selected for a relatively low lignin content, gave an 89.7% yield of cellulose conversion to glucose at 40 FPU/g glucan versus a 68.3% yield from a comparably treated bagasse from the high lignin bred line. A lower enzyme loading of 5 FPU/g glucan (equivalent to 3.2 FPU/g total solids) resulted in 31.4% and 21.9% conversion yields, respectively, for low and high lignin samples, suggesting the significance of lignin content in the saccharification process. Further increases in the enzymatic conversion of cellulose to glucose were achieved when the bagasse sample was pre-incubated with a lignin blocking agent, e.g., bovine serum albumin (50 mg BSA/g glucan) at 50 °C for 1 h prior to an actual saccharification. In this work, we have demonstrated that even relatively small differences in lignin content can result in considerably increased sugar production, which supports the dissimilarity of bagasse lignin content and its effects on cellulose digestibility. The increased glucose yields with the addition of BSA helped to decrease the inhibition of non-productive absorption of cellulose enzymes onto lignin and solid residual lignin fractions.

The Effect of Becoming Disabled on the Subjective Well-Being of Religious and Nonreligious People.

Positive associations between religiosity and subjective well-being (SWB) have been found in a multitude of studies. However, there has been little effort in documenting the role that religion plays in helping people during the onset of adverse circumstances in their lives. This study investigates the effect of religion on the SWB of the disabled. We utilized secondary data from the Korean Longitudinal Study of Aging from 2006 to 2016 with a sample size of 36,484. Starting with nondisabled participants, we applied a difference-in-differences approach to a fixed-effects model and compared the magnitude of the decrease in SWB resulting from disability between religious individuals and their nonreligious counterparts. The empirical results show that following a religion increased SWB by 0.94 (p < 0.01) and the onset of a disability reduced SWB by 3.57 (p < 0.01) out of 100. Furthermore, there is a significant gap in happiness levels between religious and nonreligious individuals when they are diagnosed with a disability. Becoming disabled reduces SWB for nonreligious people more than that for religious people by 2.62 (p < 0.01). This study confirms that following a religion helps people cope with adverse circumstances such as the onset of a disability.

Centrifuge: rapid and sensitive classification of metagenomic sequences.

Centrifuge is a novel microbial classification engine that enables rapid, accurate, and sensitive labeling of reads and quantification of species on desktop computers. The system uses an indexing scheme based on the Burrows-Wheeler transform (BWT) and the Ferragina-Manzini (FM) index, optimized specifically for the metagenomic classification problem. Centrifuge requires a relatively small index (4.2 GB for 4078 bacterial and 200 archaeal genomes) and classifies sequences at very high speed, allowing it to process the millions of reads from a typical high-throughput DNA sequencing run within a few minutes. Together, these advances enable timely and accurate analysis of large metagenomics data sets on conventional desktop computers. Because of its space-optimized indexing schemes, Centrifuge also makes it possible to index the entire NCBI nonredundant nucleotide sequence database (a total of 109 billion bases) with an index size of 69 GB, in contrast to k-mer-based indexing schemes, which require far more extensive space.

KDM2B is a histone H3K79 demethylase and induces transcriptional repression via sirtuin-1-mediated chromatin silencing.

The methylation of histone H3 lysine 79 (H3K79) is an active chromatin marker and is prominent in actively transcribed regions of the genome; however, demethylase of H3K79 remains unknown despite intensive research. Here, we show that KDM2B, also known as FBXL10 and a member of the Jumonji C family of proteins known for its histone H3K36 demethylase activity, is a di- and trimethyl H3K79 demethylase. We demonstrate that KDM2B induces transcriptional repression of HOXA7 and MEIS1 via occupancy of promoters and demethylation of H3K79. Furthermore, genome-wide analysis suggests that H3K79 methylation levels increase when KDM2B is depleted, which indicates that KDM2B functions as an H3K79 demethylase in vivo. Finally, stable KDM2B-knockdown cell lines exhibit displacement of NAD+-dependent deacetylase sirtuin-1 (SIRT1) from chromatin, with concomitant increases in H3K79 methylation and H4K16 acetylation. Our findings identify KDM2B as an H3K79 demethylase and link its function to transcriptional repression via SIRT1-mediated chromatin silencing.-Kang, J.-Y., Kim, J.-Y., Kim, K.-B., Park, J. W., Cho, H., Hahm, J. Y., Chae, Y.-C., Kim, D., Kook, H., Rhee, S., Ha, N.-C., Seo, S.-B. KDM2B is a histone H3K79 demethylase and induces transcriptional repression via sirtuin-1-mediated chromatin silencing.

Effect of Inflow Arterial Calcification on Arteriovenous Fistula Maturation.

BACKGROUND: The aim of this study is to investigate the effect of preexisting calcification in the inflow artery on maturation and flow volume of an arteriovenous fistula (AVF). METHODS: Patients who underwent AVF creation for hemodialysis were prospectively recruited between March and November 2017. On preoperative duplex ultrasound, calcification in the arterial media within 5 cm of the planned anastomosis area was assessed. Clinical maturation was defined as the successful use of the fistula for ≥75% of the dialysis sessions during a month within 6 months after surgery. Radiological maturation was defined as a venous diameter of ≥0.4 cm and a flow volume of ≥500 mL/min. Flow volumes of the inflow artery and the cephalic vein were measured at 6 and 12 weeks after AVF creation. RESULTS: Eighteen patients with calcification and 29 patients without calcification were enrolled in this study. There was no significant difference in the clinical and radiological maturation between the groups. The flow volume of the inflow artery, measured at 6 weeks postoperatively, was significantly higher in the noncalcification group than in the calcification group (P = 0.042). The flow volume of the inflow artery in the noncalcification group was increased at 12 weeks postoperatively (P = 0.091). Flow volume of the vein was higher in the noncalcification group than in the calcification group, although it did not reach statistical significance. CONCLUSIONS: In conclusions, preexisting arterial calcification did not adversely affect the AVF maturation. However, arterial calcification correlated with the flow volume of the inflow artery of AVF.

Key Age-Imposed Signaling Changes That Are Responsible for the Decline of Stem Cell Function.

This chapter analyzes recent developments in the field of signal transduction of ageing with the focus on the age-imposed changes in TGF-beta/pSmad, Notch, Wnt/beta-catenin, and Jak/Stat networks. Specifically, this chapter delineates how the above-mentioned evolutionary-conserved morphogenic signaling pathways operate in young versus aged mammalian tissues, with insights into how the age-specific broad decline of stem cell function is precipitated by the deregulation of these key cell signaling networks. This chapter also provides perspectives onto the development of defined therapeutic approaches that aim to calibrate intensity of the determinant signal transduction to health-youth, thereby rejuvenating multiple tissues in older people.