More than causing (epi)genomic instability: emerging physiological implications of transposable element modulation.

Transposable elements (TEs) initially attracted attention because they comprise a major portion of the genomic sequences in plants and animals. TEs may jump around the genome and disrupt both coding genes as well as regulatory sequences to cause disease. Host cells have therefore evolved various epigenetic and functional RNA-mediated mechanisms to mitigate the disruption of genomic integrity by TEs. TE associated sequences therefore acquire the tendencies of attracting various epigenetic modifiers to induce epigenetic alterations that may spread to the neighboring genes. In addition to posting threats for (epi)genome integrity, emerging evidence suggested the physiological importance of endogenous TEs either as cis-acting control elements for controlling gene regulation or as TE-containing functional transcripts that modulate the transcriptome of the host cells. Recent advances in long-reads sequence analysis technologies, bioinformatics and genetic editing tools have enabled the profiling, precise annotation and functional characterization of TEs despite their challenging repetitive nature. The importance of specific TEs in preimplantation embryonic development, germ cell differentiation and meiosis, cell fate determination and in driving species specific differences in mammals will be discussed.

Variants in Maternal Effect Genes and Relaxed Imprinting Control in a Special Placental Mesenchymal Dysplasia Case with Mild Trophoblast Hyperplasia.

Placental mesenchymal dysplasia (PMD) and partial hydatidiform mole (PHM) placentas share similar characteristics, such as placental overgrowth and grape-like placental tissues. Distinguishing PMD from PHM is critical because the former can result in normal birth, while the latter diagnosis will lead to artificial abortion. Aneuploidy and altered dosage of imprinted gene expression are implicated in the pathogenesis of PHM and also some of the PMD cases. Diandric triploidy is the main cause of PHM, whereas mosaic diploid androgenetic cells in the placental tissue have been associated with the formation of PMD. Here, we report a very special PMD case also presenting with trophoblast hyperplasia phenotype, which is a hallmark of PHM. This PMD placenta has a normal biparental diploid karyotype and is functionally sufficient to support normal fetal growth. We took advantage of this unique case to further dissected the potential common etiology between these two diseases. We show that the differentially methylated region (DMR) at NESP55, a secondary DMR residing in the GNAS locus, is significantly hypermethylated in the PMD placenta. Furthermore, we found heterozygous mutations in NLRP2 and homozygous variants in NLRP7 in the mother's genome. NLRP2 and NLRP7 are known maternal effect genes, and their mutation in pregnant females affects fetal development. The variants/mutations in both genes have been associated with imprinting defects in mole formation and potentially contributed to the mild abnormal imprinting observed in this case. Finally, we identified heterozygous mutations in the X-linked ATRX gene, a known maternal-zygotic imprinting regulator in the patient. Overall, our study demonstrates that PMD and PHM may share overlapping etiologies with the defective/relaxed dosage control of imprinted genes, representing two extreme ends of a spectrum.

The Inhibitory Effect of Extra Virgin Olive Oil and Its Active Compound Oleocanthal on Prostaglandin-Induced Uterine Hypercontraction and Pain-Ex Vivo and In Vivo Study.

Primary dysmenorrhea is a common occurrence in adolescent women and is a type of chronic inflammation. Dysmenorrhea is due to an increase in oxidative stress, which increases cyclooxygenase-2 (COX-2) expression, increases the concentration of prostaglandin F2α (PGF2α), and increases the calcium concentration in uterine smooth muscle, causing excessive uterine contractions and pain. The polyphenolic compound oleocanthal (OC) in extra virgin olive oil (EVOO) has been shown to have an anti-inflammatory and antioxidant effect. This study aimed to investigate the inhibitory effect of extra virgin olive oil and its active ingredient oleocanthal (OC) on prostaglandin-induced uterine hyper-contraction, its antioxidant ability, and related mechanisms. We used force-displacement transducers to calculate uterine contraction in an ex vivo study. To analyze the analgesic effect, in an in vivo study, we used an acetic acid/oxytocin-induced mice writhing model and determined uterus contraction-related signaling protein expression. The active compound OC inhibited calcium/PGF2α-induced uterine hyper-contraction. In the acetic acid and oxytocin-induced mice writhing model, the intervention of the EVOO acetonitrile layer extraction inhibited pain by inhibiting oxidative stress and the phosphorylation of the protein kinase C (PKC)/extracellular signal-regulated kinases (ERK)/ myosin light chain (MLC) signaling pathway. These findings supported the idea that EVOO and its active ingredient, OC, can effectively decrease oxidative stress and PGF2α-induced uterine hyper-contraction, representing a further treatment for dysmenorrhea.

Transient DNMT3L Expression Reinforces Chromatin Surveillance to Halt Senescence Progression in Mouse Embryonic Fibroblast.

Global heterochromatin reduction, which is one of the hallmarks of senescent cells, is associated with reduced transposable element repression and increased risk of chromatin instability. To ensure genomic integrity, the irreparable cells in a population exit permanently from the cell cycle, and this process is termed "senescence." However, senescence only blocks the expansion of unwanted cells, and the aberrant chromatin of senescent cells remains unstable. Serendipitously, we found that the transient ectopic expression of a repressive epigenetic modulator, DNA methyltransferase 3-like (DNMT3L) was sufficient to delay the premature senescence progression of late-passage mouse embryonic fibroblasts (MEFs) associated with a tightened global chromatin structure. DNMT3L induces more repressive H3K9 methylation on endogenous retroviruses and downregulates the derepressed transposons in aging MEFs. In addition, we found that a pulse of ectopic DNMT3L resulted in the reestablishment of H3K27me3 on polycomb repressive complex 2 (PRC2)-target genes that were derepressed in old MEFs. We demonstrated that ectopic DNMT3L interacted with PRC2 in MEFs. Our data also suggested that ectopic DNMT3L might guide PRC2 to redress deregulated chromatin regions in cells undergoing senescence. This study might lead to an epigenetic reinforcement strategy for overcoming aging-associated epimutation and senescence.

Dynamics of alternative polyadenylation in human preimplantation embryos.

Alternative polyadenylation (APA) affects the length of the 3' untranslated region (3'-UTR) and the regulation of microRNAs. Previous studies have shown that cancer cells tend to have shorter 3'-UTRs than normal cells. A plausible explanation for this is that it enables cancer cells to escape the regulation of microRNAs. Here, we extend this concept to an opposing context: changes in 3'-UTR length in the development of the human preimplantation embryo. Unlike cancer cells, during early development 3'-UTRs tended to become longer, and gene expression was negatively correlated with 3'-UTR length. Moreover, our functional enrichment results showed that length changes are part of the development mechanism. We also investigated the analogy of 3'-UTR length variation with respect to lncRNAs and found that, similarly, lncRNA length tended to increase during embryo development.

Diversity and enterotype in gut bacterial community of adults in Taiwan.

BACKGROUND: Gastrointestinal microbiota, particularly gut microbiota, is associated with human health. The biodiversity of gut microbiota is affected by ethnicities and environmental factors such as dietary habits or medicine intake, and three enterotypes of the human gut microbiome were announced in 2011. These enterotypes are not significantly correlated with gender, age, or body weight but are influenced by long-term dietary habits. However, to date, only two enterotypes (predominantly consisting of Bacteroides and Prevotella) have shown these characteristics in previous research; the third enterotype remains ambiguous. Understanding the enterotypes can improve the knowledge of the relationship between microbiota and human health. RESULTS: We obtained 181 human fecal samples from adults in Taiwan. Microbiota compositions were analyzed using next-generation sequencing (NGS) technology, which is a culture-independent method of constructing microbial community profiles by sequencing 16S ribosomal DNA (rDNA). In these samples, 17,675,898 sequencing reads were sequenced, and on average, 215 operational taxonomic units (OTUs) were identified for each sample. In this study, the major bacteria in the enterotypes identified from the fecal samples were Bacteroides, Prevotella, and Enterobacteriaceae, and their correlation with dietary habits was confirmed. A microbial interaction network in the gut was observed on the basis of the amount of short-chain fatty acids, pH value of the intestine, and composition of the bacterial community (enterotypes). Finally, a decision tree was derived to provide a predictive model for the three enterotypes. The accuracies of this model in training and independent testing sets were 97.2 and 84.0%, respectively. CONCLUSIONS: We used NGS technology to characterize the microbiota and constructed a predictive model. The most significant finding was that Enterobacteriaceae, the predominant subtype, could be a new subtype of enterotypes in the Asian population.

Unveiling MYCN regulatory networks in neuroblastoma via integrative analysis of heterogeneous genomics data.

MYCN, an oncogenic transcription factor of the Myc family, is a major driver of neuroblastoma tumorigenesis. Due to the difficulty in drugging MYCN directly, revealing the molecules in MYCN regulatory networks will help to identify effective therapeutic targets for neuroblastoma therapy. Here we perform ChIP-sequencing and small RNA-sequencing of neuroblastoma cells to determine the MYCN-binding sites and MYCN-associated microRNAs, and integrate various types of genomic data to construct MYCN regulatory networks. The overall analysis indicated that MYCN-regulated genes were involved in a wide range of biological processes and could be used as signatures to identify poor-prognosis MYCN-non-amplified patients. Analysis of the MYCN binding sites showed that MYCN principally served as an activator. Using a computational approach, we identified 32 MYCN co-regulators, and some of these findings are supported by previous studies. Moreover, we investigated the interplay between MYCN transcriptional and microRNA post-transcriptional regulations and identified several microRNAs, such as miR-124-3p and miR-93-5p, which may significantly contribute to neuroblastoma pathogenesis. We also found MYCN and its regulated microRNAs acted together to repress the tumor suppressor genes. This work provides a comprehensive view of MYCN regulations for exploring therapeutic targets in neuroblastoma, as well as insights into the mechanism of neuroblastoma tumorigenesis.

Oxypurinol-Specific T Cells Possess Preferential TCR Clonotypes and Express Granulysin in Allopurinol-Induced Severe Cutaneous Adverse Reactions.

Allopurinol, a first-line drug for treating gout and hyperuricemia, is one of the leading causes of severe cutaneous adverse reactions (SCARs). To investigate the molecular mechanism of allopurinol-induced SCAR, we enrolled 21 patients (13 Stevens-Johnson syndrome (SJS)/toxic epidermal necrolysis (TEN) and 8 drug reaction with eosinophilia and systemic symptoms (DRESS)), 11 tolerant controls, and 23 healthy donors. We performed in vitro T-cell activation assays by culturing peripheral blood mononuclear cells (PBMCs) with allopurinol, oxypurinol, or febuxostat and measuring the expression of granulysin and IFN-γ in the supernatants of cultures. TCR repertoire was investigated by next-generation sequencing. Oxypurinol stimulation resulted in a significant increase in granulysin in the cultures of blood samples from SCAR patients (n=14) but not tolerant controls (n=11) or healthy donors (n=23). Oxypurinol induced T-cell response in a concentration- and time-dependent manner, whereas allopurinol or febuxostat did not. T cells from patients with allopurinol-SCAR showed no crossreactivity with febuxostat. Preferential TCR-V-ß usage and clonal expansion of specific CDR3 (third complementarity-determining region) were found in the blister cells from skin lesions (n=8) and oxypurinol-activated T-cell cultures (n=4) from patients with allopurinol-SCAR. These data suggest that, in addition to HLA-B*58:01, clonotype-specific T cells expressing granulysin upon oxypurinol induction participate in the pathogenesis of allopurinol-induced SCAR.

Revealing the anti-tumor effect of artificial miRNA p-27-5p on human breast carcinoma cell line T-47D.

microRNAs (miRNAs) cause mRNA degradation or translation suppression of their target genes. Previous studies have found direct involvement of miRNAs in cancer initiation and progression. Artificial miRNAs, designed to target single or multiple genes of interest, provide a new therapeutic strategy for cancer. This study investigates the anti-tumor effect of a novel artificial miRNA, miR P-27-5p, on breast cancer. In this study, we reveal that miR P-27-5p downregulates the differential gene expressions associated with the protein modification process and regulation of cell cycle in T-47D cells. Introduction of this novel artificial miRNA, miR P-27-5p, into breast cell lines inhibits cell proliferation and induces the first "gap" phase (G1) cell cycle arrest in cancer cell lines but does not affect normal breast cells. We further show that miR P-27-5p targets the 3'-untranslated mRNA region (3'-UTR) of cyclin-dependent kinase 4 (CDK4) and reduces both the mRNA and protein level of CDK4, which in turn, interferes with phosphorylation of the retinoblastoma protein (RB1). Overall, our data suggest that the effects of miR p-27-5p on cell proliferation and G1 cell cycle arrest are through the downregulation of CDK4 and the suppression of RB1 phosphorylation. This study opens avenues for future therapies targeting breast cancer.

Predicting essential genes based on network and sequence analysis.

Essential genes are indispensable to the viability of an organism. Identification and analysis of essential genes is key to understanding the systems level organization of living cells. On the other hand, the ability to predict these genes in pathogens is of great importance for directed drug development. Global analysis of protein interaction networks provides an effective way to elucidate the relationships between genes. It has been found that essential genes tend to be highly connected and generally have more interactions than nonessential ones. With recent large-scale identifications of essential genes and protein-protein interactions in Saccharomyces cerevisiae and Escherichia coli, we have systematically investigated the topological properties of essential and nonessential genes in the protein-protein interaction networks. Essential genes tend to play topologically more important roles in protein interaction networks. Many topological features were found to be statistically discriminative between essential and nonessential genes. In addition, we have also examined sequence properties such as open reading frame length, strand, and phyletic retention for their association with the gene essentiality. Employing the topological features in the protein interaction network and the sequence properties, we have built a machine learning classifier capable of predicting essential genes. Computational prediction of essential genes circumvents expensive and difficult experimental screens and will help antimicrobial drug development.