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Background: PURA-related neurodevelopmental disorders (PURA-NDDs) include 5q31.3 deletion syndrome and PURA syndrome. PURA-NDDs are characterized by neonatal hypotonia, moderate to severe global developmental delay/intellectual disability (GDD/ID), facial dysmorphism, epileptic seizures, nonepileptic movement disorders, and ophthalmological problems. PURA-NDDs have recently been identified and underestimated in neurodevelopmental cohorts, but their diagnosis is still challenging. We retrospectively reviewed the clinical characteristics, genetic spectrum, and diagnostic journey of patients with PURA-NDDs.Results: We report 2 patients with 5q31.3 microdeletion and 5 with PURA pathogenic variants. They demonstrated hypotonia (7/7, 100%), feeding difficulties (4/5, 80%), and respiratory problems (4/7, 57%) in the neonatal period. All of them had severe GDD/ID and could not achieve independent waking and verbal responses. Distinctive facial features of open-tented upper vermilion, long philtrum, and anteverted nares and poor visual fixation and tracking with or without nystagmus were most commonly found (5/7, 71.4%). There were no significant differences in clinical phenotypes between 5q31.3 microdeletion syndrome and PURA syndrome. PURA-NDDs need to be considered as a differential diagnosis in individuals who show severe hypotonia, including feeding difficulties since birth and severe developmental retardation with distinctive facial and ophthalmological features.Conclusions: Our data expands the phenotypic and genetic spectrum of PURA-NDD. Next-generation sequencing methods based on the detailed phenotypic evaluation would shorten the diagnostic delay and would help this rare disorder become a recognizable cause of neurodevelopmental delay.

Prediabetes (PD) is a high-risk state of developing type 2 diabetes, and cardiovascular and metabolic diseases. Metabolomics-based biomarker studies can provide advanced opportunities for prediction of PD over the conventional methods. Here, we aimed to identify metabolic markers and verify their abilities to predict PD, as compared to the performance of the traditional clinical risk factor (CRF) and previously reported metabolites in other population-based studies. Targeted metabolites quantification was performed in 1723 participants in the Korea Association REsource (KARE) cohort, from which 500 normal individuals were followed up for 6 years. We selected 12 significant metabolic markers, including five amino acids, four glycerophospholipids, two sphingolipids, and one acylcarnitine, at baseline, resulting in a predicted incidence of PD with an area under the curve (AUC) of 0.71 during follow-up. The performance of these metabolic markers compared to that of fasting glucose was significantly higher in obese patients (body mass index: BMI ≥ 25 kg/m2, 0.79 vs. 0.58, P

Metabolome-genome wide association studies (mGWASs) are useful for understanding the genetic regulation of metabolites in complex diseases, including type 2 diabetes (T2D). Numerous genetic variants associated with T2D-related metabolites have been identified in previous mGWASs; however, these analyses seem to have difficulty in detecting the genetic variants with functional effects. An exome array focussed on potentially functional variants is an alternative platform to obtain insight into the genetics of biochemical conversion processes. In the present study, we performed an mGWAS using 27,140 non-synonymous variants included in the Illumina HumanExome BeadChip and nine T2D-related metabolites identified by a targetted metabolomics approach to evaluate 2,338 Korean individuals from the Korea Association REsource (KARE) cohort. A linear regression analysis controlling for age, sex, BMI, and T2D status as covariates was performed to identify novel non-synonymous variants associated with T2D-related metabolites. We found significant associations between glycine and CPS1 (rs1047883) and PC ae C36:0 and CYP4F2 (rs2108622) variants (P

Background: Prediabetes (PD) is a high-risk state for type 2 diabetes (T2D), but the risk of progression from PD to T2D can be prevented by early diagnosis and intervention. Metabolomics studies suggest that metabolic biomarkers can shed insight into etiological mechanisms and improve the accuracy of prediction of disease onset. Objectives: We aimed to identify serum metabolic biomarkers and verify their predictive performance for PD in the Korean population, as compared to the performance of known clinical risk factor (CRF) as well as previously reported metabolites (PRM) in other population studies. Methods: A targeted metabolomics was carried out to quantify serum metabolites for 1,723 participants from the Korea Association REsource (KARE) cohort from which 500 individuals were followed-up to 6-years. PD-related metabolites were identified at baseline by statistical methods including multivariable regression analysis, and tested their association to incidence of PD during follow-up. Results: The 12 metabolites were significantly altered in PD (adjusted P= 25 kg/m2, 0.79 vs. 0.58, P Conclusion: Our results revealed that the novel metabolic markers are not only associated with the risk of PD, but also improve the prediction performance in combination with classical approaches. These findings may help understanding causality of diabetes progression and developing preventive strategies for T2D. Disclosure H. Lee: None. T. Park: None. B. Kim: None.

Histone residues can serve as platforms for specific regulatory function. Here we constructed a map of regulatory associations between histone residues and a wide spectrum of chromatin regulation factors based on gene expression changes by histone point mutations in Saccharomyces cerevisiae. Detailed analyses of this map revealed novel associations. Regarding the modulation of H3K4 and K36 methylation by Set1, Set2, or Jhd2, we proposed a role for H4K91 acetylation in early Pol II elongation, and for H4K16 deacetylation in late elongation and crosstalk with H3K4 demethylation for gene silencing. The association of H3K56 with nucleosome positioning suggested that this lysine residue and its acetylation might contribute to nucleosome mobility for transcription activation. Further insights into chromatin regulation are expected from this approach.

Copy number variation is a well-known genetic variation. microRNAs (miRNAs/miRs) are non-coding RNAs that mediate gene expression by regulating target mRNAs. In the present study, copy number deletions encompassing miRNA coding regions were investigated to determine the association between the deletion of miRNA and its phenotypic effects. A total of 38 human miRNAs in copy number variants were identified and miR-650, which is functional in the human osteosarcoma MG-63 cell line, was selected. Overexpression of miR-650 decreased the expression of inhibitor of growth family member 4 (ING4) in the MG-63 cells and increased interleukin (IL)6 transcription, as well as IL6 secretion in IL1B-stimulated cells. Furthermore, miR-650 downregulated the amount of nuclear factor of κ light polypeptide gene enhancer in B cells inhibitor α and increased the transcriptional activity of nuclear factor (NF)κB. Downregulation of ING4 also increased the production of IL6, similar to miR-650 overexpression. Taken together, these data indicate that miR-650 plays a significant role in the production of IL6 by regulating ING4 expression and NFκB signaling in IL1B-stimulated MG-63 osteosarcoma cells.

Type 2 diabetes (T2D) is a multifactorial disease resulting from a complex interaction between environmental and genetic risk factors. Metabolomics provide a logical framework that reflects the functional endpoints of biological processes being triggered by genetic information and various external influences. Identification of metabolite biomarkers can shed insight into etiological pathways and improve the prediction of disease risk. Here, we aimed to identify serum metabolites as putative biomarkers for T2D and their association with genetic variants in the Korean population. A targeted metabolomics approach was employed to quantify serum metabolites for 2240 participants in the Korea Association REsource (KARE) cohort. T2D-related metabolites were identified by statistical methods including multivariable linear and logistic regression, and were independently replicated in the Cooperative Health Research in the Region of Augsburg (KORA) cohort. Additionally, by combining a genome wide association study (GWAS) with metabolomics, genetic variants associated with the identified T2D-related metabolites were uncovered. 123 metabolites were quantified from fasting serum samples and four metabolites, hexadecanoylcarnitine (C16), glycine, lysophosphatidylcholine acyl C18:2 (lysoPC a C18:2), and phosphatidylcholine acyl-alkyl C36:0 (PC ae C36:0), were significantly altered in T2D compared to non-T2D subjects (after the Bonferroni correction for multiple testing with P

The single nucleotide polymorphism rs9939609 of the gene FTO, which encodes fat mass and obesity–associated protein, is strongly associated with obesity and type 2 diabetes (T2D) in multiple populations; however, the underlying mechanism of this association is unclear. The present study aimed to investigate FTO genotype–dependent metabolic changes in obesity and T2D. To elucidate metabolic dysregulation associated with disease risk genotype, genomic and metabolomic datasets were recruited from 2,577 participants of the Korean Association REsource (KARE) cohort, including 40 homozygous carriers of the FTO risk allele (AA), 570 heterozygous carriers (AT), and 1,967 participants carrying no risk allele (TT). A total of 134 serum metabolites were quantified using a targeted metabolomics approach. Through comparison of various statistical methods, seven metabolites were identified that are significantly altered in obesity and T2D based on the FTO risk allele (adjusted p < 0.05). These identified metabolites are relevant to phosphatidylcholine metabolic pathway, and previously reported to be metabolic markers of obesity and T2D. In conclusion, using metabolomics with the information from genome-wide association studies revealed significantly altered metabolites depending on the FTO genotype in complex disorders. This study may contribute to a better understanding of the biological mechanisms linking obesity and T2D.

α-Viniferin, an oligostilbene of trimeric resveratrol, has been reported to have antiinflammatory potential in carrageenin-induced paw edema or adjuvant-induced arthritis in animal models. However, little is known about the molecular basis. In this study, α-viniferin at 3 – 10 μM dose-dependently inhibited interferon (IFN)-γ–induced Ser727 phosphorylation of the signal transducer and activation of transcription-1 (STAT-1), a pivotal transcription factor controlling IFN-γ–targeted genes, in RAW 264.7 macrophages, and also IFN-γ–induced activation of the extracellular signal-regulated kinase (ERK)-1, a protein kinase upstream of the Ser727 phosphorylation of STAT-1. However, α-viniferin, only at a higher concentration of 10 μM, inhibited Janus kinase 2–mediated Tyr701 phosphorylation of STAT-1 in the cells. To understand STAT-1–dependent inflammatory responses, we quantified nitric oxide (NO) or chemokines. α-Viniferin at 3 – 10 μM dose-dependently inhibited IFN-γ–induced production of NO, IFN-γ–inducible protein-10 (IP-10), or the monokine induced by IFN-γ (MIG) in RAW 264.7 cells and also that of NO in primary macrophages-derived from C57BL/6 mice. Furthermore, α-viniferin diminished IFN-γ–induced protein levels of inducible NO synthase (iNOS), attenuated mRNA levels of iNOS, IP-10, or MIG as well as inhibited promoter activity of the iNOS gene. In conclusion, this study proposes an antiinflammatory mechanism of α-viniferin, down-regulating STAT-1–inducible inflammatory genes via inhibiting ERK-mediated STAT-1 activation in IFN-γ–stimulated macrophages. Keywords:: α-viniferin, anti-inflammation, signal transducer and activation of transcription-1 (STAT-1) activation, interferon (IFN)-γ, macrophage

Background OA is a complex disease caused by environmental and genetic risk factors. The purpose of this study is to identify candidate copy number variations (CNVs) associated with OA. Methods We performed a genome-wide association study of CNV to identify potential loci that confer susceptibility to or protection from OA. CNV genotyping was conducted using NimbleGen HD2 3 × 720K comparative hybridization array and included samples from 371 OA patients and 467 healthy controls. The putative CNV regions identified were confirmed with a TaqMan assay. Results We identified six genomic regions associated with OA encompassing CNV loci. None of six loci had previously been reported in genome-wide association studies with OA, although a genetic analysis suggested that they have functional effects. The protein product of a candidate risk gene for obesity, TNKS, targets Wnt inhibition, and this gene was significantly associated with hand and knee OA. Copy number deletion on TNKS was associated with a 1.37-fold decreased risk for OA. In addition, CA10, which shows a strong association with osteoporosis, was also significant in our study. Copy number deletion on this gene was associated with a 1.69-fold decreased risk for OA. Conclusion We identified several CNV loci that may contribute to OA susceptibility in Koreans. Further functional investigations of these genes are warranted to fully characterize their putative association. Electronic supplementary material The online version of this article (doi:10.1186/s12891-015-0531-4) contains supplementary material, which is available to authorized users.

Migration is an essential feature of metastatic cancer cells. To understand how motility is regulated in hepatocellular carcinoma, we analyzed gene expression profiles of mouse model cell lines we established from transgenic mice carrying SV40 large T antigen. A non-motile HC9 cell line was isolated from mouse liver tumors, and two additional cell lines, HCM1 and HCM4, were derived from HC9 cells. We found that both HCM1 and HCM4 cells were substantially more migratory than HC9, and that HCM1 generated tumor nodules in nude mice. In contrast to HCM4 cells that exhibited mesenchymal cell-type gene expression similar to HC9 cells, HCM1 cells appeared to have undergone a mesenchymal-amoeboidal transition. Thus, HCM1 and HCM4 cells have distinct migration and gene expression patterns, and together with HC9 cells, they can serve as model cell lines for understanding how migration is acquired and controlled in hepatocellular carcinoma.

Phosphorylation of the histone variant H2AX forms γ-H2AX that marks DNA double-strand break (DSB). Here, we generated the sequencing-based maps of H2AX and γ-H2AX positioning in resting and proliferating cells before and after ionizing irradiation. Genome-wide locations of possible endogenous and exogenous DSBs were identified based on γ-H2AX distribution in dividing cancer cells without irradiation and that in resting cells upon irradiation, respectively. γ-H2AX-enriched regions of endogenous origin in replicating cells included sub-telomeres and active transcription start sites, apparently reflecting replication- and transcription-mediated stress during rapid cell division. Surprisingly, H2AX itself, prior to phosphorylation, was specifically located at these endogenous hotspots. This phenomenon was only observed in dividing cancer cells but not in resting cells. Endogenous H2AX was concentrated on the transcription start site of actively transcribed genes but was irrelevant to pausing of RNA polymerase II (pol II), which precisely coincided with γ-H2AX of endogenous origin. γ-H2AX enrichment upon irradiation also coincided with actively transcribed regions, but unlike endogenous γ-H2AX, it extended into the gene body and was not specifically concentrated on the pausing site of pol II. Sub-telomeres were less responsive to external DNA damage than to endogenous stress. Our findings provide insight into DNA repair programs of cancer and may have implications for cancer therapy.

International audience; The Wnt/β-catenin pathway controls cell proliferation, death and differentiation. Several families of extracellular proteins can antagonize Wnt/β-catenin signaling, including the decoy receptors known as secreted frizzled related proteins (SFRPs), which have a cysteine-rich domain (CRD) structurally similar to the extracellular Wnt-binding domain of the frizzled receptors. SFRPs inhibit Wnt signaling by sequestering Wnts through the CRD or by forming inactive complexes with the frizzled receptors. Other endogenous molecules carrying frizzled CRDs inhibit Wnt signaling, such as V3Nter, which is proteolytically derived from the cell surface component collagen XVIII and contains a biologically active frizzled domain (FZC18) inhibiting in vivo cell proliferation and tumor growth in mice. We recently showed that FZC18 expressing cells deliver short-range signals to neighboring cells, decreasing their proliferation in vitro and in vivo through the Wnt/β-catenin signaling pathway. Here, using low concentrations of soluble FZC18 and Wnt3a, we show that they physically interact in a cell-free system. In addition, soluble FZC18 binds the frizzled 1 and 8 receptors' CRDs, reducing cell sensitivity to Wnt3a. Conversely, inhibition of Wnt/β-catenin signaling was partially rescued by the expression of full-length frizzled 1 and 8 receptors, but enhanced by the expression of a chimeric cell-membrane-tethered frizzled 8 CRD. Moreover, soluble, partially purified recombinant FZC18_CRD inhibited Wnt3a-induced β-catenin activation. Taken together, the data indicate that collagen XVIII-derived frizzled CRD shifts Wnt sensitivity of normal cells to a lower pitch and controls their growth.

Myeloid differentiation 2 (MD-2) recognizes endotoxin lipopolysaccharide (LPS), which is required for Toll-like receptor 4 (TLR4) activity. MD-2 represents a more attractive therapeutic target than TLR4 for intervention in severe inflammatory disorders due to microbial infection. Here, we suggest MD-2 as a molecular target of nonlipid chalcone in the inhibition of LPS-induced cellular inflammation. A chalcone derivative, 2',4-dihydroxy-6'-isopentyloxychalcone (JSH) competitively displaced LPS from MD-2, and was fitted into the ligand-binding site on the crystal structure of MD-2 under the most energetically favorable simulation. JSH nullified TLR4 activation mechanism and sequentially inhibited nuclear factor-κB (NF-κB) activation that involves the phosphorylation and degradation of inhibitory κBs and the nuclear import and transcriptional activity of NF-κB in LPS-activated macrophages. Moreover, JSH suppressed NF-κB-target inflammatory genes such as inducible nitric oxide synthase, cyclooxygenase-2, interleukin-1β (IL-1β) and IL-6. Taken together, this study assigns the chalcone structure as an LPS antagonist binding to MD-2 with therapeutic potential against inflammatory conditions.

Cinnamaldehydes have been reported to induce apoptosis in human carcinomas through the generation of reactive oxygen species (ROS). 2'-benzoyloxycinnamaldehyde (BCA) has been reported to inhibit tumor formation in H-ras12V transgenic mice. To see the antitumor effects of BCA, BCA was administrated intraperitoneally (50 mg/kg) to H-ras12V transgenic mice for 3 wk, and it was found that the hepatic tumor volume and the total number of tumors were decreased in BCA-treated mice as compared to control H-ras12V transgenic mice. To identify possible target genes responsible for BCA antitumor effects in H-ras12V transgenic mice, cDNA microarray analyses were performed comparing gene expression between BCA treated and control transgenic mice. We found that 42 genes were downregulated, and 40 genes were upregulated in the BCA-treated transgenic mice. The downregulated genes included several genes involved in ROS regulation and immune response (aconitase, metallothionein-1, metallothionein-2, and purine nucleoside phosphorylase). The expression of ROS-related genes, metallothionein 1 and metallothionein 2, was decreased more than twofold with BCA treatment (P < 0.001). It was confirmed by RT-PCR and immunohistochemical analyses. The inhibition of tumor formation and growth in H-ras12V transgenic mice by BCA was mediated through inhibition of the expression of the ROS scavengers metallothionein 1 and metallothionein 2.

We generated gene expression data from the tissues of 50 gastric cancer patients, and applied meta-analysis and gene set analysis to this data and three other stomach cancer gene expression data sets to define the gene expression changes in gastric tumors. By meta-analysis we identified genes consistently changed in gastric carcinomas, while gene set analysis revealed consistently changed biological themes. Genes and gene sets involved in digestion, fatty acid metabolism, and ion transport were consistently down-regulated in gastric carcinomas, while those involved in cellular proliferation, cell cycle, and DNA replication were consistently up-regulated. We also found significant differences between the genes and gene sets expressed in diffuse and intestinal type gastric carcinoma. By gene set analysis of cytogenetic bands, we identified many chromosomal regions with possible gross chromosomal changes (amplifications or deletions). Similar analysis of transcription factor binding sites (TFBSs), revealed transcription factors that may have caused the observed gene expression changes in gastric carcinomas, and we confirmed the overexpression of one of these, E2F1, in many gastric carcinomas by tissue array and immunohistochemistry. We have incorporated the results of our meta- and gene set analyses into a web accessible database (http://human-genome.kribb.re.kr/stomach/).

The activity of beta-catenin/TCF, the key component of Wnt signaling pathway, is frequently deregulated in HCC, resulting in the activation of genes whose dysregulation has significant consequences on tumor development. Therefore, identifying the target genes of Wnt signaling is important for understanding beta-catenin-mediated carcinogenesis. We analyzed the transcriptome profile of human hepatoma cell lines using cDNA microarrays representing 15,127 unique, liver-enriched gene loci to identify the target genes of beta-catenin-mediated transcription (p

Trichostatin A (TSA) and S-adenosyl-L-homocysteine (AdoHcy) have been reported to affect histone modifications. To investigate the effects of two drugs that can reportedly affect chromatin remodeling, we analyzed the gene expression profiles of TSA and AdoHcy in a gastric cancer cell line using 14 K cDNA microarray. The significant analysis of microarray (SAM) identified 98 and 43 differentially expressed genes in TSA and AdoHcy treated sets, respectively, and selected genes were functionally classified. In the gastric cancer cell line, genes related to cell communication, cell growth/maintenance, and morphogenesis were highly expressed with TSA, and genes with cell growth/maintenance, metabolism, oxidoreductase activity were upregulated with AdoHcy. Genes downregulated with TSA included those controlling the cell cycle, cell growth/proliferation, DNA binding, and metabolism, whereas genes involved in calcium signaling, cell growth/proliferation, and metabolism were downregulated with AdoHcy. Furthermore, we identified the genes commonly expressed in both drug treatments. Compared to TSA, AdoHcy did not induce apoptosis in the SNU-16 gastric cancer cell line, and RT-PCR was performed for selective genes to confirm the microarray data. This gene expression profile analysis with TSA and AdoHcy should contribute to a greater understanding of the molecular mechanism of chromatin remodeling and cancer, and provide candidate genes for further studies involving the roles of histone modifications in gastric cancer.

Chromatin regulation underlies a variety of DNA metabolism processes, including transcription, recombination, repair, and replication. To perform a quantitative genetic analysis of chromatin accessibility, we obtained open chromatin profiles across 96 genetically different yeast strains by FAIRE (formaldehyde-assisted isolation of regulatory elements) assay followed by sequencing. While 5∼10% of open chromatin region (OCRs) were significantly affected by variations in their underlying DNA sequences, subtelomeric areas as well as gene-rich and gene-poor regions displayed high levels of sequence-independent variation. We performed quantitative trait loci (QTL) mapping using the FAIRE signal for each OCR as a quantitative trait. While individual OCRs were associated with a handful of specific genetic markers, gene expression levels were associated with many regulatory loci. We found multi-target trans-loci responsible for a very large number of OCRs, which seemed to reflect the widespread influence of certain chromatin regulators. Such regulatory hotspots were enriched for known regulatory functions, such as recombinational DNA repair, telomere replication, and general transcription control. The OCRs associated with these multi-target trans-loci coincided with recombination hotspots, telomeres, and gene-rich regions according to the function of the associated regulators. Our findings provide a global quantitative picture of the genetic architecture of chromatin regulation., Author Summary Quantitative trait loci (QTL) mapping is a genetic approach that allows the identification of genetic factors underlying a phenotype of interest. Genomic technologies such as DNA microarray and next-generation sequencing provide data that can be used for the analysis of multiple molecular phenotypes. For example, the expression levels of thousands of genes can be associated with subject-specific genome-wide genetic information in expression QTL mapping. Similarly, the genetic regulation of transcription factor binding or epigenetic mechanisms such as DNA methylation or chromatin structure has begun to be investigated. In particular, the mechanisms controlling chromatin accessibility have attracted special interest due to their importance in a variety of DNA regulation processes including recombination, repair, replication, and transcription. In this work, we sought to dissect the genetic architecture of chromatin accessibility regulation by harnessing the power of genetic and genomic techniques. By analyzing open (accessible) chromatin maps of multiple yeast individuals in association with their genetic backgrounds, we were able to characterize the regulatory structure of chromatin traits versus that of gene expression. Importantly, we observed that the genetic loci responsible for multiple open chromatin regions were enriched for known regulatory factors.

Previous studies have shown that chalcone has beneficial effects with antioxidant and anti-inflammatory activities. However, molecular basis for these effects not yet fully understood. In this study, a chalcone derivative, 2’, 4’-dihydroxy-6’-isopentyloxy chalcone (JSH 4-4) was discovered as an inhibitor of NF-kB activation. To demonstrate molecular target affected by JSH 4-4, we examined a homotypic interaction of TLR4, using Ba/F3 cells stably expressing TLR4-Flag, TLR4-GFP CD14 and MD-2, seperately. As a result, JSH 4-4 inhibited the LPS-induced TLR4 oligomerization. Moreover, we demonstrated that JSH 4-4 inhibit LPS-induced NF-kB activation via targeting LPS-binding site of MD-2. This mechanism of action could contribute to suppressive effect of JSH 4-4 on NF-kB-regulated production of inflammatory mediators such as nitric oxide, COX-2 and cytokines. TLR4-mediated NF-kB signalling occurs following sequential activation of IRAK family and TAK1. JSH 4-4 inhibited LPS-induced phosphorylation of IRAK-1 and TAK1. In addition, JSH 4-4 significantly inhibited IKK activity and phosphorylation of IkBa, resulting in sequential prevention of downstream events, including degradation of IkBs, nuclear translocation of NF-kB p65. In conclusion, JSH 4-4 inhibit NF-kB-regulated inflammatory mediators via targeting the receptor complex MD2-TLR4 in macrophages, thus JSH 4-4 might be considered as a potential agent for the treatment of NF-kB-associated inflammatory diseases.