Epigenetic modifications are essential regulators of biological processes. Decreased DNA methylation of OAS2 (2'-5'-Oligoadenylate Synthetase 2), encoding an antiviral protein, has been seen in psoriasis. To provide further insight into the epigenetic regulation of OAS2, we performed pyrosequencing to detect OAS2 DNA methylation status at 11 promoter and first exon located CpG sites in psoriasis (n = 12) and two common subtypes of squamous cell carcinoma (SCC) of the head and neck: tongue (n = 12) and tonsillar (n = 11). Compared to corresponding controls, a general hypomethylation was seen in psoriasis. In tongue and tonsillar SCC, hypomethylation was found at only two CpG sites, the same two sites that were least demethylated in psoriasis. Despite differences in the specific residues targeted for methylation/demethylation, OAS2 expression was upregulated in all conditions and correlations between methylation and expression were seen in psoriasis and tongue SCC. Distinctive methylation status at four successively located CpG sites within a genomic area of 63 bp reveals a delicately integrated epigenetic program and indicates that detailed analysis of individual CpGs provides additional information into the mechanisms of epigenetic regulation in specific disease states. Methylation analyses as clinical biomarkers need to be tailored according to disease-specific sites.

• MeSH
• 2',5'-oligoadenylátsynthetasa genetika   metabolismus   MeSH
• CpG ostrůvky * MeSH
• epigeneze genetická * MeSH
• lidé MeSH
• metylace DNA MeSH
• nádory hlavy a krku genetika   metabolismus   patologie   MeSH
• nádory jazyka genetika   metabolismus   patologie   MeSH
• promotorové oblasti (genetika) MeSH
• psoriáza genetika   metabolismus   patologie   MeSH
• retrospektivní studie MeSH
• spinocelulární karcinom genetika   metabolismus   patologie   MeSH
• studie případů a kontrol MeSH
• tonzilární nádory genetika   metabolismus   patologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 2',5'-oligoadenylátsynthetasa MeSH
• OAS2 protein, human MeSH Prohlížeč

Unmethylated CpG islands are known to keep adjacent promoters transcriptionally active. In the CpG island adjacent to the adenosine phosphoribosyltransferase gene, the protection against transcriptional silencing can be attributed to the short CpG-rich core element containing Sp1 binding sites. We report here the insertion of this CpG island core element, IE, into the long terminal repeat of a retroviral vector derived from Rous sarcoma virus, which normally suffers from progressive transcriptional silencing in mammalian cells. IE insertion into a specific position between enhancer and promoter sequences led to efficient protection of the integrated vector from silencing and gradual CpG methylation in rodent and human cells. Individual cell clones with IE-modified reporter vectors display high levels of reporter expression for a sustained period and without substantial variegation in the cell culture. The presence of Sp1 binding sites is important for the protective effect of IE, but at least some part of the entire antisilencing capacity is maintained in IE with mutated Sp1 sites. We suggest that this strategy of antisilencing protection by the CpG island core element may prove generally useful in retroviral vectors.

• MeSH
• biologické modely MeSH
• CpG ostrůvky * MeSH
• genetická transkripce * MeSH
• koncové repetice MeSH
• lidé MeSH
• mutace MeSH
• průtoková cytometrie MeSH
• ptačí sarkom genetika   virologie   MeSH
• ptáci MeSH
• reportérové geny MeSH
• transkripční faktor Sp1 metabolismus   MeSH
• umlčování genů * MeSH
• vazebná místa MeSH
• virus ptačí leukózy metabolismus   MeSH
• virus Rousova sarkomu metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• transkripční faktor Sp1 MeSH

Accurate assessment of neuroblastoma outcome prediction remains challenging. Therefore, this study aims at establishing novel prognostic tumor DNA methylation biomarkers. In total, 396 low- and high-risk primary tumors were analyzed, of which 87 were profiled using methyl-CpG-binding domain (MBD) sequencing for differential methylation analysis between prognostic patient groups. Subsequently, methylation-specific PCR (MSP) assays were developed for 78 top-ranking differentially methylated regions and tested on two independent cohorts of 132 and 177 samples, respectively. Further, a new statistical framework was used to identify a robust set of MSP assays of which the methylation score (i.e. the percentage of methylated assays) allows accurate outcome prediction. Survival analyses were performed on the individual target level, as well as on the combined multimarker signature. As a result of the differential DNA methylation assessment by MBD sequencing, 58 of the 78 MSP assays were designed in regions previously unexplored in neuroblastoma, and 36 are located in non-promoter or non-coding regions. In total, 5 individual MSP assays (located in CCDC177, NXPH1, lnc-MRPL3-2, lnc-TREX1-1 and one on a region from chromosome 8 with no further annotation) predict event-free survival and 4 additional assays (located in SPRED3, TNFAIP2, NPM2 and CYYR1) also predict overall survival. Furthermore, a robust 58-marker methylation signature predicting overall and event-free survival was established. In conclusion, this study encompasses the largest DNA methylation biomarker study in neuroblastoma so far. We identified and independently validated several novel prognostic biomarkers, as well as a prognostic 58-marker methylation signature.

• Klíčová slova
• DNA methylation, biomarker, neuroblastoma, prognosis,
• MeSH
• biologické markery analýza   MeSH
• CpG ostrůvky genetika   MeSH
• DNA nádorová genetika   MeSH
• kohortové studie MeSH
• kojenec MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé MeSH
• metylace DNA * MeSH
• nádorové buňky kultivované MeSH
• neuroblastom diagnóza   genetika   MeSH
• prognóza MeSH
• staging nádorů MeSH
• vazebná místa MeSH
• výpočetní biologie MeSH
• Check Tag
• kojenec MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• biologické markery MeSH
• DNA nádorová MeSH

Mutations can be induced by environmental factors but also arise spontaneously during DNA replication or due to deamination of methylated cytosines at CpG dinucleotides. Sites where mutations occur with higher frequency than would be expected by chance are termed hotspots while sites that contain mutations rarely are termed coldspots. Mutations are permanently scanned and repaired by repair systems. Among them, the mismatch repair targets base pair mismatches, which are discriminated from canonical base pairs by probing altered elasticity of DNA. Using biased molecular dynamics simulations, we investigated the elasticity of coldspots and hotspots motifs detected in human genes associated with inherited disorders, and also of motifs with Czech population hotspots and de novo mutations. Main attention was paid to mutations leading to G/T and A+/C pairs. We observed that hotspots without CpG/CpHpG sequences are less flexible than coldspots, which indicates that flexible sequences are more effectively repaired. In contrary, hotspots with CpG/CpHpG sequences exhibited increased flexibility as coldspots. Their mutability is more likely related to spontaneous deamination of methylated cytosines leading to C > T mutations, which are primarily targeted by base excision repair. We corroborated conclusions based on computer simulations by measuring melting curves of hotspots and coldspots containing G/T mismatch.

• Klíčová slova
• DNA bending, Muts protein, free energy calculations, hotspots–coldspots, mutations,
• MeSH
• CpG ostrůvky MeSH
• DNA chemie   genetika   MeSH
• lidé MeSH
• mutace * MeSH
• nukleotidové motivy * MeSH
• simulace molekulární dynamiky * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA MeSH

BACKGROUND: Long-term environmental exposure to metals leads to epigenetic changes and may increase risks to human health. The relationship between the type and level of metal exposure and epigenetic changes in subjects exposed to high concentrations of metals in the environment is not yet clear. The aim of our study is to find the possible association of environmental long-term exposure to metals with DNA methylation changes of genes related to immune response and carcinogenesis. We investigated the association of plasma levels of 21 essential and non-essential metals detected by ICP-MS and the methylation level of 654 CpG sites located on NFKB1, CDKN2A, ESR1, APOA5, IGF2 and H19 genes assessed by targeted bisulfite sequencing in a cohort of 40 subjects living near metal mining area and 40 unexposed subjects. Linear regression was conducted to find differentially methylated positions with adjustment for gender, age, BMI class, smoking and metal concentration. RESULTS: In the metal-exposed group, five CpGs in the NFKB1 promoter region were hypomethylated compared to unexposed group. Four differentially methylated positions (DMPs) were associated with multiple metals, two of them are located on NFKB1 gene, and one each on CDKN2A gene and ESR1 gene. Two DMPs located on NFKB1 (chr4:102500951, associated with Be) and IGF2 (chr11:2134198, associated with U) are associated with specific metal levels. The methylation status of the seven CpGs located on NFKB1 (3), ESR1 (2) and CDKN2A (2) positively correlated with plasma levels of seven metals (As, Sb, Zn, Ni, U, I and Mn). CONCLUSIONS: Our study revealed methylation changes in NFKB1, CDKN2A, IGF2 and ESR1 genes in individuals with long-term human exposure to metals. Further studies are needed to clarify the effect of environmental metal exposure on epigenetic mechanisms and pathways involved.

• Klíčová slova
• CDKN2A, DNA methylation, ESR1, Environmental exposure, IGF2, NFKB1, Toxic metal, Uranium,
• MeSH
• epigeneze genetická * MeSH
• karcinogeneze genetika   MeSH
• kovy * škodlivé účinky   MeSH
• lidé MeSH
• metylace DNA * MeSH
• NF-kappa B - podjednotka p50 genetika   MeSH
• tumor supresorové geny MeSH
• vystavení vlivu životního prostředí * škodlivé účinky   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kovy * MeSH
• NF-kappa B - podjednotka p50 MeSH
• NFKB1 protein, human MeSH Prohlížeč

BACKGROUND: DNA methylation is likely a key mechanism regulating changes in gene transcription in traits that show temporal fluctuations in response to environmental conditions. To understand the transcriptional role of DNA methylation we need simultaneous within-individual assessment of methylation changes and gene expression changes over time. Within-individual repeated sampling of tissues, which are essential for trait expression is, however, unfeasible (e.g. specific brain regions, liver and ovary for reproductive timing). Here, we explore to what extend between-individual changes in DNA methylation in a tissue accessible for repeated sampling (red blood cells (RBCs)) reflect such patterns in a tissue unavailable for repeated sampling (liver) and how these DNA methylation patterns are associated with gene expression in such inaccessible tissues (hypothalamus, ovary and liver). For this, 18 great tit (Parus major) females were sacrificed at three time points (n = 6 per time point) throughout the pre-laying and egg-laying period and their blood, hypothalamus, ovary and liver were sampled. RESULTS: We simultaneously assessed DNA methylation changes (via reduced representation bisulfite sequencing) and changes in gene expression (via RNA-seq and qPCR) over time. In general, we found a positive correlation between changes in CpG site methylation in RBCs and liver across timepoints. For CpG sites in close proximity to the transcription start site, an increase in RBC methylation over time was associated with a decrease in the expression of the associated gene in the ovary. In contrast, no such association with gene expression was found for CpG site methylation within the gene body or the 10 kb up- and downstream regions adjacent to the gene body. CONCLUSION: Temporal changes in DNA methylation are largely tissue-general, indicating that changes in RBC methylation can reflect changes in DNA methylation in other, often less accessible, tissues such as the liver in our case. However, associations between temporal changes in DNA methylation with changes in gene expression are mostly tissue- and genomic location-dependent. The observation that temporal changes in DNA methylation within RBCs can relate to changes in gene expression in less accessible tissues is important for a better understanding of how environmental conditions shape traits that temporally change in expression in wild populations.

• Klíčová slova
• Accessible and inaccessible tissues, DNA methylation, Great tit, RNA expression, Tissue-specific and tissue-general temporal changes,
• MeSH
• CpG ostrůvky MeSH
• metylace DNA MeSH
• Passeriformes * genetika   MeSH
• RNA MeSH
• rozmnožování MeSH
• zpěvní ptáci * genetika   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• RNA MeSH

Toll-like receptor (TLR) signaling relies on Toll/interleukin-1 receptor homology (TIR) domain-containing adaptor proteins that recruit downstream signaling molecules to generate tailored immune responses. In addition, the palmitoylated transmembrane adaptor protein family member Scimp acts as a non-TIR-containing adaptor protein in macrophages, scaffolding the Src family kinase Lyn to enable TLR phosphorylation and proinflammatory signaling responses. Here we report the existence of a smaller, naturally occurring translational variant of Scimp (Scimp TV1), which is generated through leaky scanning and translation at a downstream methionine. Scimp TV1 also scaffolds Lyn, but in contrast to full-length Scimp, it is basally rather than lipopolysaccharide (LPS)-inducibly phosphorylated. Macrophages from mice that selectively express Scimp TV1, but not full-length Scimp, have impaired sustained LPS-inducible cytokine responses. Furthermore, in granulocyte macrophage colony-stimulating factor-derived myeloid cells that express high levels of Scimp, selective overexpression of Scimp TV1 enhances CpG DNA-inducible cytokine production. Unlike full-length Scimp that localizes to the cell surface and filopodia, Scimp TV1 accumulates in intracellular compartments, particularly the Golgi. Moreover, this variant of Scimp is not inducibly phosphorylated in response to CpG DNA, suggesting that it may act via an indirect mechanism to enhance TLR9 responses. Our findings thus reveal the use of alternative translation start sites as a previously unrecognized mechanism for diversifying TLR responses in the innate immune system.

• Klíčová slova
• Adaptor protein, CpG DNA, Toll-like receptor, alternative translation start site, macrophage, protein translation,
• MeSH
• DNA metabolismus   MeSH
• makrofágy metabolismus   MeSH
• myši MeSH
• signální transdukce * MeSH
• skupina kinas odvozených od src-genu metabolismus   MeSH
• toll-like receptory * metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA MeSH
• skupina kinas odvozených od src-genu MeSH
• toll-like receptory * MeSH

The nucleotide sequence of an 8.2-kb DNA fragment from the 5' proximal part of the chicken myb proto-oncogene spanning 1761 nucleotides upstream and 6436 nucleotides downstream from a presumed c-myb initiation codon was determined. A 3.3-kb G + C-rich region found in this sequence had also other features characterizing CpG islands, i.e. no CpG underrepresentation and lack of CpG methylation. In haematopoietic tissues c-myb mRNA synthesis starts in two major regions of the CpG island, namely 98 to 108 and 143 to 145 nucleotides upstream from the c-myb initiation codon. These two regions are in or close to the 124-bp evolutionarily conserved element located in the middle part of the CpG island. No alternative splicing of the 5' end of c-myb mRNA suggested earlier (1,2) was observed. The c-myb promoter contains neither TATA nor CAAT box-like structures at the usual positions. Instead, numerous potential Sp1 factor binding sites were found both upstream and downstream from the transcription initiation sites. Moreover, consensus v-myb protein DNA-binding sites were revealed in the promoter region and in sequences downstream from it.

• MeSH
• dinukleosidfosfáty analýza   MeSH
• endonukleasy specifické pro jednořetězcové nukleové kyseliny MeSH
• endonukleasy MeSH
• erytrocyty analýza   MeSH
• genetická transkripce * MeSH
• klonování DNA MeSH
• kur domácí MeSH
• molekulární sekvence - údaje MeSH
• myši MeSH
• orgánová specificita MeSH
• promotorové oblasti (genetika) MeSH
• protoonkogenní proteiny c-myb MeSH
• protoonkogenní proteiny genetika   MeSH
• protoonkogeny * MeSH
• restrikční mapování MeSH
• RNA čepičky genetika   MeSH
• sekvence nukleotidů MeSH
• sekvenční homologie nukleových kyselin MeSH
• thymus analýza   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• cytidylyl-3'-5'-guanosine MeSH Prohlížeč
• dinukleosidfosfáty MeSH
• endonukleasy specifické pro jednořetězcové nukleové kyseliny MeSH
• endonukleasy MeSH
• protoonkogenní proteiny c-myb MeSH
• protoonkogenní proteiny MeSH
• RNA čepičky MeSH

Overwhelming evidence supports the theory that inflammatory bowel disease (IBD) is caused by a complex interplay between genetic predispositions of multiple genes, combined with an abnormal interaction with environmental factors. It is becoming apparent that epigenetic factors can have a significant contribution in the pathogenesis of disease. Changes in the methylation state of IBD-associated genes could significantly alter levels of gene expression, potentially contributing to disease onset and progression. We have explored the role of DNA methylation in IBD pathogenesis. DNA methylation profiles (1505 CpG sites of 807 genes) of matched diseased (n = 26) and non-diseased (n = 26) intestinal tissues from 26 patients with IBD [Crohn's disease (CD) n = 9, ulcerative colitis (UC) n = 17] were profiled using the GoldenGate™ methylation assay. After an initial identification of a panel of 50 differentially methylated CpG sites from a training set (14 non-diseased and 14 diseased tissues) and subsequent validation with a testing set (12 non-diseased and 12 diseased tissues), we identified seven CpG sites that are differentially methylated in intestinal tissues of IBD patients. We have also identified changes in DNA methylation associated with the two major IBD subtypes, CD and UC. This study reports IBD-associated changes in DNA methylation in intestinal tissue, which may be disease subtype-specific.

• MeSH
• CpG ostrůvky MeSH
• Crohnova nemoc genetika   MeSH
• epigeneze genetická MeSH
• idiopatické střevní záněty genetika   MeSH
• lidé MeSH
• metylace DNA * MeSH
• shluková analýza MeSH
• stanovení celkové genové exprese MeSH
• střevní sliznice metabolismus   MeSH
• ulcerózní kolitida genetika   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The transcriptional activity of an integrated retroviral copy strongly depends on the adjacent host-cell DNA at the site of integration. Transcribed DNA loci as well as cis-acting sequences like enhancers or CpG islands usually permit expression of nearby integrated proviruses. In contrast, proviruses residing close to cellular silencers tend to transcriptional silencing and CpG methylation. Little is known, however, about the influence of provirus integration on the target sequence in the host genome. Here, we report interesting features of a simplified Rous sarcoma virus integrated into a non-transcribed hypermethylated DNA sequence in the Syrian hamster genome. After integration, CpG methylation of this sequence has been lost almost completely and hypomethylated DNA permits proviral transcription and hamster cell transformation by the proviral v-src oncogene. This, however, is not a stable state, and non-transformed revertants bearing transcriptionally silenced proviruses segregate with a high rate. The provirus silencing is followed by DNA methylation of both provirus regulatory regions and adjacent cellular sequences. This CpG methylation is very dense and resistant to the demethylation effects of 5-aza-2(')-deoxycytidine and/or trichostatin A. Our description exemplifies the capacity of retroviruses/retroviral vectors to overcome, at least transiently, negative position effects of DNA methylation at the site of integration.

• MeSH
• agar farmakologie   MeSH
• azacytidin analogy a deriváty   farmakologie   MeSH
• buněčné linie MeSH
• CpG ostrůvky MeSH
• decitabin MeSH
• genetická transkripce MeSH
• geny src genetika   MeSH
• histondeacetylasy metabolismus   MeSH
• inhibitory enzymů farmakologie   MeSH
• inhibitory syntézy proteinů farmakologie   MeSH
• integrace viru * MeSH
• koncové repetice MeSH
• křečci praví MeSH
• křeček rodu Mesocricetus MeSH
• kyseliny hydroxamové farmakologie   MeSH
• metylace DNA * MeSH
• modely genetické MeSH
• molekulární sekvence - údaje MeSH
• nádorové buněčné linie MeSH
• Retroviridae genetika   MeSH
• siřičitany farmakologie   MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• agar MeSH
• azacytidin MeSH
• decitabin MeSH
• histondeacetylasy MeSH
• inhibitory enzymů MeSH
• inhibitory syntézy proteinů MeSH
• kyseliny hydroxamové MeSH
• siřičitany MeSH
• trichostatin A MeSH Prohlížeč