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Aim: Fetal skin is known to heal without scarring. In mice, the phenomenon is observed until the 16–17 day of gestation – the day of transition from scarless to normal healing. The study aims to identify key methylome and transcriptome changes following the transition. Materials & methods: Methylome and transcriptome profiles were analyzed in murine dorsal skin using microarray approach. Results & conclusion: The genes associated with inflammatory response and hyaluronate degradation showed increased DNA methylation before the transition, while those involved in embryonic morphogenesis, neuron differentiation and synapse functions did so after. A number of the methylome alterations were retained until adulthood and correlated with gene expression, while the functional associations imply that scarless healing depends on epigenetic regulation.

The molecular bases of miR-182 deregulation in epithelial ovarian cancers (EOCs) remain unknown and its diagnostic or prognostic role in EOCs is still unclear. We performed miR-182 expression analysis using a microarray approach and real-time PCR (qPCR). We also used array comparative genomic hybridization and methylated DNA immunoprecipitation to study copy number changes and methylation aberrations within coding locus/promoter sequences of miR-182 in EOC tissues, respectively. We have found that miR-182 expression is significantly increased in EOC (P

Somatic mosaicism for DNA copy-number alterations (SMC-CNAs) is defined as gain or loss of chromosomal segments in somatic cells within a single organism. As cells harboring SMC-CNAs can undergo clonal expansion, it has been proposed that SMC-CNAs may contribute to the predisposition of these cells to genetic disease including cancer. Herein, the gross genomic alterations (>500 kbp) were characterized in uninvolved mammary glandular tissue from 59 breast cancer patients and matched samples of primary tumors and lymph node metastases. Array-based comparative genomic hybridization showed 10% (6/59) of patients harbored one to 359 large SMC-CNAs (mean: 1,328 kbp; median: 961 kbp) in a substantial portion of glandular tissue cells, distal from the primary tumor site. SMC-CNAs were partially recurrent in tumors, albeit with considerable contribution of stochastic SMC-CNAs indicating genomic destabilization. Targeted resequencing of 301 known predisposition and somatic driver loci revealed mutations and rare variants in genes related to maintenance of genomic integrity: BRCA1 (p.Gln1756Profs*74, p.Arg504Cys), BRCA2 (p.Asn3124Ile), NCOR1 (p.Pro1570Glnfs*45), PALB2 (p.Ser500Pro), and TP53 (p.Arg306*). Co-occurrence of gross SMC-CNAs along with point mutations or rare variants in genes responsible for safeguarding genomic integrity highlights the temporal and spatial neoplastic potential of uninvolved glandular tissue in breast cancer patients.

Aim: We aimed to identify the pivotal differences in the DNA methylation profiles between the regeneration capable MRL/MpJ mouse and reference mouse strains. Materials & methods: Global DNA methylation profiling was performed in ear pinnae, bone marrow, spleen, liver and heart from uninjured adult females of the MRL/MpJ and C57BL/6J and BALB/c. Results & conclusion: A number of differentially methylated regions (DMRs) distinguishing between the MRL/MpJ mouse and both references were identified. In the ear pinnae, the DMRs were enriched in genes associated with development, inflammation and apoptosis, and in binding sites of transcriptional modulator Smad1. Several DMRs overlapped previously mapped quantitative trait loci of regenerative capability. The results suggest potential epigenetic determinants of regenerative phenomenon.

Epigenetic regulation plays essential role in cell differentiation and dedifferentiation, which are the intrinsic processes involved in regeneration. To investigate the epigenetic basis of regeneration capacity, we choose DNA methylation as one of the most important epigenetic mechanisms and the MRL/MpJ mouse as a model of mammalian regeneration known to exhibit enhanced regeneration response in different organs. We report the comparative analysis of genomic DNA methylation profiles of the MRL/MpJ and the control C57BL/6J mouse. Methylated DNA immunoprecipitation followed by microarray analysis using the Nimblegen '3 × 720 K CpG Island Plus RefSeq Promoter' platform was applied in order to carry out genome-wide DNA methylation profiling covering 20 404 promoter regions. We identified hundreds of hypo- and hypermethylated genes and CpG islands in the heart, liver, and spleen, and 37 of them in the three tissues. Decreased inter-tissue diversification and the shift of DNA methylation balance upstream the genes distinguish the genomic methylation patterns of the MRL/MpJ mouse from the C57BL/6J. Homeobox genes and a number of other genes involved in embryonic morphogenesis are significantly overrepresented among the genes hypomethylated in the MRL/MpJ mouse. These findings indicate that epigenetic patterning might be a likely molecular basis of regeneration capability in the MRL/MpJ mouse.

The molecular bases of miR-182 deregulation in epithelial ovarian cancers (EOCs) remain unknown and its diagnostic or prognostic role in EOCs is still unclear. We performed miR-182 expression analysis using a microarray approach and real-time PCR (qPCR). We also used array comparative genomic hybridization and methylated DNA immunoprecipitation to study copy number changes and methylation aberrations within coding locus/promoter sequences of miR-182 in EOC tissues, respectively. We have found that miR-182 expression is significantly increased in EOC (P 0.00001) and that higher miR-182 expression in EOC is linked with significantly shorter overall survival (P = 0.026). The methylation of miR-182 promoter was significantly associated with lower miR-182 expression in EOC tissues (P = 0.045). miR-182 over-expression is connected with copy number (CN) gains of this miRNA coding sequences in EOC (P = 0.002), and the number of PRDM5 copies is significantly and inversely correlated with miR-182 expression evaluated by qPCR (R = -0.615, P = 0.009). We conclude that the aberrant miR-182 expression in EOC may be due to CN gains within its coding locus. The miR-182 promoter is rarely methylated in EOC, and its methylation status is associated with lower miR-182 expression. Deletion of the PRDM5 locus may play a supportive role in miR-182 overexpression in EOC. miR-182 is an unfavorable prognostic factor in EOC. © 2016 Wiley Periodicals, Inc.

Reuse of materials in DNA hybridization-based methods has been known since the advent of Southern membranes. Array-based comparative genomic hybridization is essentially Southern hybridization with multiple probes immobilized on a solid surface. We show that comparative genomic hybridization microarrays fabricated with maskless array synthesizer technology can be used up to four times with the application of 1,3-dimethylurea as an array-stripping agent. We reproducibly detected chromosomal aberrations (0.6–22.4 Mb in size) in four hybridization rounds using regenerated microarray slides. We also demonstrated that regenerated arrays can detect smaller alterations (16–200 kbp), such as common copy number variants, as well as complex aberration profiles in tumor DNA.

Background Signal transducer and activator of transcription 3 (STAT3) is constitutively activated in many human tumors, including gliomas, and regulates the expression of genes implicated in proliferation, survival, apoptosis, angiogenesis and immune regulation. Only a small fraction of those genes has been proven to be direct STAT3 targets. In gliomas, STAT3 can play tumor suppressive or oncogenic roles depending on the tumor genetic background with target genes being largely unknown. Results We used chromatin immunoprecipitation, promoter microarrays and deep sequencing to assess the genome-wide occupancy of phospho (p)-Stat3 and epigenetic modifications of H3K4me3 and H3ac in C6 glioma cells. This combined assessment identified a list of 1200 genes whose promoters have both Stat3 binding sites and epigenetic marks characteristic for actively transcribed genes. The Stat3 and histone markings data were also intersected with a set of microarray data from C6 glioma cells after inhibition of Jak2/Stat3 signaling. Subsequently, we found 284 genes characterized by p-Stat3 occupancy, activating histone marks and transcriptional changes. Novel genes were screened for their potential involvement in oncogenesis, and the most interesting hits were verified by ChIP-PCR and STAT3 knockdown in human glioma cells. Conclusions Non-random association between silent genes, histone marks and p-Stat3 binding near transcription start sites was observed, consistent with its repressive role in transcriptional regulation of target genes in glioma cells with specific genetic background.

Most neurological diseases are associated with chronic inflammation initiated by the activation of microglia, which produce cytotoxic and inflammatory factors. Signal transducers and activators of transcription (STATs) are potent regulators of gene expression but contribution of particular STAT to inflammatory gene expression and STAT-dependent transcriptional networks underlying brain inflammation need to be identified. In the present study, we investigated the genomic distribution of Stat binding sites and the role of Stats in the gene expression in lipopolysaccharide (LPS)-activated primary microglial cultures. Integration of chromatin immunoprecipitation-promoter microarray data and transcriptome data revealed novel Stat-target genes including Jmjd3, Ccl5, Ezr, Ifih1, Irf7, Uba7, and Pim1. While knockdown of individual Stat had little effect on the expression of tested genes, knockdown of both Stat1 and Stat3 inhibited the expression of Jmjd3 and inflammatory genes. Transcriptional regulation of Jmjd3 by Stat1 and Stat3 is a novel mechanism crucial for launching inflammatory responses in microglia. The effects of Jmjd3 on inflammatory gene expression were independent of its H3K27me3 demethylase activity. Forced expression of constitutively activated Stat1 and Stat3 induced the expression of Jmjd3, inflammation-related genes, and the production of pro-inflammatory cytokines as potently as lipopolysacharide. Gene set enrichment and gene function analysis revealed categories linked to the inflammatory response in LPS and Stat1C + Stat3C groups. We defined upstream pathways that activate STATs in response to LPS and demonstrated contribution of Tlr4 and Il-6 and interferon-γ signaling. Our findings define novel direct transcriptional targets of Stat1 and Stat3 and highlight their contribution to inflammatory gene expression. Key Message Combined analysis of genomic Stat occupancy and transcriptome revealed novel Stat target genes in LPS-induced microglia. Jmjd3 transcription factor is a novel transcriptional target of Stat1 and Stat3. Stat1 and Stat3 cooperate with Jmjd3 to induce the expression of pro-inflammatory genes. Constitutively active Stat1 and Stat3 fully mimic the LPS-induced upregulation of inflammatory genes and secretion of cytokines. Electronic supplementary material The online version of this article (doi:10.1007/s00109-013-1090-5) contains supplementary material, which is available to authorized users.

Structural variations are among the most frequent interindividual genetic differences in the human genome. The frequency and distribution of de novo somatic structural variants in normal cells is, however, poorly explored. Using age-stratified cohorts of 318 monozygotic (MZ) twins and 296 single-born subjects, we describe age-related accumulation of copy-number variation in the nuclear genomes in vivo and frequency changes for both megabase- and kilobase-range variants. Megabase-range aberrations were found in 3.4% (9 of 264) of subjects ≥60 years old; these subjects included 78 MZ twin pairs and 108 single-born individuals. No such findings were observed in 81 MZ pairs or 180 single-born subjects who were ≤55 years old. Recurrent region- and gene-specific mutations, mostly deletions, were observed. Longitudinal analyses of 43 subjects whose data were collected 7-19 years apart suggest considerable variation in the rate of accumulation of clones carrying structural changes. Furthermore, the longitudinal analysis of individuals with structural aberrations suggests that there is a natural self-removal of aberrant cell clones from peripheral blood. In three healthy subjects, we detected somatic aberrations characteristic of patients with myelodysplastic syndrome. The recurrent rearrangements uncovered here are candidates for common age-related defects in human blood cells. We anticipate that extension of these results will allow determination of the genetic age of different somatic-cell lineages and estimation of possible individual differences between genetic and chronological age. Our work might also help to explain the cause of an age-related reduction in the number of cell clones in the blood; such a reduction is one of the hallmarks of immunosenescence. © 2012 The American Society of Human Genetics.

The sporulation process is a complex genetic developmental program leading to profound changes in global gene expression profile. In this work, we have applied genome-wide microarray approach for transcriptional profiling of Bacillus subtilis strain lacking a gene coding for PrpE protein phosphatase. This protein was previously shown to be involved in the regulation of germination of B. subtilis spores. Moreover, the deletion of prpE gene resulted in changing the resistance properties of spores. Our results provide genome-wide insight into the influence of this protein phosphatase on the physiology of B. subtilis cells. Although the precise role of PrpE in shaping the observed phenotype of ΔprpE mutant strain still remains beyond the understanding, our experiments brought observations of possible indirect implication of this protein in the regulation of cell motility and chemotaxis, as well as the development of competence. Surprisingly, prpE-deleted cells showed elevated level of general stress response, which turned out to be growth medium specific. Electronic supplementary material The online version of this article (doi:10.1007/s00438-013-0763-7) contains supplementary material, which is available to authorized users.

Background The MRL/MpJ mouse is a laboratory inbred strain known for regenerative abilities which are manifested by scarless closure of ear pinna punch holes. Enhanced healing responses have been reported in other organs. A remarkable feature of the strain is that the adult MRL/MpJ mouse retains several embryonic biochemical characteristics, including increased expression of stem cell markers. Results We explored the transcriptome of the MRL/MpJ mouse in the heart, liver, spleen, bone marrow and ears. We used two reference strains, thus increasing the chances to discover the genes responsible for the exceptional properties of the regenerative strain. We revealed several distinctive characteristics of gene expression patterns in the MRL/MpJ mouse, including the repression of immune response genes, the up-regulation of those associated with retinol metabolism and PPAR signalling, as well as differences in expression of the genes engaged in wounding response. Another crucial finding is that the gene expression patterns in the adult MRL/MpJ mouse and murine neonates share a number of parallels, which are also related to immune and wounding response, PPAR pathway, and retinol metabolism. Conclusions Our results indicate the significance of retinol signalling and neonatal transcriptomic relics as the distinguishing features of the MRL/MpJ mouse. The possibility that retinoids could act as key regulatory molecules in this regeneration model brings important implications for regenerative medicine. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-2075-2) contains supplementary material, which is available to authorized users.

Background The neonatal murine heart is able to regenerate after severe injury; this capacity however, quickly diminishes and it is lost within the first week of life. DNA methylation is an epigenetic mechanism which plays a crucial role in development and gene expression regulation. Under investigation here are the changes in DNA methylation and gene expression patterns which accompany the loss of regenerative potential. Results The MeDIP-chip (methylated DNA immunoprecipitation microarray) approach was used in order to compare global DNA methylation profiles in whole murine hearts at day 1, 7, 14 and 56 complemented with microarray transcriptome profiling. We found that the methylome transition from day 1 to day 7 is characterized by the excess of genomic regions which gain over those that lose DNA methylation. A number of these changes were retained until adulthood. The promoter genomic regions exhibiting increased DNA methylation at day 7 as compared to day 1 are significantly enriched in the genes critical for heart maturation and muscle development. Also, the promoter genomic regions showing an increase in DNA methylation at day 7 relative to day 1 are significantly enriched with a number of transcription factors binding motifs including those of Mfsd6l, Mef2c, Meis3, Tead4, and Runx1. Conclusions The results indicate that the extensive alterations in DNA methylation patterns along the development of neonatal murine hearts are likely to contribute to the decline of regenerative capabilities observed shortly after birth. This conclusion is supported by the evidence that an increase in DNA methylation in the neonatal murine heart from day 1 to day 7 occurs in the promoter regions of genes playing important roles in cardiovascular system development. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-2545-1) contains supplementary material, which is available to authorized users.