Increasing levels of tissue hypoxia have been reported as a natural feature of the aging prostate gland and may be a risk factor for the development of prostate cancer. In this study, we have used ...PwR-1E benign prostate epithelial cells and an equivalently aged hypoxia-adapted PwR-1E sub-line to identify phenotypic and epigenetic consequences of chronic hypoxia in prostate cells. We have identified a significantly altered cellular phenotype in response to chronic hypoxia as characterized by increased receptor-mediated apoptotic resistance, the induction of cellular senescence, increased invasion and the increased secretion of IL-1β, IL6, IL8 and TNFα cytokines. In association with these phenotypic changes and the absence of HIF-1α protein expression, we have demonstrated significant increases in global levels of DNA methylation and H3K9 histone acetylation in these cells, concomitant with the increased expression of DNA methyltransferase DMNT3b and gene-specific changes in DNA methylation at key imprinting loci. In conclusion, we have demonstrated a genome-wide adjustment of DNA methylation and histone acetylation under chronic hypoxic conditions in the prostate. These epigenetic signatures may represent an additional mechanism to promote and maintain a hypoxic-adapted cellular phenotype with a potential role in tumour development.
We present an analysis of chromatin fiber structure across the human genome. Compact and open chromatin fiber structures were separated by sucrose sedimentation and their distributions analyzed by ...hybridization to metaphase chromosomes and genomic microarrays. We show that compact chromatin fibers originate from some sites of heterochromatin (C-bands), and G-bands (euchromatin). Open chromatin fibers correlate with regions of highest gene density, but not with gene expression since inactive genes can be in domains of open chromatin, and active genes in regions of low gene density can be embedded in compact chromatin fibers. Moreover, we show that chromatin fiber structure impacts on further levels of chromatin condensation. Regions of open chromatin fibers are cytologically decondensed and have a distinctive nuclear organization. We suggest that domains of open chromatin may create an environment that facilitates transcriptional activation and could provide an evolutionary constraint to maintain clusters of genes together along chromosomes.
Replication timing of the human genome Woodfine, Kathryn; Fiegler, Heike; Beare, David M. ...
Human molecular genetics,
01/2004, Letnik:
13, Številka:
2
Journal Article
Recenzirano
Odprti dostop
We have developed a directly quantitative method utilizing genomic clone DNA microarrays to assess the replication timing of sequences during the S phase of the cell cycle. The genomic resolution of ...the replication timing measurements is limited only by the genomic clone size and density. We demonstrate the power of this approach by constructing a genome-wide map of replication timing in human lymphoblastoid cells using an array with clones spaced at 1 Mb intervals and a high-resolution replication timing map of 22q with an array utilizing overlapping sequencing tile path clones. We show a positive correlation, both genome-wide and at a high resolution, between replication timing and a range of genome parameters including GC content, gene density and transcriptional activity.
Replication timing of the human genome Woodfine, Kathryn; Fiegler, Heike; Beare, David M. ...
Human molecular genetics,
03/2004, Letnik:
13, Številka:
5
Journal Article
Replication Timing of Human Chromosome 6 Woodfine, Kathryn; Beare, David M.; Ichimura, Koichi ...
Cell cycle (Georgetown, Tex.),
20/1/1/, Letnik:
4, Številka:
1
Journal Article
Recenzirano
Odprti dostop
Genomic microarrays have been used to assess DNA replication timing in a variety of eukaryotic organisms1-3. A replication timing map of the human genome has already been published at a 1Mb ...resolution3. Here we describe how the same method can be used to assess the replication timing of chromosome 6 with a greater resolution using an array of overlapping tile path clones. We report the replication timing map of the whole of chromosome 6 in general, and the MHC region in particular. Positive correlations are observed between replication timing and a number of genomic features including GC content, repeat content and transcriptional activity.
The 22q11 deletion syndrome (22q11DS) is a developmental syndrome comprising of heart, palate, thymus and parathyroid glands defects. Individuals with 22q11DS usually carry a 1.5- to 3-Mb ...heterozygous deletion on chromosome 22q11.2. However, there are many patients with features of 22q11DS without a known cause from conventional karyotype and FISH analysis. Six patients with features of 22q11DS, a normal chromosomal and FISH 22q11 analysis, were selected for investigation by microarray genomic comparative hybridisation (array CGH). Array-CGH is a powerful technology enabling detection of submicroscopic chromosome duplications and deletions by comparing a differentially labelled test sample to a control. The samples are co-hybridised to a microarray containing genomic clones and the resulting ratio of fluorescence intensities on each array element is proportional to the DNA copy number difference. No chromosomal changes were detected by hybridisation to a high resolution array representing chromosome 22q. However, one patient was found to have a 6-Mb deletion on 5q11.2 detected by a whole genome 1-Mb array. This deletion was confirmed with fluorescence in-situ hybridisation (FISH) and microsatellite marker analysis. It is the first deletion described in this region. The patient had tetralogy of Fallot, a bifid uvula and velopharyngeal insufficiency, short stature, learning and behavioural difficulties. This case shows the increased sensitivity of array CGH over detailed karyotype analysis for detection of chromosomal changes. It is anticipated that array CGH will improve the clinician's capacity to diagnose congenital syndromes with an unknown aetiology.
DNA microarrays are increasingly being used to investigate the functional role of chromatin. These studies are enhanced by the development of high-resolution arrays covering either the whole genome ...or specific regions of selected chromosomes with large insert clones, PCR products or oligonucleotides of around 100 bp or less. In combination with chromatin immunoprecipitation, this approach allows identification of protein binding for transcription factors, proteins involved in DNA replication and repair as well as sites of chromatin modification. Furthermore, by application of S phase fractions to genomic microarrays, replication timing can be estimated. Thus, microarrays can provide new information about chromosome structure and gene regulation.
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