CpG islands, genes and isochores in the genomes of vertebrates

We have shown that human genes associated with CpG islands increase in number as they increase in % of guanine + cytosine (GC) levels, and that most genes associated with CpG islands are located in the GC-richest compartment of the human genome. This is an independent confirmation of the concentration gradient of CpG islands (detected as HpaII tiny fragments, or HTF) which was demonstrated in the genome of warm-blooded vertebrates [A?ssani and Bernardi, Gene 106 (1991) 173-183]. We then reassessed the location of CpG islands using the data currently available and confirmed that CpG islands are most frequently located in the 5'-flanking sequences of genes and that they overlap genes to variable extents. We have shown that such extents increase with the increasing GC levels of genes, the GC-richest genes being completely included in CpG islands. Under such circumstances, we have investigated the properties of the 'extragenic' CpG islands located in the 5'-flanking segments of homologous genes from both warm- and cold-blooded vertebrates. We have confirmed that, in cold-blooded vertebrates, CpG islands are often absent; when present, they have lower GC and CpG levels; the latter attain, however, statistically expected values. Finally, we have shown that CpG doublets increase with the increasing GC of exons, introns and intergenic sequences (including 'extragenic' CpG islands) in the genomes from both warm- and cold-blooded vertebrates. The correlations found are the same for both classes of vertebrates, and are similar for exons, introns and intergenic sequences (including 'extragenic' CpG islands). The findings just outlined indicate that the origin and evolution of CpG islands in the vertebrate genome are associated with compositional transitions (GC increases) in genes and isochores.     

Gene-associated CpG islands in plants as revealed by analyses of genomic sequences

We screened plant genome sequences, primarily from rice and Arabidopsis thaliana, for CpG islands, and identified DNA segments rich in CpG dinucleotides within these sequences. These CpG-rich clusters appeared in the analysed sequences as discrete peaks and occurred at the frequencies of one per 4.7 kb in rice and one per 4.0 kb in A. thaliana. In rice and A. thaliana, most of the CpG-rich clusters were associated with genes, which suggests that these clusters are useful landmarks in genome sequences for identifying genes in plants with small genomes. In contrast, in plants with larger genomes, only a few of the clusters were associated with genes. These plant CpG-rich clusters satisfied the criteria used for identifying human CpG islands, which suggests that these CpG clusters may be regarded as plant CpG islands. The position of each island relative to the 5'-end of its associated gene varied considerably. Genes in the analysed sequences were grouped into five classes according to the position of the CpG islands within their associated genes. A large proportion of the genes belonged to one of two classes, in which a CpG island occurred near the 5'-end of the gene or covered the whole gene region. The position of a plant CpG island within its associated gene appeared to be related to the extent of tissue-specific expression of the gene; the CpG islands of most of the widely expressed rice genes occurred near the 5'-end of the genes.     

Alu-PCR Approach to Isolating NotI-Linking Clones from the 3p14-p21 Region Frequently Deleted in Renal Cell Carcinoma

In the mammalian genome CpG islands are associated with functional genes and cloning of these islands could be an alternative approach for cloning functional genes. Recently we have developed a new approach for cloning CpG islands and constructing NotI linking libraries. We have initiated the construction of a NotI restriction map for chromosome 3, especially focusing on the rearrangements in the 3p14-p21 region, which are associated with different malignancies. CpG islands from this region are useful for isolation of candidate tumor suppressor genes that map to this region and for isolating NotI-linking clones from 3p14-p21 for mapping purposes. Here we suggest a modification of Alu-PCR as an approach to isolating Not I sites (e.g., CpG islands) from defined regions of the chromosome. Instead of using whole chromosomal DNA for Alu-PCR, we have used representative NotI-linking libraries from hybrid cell lines containing either whole or deleted human chromosome 3 (MCH903.1 and MCH924.4, respectively). This decreases the complexity of the Alu-PCR products 10-100 times compared to the whole human genome. Using this modification, we can isolate NotI-linking clones, which are natural markers on the chromosome, rather than random genomic fragments. Among eight clones selected by this method, seven were from the region deleted in MCH924.4. The results clearly demonstrate the feasibility of Alu-PCR for isolating CpG islands from defined regions of the genome.     

Particle swarm optimization with reinforcement learning for the prediction of CpG islands in the human genome

Background

Regions with abundant GC nucleotides, a high CpG number, and a length greater than 200 bp in a genome are often referred to as CpG islands. These islands are usually located in the 5′ end of genes. Recently, several algorithms for the prediction of CpG islands have been proposed.

Methodology/Principal Findings

We propose here a new method called CPSORL to predict CpG islands, which consists of a complement particle swarm optimization algorithm combined with reinforcement learning to predict CpG islands more reliably. Several CpG island prediction tools equipped with the sliding window technique have been developed previously. However, the quality of the results seems to rely too much on the choices that are made for the window sizes, and thus these methods leave room for improvement.

Conclusions/Significance

Experimental results indicate that CPSORL provides results of a higher sensitivity and a higher correlation coefficient in all selected experimental contigs than the other methods it was compared to (CpGIS, CpGcluster, CpGProd and CpGPlot). A higher number of CpG islands were identified in chromosomes 21 and 22 of the human genome than with the other methods from the literature. CPSORL also achieved the highest coverage rate (3.4%). CPSORL is an application for identifying promoter and TSS regions associated with CpG islands in entire human genomic. When compared to CpGcluster, the islands predicted by CPSORL covered a larger region in the TSS (12.2%) and promoter (26.1%) region. If Alu sequences are considered, the islands predicted by CPSORL (Alu) covered a larger TSS (40.5%) and promoter (67.8%) region than CpGIS. Furthermore, CPSORL was used to verify that the average methylation density was 5.33% for CpG islands in the entire human genome.     

CpG islands: features and distribution in the genomes of vertebrates

We have investigated the distribution of unmethylated CpG islands in vertebrate genomes fractionated according to their base composition. Genomes from warm-blooded vertebrates (man, mouse and chicken) are characterized by abundant CpG islands, whose frequency increases in DNA fractions of increasing % of guanine + cytosine; % G + C (GC), in parallel with the distribution of genes and CpG doublets. Small, yet significant, differences in the distribution of CpG islands were found in the three genomes. In contrast, genomes from cold-blooded vertebrates (two reptiles, one amphibian, and two fishes) were characterized by an extreme scarcity or absence of CpG islands (detected in these experiments as HpaII tiny fragments or HTF). CpG islands associated with homologous genes from cold- and warm-blooded vertebrates were then compared by analyzing CpG frequencies, GC levels, HpaII sites, rare-cutter sites and G/C boxes (GGGGCGGGGC and closely related motifs) in sequences available in gene banks. Small, yet significant, differences were again detected among the CpG islands associated with homologous genes from warm-blooded vertebrates, in that CpG islands associated with mouse or rat genes often showed low CpG and/or GC levels, as well as low numbers of HpaII sites, rare-cutter sites and G/C boxes, compared to homologous human genes; more rarely, CpG islands were just absent. As far as cold-blooded vertebrates were concerned, a number of genes showed CpG islands, which exhibited a much lower frequency of CpG doublets than that found in CpG islands of warm-blooded vertebrates, but still approached the statistically expected frequency; none of the other features of CpG islands associated with genes from warm-blooded vertebrates were present. Other genes did not show any associated CpG islands, unlike their homologues from warm-blooded vertebrates.     

CpG islands of the X chromosome are gene associated.

Unmethylated CpG rich islands are a feature of vertebrate DNA: they are associated with housekeeping and many tissue specific genes. CpG islands on the active X chromosome of mammals are also unmethylated. However, islands on the inactive X chromosome are heavily methylated. We have identified a CpG island in the 5' region of the G6PD gene, and two islands forty Kb 3' from the G6PD gene, on the human X chromosome. Expression of the G6PD gene is associated with concordant demethylation of all three CpG islands. We have shown that one of the two islands is in the promoter region of a housekeeping gene, GdX. In this paper we show that the second CpG island is also associated with a gene, P3. The P3 gene has no homology to previously described genes. It is a single copy, 4 kb gene, conserved in evolution, and it has the features of a housekeeping two genes is within the CpG island and that sequences in the islands have promoter function.     

Methylation detection oligonucleotide microarray analysis: a high-resolution method for detection of CpG island methylation

Methylation of CpG islands associated with genes can affect the expression of the proximal gene, and methylation of non-associated CpG islands correlates to genomic instability. This epigenetic modification has been shown to be important in many pathologies, from development and disease to cancer. We report the development of a novel high-resolution microarray that detects the methylation status of over 25 000 CpG islands in the human genome. Experiments were performed to demonstrate low system noise in the methodology and that the array probes have a high signal to noise ratio. Methylation measurements between different cell lines were validated demonstrating the accuracy of measurement. We then identified alterations in CpG islands, both those associated with gene promoters, as well as non-promoter-associated islands in a set of breast and ovarian tumors. We demonstrate that this methodology accurately identifies methylation profiles in cancer and in principle it can differentiate any CpG methylation alterations and can be adapted to analyze other species.