Isochore Boundaries Research Articles

Prediction of replication time zones at single nucleotide resolution in the human genome

The human genome is structured at multiple levels: it is organized into a series of replication time zones, and meanwhile it is composed of isochores. Accumulating evidence suggests a match between these two genome features. Based on newly developed software GC-Profile, we obtained a complete coverage of the human genome by 3198 isochores with boundaries at single nucleotide resolution. Interestingly, the experimentally confirmed replication timing sites in the regions of 1p36.1, 6p21.32, 17q11.2 and 22q12.1 nearly all coincide with the determined isochore boundaries. The precise boundaries of the 3198 isochores are available via the website: http://tubic.tju.edu.cn/isomap/.

Biological Implications of Isochore Boundaries in the Human Genome

The human genome is composed of large sequence segments with fairly homogeneous GC content, namely isochores, which have been linked to many important functions; biological implications of most isochore boundaries, however, remain elusive, partly due to the difficulty in determining these boundaries at high resolution. Using the segmentation algorithm based on the quadratic divergence, we re-determined all 79 boundaries of previously identified human isochores at single-nucleotide resolution, and then compared the boundary coordinates with other genome features. We found that 55.7% of isochore boundaries coincide with termini of repeat elements; 45.6% of isochore boundaries coincide with termini of highly conserved sequences based on alignment of 17 vertebrate genomes, i.e., the highly conserved genome sequence switches to a less or non-conserved one at the isochore boundary; some isochore boundaries coincide with abrupt change of CpG island distribution (note that one boundary can associate with more than one genome feature). In addition, sequences around isochore boundaries are highly conserved. It seems reasonable to deduce that the boundaries of all the isochores studied here would be replication timing sites in the human genome. These results suggest possible key roles of the isochore boundaries and may further our understanding of the human genome organization.

Assignment of isochores for all completely sequenced vertebrate genomes using a consensus

A new consensus isochore assignment method and a database of isochore maps for all completely sequenced vertebrate genomes are presented.

Isochores and replication time zones: a perfect match

The mammalian genome is not a random sequence but shows a specific, evolutionarily conserved structure that becomes manifest in its isochore pattern. Isochores, i.e. stretches of DNA with a distinct sequence composition and thus a specific GC content, cause the chromosomal banding pattern. This fundamental level of genome organization is related to several functional features like the replication timing of a DNA sequence. GC richness of genomic regions generally corresponds to an early replication time during S phase. Recently, we demonstrated this interdependency on a molecular level for an abrupt transition from a GC-poor isochore to a GC-rich one in the NF1 gene region; this isochore boundary also separates late from early replicating chromatin. Now, we analyzed another genomic region containing four isochores separated by three sharp isochore transitions. Again, the GC-rich isochores were found to be replicating early, the GC-poor isochores late in S phase; one of the replication time zones was discovered to consist of one single replicon. At the boundaries between isochores, that all show no special sequence elements, the replication machinery stopped for several hours. Thus, our results emphasize the importance of isochores as functional genomic units, and of isochore transitions as genomic landmarks with a key function for chromosome organization and basic biological properties.

Identification of isochore boundaries in the human genome using the technique of wavelet multiresolution analysis

Incorporated with the Z curve method, the technique of wavelet multiresolution (also known as multiscale) analysis has been proposed to identify the boundaries of isochores in the human genome. The human MHC sequence and the longest contigs of human chromosomes 21 and 22 are used as examples. The boundary between the isochores of Class III and Class II in the MHC sequence has been detected and found to be situated at the position 2,490,368 bp. This result is in good agreement with the experimental evidence. An isochore with a length of about 7 Mb in chromosome 21 has been identified and found to be gene- and Alu-poor. We have also found that the G + C content of chromosome 21 is more homogeneous than that of chromosome 22. Compared with the window-based methods, the present method has the highest resolution for identifying the boundaries of isochores, even at a scale of single base. Compared with the entropic segmentation method, the present method has the merits of more intuitiveness and less calculations. The important conclusion drawn in this study is that the segmentation points, at which the G + C content undergoes relatively dramatic changes, do exist in the human genome. These ‘singularity’ points may be considered to be candidates of isochore boundaries in the human genome. The method presented is a general one and can be used to analyze any other genomes.

Isochore chromosome maps of the human genome

The human genome is a mosaic of isochores, which are long DNA segments (≫300 kbp) relatively homogeneous in G+C. Human isochores were first identified by density-gradient ultracentrifugation of bulk DNA, and differ in important features, e.g. genes are found predominantly in the GC-richest isochores. Here, we use a reliable segmentation method to partition the longest contigs in the human genome draft sequence into long homogeneous genome regions (LHGRs), thereby revealing the isochore structure of the human genome. The advantages of the isochore maps presented here are: (1) sequence heterogeneities at different scales are shown in the same plot; (2) pair-wise compositional differences between adjacent regions are all statistically significant; (3) isochore boundaries are accurately defined to single base pair resolution; and (4) both gradual and abrupt isochore boundaries are simultaneously revealed. Taking advantage of the wide sample of genome sequence analyzed, we investigate the correspondence between LHGRs and true human isochores revealed through DNA centrifugation. LHGRs show many of the typical isochore features, mainly size distribution, G+C range, and proportions of the isochore classes. The relative density of genes, Alu and long interspersed nuclear element repeats and the different types of single nucleotide polymorphisms on LHGRs also coincide with expectations in true isochores. Potential applications of isochore maps range from the improvement of gene-finding algorithms to the prediction of linkage disequilibrium levels in association studies between marker genes and complex traits. The coordinates for the LHGRs identified in all the contigs longer than 2 Mb in the human genome sequence are available at the online resource on isochore mapping: http://bioinfo2.ugr.es/isochores.

Isochore chromosome maps of eukaryotic genomes

Analytical DNA ultracentrifugation revealed that eukaryotic genomes are mosaics of isochores: long DNA segments (≫300 kb on average) relatively homogeneous in G+C. Important genome features are dependent on this isochore structure, e.g. genes are found predominantly in the GC-richest isochore classes. However, no reliable method is available to rigorously partition the genome sequence into relatively homogeneous regions of different composition, thereby revealing the isochore structure of chromosomes at the sequence level. Homogeneous regions are currently ascertained by plain statistics on moving windows of arbitrary length, or simply by eye on G+C plots. On the contrary, the entropic segmentation method is able to divide a DNA sequence into relatively homogeneous, statistically significant domains. An early version of this algorithm only produced domains having an average length far below the typical isochore size. Here we show that an improved segmentation method, specifically intended to determine the most statistically significant partition of the sequence at each scale, is able to identify the boundaries between long homogeneous genome regions displaying the typical features of isochores. The algorithm precisely locates classes II and III of the human major histocompatibility complex region, two well-characterized isochores at the sequence level, the boundary between them being the first isochore boundary experimentally characterized at the sequence level. The analysis is then extended to a collection of human large contigs. The relatively homogeneous regions we find show many of the features (G+C range, relative proportion of isochore classes, size distribution, and relationship with gene density) of the isochores identified through DNA centrifugation. Isochore chromosome maps, with many potential applications in genomics, are then drawn for all the completely sequenced eukaryotic genomes available.

Crossing over a boundary

Where does recombination occur in the human genome? If it is not uniform, as it appears not to be, are there any regularities in its distribution? In the human genome the GC content tends to be consistent over large domains (upwards of 300 kb), the ‘isochores’, and the variation in GC content is also thought to reflect differences in the recombination rate. The mean GC content of individual autosomes correlates with the density of chiasmata. At a higher resolution, chiasmata tend to be found in R and T bands rather than G bands. G bands contain most GC-poor isochores, whereas T bands, a subset of the R bands, are the only bands where the GC-richest isochores are found. But is there a strong connection between isochore GC content and recombination? What, for example, if one looks across a boundary between isochores: is there a step up in the recombination rate as one moves to the higher GC content?Eisenbarth et al.1xAn isochore transition in the NF1 gene region coincides with a switch in the extent of linkage disequilibrium. Eisenbarth, I. et al. Am. J. Hum. Genet. 2000; 67: 873–880Abstract | Full Text | Full Text PDF | PubMed | Scopus (52)See all References1 are the first to look at this issue, examining the region around the NF1 gene on 17q11.2. They examined 93 individuals selected at random and characterized eight polymorphic markers over ∼300 kb. From haplotype frequencies they could calculate the degree of linkage disequilibrium, and from assumptions about the age of the variants they could estimate the recombination rate between markers. Knowing the genomic distances allowed them to estimate the local recombination rate per megabase.The NF1 gene resides in a GC-poor region and over the ∼200 kb surrounding the gene there is very strong linkage disequilibrium. Eisenbarth et al. estimate the recombination frequency to be only 0.0001 per Mb in this region (assuming variants are about 5000–10 000 generations old). By contrast, in the neighbouring GC-rich isochore, there is no linkage disequilibrium between markers 26-kb apart. The recombination rate is about two orders of magnitude higher. Plotting the recombination rate against the GC content shows both a sharp transition in GC content between isochores and a sharp transition, coincident with the isochore boundary, in the local recombination rate.If isochore boundaries are typically also recombination-rate boundaries, this has major implications for those searching for genes associated with disease. The methods typically rely on non-random associations between alleles (i.e. linkage disequilibrium), but this is broken down by recombination. The number of polymorphic markers that will be needed in a given genomic region increases, therefore, as the local recombination rate goes up. Finding genes in GC-rich regions could then prove harder than previously thought.

Gene organisation, sequence variation and isochore structure at the centromeric boundary of the human MHC

We have mapped and sequenced the region immediately centromeric of the human major histocompatibility complex (MHC). A cluster of 13 genes/pseudogenes was identified in a 175 kb PAC linking the TAPASIN locus with the class II region. It includes two novel human genes ( BING4 and SACM2L) and a thus far unnoticed human leucocyte antigen (HLA) class II pseudogene, termed HLA-DPA3. Analysis of the G+C content revealed an isochore boundary which, together with the previously reported telomeric boundary, defines the MHC class II region as one of the first completely sequenced isochores in the human genome. Comparison of the sequence with limited sequence from other cell lines shows that the high sequence variation found within the classical class II region extends beyond the identified isochore boundary leading us to propose the concept of an “extended MHC”. By comparative analysis, we have precisely identified the mouse/human synteny breakpoint at the centromeric end of the extended MHC class II region between the genes HSET and PHF1.