Cross chromosomal similarity for DNA sequence compression

Current DNA compression algorithms work by finding similar repeated regions within the DNA sequence and then encoding these regions together to achieve compression. Our study on chromosome sequence similarity reveals that the length of similar repeated regions within one chromosome is about 4.5% of the total sequence length. The compression gain is often not high because of these short lengths. It is well known that similarity exist among different regions of chromosome sequences. This implies that similar repeated sequences are found among different regions of chromosome sequences. Here, we study cross-chromosomal similarity for DNA sequence compression. The length and location of similar repeated regions among the sixteen chromosomes of S. cerevisiae are studied. It is found that the average percentage of similar subsequences found between two chromosome sequences is about 10% in which 8% comes from cross-chromosomal prediction and 2% from self-chromosomal prediction. The percentage of similar subsquences is about 18% in which only 1.2% comes from self-chromosomal prediction while the rest is from cross-chromosomal prediction among the 16 chromosomes studied. This suggests the importance of cross-chromosomal similarities in addition to self-chromosomal similarities in DNA sequence compression. An additional 23% of storage space could be reduced on average using self-chromosomal and cross-chromosomal predictions in compressing the 16 chromosomes of S. cerevisiae.     

Linearization and integration of DNA into cells preferentially occurs at intrinsically curved regions from human LINE-1 repetitive element.

A bent DNA library was constructed from human genomic DNA, from which a new clone belonging to the human LINE-1 sequence family was isolated and characterized. This clone, with a length of 378 base pairs and termed HBC-1 (human bent clone-1), contained an intrinsically occurring curved DNA structure. By permutation analysis, the center of curvature of this fragment was mapped onto the nucleotide position 886 from the 5' terminus of the complete LINE-1 sequence. Reporter plasmids, which contain HBC-1, were effectively integrated into human chromosome, indicating that the bent DNA structure provides a preferential donor site for the integration of human LINE-1 sequences. The present finding may provide an explanation as to why some inactivated LINE-1 sequences on human chromosomes carry the deletion at their 5' termini.     

High-resolution characterization of CPD hotspot formation in human fibroblasts

Repair of DNA lesions must occur within the chromatin landscape and is associated with alterations in histone modifications and nucleosome rearrangement. To directly associate these chromatin features with DNA damage and repair, it is necessary to be able to map DNA adducts. We have developed a cyclobutane pyrimidine dimer (CPD)-specific immunoprecipitation method and mapped ultraviolet damage hotspots across human chromosomes 1 and 6. CPD hotspots occur almost equally in genic and intergenic regions. However, these hotspots are significantly more prevalent adjacent to repeat elements, especially Alu repeats. Nucleosome mapping studies indicate that nucleosomes are consistently positioned at Alu elements where CPD hotspots form, but by 2 h post-irradiation, these same regions are significantly depleted of nucleosomes. These results indicate that nucleosomes associated with hotspots of CPD formation are readily rearranged, potentially making them accessible to DNA repair machinery. Our results represent the first chromosome scale map of ultraviolet-induced DNA lesions in the human genome, and reveal the sequence features and dynamic chromatin changes associated with CPD hotspots.     

Complex telomere-associated repeat units in members of the genus Chironomus evolve from sequences similar to simple telomeric repeats.

The dipteran Chironomus tentans has complex tandemly repeated 350-bp DNA sequences at or near the chromosome ends. As in Drosophila melanogaster, short simple repeats with cytosines and guanines in different strands have never been observed. We were therefore interested in learning whether the Chironomus repeats could have evolved from simple sequence telomeric DNA, which might suggest that they constitute a functional equivalent. We screened for repeat units with evolutionarily ancient features within the tandem arrays and recovered two clones with a less-evolved structure. Sequence analysis reveals that the present-day 350-bp unit probably evolved from a simpler 165-bp unit through the acquisition of transposed sequences. The 165-bp unit contains DNA with a highly biased distribution of cytosine and guanine between the two strands, although with the ratios inverted in two minor parts of the repeat. It is largely built up of short degenerate subrepeats for which most of the sequence can be reconstructed. The consensus for the subrepeat sequence is similar to the simple telomeric repeat sequences of several kinds of eukaryotes. We propose that the present-day unit has evolved from telomeric, simple sequence, asymmetric DNA from which it has retained some original sequence features and possibly functions.     

Entropic Profiles of DNA Sequences Through Chaos-game-derived Images

A new method to determine entropic profiles in DNA sequences is presented. It is based on the chaos-game representation (CGR) of gene structure, a technique which produces a fractal-like picture of DNA sequences. First, the CGR image was divided into squares 4^-m in size (m being the desired resolution), and the point density counted. Second, appropriate intervals were adjusted, and then a histogram of densities was prepared. Third, Shannon's formula was applied to the probability-distribution histogram, thus obtaining a new entropic estimate for DNA sequences, the histogram entropy , a measurement that goes with the level of constraints on the DNA sequence. Lastly, the entropic profile for the sequence was drawn, by considering the entropies at each resolution level, thus providing a way to summarize the complexity of large genomic regions or even entire genomes at different resolution levels. The application of the method to DNA sequences reveals that entropic profiles obtained in this way, as opposed to previously published ones, clearly discriminate between random and natural DNA sequences. Entropic profiles also show a different degree of variability within and between genomes. The results of these analyses are discussed in relation both to the genome compartmentalization in vertebrates and to the differential action of compositional and/or functional constraints on DNA sequences.     

Genetic and physical maps of the Bacillus subtilis chromosome

Sequencing of the complete Bacillus subtilis chromosome revealed the presence of approximately 4100 genes, 1000 of which were previously identified and mapped by classical genetic crosses. Comparison of these experimentally determined positions to those derived from the nucleotide sequence showed discrepancies reaching up to 24 degrees (approximately 280 kb). The size of these discrepancies as a function of their position along the chromosome is not random but, apparently, reveals some periodicity. Our analyses demonstrate that the discrepancies can be accounted for by inaccurate positioning of the early reference markers with respect to which all subsequently identified loci were mapped by transduction and transformation. We conclude (i) that specific DNA sequences, such as recombination hotspots or presence of heterologous DNA, had no detectable effect on the results obtained by classical mapping, and (ii) that PBS1 transduction appears to be an accurate and unbiased mapping method in B. subtilis.     

Chromosome Banding and DNA Methylation Patterns, Chromatin Organisation and Nuclear DNA Content in Zingeria biebersteiniana

Chromosome structure and chromatin organisation of a two-chromosome model cereal Zingeria biebersteiniana (Claus) P. Smirnov were studied: nuclear DNA content was determined by microdensitometric analysis after Feulgen staining; Feulgen absorption at different thresholds of absorbance in interphase nuclei also provided evidence on the organisation of chromatin, allowing quantitative estimation of condensed chromatin within interphasic nucleus. The DNA methylation pattern of Z. biebersteiniana metaphase chromosomes was examined with a specific monoclonal antibody. 5-methyl-cytosine residues are present in several chromosome sites and differences may be present between corresponding regions of homologues. Chromosome banding pattern reveals large bands in the centromeric regions of each chromosome, showing constitutive heterochromatin; by fluorochromes staining pericentromeric blocks are evidenced. After the cold and 9-aminoacridine pre-treatments and after aceto-carmine and aceto-orceine staining, respectively, the metaphase chromosomes were analysed by image analysis system revealing a segmentation of the chromosome body that resembles Giemsa/Reverse banding in animal chromosomes. This revised version was published online in July 2006 with corrections to the Cover Date.     

Organization of chromosome ends in Ustilago maydis. RecQ-like helicase motifs at telomeric regions.

In this study we have established the structure of chromosome ends in the basidiomycete fungus Ustilago maydis. We isolated and characterized several clones containing telomeric regions and found that as in other organisms, they consist of middle repeated DNA sequences. Two principal types of sequence were found: UTASa was highly conserved in nucleotide sequence and located almost exclusively at the chromosome ends, and UTASb was less conserved in nucleotide sequence than UTASa and found not just at the ends but highly interspersed throughout the genome. Sequence analysis revealed that UTASa encodes an open reading frame containing helicase motifs with the strongest homology to RecQ helicases; these are DNA helicases whose function involves the maintenance of genome stability in Saccharomyces cerevisiae and in humans, and the suppression of illegitimate recombination in Escherichia coli. Both UTASa and UTASb contain a common region of about 300 bp located immediately adjacent to the telomere repeats that are also found interspersed in the genome. The analysis of the chromosome ends of U. maydis provides information on the general structure of chromosome ends in eukaryotes, and the putative RecQ helicase at UTASa may reveal a novel mechanism for the maintenance of chromosome stability.     

The specific organisation of satellite DNA sequences on the X-chromosome of Mus musculus: partial independence of chromosome evolution

DNA was isolated from a chinese hamster/mouse hybrid cell line containing a single mouse chromosome, the X-chromosome, and digested with a variety of restriction endonucleases known to cut mouse satellite DNA. After agarose gel electrophoresis and transfer to nitrocellulose, hybridisation was carried out to a radioactive mouse satellite DNA probe. In this manner the organisation of satellite sequences at an individual chromosome was determined. We have found that the organisation of centromeric satellite DNA sequences on the mouse X-chromosome differs from that of other chromosomes in the complement. The nature of the differences suggests features of evolution of highly repeated sequences within a karyotype.     

Alpha satellite DNAs on chromosomes 10 and 12 are both members of the dimeric suprachromosomal subfamily, but display little identity at the nucleotide sequence level

We have investigated the organization and complexity of alpha satellite DNA on chromosomes 10 and 12 by restriction endonuclease mapping, in situ hybridization (ISH), and DNA-sequencing methods. Alpha satellite DNA on both chromosomes displays a basic dimeric organization, revealed as a 6- and an 8-mer higher-order repeat (HOR) unit on chromosome 10 and as an 8-mer HOR on chromosome 12. While these HORs show complete chromosome specificity under high-stringency ISH conditions, they recognize an identical set of chromosomes under lower stringencies. At the nucleotide sequence level, both chromosome 10 HORs are 50% identical to the HOR on chromosome 12 and to all other alpha satellite DNA sequences from the in situ cross-hybridizing chromosomes, with the exception of chromosome 6. An 80% identity between chromosome 6- and chromosome 10-derived alphoid sequences was observed. These data suggest that the alphoid DNA on chromosomes 6 and 10 may represent a distinct subclass of the dimeric subfamily. These sequences are proposed to be present, along with the more typical dimeric alpha satellite sequences, on a number of different human chromosomes.     

Theoretical analysis of mutation hotspots and their DNA sequence context specificity

Mutation frequencies vary significantly along nucleotide sequences such that mutations often concentrate at certain positions called hotspots. Mutation hotspots in DNA reflect intrinsic properties of the mutation process, such as sequence specificity, that manifests itself at the level of interaction between mutagens, DNA, and the action of the repair and replication machineries. The hotspots might also reflect structural and functional features of the respective DNA sequences. When mutations in a gene are identified using a particular experimental system, resulting hotspots could reflect the properties of the gene product and the mutant selection scheme. Analysis of the nucleotide sequence context of hotspots can provide information on the molecular mechanisms of mutagenesis. However, the determinants of mutation frequency and specificity are complex, and there are many analytical methods for their study. Here we review computational approaches for analyzing mutation spectra (distribution of mutations along the target genes) that include many mutable (detectable) positions. The following methods are reviewed: derivation of a consensus sequence, application of regression approaches to correlate nucleotide sequence features with mutation frequency, mutation hotspot prediction, analysis of oligonucleotide composition of regions containing mutations, pairwise comparison of mutation spectra, analysis of multiple spectra, and analysis of "context-free" characteristics. The advantages and pitfalls of these methods are discussed and illustrated by examples from the literature. The most reliable analyses were obtained when several methods were combined and information from theoretical analysis and experimental observations was considered simultaneously. Simple, robust approaches should be used with small samples of mutations, whereas combinations of simple and complex approaches may be required for large samples. We discuss several well-documented studies where analysis of mutation spectra has substantially contributed to the current understanding of molecular mechanisms of mutagenesis. The nucleotide sequence context of mutational hotspots is a fingerprint of interactions between DNA and DNA repair, replication, and modification enzymes, and the analysis of hotspot context provides evidence of such interactions.     

Hypermethylation of human DNA sequences in embryonal carcinoma cells and somatic tissues but not in sperm.

Certain human DNA sequences are much less methylated at CpG sites in sperm than in various adult somatic tissues. The DNA of term placenta displays intermediate levels of methylation at these sequences (Sp-0.3 sequences). We report here that pluripotent embryonal carcinoma (EC) cells derived from testicular germ cell tumors are hypermethylated at the three previously cloned Sp-0.3 sequences and seven newly isolated sequences that exhibit sperm-specific hypomethylation. In contrast to their hypermethylation in EC cells, the Sp-0.3 sequences are hypomethylated in a line of yolk sac carcinoma cells, which like placenta, represent an extraembryonic lineage. These DNA sequences, therefore, appear to be subject to coordinate changes in their methylation during differentiation, probably early in embryogenesis, despite their diversity in copy number (1 to 10(4] and primary structure. Two of these Sp-0.3 sequences are highly homologous to DNA sequences in human chromosomal regions that might be recombination hotspots, namely, a cryptic satellite DNA sequence at a fragile site and the downstream region of the beta-globin gene cluster.     

A novel autonomously replicating sequence (ARS) for multiple integration in the yeast Hansenula polymorpha DL-1.

Several autonomously replicating sequences of Hansenula polymorpha DL-1 (HARSs) with the characteristics of tandem integration were cloned by an enrichment procedure and analyzed for their functional elements to elucidate the mechanism of multiple integration in tandem repeats. All plasmids harboring newly cloned HARSs showed a high frequency of transformation and were maintained episomally before stabilization. After stabilization, the transforming DNA was stably integrated into the chromosome. HARS36 was selected for its high efficiency of transformation and tendency for integration. Several tandemly repeated copies of the transforming plasmid containing HARS36 (pCE36) integrated into the vicinity of the chromosomal end. Bal 31 digestion of the total DNA from the integrants followed by Southern blotting generated progressive shortening of the hybridization signal, indicating the telomeric localization of the transforming plasmids on the chromosome. The minimum region of HARS36 required for its HARS activity was analyzed by deletion analyses. Three important regions, A, B, and C, for episomal replication and integration were detected. Analysis of the DNA sequences of regions A and B required for the episomal replication revealed that region A contained several AT-rich sequences that showed sequence homology with the ARS core consensus sequence of Saccharomyces cerevisiae. Region B contained two directly repeated sequences which were predicted to form a bent DNA structure. Deletion of the AT-rich core in region A resulted in a complete loss of ARS activity, and deletion of the repeated sequences in region B greatly reduced the stability of the transforming plasmid and resulted in retarded cell growth. Region C was required for the facilitated chromosomal integration of transforming plasmids.     

A symmetrical six-base-pair target site sequence determines Tn10 insertion specificity

Transposon Tn10 inserts at many sites in the bacterial chromosome, but preferentially inserts at particular hotspots. We believe we have identified the target DNA signal responsible for this specificity. We have determined the DNA sequences of 11 Tn10 insertion sites and identified a particular 6 base pair (bp) symmetrical consensus sequence (GCTNAGC) common to those sites. The sequences at some sites differ from the consensus sequence but only in limited and well defined ways. The sequences at some sites differ from the consensus sequence than do sequences at other sites, and the consensus sequence and closely related sequences are generally absent from potential target regions where Tn10 is known not to insert. Other aspects of the target DNA can significantly influence the efficiency with which a particular target site sequence is used. The 6 bp consensus sequence is symmetrically located within the 9 bp target DNA sequence that is cleaved and duplicated during Tn10 insertion. This juxtaposition of recognition and cleavage sites plus the symmetry of the perfect consensus sequence suggest that the target DNA may be both recognized and cleaved by the symmetrically disposed subunits of a single protein, as suggested for type II restriction endonucleases. There is plausible homology between the consensus sequence and the very ends of Tn10, compatible with recognition of transposon ends and target DNA by the same protein. The sequences of actual insertion sites deviate from the perfect consensus sequence in a way which suggests that the 6 bp specificity determinant may be recognized through protein-DNA contacts along the major groove of the DNA double helix.     

A Musca domestica satellite sequence detects individual polymorphic regions in insect genome.

A recombinant plasmid, pHMd.24, was constructed which contains an array of a 24 bp sequence from the genome of the Musca domestica. This sequence hybridizes to sites occurring at various genomic locations in different housefly strains. The pHMd.24 insert has a structural organisation similar to satellite DNA sequences and hybridizes to homologous repeated sequences representing a small fraction of the M. domestica genome. Interestingly, this clone reveals polymorphic fragments from DNA isolated from single houseflies, thus generating an insect DNA fingerprint. In addition, the segregation of the fragments detected by hybridisation in a M. domestica pedigree indicates a transmission that follows mendelian inheritance.