Patterns of insertions and their covariation with substitutions in the rat, mouse, and human genomes

The rates at which human genomic DNA changes by neutral substitution and insertion of certain families of transposable elements covary in large, megabase-sized segments. We used the rat, mouse, and human genomic DNA sequences to examine these processes in more detail in comparisons over both shorter (rat-mouse) and longer (rodent-primate) times, and demonstrated the generality of the covariation. Different families of transposable elements show distinctive insertion preferences and patterns of variation with substitution rates. SINEs are more abundant in GC-rich DNA, but the regional GC preference for insertion (monitored in young SINEs) differs between rodents and humans. In contrast, insertions in the rodent genomes are predominantly LINEs, which prefer to insert into AT-rich DNA in all three mammals. The insertion frequency of repeats other than SINEs correlates strongly positively with the frequency of substitutions in all species. However, correlations with SINEs show the opposite effects. The correlations are explained only in part by the GC content, indicating that other factors also contribute to the inherent tendency of DNA segments to change over evolutionary time.     

Reconstructing the genomic architecture of ancestral mammals: lessons from human, mouse, and rat genomes

Recent analysis of genome rearrangements in human and mouse genomes revealed evidence for more rearrangements than thought previously and shed light on previously unknown features of mammalian evolution, like breakpoint reuse and numerous microrearrangements. However, two-way analysis cannot reveal the genomic architecture of ancestral mammals or assign rearrangement events to different lineages. Thus, the "original synteny" problem introduced by Nadeau and Sankoff previously, remains unsolved, as at least three mammalian genomes are required to derive the ancestral mammalian karyotype. We show that availability of the rat genome allows one to reconstruct a putative genomic architecture of the ancestral murid rodent genome. This reconstruction suggests that this ancestral genome retained many previously postulated chromosome associations in the placental ancestor and reveals others that were beyond the resolution of cytogenetic, radiation hybrid mapping, and chromosome painting techniques. Three-way analysis of rearrangements leads to a reliable reconstruction of the genomic architecture of specific regions in the murid ancestor, including the X chromosome, and for the first time allows one to assign major rearrangement events to one of human, mouse, and rat lineages. Our analysis implies that the rate of rearrangements is much higher in murid rodents than in the human lineage and confirms the existence of rearrangement hot-spots in all three lineages.     

Low conservation of alternative splicing patterns in the human and mouse genomes

Alternative splicing has recently emerged as a major mechanism of generating protein diversity in higher eukaryotes. We compared alternative splicing isoforms of 166 pairs of orthologous human and mouse genes. As the mRNA and EST libraries of human and mouse are not complete and thus cannot be compared directly, we instead analyzed whether known cassette exons or alternative splicing sites from one genome are conserved in the other genome. We demonstrate that about half of the analyzed genes have species-specific isoforms, and about a quarter of elementary alternatives are not conserved between the human and mouse genomes. The detailed results of this study are available at www.ig-msk.ru:8005/HMG_paper.     

Homologous recombination and dynamics of rhizobial genomes

The nitrogen-fixing bacteria commonly known as rhizobia are attractive organisms due to their symbiotic association with legume plants. Their genomes contain a large number of redundant genetic elements. These reiterations might participate in homologous recombination events and lead to diverse genomic rearrangements. Here we analyze the role of homologous recombination in the dynamics of these bacterial genomes, as well as its possible biological consequences.     

Comparative gene prediction in human and mouse

The completion of the sequencing of the mouse genome promises to help predict human genes with greater accuracy. While current ab initio gene prediction programs are remarkably sensitive (i.e., they predict at least a fragment of most genes), their specificity is often low, predicting a large number of false-positive genes in the human genome. Sequence conservation at the protein level with the mouse genome can help eliminate some of those false positives. Here we describe SGP2, a gene prediction program that combines ab initio gene prediction with TBLASTX searches between two genome sequences to provide both sensitive and specific gene predictions. The accuracy of SGP2 when used to predict genes by comparing the human and mouse genomes is assessed on a number of data sets, including single-gene data sets, the highly curated human chromosome 22 predictions, and entire genome predictions from ENSEMBL. Results indicate that SGP2 outperforms purely ab initio gene prediction methods. Results also indicate that SGP2 works about as well with 3x shotgun data as it does with fully assembled genomes. SGP2 provides a high enough specificity that its predictions can be experimentally verified at a reasonable cost. SGP2 was used to generate a complete set of gene predictions on both the human and mouse by comparing the genomes of these two species. Our results suggest that another few thousand human and mouse genes currently not in ENSEMBL are worth verifying experimentally.     

The organization and rate of evolution of wheat genomes are correlated with recombination rates along chromosome arms

Genes detected by wheat expressed sequence tags (ESTs) were mapped into chromosome bins delineated by breakpoints of 159 overlapping deletions. These data were used to assess the organizational and evolutionary aspects of wheat genomes. Relative gene density and recombination rate increased with the relative distance of a bin from the centromere. Single-gene loci present once in the wheat genomes were found predominantly in the proximal, low-recombination regions, while multigene loci tended to be more frequent in distal, high-recombination regions. One-quarter of all gene motifs within wheat genomes were represented by two or more duplicated loci (paralogous sets). For 40 such sets, ancestral loci and loci derived from them by duplication were identified. Loci derived by duplication were most frequently located in distal, high-recombination chromosome regions whereas ancestral loci were most frequently located proximal to them. It is suggested that recombination has played a central role in the evolution of wheat genome structure and that gradients of recombination rates along chromosome arms promote more rapid rates of genome evolution in distal, high-recombination regions than in proximal, low-recombination regions.     

Genome rearrangements in mammalian evolution: lessons from human and mouse genomes

Although analysis of genome rearrangements was pioneered by Dobzhansky and Sturtevant 65 years ago, we still know very little about the rearrangement events that produced the existing varieties of genomic architectures. The genomic sequences of human and mouse provide evidence for a larger number of rearrangements than previously thought and shed some light on previously unknown features of mammalian evolution. In particular, they reveal that a large number of microrearrangements is required to explain the differences in draft human and mouse sequences. Here we describe a new algorithm for constructing synteny blocks, study arrangements of synteny blocks in human and mouse, derive a most parsimonious human-mouse rearrangement scenario, and provide evidence that intrachromosomal rearrangements are more frequent than interchromosomal rearrangements. Our analysis is based on the human-mouse breakpoint graph, which reveals related breakpoints and allows one to find a most parsimonious scenario. Because these graphs provide important insights into rearrangement scenarios, we introduce a new visualization tool that allows one to view breakpoint graphs superimposed with genomic dot-plots.     

Cross contamination of the genomes in human/hamster cell hybrids by multiple short recombination events

We have isolated sites of de novo rearrangements from interspecific cell hybrids. Of 147,000 clones screened from a human/hamster hybrid genomic library, 14 clones were found with homology to both human and hamster repetitive DNA sequences. Five of these clones contained recombination events involving less than 13 kb of DNA, three with human DNA recombined into a section of hamster DNA, and two with hamster DNA recombined into human DNA. None of the clones involving human L1 sequences were found to be de novo transposition events, but simply short recombination or insertion events. Considering the apparent random nature of these events, they are likely to involve unique as well as repetitive sequences and also involve integration into the homologous as well as heterologous chromosome sets. These results suggest that the chromosome sets in somatic cell hybrids may be randomly contaminated with small DNA segments derived from either set of chromosomes.     

Comparative analysis of the alpha-like globin clusters in mouse, rat, and human chromosomes indicates a mechanism underlying breaks in conserved synteny

We have sequenced and fully annotated a 65,871-bp region of mouse Chromosome 17 including the Hba-ps4 α-globin pseudogene. Comparative sequence analysis with the functional α-globin loci at human Chromosome 16p13.3 and mouse Chromosome 11 shows that this segment of mouse Chromosome 17 contains a group of three α-like pseudogenes (Hba-psm–Hba-ps4–Hba-q3), similar to the duplicated sets found at the functional mouse cluster on Chromosome 11. In addition, exons 7 to 12 of the mLuc7L gene are present just downstream from the pseudogene cluster, indicating that this clone contains the region in which human 16p13.3 switches in synteny between mouse Chromosomes 11 and 17. Comparison of the sequences around the α-like clusters on the two mouse chromosomes reveals the presence of conserved tandem repeats. We propose that these repetitive elements have played a role in the fragmentation of the mouse α cluster during evolution.     

Comparative genomics of the human,macaque and mouse major histocompatibility complex

The MHC is a highly polymorphic genomic region that encodes the transplantation and immune regulatory molecules. It receives special attention for genetic investigation because of its important role in the regulation of innate and adaptive immune responses and its strong association with numerous infectious and/or autoimmune diseases. The MHC locus was first discovered in the mouse and for the past 50 years it has been studied most intensively in both mice and humans. However, in recent years the macaque species have emerged as some of the more important and advanced experimental animal models for biomedical research into MHC with important human immunodeficiency virus/simian immunodeficiency virus and transplantation studies undertaken in association with precise MHC genotyping and haplotyping methods using Sanger sequencing and next‐generation sequencing. Here, in this special issue on ‘Macaque Immunology’ we provide a short review of the genomic similarities and differences among the human, macaque and mouse MHC class I and class II regions, with an emphasis on the association of the macaque class I region with MHC polymorphism, haplotype structure and function.     

Comparative ultrastructural morphometric analysis of atrial specific granules in the bat,mouse, and rat

Size, incidence, and volume density of atrial specific granules (ASG) in right atrial cells from five animals each of the rat (average weight 210 g), mouse (average weight 28 g), fruit-eating bat Megaloglossus woermanni (BMW;average weight 35 g), and the insect-eating bat Pipistrellus pipistrellus (BPP; average weight 6 g) have been compared via ultrastructural morphometry. In all three parameters of granule measurement, significantly higher figures were obtained in the rodents than in the bats. However, between the rat and the mouse, as also between the two species of bats, no significant differences were noted in any of the measurements. These results therefore do not support the prevalent view that the number and size of the granules decrease with increase in size of the animal species. The low content of ASG in atrial cells of the bats is probably an indication of low demand for the natriuretic hormone of the granule, because, in such animals, and particularly in flight, conservation of fluid and electrolytes is of paramount importance. This suggests that granule content is adapted to fluid and electrolyte regulation in relation to the functional capacity of the animal. We also observed ASG-like sgtructures in endothelial cells of capillaries of bat tissue but not in rodents. The function of these granules or whether or not they represent atrial specific ones is not clear from the present study. © Wiley-Liss, Inc.     

Comparison of the genomes of human and mouse lays the foundation of genome zoology

The extensive similarities between the genomes of human and model organisms are the foundation of much of modern biology, with model organism experimentation permitting valuable insights into biological function and the aetiology of human disease. In contrast, differences among genomes have received less attention. Yet these can be expected to govern the physiological and morphological distinctions apparent among species, especially if such differences are the result of evolutionary adaptation. A recent comparison of the draft sequences of mouse and human genomes has shed light on the selective forces that have predominated in their recent evolutionary histories. In particular, mouse-specific clusters of homologues associated with roles in reproduction, immunity and host defence appear to be under diversifying positive selective pressure, as indicated by high ratios of non-synonymous to synonymous substitution rates. These clusters are also frequently punctuated by homologous pseudogenes. They thus have experienced numerous gene death, as well as gene birth, events. These regions appear, therefore, to have borne the brunt of adaptive evolution that underlies physiological and behavioural innovation in mice. We predict that the availability of numerous animal genomes will give rise to a new field of genome zoology in which differences in animal physiology and ethology are illuminated by the study of genomic sequence variations.     

Termini of human chromosomes display elevated rates of mitotic recombination

The strand-specific in situ hybridization technique of CO-FISH was used to probe telomeres of human mitotic cells in order to determine the spontaneous frequency of crossover. This approach allowed the detection of recombinational crossovers occurring anywhere along the length of individual chromosomes, including reciprocal events taking place between sister chromatids. Although the process of sister chromatid exchange (SCE) is the most prominent type of recombination in somatic mammalian cells, our results show that SCEs accounted for less than a third of the recombinational events revealed by CO-FISH. It is concluded that chromosomal regions near the termini of chromosome arms undergo extraordinarily high rates of spontaneous recombination, producing terminal crossovers whose small size precludes detection by standard cytogenetic methods. That similar results were observed for transformed epithelial cells, as well as primary fibroblasts, suggests that the phenomenon is a common characteristic of human cells. These findings are noteworthy because, although telomeric and subtelomeric DNA is known to be preferentially involved in certain types of recombination, the tips of somatic mammalian chromosomes have not previously been identified as preferred sites for crossover. Implications of these results are discussed in terms of limitations imposed on CO-FISH for its proposed use in directional hybridization mapping.     

Comparative analysis of the genomes of intestinal spirochetes of human and animal origin

The aim of the present work was to compare the genomes of 21 strains of intestinal spirochetes, which were isolated from patients suffering intestinal disorders, with those of Treponema hyodysenteriae (strain P18), the known etiological agent of swine dysentery (bloody scours), and of a nonpathogenic strain (M1) of Treponema innocens. The percent guanine-plus-cytosine value of the 23 DNAs was found to be 25.5 to 30.1, as determined by a double-labeling procedure based on nick-translation by DNA polymerase I. The genome size of two spirochetal strains, of human and porcine origin, was found to be similar (4 x 10(6) base pairs) and close to that of the reference bacterium Escherichia coli (4.2 x 10(6) base pairs). Restriction analysis showed the presence of two modified bases in spirochetal DNA. Methyladenine was present in the GATC sequence of DNA from 15 spirochetes of human origin, and methylcytosine was present in several sequences occurring in all strains. The DNA of T. hyodysenteriae displayed a 30 to 100% homology with respect to that of 21 spirochetes from humans, thus suggesting the occurrence of a genetic heterogeneity in the latter group. These data indicate that the intestinal spirochetes analyzed in the present work are related; hence there is a possibility of domestic animals being reservoirs of microorganisms pathogenic for humans. A classification of intestinal treponemes into subgroups has been proposed on the basis of restriction analysis and hybridization experiments.     

Comparative sequence analysis of duck and human hepatitis B virus genomes

We have cloned and sequenced an infectious, functionally active genome of a duck hepatitis B virus (DHBV). It is 3,021 base pairs (bp) in length and shows little DNA sequence homology to the genome of human hepatitis B virus (HBV). However, the amino acid sequences of predicted viral gene products are similar between DHBV and HBV, and the genome organization present in DHBV reflects that of HBV. As in the mammalian virus the long minus strand of the DHBV genome encodes three long overlapping reading frames designated as P, S, and C. The fourth open reading frame, termed X, is absent in DHBV. A comparison with a sequence of a second DHBV isolate [Mandart et al, Journal of Virology 49:782-792, 1984] revealed a nucleotide sequence variation of 5.6% and confirmed the presented overall gene organization of DHBV.     

Automated whole-genome multiple alignment of rat, mouse, and human

We have built a whole-genome multiple alignment of the three currently available mammalian genomes using a fully automated pipeline that combines the local/global approach of the Berkeley Genome Pipeline and the LAGAN program. The strategy is based on progressive alignment and consists of two main steps: (1) alignment of the mouse and rat genomes, and (2) alignment of human to either the mouse-rat alignments from step 1, or the remaining unaligned mouse and rat sequences. The resulting alignments demonstrate high sensitivity, with 87% of all human gene-coding areas aligned in both mouse and rat. The specificity is also high: <7% of the rat contigs are aligned to multiple places in human, and 97% of all alignments with human sequence >100 kb agree with a three-way synteny map built independently, using predicted exons in the three genomes. At the nucleotide level <1% of the rat nucleotides are mapped to multiple places in the human sequence in the alignment, and 96.5% of human nucleotides within all alignments agree with the synteny map. The alignments are publicly available online, with visualization through the novel Multi-VISTA browser that we also present.     

Integration of the rat recombination and EST maps in the rat genomic sequence and comparative mapping analysis with the mouse genome

Inbred strains of the laboratory rat are widely used for identifying genetic regions involved in the control of complex quantitative phenotypes of biomedical importance. The draft genomic sequence of the rat now provides essential information for annotating rat quantitative trait locus (QTL) maps. Following the survey of unique rat microsatellite (11,585 including 1648 new markers) and EST (10,067) markers currently available, we have incorporated a selection of 7952 rat EST sequences in an improved version of the integrated linkage-radiation hybrid map of the rat containing 2058 microsatellite markers which provided over 10,000 potential anchor points between rat QTL and the genomic sequence of the rat. A total of 996 genetic positions were resolved (avg. spacing 1.77 cM) in a single large intercross and anchored in the rat genomic sequence (avg. spacing 1.62 Mb). Comparative genome maps between rat and mouse were constructed by successful computational alignment of 6108 mapped rat ESTs in the mouse genome. The integration of rat linkage maps in the draft genomic sequence of the rat and that of other species represents an essential step for translating rat QTL intervals into human chromosomal targets.     

Bleomycin sulfate-induced micronuclei in human,rat, and mouse peripheral blood lymphocytes

The sensitivity to micronucleus (MN) induction of human, mouse, and rat peripheral blood lymphocytes (PBLs) exposed to bleomycin sulfate (BLM) in vitro was compared in cytochalasin B-induced binucleated (BN) cells. For the PBLs of each species, either 0, 5, 10, 20, 40, 60, 80, or 160 μg/ml BLM was added to 5 ml aliquots of whole blood for 4 hr at 37°C in a 5% CO₂ atmosphere. Leukocytes were isolated on a density gradient and cultured in the presence of phytohemagglutinin to stimulate blastogenesis, and cytochalasin B was added to each culture at 21 hr postinitiation to prevent cytokinesis. A total of 4,000 BNs/concentration/species was analyzed for MN in two independent experiments. In addition, multiple-MN-BNs were quantitated, and the nucleation index was determined. Significant increases both in total MN-BNs and multiple-MN-BNs were observed at all concentrations in all species. All three species concentration-response curves gave good fits (r² values from 0.87 to 0.95) to either a linear or a square root model (y = mx + b or y = m[x]^0.5 + b, respectively; where y = the percentage of MN-BN, m is the slope, and b is the y-intercept). The MN induction in the human and rat PBLs was not statistically different, but both were significantly less sensitive than the response shown by the BLM-exposed mouse PBLs. This difference in MN susceptibility was observed only at BLM test concentrations ≧ 20 μg/ml. The nucleation index was significantly decreased in all species at either 80 or 160 μg/ml. © 1995 Wiley-Liss, Inc.