Showing papers on "Chromosome published in 2000"
TL;DR: The V. cholerae genomic sequence provides a starting point for understanding how a free-living, environmental organism emerged to become a significant human bacterial pathogen.
Abstract: Here we determine the complete genomic sequence of the Gram negative, g-Proteobacterium Vibrio cholerae El Tor N16961 to be 4,033,460 base pairs (bp). The genome consists of two circular chromosomes of 2,961,146 bp and 1,072,314 bp that together encode 3,885 open reading frames. The vast majority of recognizable genes for essential cell functions (such as DNA replication, transcription, translation and cell-wall biosynthesis) and pathogenicity (for example, toxins, surface antigens and adhesins) are located on the large chromosome. In contrast, the small chromosome contains a larger fraction (59%) of hypothetical genes compared with the large chromosome (42%), and also contains many more genes that appear to have origins other than the g-Proteobacteria. The small chromosome also carries a gene capture system (the integron island) and host ‘addiction’ genes that are typically found on plasmids; thus, the small chromosome may have originally been a megaplasmid that was captured by an ancestral Vibrio species. The V. cholerae genomic sequence provides a starting point for understanding how a free-living, environmental organism emerged to become a significant human bacterial pathogen.
1,785 citations
TL;DR: In this article, the authors reported the sequence and gene catalogue of the long arm of chromosome 21 and sequenced 33,546,361 base pairs (bp) of DNA with very high accuracy, the largest contig being 25,491,867 bp.
Abstract: Chromosome 21 is the smallest human autosome. An extra copy of chromosome 21 causes Down syndrome, the most frequent genetic cause of significant mental retardation, which affects up to 1 in 700 live births. Several anonymous loci for monogenic disorders and predispositions for common complex disorders have also been mapped to this chromosome, and loss of heterozygosity has been observed in regions associated with solid tumours. Here we report the sequence and gene catalogue of the long arm of chromosome 21. We have sequenced 33,546,361 base pairs (bp) of DNA with very high accuracy, the largest contig being 25,491,867 bp. Only three small clone gaps and seven sequencing gaps remain, comprising about 100 kilobases. Thus, we achieved 99.7% coverage of 21q. We also sequenced 281,116 bp from the short arm. The structural features identified include duplications that are probably involved in chromosomal abnormalities and repeat structures in the telomeric and pericentromeric regions. Analysis of the chromosome revealed 127 known genes, 98 predicted genes and 59 pseudogenes.
1,404 citations
TL;DR: A functional genomic screen to identify genes required for cell division in Caenorhabditis elegans using an in vivo time-lapse differential interference contrast microscopy assay and found that this screen provides putative gene functions for other species as well.
Abstract: Genome sequencing projects generate a wealth of information; however, the ultimate goal of such projects is to accelerate the identification of the biological function of genes. This creates a need for comprehensive studies to fill the gap between sequence and function. Here we report the results of a functional genomic screen to identify genes required for cell division in Caenorhabditis elegans. We inhibited the expression of approximately 96% of the approximately 2,300 predicted open reading frames on chromosome III using RNA-mediated interference (RNAi). By using an in vivo time-lapse differential interference contrast microscopy assay, we identified 133 genes (approximately 6%) necessary for distinct cellular processes in early embryos. Our results indicate that these genes represent most of the genes on chromosome III that are required for proper cell division in C. elegans embryos. The complete data set, including sample time-lapse recordings, has been deposited in an open access database. We found that approximately 47% of the genes associated with a differential interference contrast phenotype have clear orthologues in other eukaryotes, indicating that this screen provides putative gene functions for other species as well.
906 citations
TL;DR: Comparative analysis suggests that an excess of chromosome fissions in the tetrapod lineage may account for chromosome numbers and provides histories for several human chromosomes.
Abstract: To help understand mechanisms of vertebrate genome evolution, we have compared zebrafish and tetrapod gene maps. It has been suggested that translocations are fixed more frequently than inversions in mammals. Gene maps showed that blocks of conserved syntenies between zebrafish and humans were large, but gene orders were frequently inverted and transposed. This shows that intrachromosomal rearrangements have been fixed more frequently than translocations. Duplicated chromosome segments suggest that a genome duplication occurred in ray-fin phylogeny, and comparative studies suggest that this event happened deep in the ancestry of teleost fish. Consideration of duplicate chromosome segments shows that at least 20% of duplicated gene pairs may be retained from this event. Despite genome duplication, zebrafish and humans have about the same number of chromosomes, and zebrafish chromosomes are mosaically orthologous to several human chromosomes. Is this because of an excess of chromosome fissions in the human lineage or an excess of chromosome fusions in the zebrafish lineage? Comparative analysis suggests that an excess of chromosome fissions in the tetrapod lineage may account for chromosome numbers and provides histories for several human chromosomes.
673 citations
TL;DR: Evidence that gene silencing accompanies allopolyploidization opens new avenues to this area of research.
Abstract: Allopolyploid hybridization serves as a major pathway for plant evolution, but in its early stages it is associated with phenotypic and genomic instabilities that are poorly understood. We have investigated allopolyploidization between Arabidopsis thaliana (2 n = 2 x = 10; n , gametic chromosome number; x , haploid chromosome number) and Cardaminopsis arenosa (2 n = 4 x = 32). The variable phenotype of the allotetraploids could not be explained by cytological abnormalities. However, we found suppression of 20 of the 700 genes examined by amplified fragment length polymorphism of cDNA. Independent reverse transcription–polymerase chain reaction analyses of 10 of these 20 genes confirmed silencing in three of them, suggesting that ∼0.4% of the genes in the allotetraploids are silenced. These three silenced genes were characterized. One, called K7, is repeated and similar to transposons. Another is RAP2.1 , a member of the large APETALA2 (AP2) gene family, and has a repeated element upstream of its 5′ end. The last, L6, is an unknown gene close to ALCOHOL DEHYDROGENASE on chromosome 1. CNG DNA methylation of K7 was less in the allotetraploids than in the parents, and the element varied in copy number. That K7 could be reactivated suggests epigenetic regulation. L6 was methylated in the C. arenosa genome. The present evidence that gene silencing accompanies allopolyploidization opens new avenues to this area of research.
527 citations
TL;DR: The general usefulness of expression profiles for nearly 300 Saccharomyces cerevisiae deletion mutants, obtained recently, is demonstrated, with implications for interpreting whole-genome expression data, particularly from cells known to suffer genomic instability, such as malignant or immortalized cells.
Abstract: Expression profiling using DNA microarrays holds great promise for a variety of research applications, including the systematic characterization of genes discovered by sequencing projects. To demonstrate the general usefulness of this approach, we recently obtained expression profiles for nearly 300 Saccharomyces cerevisiae deletion mutants. Approximately 8% of the mutants profiled exhibited chromosome-wide expression biases, leading to spurious correlations among profiles. Competitive hybridization of genomic DNA from the mutant strains and their isogenic parental wild-type strains showed they were aneuploid for whole chromosomes or chromosomal segments. Expression profile data published by several other laboratories also suggest the use of aneuploid strains. In five separate cases, the extra chromosome harboured a close homologue of the deleted gene; in two cases, a clear growth advantage for cells acquiring the extra chromosome was demonstrated. Our results have implications for interpreting whole-genome expression data, particularly from cells known to suffer genomic instability, such as malignant or immortalized cells.
497 citations
TL;DR: Transcriptional up-regulation of genes in the major histocompatibility complex by interferon-gamma led to an increase in the frequency with which this large gene cluster was found on an external chromatin loop, consistent with an association between large-scale chromatin organization of specific genomic regions and their transcriptional status.
Abstract: The large-scale chromatin organization of the major histocompatibility complex and other regions of chromosome 6 was studied by three-dimensional image analysis in human cell types with major differences in transcriptional activity. Entire gene clusters were visualized by fluorescence in situ hybridization with multiple locus-specific probes. Individual genomic regions showed distinct configurations in relation to the chromosome 6 terrritory. Large chromatin loops containing several megabases of DNA were observed extending outwards from the surface of the domain defined by the specific chromosome 6 paint. The frequency with which a genomic region was observed on an external chromatin loop was cell type dependent and appeared to be related to the number of active genes in that region. Transcriptional up-regulation of genes in the major histocompatibility complex by interferon-gamma led to an increase in the frequency with which this large gene cluster was found on an external chromatin loop. Our data are consistent with an association between large-scale chromatin organization of specific genomic regions and their transcriptional status.
462 citations
TL;DR: Evidence for mating included formation of stable prototrophs from strains with complementing auxotrophic markers; these contained both MTL alleles and molecular markers from both parents and were tetraploid in DNA content and mononucleate.
Abstract: Although the diploid fungus Candida albicans , a human pathogen, has been thought to have no sexual cycle, it normally possesses mating-type–like orthologs ( MTL ) of both of the Saccharomyces cerevisiae mating-type genes ( MAT ) a and α. When strains containing only MTL a or MTL α were constructed by the loss of one homolog of chromosome 5, the site of the MTL loci, MTL a and MTL α strains mated, but like mating types did not. Evidence for mating included formation of stable prototrophs from strains with complementing auxotrophic markers; these contained both MTL alleles and molecular markers from both parents and were tetraploid in DNA content and mononucleate.
458 citations
TL;DR: The disrupted mouse Cenpa gene is disrupted and demonstrated that the gene is essential for kinetochore targeting of Cenpc and plays an early role in organizing centromeric chromatin at interphase.
Abstract: Centromere protein A (Cenpa for mouse, CENP-A for other species) is a histone H3-like protein that is thought to be involved in the nucleosomal packaging of centromeric DNA. Using gene targeting, we have disrupted the mouse Cenpa gene and demonstrated that the gene is essential. Heterozygous mice are healthy and fertile whereas null mutants fail to survive beyond 6.5 days postconception. Affected embryos show severe mitotic problems, including micronuclei and macronuclei formation, nuclear bridging and blebbing, and chromatin fragmentation and hypercondensation. Immunofluorescence analysis of interphase cells at day 5.5 reveals complete Cenpa depletion, diffuse Cenpb foci, absence of discrete Cenpc signal on centromeres, and dispersion of Cenpb and Cenpc throughout the nucleus. These results suggest that Cenpa is essential for kinetochore targeting of Cenpc and plays an early role in organizing centromeric chromatin at interphase. The evidence is consistent with the proposal of a critical epigenetic function for CENP-A in marking a chromosomal region for centromere formation.
441 citations
TL;DR: Tumors with BFB events showed a decreased elimination rate of unstable chromosome aberrations after irradiation compared with normal cells and other tumor cells, suggesting that a combination of mitotically unstable chromosomes and an elevated tolerance to chromosomal damage leads to constant genomic reorganization in many malignancies, thereby providing a flexible genetic system for clonal evolution and progression.
Abstract: It has long been known that rearrangements of chromosomes through breakage-fusion-bridge (BFB) cycles may cause variability of phenotypic and genetic traits within a cell population. Because intercellular heterogeneity is often found in neoplastic tissues, we investigated the occurrence of BFB events in human solid tumors. Evidence of frequent BFB events was found in malignancies that showed unspecific chromosome aberrations, including ring chromosomes, dicentric chromosomes, and telomeric associations, as well as extensive intratumor heterogeneity in the pattern of structural changes but not in tumors with tumor-specific aberrations and low variability. Fluorescence in situ hybridization analysis demonstrated that chromosomes participating in anaphase bridge formation were involved in a significantly higher number of structural aberrations than other chromosomes. Tumors with BFB events showed a decreased elimination rate of unstable chromosome aberrations after irradiation compared with normal cells and other tumor cells. This result suggests that a combination of mitotically unstable chromosomes and an elevated tolerance to chromosomal damage leads to constant genomic reorganization in many malignancies, thereby providing a flexible genetic system for clonal evolution and progression.
415 citations
TL;DR: It is reported that the fission yeast homolog SpCENP-A is essential for establishing centromere chromatin associated with equal chromosome segregation.
Abstract: Mammalian kinetochores contain the centromere-specific histone H3 variant CENP-A, whose incorporation into limited chromosomal regions may be important for centromere function and chromosome segregation during mitosis. However, regulation of CENP-A localization and its role have not been clear. Here we report that the fission yeast homolog SpCENP-A is essential for establishing centromere chromatin associated with equal chromosome segregation. SpCENP-A binding to the nonrepetitious inner centromeres depended on Mis6, an essential centromere connector protein acting during G1-S phase of the cell cycle. Mis6 is likely required for recruiting SpCENP-A to form proper connection of sister centromeres.
TL;DR: Used in conjunction with embryo biopsy, diagnostic CGH should allow the exclusion of a proportion of embryos that appear normal but that have a poor probability of survival and, therefore, may improve the implantation rate after in vitro fertilization.
Abstract: Karyotypic studies of aborted fetuses have been used to draw the inference that the proportion of conceptuses with chromosome abnormalities is very high. Fluorescent in situ hybridization (FISH) studies of blastomeres from early cleavage embryos have provided some support for this inference but they are limited to the study of a few chromosomes. We describe the novel application of comparative genomic hybridization (CGH) to the study of numerical and structural abnormalities of single blastomeres from disaggregated 3-day-old human embryos. CGH results were obtained for 63 blastomeres from 12 embryos. Identification of all chromosomes with the exception of chromosomes 17, 19, 20 and 22 was possible. The embryos divided into four groups: (1) embryos with a normal CGH karyotype seen in all blastomeres; (2) embryos with consistent aneuploidy suggesting meiotic non-disjunction had occurred; (3) embryos that were mosaic generally with one or more cells showing aneuploidy for one or two chromosomes but some with cells showing extensive aneuploidy; and (4) one embryo with extensive aneuploidy in all blastomeres. The extensive aneuploidy in group 4 is interpreted as corresponding to the random aneuploidy seen in "chaotic" embryos reported by using interphase FISH. Partial chromosome loss and gain following chromosome breakage was observed in one embryo. Our analysis provides basic biological information on the occurrence of constitutional and post-zygotic chromosome abnormalities in early human embryos. Used in conjunction with embryo biopsy, diagnostic CGH should allow the exclusion of a proportion of embryos that appear normal but that have a poor probability of survival and, therefore, may improve the implantation rate after in vitro fertilization.
TL;DR: During interphase in the budding yeast, Saccharomyces cerevisiae, centromere clustering is reduced by the ndc10 mutation, which affects a kinetochore protein, and by the microtubule poison nocodazole, which suggests that clustered is actively maintained or enforced by the association of centromeres with microtubules throughout interphase.
Abstract: During interphase in the budding yeast, Saccharomyces cerevisiae, centromeres are clustered near one pole of the nucleus as a rosette with the spindle pole body at its hub. Opposite to the centromeric pole is the nucleolus. Chromosome arms extend outwards from the centromeric pole and are preferentially directed towards the opposite pole. Centromere clustering is reduced by the ndc10 mutation, which affects a kinetochore protein, and by the microtubule poison nocodazole. This suggests that clustering is actively maintained or enforced by the association of centromeres with microtubules throughout interphase. Unlike the Rabl-orientation known from many higher eukaryotes, centromere clustering in yeast is not only a relic of anaphase chromosome polarization, because it can be reconstituted without the passage of cells through anaphase. Within the rosette, homologous centromeres are not arranged in a particular order that would suggest somatic pairing or genome separation.
TL;DR: The mapping of new Mcd1p-binding sites (cohesin-associated regions [CARs]) in single-copy sequences of several chromosomes establish their spacing, their sequestration to intergenic regions, and their association with AT-rich sequences as general genomic properties of CARs.
Abstract: We identified the chromosomal addresses of a cohesin subunit, Mcd1p, in vivo by chromatin immunoprecipitation coupled with high resolution PCR-based chromosomal walking. The mapping of new Mcd1p-binding sites (cohesin-associated regions [CARs]) in single-copy sequences of several chromosomes establish their spacing ( approximately 9 kb), their sequestration to intergenic regions, and their association with AT-rich sequences as general genomic properties of CARs. We show that cohesins are not excluded from telomere proximal regions, and the enrichment of cohesins at the centromere at mitosis reflects de novo loading. The average size of a CAR is 0.8-1.0 kb. They lie at the boundaries of transcriptionally silenced regions, suggesting they play a direct role in defining the silent chromatin domain. Finally, we identify CARs in tandem (rDNA) and interspersed repetitive DNA (Ty2 and subtelomeric repeats). Each 9-kb rDNA repeat has a single CAR proximal to the 5S gene. Thus, the periodicity of CARs in single-copy regions and the rDNA repeats is conserved. The presence and spacing of CARs in repetitive DNA has important implications for genomic stability and chromosome packaging/condensation.
TL;DR: It is demonstrated here that global organization within the G(1) interphase nucleus is affected by one of the most fundamental physical quantities-chromosome size or mass-and two biophysical models are proposed, a volume exclusion model and a mitotic preset model, to explain the finding.
TL;DR: The results suggest that decondensation of 1qh and 16qh often leads to unresolved Holliday junctions, chromosome breakage, arm missegregation, and the formation of multiradials that may yield more stable chromosomal abnormalities, such as translocations.
Abstract: The ICF syndrome (immunodeficiency, centromeric region instability, facial anomalies) is a unique DNA methylation deficiency disease diagnosed by an extraordinary collection of chromosomal anomalies specifically in the vicinity of the centromeres of chromosomes 1 and 16 (Chr1 and Chr16) in mitogen-stimulated lymphocytes. These aberrations include decondensation of centromere-adjacent (qh) heterochromatin, multiradial chromosomes with up to 12 arms, and whole-arm deletions. We demonstrate that lymphoblastoid cell lines from two ICF patients exhibit these Chr1 and Chr16 anomalies in 61% of the cells and continuously generate 1qh or 16qh breaks. No other consistent chromosomal abnormality was seen except for various telomeric associations, which had not been previously noted in ICF cells. Surprisingly, multiradials composed of arms of both Chr1 and Chr16 were favored over homologous associations and cells containing multiradials with 3 or >4 arms almost always displayed losses or gains of Chr1 or Chr16 arms from the metaphase. Our results suggest that decondensation of 1qh and 16qh often leads to unresolved Holliday junctions, chromosome breakage, arm missegregation, and the formation of multiradials that may yield more stable chromosomal abnormalities, such as translocations. These cell lines maintained the abnormal hypomethylation in 1qh and 16qh seen in ICF tissues. The ICF-specific hypomethylation occurs in only a small percentage of the genome, e.g., ICF brain DNA had 7% less 5-methylcytosine than normal brain DNA. The ICF lymphoblastoid cell lines, therefore, retain not only the ICF-specific pattern of chromosome rearrangements, but also of targeted DNA hypomethylation. This hypomethylation of heterochromatic DNA sequences is seen in many cancers and may predispose to chromosome rearrangements in cancer as well as in ICF.
TL;DR: Because the protein composition of the resulting translocation lines is identical to that of normal wheat, it is believed that these manipulations could eliminate the quality defect associated with the 1RS.
Abstract: Centric translocations of the short arm of rye (Secale cereale L.) chromosome 1R are useful in wheat (Triticum aestivum L.) breeding because they confer resistance to several pests and diseases and improve yield. Their major disadvantage is in reduced bread making quality. To remedy this defect, rye chromosome arm 1RS in translocations 1RS.1BL and 1RS.1DL was induced by the ph1b mutation to recombine with the short arms of wheat group-1 chromosomes. Among 20 234 progeny screened, 139 primary recombinant chromosomes were recovered including 103 with 1BS, 22 with 1AS and 14 with 1DS. The Gli-1/Glu-3 loci of wheat were non-homoeoallelic to the Sec-1 locus of rye and were separated by about a 13-cM-long segment, which on the rye chromosome contained disease resistance loci Pm8, Lr26, Sr31, and Yr9. Pairs of primary recombinants 1RS-1BS with breakpoints flanking the storage protein loci were intercrossed and two types of secondary recombinant chromosomes 1RS were produced: a group of over 30 chromosomes where the Sec-1 locus was replaced by segments of 1BS of various lengths, and two chromosomes where 1.4- and 3.2-cM segments of 1BS introduced the Gli-1/Glu-3 loci. Selected chromosomes from each class were allowed to recombine within the shared segments of 1RS separating the intercalary wheat segments and two tertiary recombinant chromosomes were recovered. Cytologically, these chromosomes appear as normal 1RS arms but each has two intercalary segments of 1BS: one introducing the Gli-1/Glu-3 loci and the second one removing the Sec-1 locus. Because the protein composition of the resulting translocation lines is identical to that of normal wheat, it is believed that these manipulations could eliminate the quality defect associated with the 1RS.1BL translocation.
TL;DR: It is concluded that any chromosomal rearrangement can be made in ES cells with the Cre-loxP strategy provided that it does not affect cell viability.
Abstract: Chromosomal rearrangements are important resources for genetic studies. Recently, a Cre-loxP-based method to introduce defined chromosomal rearrangements (deletions, duplications, and inversions) into the mouse genome (chromosome engineering) has been established. To explore the limits of this technology systematically, we have evaluated this strategy on mouse chromosome 11. Although the efficiency of Cre-loxP-mediated recombination decreases with increasing genetic distance when the two endpoints are on the same chromosome, the efficiency is not limiting even when the genetic distance is maximized. Rearrangements encompassing up to three quarters of chromosome 11 have been constructed in mouse embryonic stem (ES) cells. While larger deletions may lead to ES cell lethality, smaller deletions can be produced very efficiently both in ES cells and in vivo in a tissue- or cell-type-specific manner. We conclude that any chromosomal rearrangement can be made in ES cells with the Cre-loxP strategy provided that it does not affect cell viability. In vivo chromosome engineering can be potentially used to achieve somatic losses of heterozygosity in creating mouse models of human cancers.
TL;DR: Data suggest allelic loss of chromosome 1p in patients with oligodendroglial neoplasms predicts longer progression-free survival among patients receiving radiotherapy +/- chemotherapy as part of their initial treatment.
Abstract: Introduction: Allelic loss of the short arm of chromosome 1 predicts radiographic response to chemotherapy and long overall survival times in patients with anaplastic oligodendrogliomas. Using a database of patients with oligodendrogliomas in whom chromosome 1p status was known, we explored whether allelic loss of 1p also predicted longer duration of tumor control when radiotherapy was part of the initial treatment of these patients. Materials and Methods: We measured progression-free survival following radiotherapy in a cohort of patients with World Health Organization (WHO) Grade II and WHO Grade III oligodendrogliomas. The effects on progression-free survival of patient age, Karnofsky performance score (KPS), tumor grade when irradiated and chromosome 1p status were examined by univariate and multivariate statistical analyses. For the subset of patients with newly diagnosed anaplastic oligodendrogliomas, relationships between use of chemotherapy, chromosome 1p status and progression-free survival were also examined. Results: Fifty-five patients (29 male, 26 female; ages 18–75 years; median, 44 years; KPS 50–90, median 80) were irradiated for either a WHO Grade II ( n = 19) or Grade III ( n = 36) oligodendroglioma. Twenty-eight patients had chemotherapy immediately prior to radiotherapy, and 27 had chemotherapy at progression following radiotherapy. The median radiation dose was 54 Gy in 30 fractions. Loss of heterozygosity (LOH) at chromosome 1p was evident in 36 tumors and absent in 19. Overall median progression-free survival after radiotherapy was 40.4 months. Median progression-free survival was 55.0 months for patients whose tumors harbored 1p loss vs. 6.2 months for those patients whose tumors retained both copies of chromosome 1p ( p p = 0.004). Conclusion: These data suggest allelic loss of chromosome 1p in patients with oligodendroglial neoplasms predicts longer progression-free survival among patients receiving radiotherapy ± chemotherapy as part of their initial treatment. Chromosome 1p loss may be an important stratification variable in future therapeutic trials of oligodendroglioma.
TL;DR: It seems realistic to propose construction of large-insert chromosome-specific DNA libraries in wheat based on results of improved procedure for preparation of chromosome suspensions and suitability of flow-sorted chromosomes for physical mapping and for construction of small-insert DNA libraries.
Abstract: The aim of this study was to develop an improved procedure for preparation of chromosome suspensions, and to evaluate the potential of flow cytometry for chromosome sorting in wheat. Suspensions of intact chromosomes were prepared by mechanical homogenization of synchronized root tips after mild fixation with formaldehyde. Histograms of relative fluorescence intensity (flow karyotypes) obtained after the analysis of DAPI-stained chromosomes were characterized and the chromosome content of all peaks on wheat flow karyotype was determined for the first time. Only chromosome 3B could be discriminated on flow karyotypes of wheat lines with standard karyotype. Remaining chromosomes formed three composite peaks and could be sorted only as groups. Chromosome 3B could be sorted at purity >95% as determined by microscopic evaluation of sorted fractions that were labeled using C-PRINS with primers for GAA microsatellites and for Afa repeats, respectively. Chromosome 5BL/7BL could be sorted in two wheat cultivars at similar purity, indicating a potential of various wheat stocks for sorting of other chromosome types. PCR with chromosome-specific primers confirmed the identity of sorted fractions and suitability of flow-sorted chromosomes for physical mapping and for construction of small-insert DNA libraries. Sorted chromosomes were also found suitable for the preparation of high-molecular-weight DNA. On the basis of these results, it seems realistic to propose construction of large-insert chromosome-specific DNA libraries in wheat. The availability of such libraries would greatly simplify the analysis of the complex wheat genome.
TL;DR: DNA technology, especially allelic imbalance (loss of heterozygosity) studies have identified chromosomal changes in oral carcinoma and head and neck squamous cell carcinoma, suggestive of the involvement of tumour suppressor genes (TSGs), particularly in chromosomes 3, 9, 11 and 17.
TL;DR: It is demonstrated that fluorescence in situ hybridization (FISH) signals derived from bacterial artificial chromosomes (BACs) can be used as chromosome-specific cytogenetic DNA markers for chromosome identification in potato.
Abstract: Reliable and easy to use techniques for chromosome identification are critical for many aspects of cytogenetic research. Unfortunately, such techniques are not available in many plant species, especially those with a large number of small chromosomes. Here we demonstrate that fluorescence in situ hybridization (FISH) signals derived from bacterial artificial chromosomes (BACs) can be used as chromosome-specific cytogenetic DNA markers for chromosome identification in potato. We screened a potato BAC library using genetically mapped restriction fragment length polymorphism markers as probes. The identified BAC clones were then labeled as probes for FISH analysis. A set of 12 chromosome-specific BAC clones were isolated and the FISH signals derived from these BAC clones serve as convenient and reliable cytological markers for potato chromosome identification. We mapped the 5S rRNA genes, the 45S rRNA genes, and a potato late blight resistance gene to three specific potato chromosomes using the chromosome-specific BAC clones.
TL;DR: The C-band distribution patterns of 105 angiosperm species were compared and showed that heterochromatin was preferentially located in similar chromosome regions, regardless of the distance from the centromere.
Abstract: The C-band distribution patterns of 105 angiosperm species were compared to identify general patterns or preferential sites for heterochromatin. The base-specific fluorochrome reaction of heterochromatin for 58 of these species and the role played by the average chromosome size in band distribution were also considered. The results showed that heterochromatin was preferentially located in similar chromosome regions, regardless of the distance from the centromere. This trend results in generalized bands, with heterochromatin distribution being identical in most chromosomes of a karyotype. Such bands very often displayed the same fluorochrome reaction, suggesting possible repeat transfer between non-homologous sites. Chromosome size may also play a role in heterochromatin location, since proximal bands were much more common in small-sized chromosomes.
TL;DR: The stable minichromosome vector allowed a 10 Mb-sized region of the mitotically unstable human chromosome 22 to be stably maintained in mouse embryonic stem (ES) cells, and in mice.
Abstract: For introducing regions of human chromosomes greater than a megabase into cells or animals, we have developed a chromosome-cloning system in which defined regions of human chromosomes can be cloned into a stable human minichromosome vector in homologous recombination-proficient chicken DT40 cells. The stable minichromosome vector allowed a 10 Mb-sized region of the mitotically unstable human chromosome 22 to be stably maintained in mouse embryonic stem (ES) cells, and in mice. Furthermore, we demonstrated functional expression of human genes from the HAC in mice. This study describes a stable cloning and expression system for greater than megabase-sized regions of human chromosomes.
Max Planck Society1, University of Padua2, University of Valencia3, University of Perpignan4, Université catholique de Louvain5, University of Amsterdam6, University of Grenoble7, Spanish National Research Council8, University of Naples Federico II9, J. Craig Venter Institute10, University of Chicago11, Celera Corporation12
TL;DR: In this paper, the authors present the sequence of chromosome 3, organized into four sequence segments (contigs), and the two largest (13.5 and 9.2 Mb) correspond to the top (long) and bottom (short) arms of the chromosome 3 and two small contigs are located in the genetically defined centromere.
Abstract: Arabidopsis thaliana is an important model system for plant biologists. In 1996 an international collaboration (the Arabidopsis Genome Initiative) was formed to sequence the whole genome of Arabidopsis and in 1999 the sequence of the first two chromosomes was reported. The sequence of the last three chromosomes and an analysis of the whole genome are reported in this issue. Here we present the sequence of chromosome 3, organized into four sequence segments (contigs). The two largest (13.5 and 9.2 Mb) correspond to the top (long) and the bottom (short) arms of chromosome 3, and the two small contigs are located in the genetically defined centromere. This chromosome encodes 5,220 of the roughly 25,500 predicted protein-coding genes in the genome. About 20% of the predicted proteins have significant homology to proteins in eukaryotic genomes for which the complete sequence is available, pointing to important conserved cellular functions among eukaryotes.
TL;DR: Analysis of the sequence of chromosome 5 yields further insights into centromere structure and the sequence determinants of heterochromatin condensation, and provides insights into the mechanisms and extent of genome evolution in plants.
Abstract: The genome of the model plant Arabidopsis thaliana has been sequenced by an international collaboration, The Arabidopsis Genome Initiative. Here we report the complete sequence of chromosome 5. This chromosome is 26 megabases long; it is the second largest Arabidopsis chromosome and represents 21% of the sequenced regions of the genome. The sequence of chromosomes 2 and 4 have been reported previously and that of chromosomes 1 and 3, together with an analysis of the complete genome sequence, are reported in this issue. Analysis of the sequence of chromosome 5 yields further insights into centromere structure and the sequence determinants of heterochromatin condensation. The 5,874 genes encoded on chromosome 5 reveal several new functions in plants, and the patterns of gene organization provide insights into the mechanisms and extent of genome evolution in plants.
TL;DR: The results suggest that heterochromatic and euchromatic domains are interspersed and closely associated within this 1.2-megabase region of the genome.
Abstract: The small fourth chromosome of Drosophila melanogaster (3.5% of the genome) presents a puzzle. Cytological analysis suggests that the bulk of the fourth, including the portion that appears banded in the polytene chromosomes, is heterochromatic; the banded region includes blocks of middle repetitious DNA associated with heterochromatin protein 1 (HP1). However, genetic screens indicate 50–75 genes in this region, a density similar to that in other euchromatic portions of the genome. Using a P element containing an hsp70-white gene and a copy of hsp26 (marked with a fragment of plant DNA designated pt), we have identified domains that allow for full expression of the white marker (R domains), and others that induce a variegating phenotype (V domains). In the former case, the hsp26-pt gene shows an accessibility and heat-shock-inducible activity similar to that seen in euchromatin, whereas in the latter case, accessibility and inducible expression are reduced to levels typical of heterochromatin. Mapping by in situ hybridization and by hybridization of flanking DNA sequences to a collection of cosmid and bacterial artificial chromosome clones shows that the R domains (euchromatin-like) and V domains (heterochromatin-like) are interspersed. Examination of the effect of genetic modifiers on the variegating transgenes shows some differences among these domains. The results suggest that heterochromatic and euchromatic domains are interspersed and closely associated within this 1.2-megabase region of the genome.
TL;DR: It is shown that the Bacillus subtilis Soj and Spo0J members of the ParAB families are responsible for the specific localization of plasmids at cell quarters in E. coli and can function as partition proteins.
Abstract: Bacterial genes required for proper partitioning consist of two transacting genes that encode proteins and a cis-acting gene that functions like a centromere. Plasmids actively partitioning by means of these genes migrate from midcell to the cell quarters and are tethered to these sites until the cells divide. Previously the partitioning genes were mainly found on plasmids and phages in Escherichia coli. However, progress in genome sequencing reveals that partitioning genes are ubiquitous in many bacterial plasmids and chromosomes. Each homologue of the two transacting genes belongs to a family, ParA or ParB. Moreover, phylogenic analysis of members of the ParA and ParB families indicates that each member falls into a chromosomal group or an extrachromosomal group. It is known that the parAB genes in the chromosomal group are located on relatively conserved chromosomal regions in several bacterial species. This suggests that the parAB genes were transferred from a chromosome to plasmids and phages, so the genes have diverged among bacterial species. To support this possibility, we show that the Bacillus subtilis Soj and Spo0J members of the ParAB families are responsible for the specific localization of plasmids at cell quarters in E. coli and can function as partition proteins. Host factors to tether actively partitioning plasmids at subcellular sites may be conserved in Gram-negative and Gram-positive bacteria so that phages and plasmids with the ParAB partitioning system can be stably inherited in host cells across bacterial species.
TL;DR: The hypothesis that chromosome reassortments that are catalyzed by aneuploidy offer a unifying explanation for the high mutation rates of aneuPLoid cancer cells and for the association of selected with unselected phenotypes, e.g., multidrug resistance.
Abstract: The mutation rates of cancer cells to drug and multidrug resistance are paradoxically high, i.e., 10−3 to 10−6, compared with those altering phenotypes of recessive genes in normal diploid cells of about 10−12. Here the hypothesis was investigated that these mutations are due to chromosome reassortments that are catalyzed by aneuploidy. Aneuploidy, an abnormal number of chromosomes, is the most common genetic abnormality of cancer cells and is known to change phenotypes (e.g., Down's syndrome). Moreover, we have shown recently that aneuploidy autocatalyzes reassortments of up to 2% per chromosome per mitosis because it unbalances spindle proteins, even centrosome numbers, via gene dosage. The hypothesis predicts that a selected phenotype is associated with multiple unselected ones, because chromosome reassortments unbalance simultaneously thousands of regulatory and structural genes. It also predicts variants of a selected phenotype based on variant reassortments. To test our hypothesis we have investigated in parallel the mutation rates of highly aneuploid and of normal diploid Chinese hamster cells to resistance against puromycin, cytosine arabinoside, colcemid, and methotrexate. The mutation rates of aneuploid cells ranged from 10−4 to 10−6, but no drug-resistant mutants were obtained from diploid cells in our conditions. Further selection increased drug resistance at similar mutation rates. Mutants selected from cloned cells for resistance against one drug displayed different unselected phenotypes, e.g., polygonal or fusiform cellular morphology, flat or three-dimensional colonies, and resistances against other unrelated drugs. Thus our hypothesis offers a unifying explanation for the high mutation rates of aneuploid cancer cells and for the association of selected with unselected phenotypes, e.g., multidrug resistance. It also predicts drug-specific chromosome combinations that could become a basis for selecting alternative chemotherapy against drug-resistant cancer.
TL;DR: In this paper, the authors showed that blocking the formation of the telomere cluster with the kms1 mutation created a disorganized chromosomal arrangement, not only for the regions proximal to the colt, but also for interstitial regions.
Abstract: A polarized chromosomal arrangement with clustered telomeres in a meiotic prophase nucleus is often called bouquet and is thought to be important for the pairing of homologous chromosomes. Fluorescence in situ hybridization in fission yeast indicated that chromosomal loci are positioned in an ordered manner as anticipated from the bouquet arrangement. Blocking the formation of the telomere cluster with the kms1 mutation created a disorganized chromosomal arrangement, not only for the regions proximal to the telomere but also for interstitial regions. The kms1 mutation also affected the positioning of a linear minichromosome. Consistent with this cytological observation, the frequency of ectopic homologous recombination between a linear minichromosome and a normal chromosome increased in the kms1 background. Intragenic recombination between allelic loci is reduced in the kms1 mutant, but those between non-allelic loci are unaffected or slightly increased. Thus, telomere-led chromosome organization facilitates homologous pairing and also restricts irregular chromosome pairing during meiosis.