Suppression of tumorigenicity in human ovarian cancer cell lines is controlled by a 2 cM fragment in chromosomal region 6q24-q25. (1/191)

Multiple distinct regions of chromosome 6 are frequently affected by losses of heterozygosity in primary human ovarian carcinomas. We introduced a normal human chromosome 6 into HEY and SKOV-3 ovarian carcinoma cell lines using microcell-mediated chromosome transfer techniques to further investigate the role of this chromosome in ovarian tumorigenesis. The exogenous chromosome was stably propagated in the recipient cells based on fluorescence in situ hybridization (FISH) analyses with a chromosome 6 painting probe. The tumorigenicity of HEY and SKOV-3 cells was completely suppressed after transfer of chromosome 6, but not after transfer of a chromosome 11q13-qter fragment used as control. Using 46 polymorphic microsatellite markers, the region bounded by D6S1649 and D6S1564 was found to be commonly deleted in HEY: chromosome 6 tumorigenic revertant clones. The boundaries of the commonly deleted region could be further narrowed down to a 2 cM (based on the Whitehead genetic map) or 0.36 megabase (based on gdb mapping data) region between D6S1637 and D6S1564 after transferring the exogenous chromosome from revertants into mouse L cells and performing allelic deletion mapping studies against this mouse background. We conclude that this region contains a tumor suppressor gene important for the control of ovarian tumor development.  (+info)

Comparison of AluI-induced frequencies of dicentrics and translocations in human lymphocytes by chromosome painting. (2/191)

It has been shown repeatedly that following irradiation of human lymphocytes in the G0 stage, more translocations are induced than dicentrics. To check the role of DNA double-strand breaks (DSB) alone for the induction of symmetrical and asymmetrical chromosome aberrations, the frequencies of induced exchange aberrations by the restriction enzyme AluI were analyzed. The enzyme was introduced into cells using the pellet pipetting technique. Frequencies of induced translocations and dicentrics were determined using a chromosome painting assay with chromosome-specific DNA libraries for chromosomes 1, 4 and X (representing 16.8% of the human genome). The number of translocations detected was approximately 3-fold higher than the number of dicentrics, indicating that the increased frequency of translocations compared with dicentrics found in irradiated human lymphocytes does not result from DNA lesions other than DSB but from differential processing of DSB.  (+info)

Analysis of bleomycin-induced chromosomal aberrations in Chinese hamster primary embryonic cells by FISH using arm-specific painting probes. (3/191)

Chinese hamster primary embryonic cells (at G1 phase) were treated with 1.0 or 3.0 microg/ml bleomycin and chromosomal aberrations in first division metaphases were analysed by fluorescence in situ hybridization (FISH) using arm-specific painting probes for chromosomes 3, 4, 8 and 9. We observed that bleomycin induced all classes of chromosome-type aberrations very efficiently. The interesting findings were: (i) the frequency of induced interstitial translocations (i.e. insertions) was approximately equal to that of reciprocal translocations; (ii) the frequency of induced pericentric inversions was higher than that of centric rings. In our earlier studies, we found that X-rays induced a low frequency of interstitial translocations in comparison with reciprocal translocations and equal frequencies of centric rings and pericentric inversions. These data suggest that bleomycin differs from X-rays with respect to the induction of some specific types of aberrations. The results of a chi2 test examining the hypothesis that formed aberrations among the chromosomes or chromosome arms are randomly distributed on the basis of their relative lengths revealed a differential involvement of these chromosomes in the aberrations following exposure to bleomycin. In general, chromosome 8 was found to be more involved in induced aberrations than expected, chromosome 4 was randomly involved, whereas chromosomes 3 and 9 were less involved. This study demonstrates the utility of arm-specific painting probes for efficient detection of a large variety of chromosomal aberrations induced by bleomycin.  (+info)

The application of comparative genomic hybridization and fluorescence in situ hybridization to the characterization of genotoxicity screening tester strains AHH-1 and MCL-5. (4/191)

AHH-1 TK+/- is a human B cell-derived lymphoblastoid cell line that constitutively expresses a high level of the cytochrome CYP1A1. The MCL-5 cell line was developed by transfection of AHH-1 with cDNAs encoding the human cytochrome P450s, CYP1A2, CYP2A6, CYP2E1, CYP3A4 and microsomal epoxide hydrolase carried in plasmids. The metabolic components of these cell lines make them a useful screening tool for use in mutagenicity studies. Although AHH-1 and MCL-5 are closely related, the two cell lines show differences which cannot be attributed to transfection. In the present study both cell lines were investigated for chromosome stability by comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH) using whole chromosome probes and telomeric probes. Amplification in chromosomes 4q, 3q and 9p was observed in both cell lines. To compare the cell lines directly, AHH-1 and MCL-5 DNAs were co-hybridized on the same metaphases using a modified CGH technique. The only difference observed between AHH-1 and MCL-5 was the degree of amplification involving the subtelomeric region of chromosome 4; the additional telomeric region (4q) was translocated onto chromosome 11 and/or chromosome X. FISH was use to show the presence of isochromosomes 3q and 9p in both cell lines with a chromosome number of 48 or higher. These data demonstrate that CGH and FISH with chromosome-specific probes are able to resolve complex karyotypes and to highlight subchromosomal regions involved in rearrangements and potential chromosome fragile sites. Analyses such as those described here may be of considerable value in the determination of the stability of a variety of the cell lines used in the mutagenicity testing of chemicals.  (+info)

Multiplex-FISH for pre- and postnatal diagnostic applications. (5/191)

For >3 decades, Giemsa banding of metaphase chromosomes has been the standard karyotypic analysis for pre- and postnatal diagnostic applications. However, marker chromosomes or structural abnormalities are often encountered that cannot be deciphered by G-banding alone. Here we describe the use of multiplex-FISH (M-FISH), which allows the visualization of the 22 human autosomes and the 2 sex chromosomes, in 24 different colors. By M-FISH, the euchromatin in marker chromosomes could be readily identified. In cases of structural abnormalities, M-FISH identified translocations and insertions or demonstrated that the rearranged chromosome did not contain DNA material from another chromosome. In these cases, deleted or duplicated regions were discerned either by chromosome-specific multicolor bar codes or by comparative genomic hybridization. In addition, M-FISH was able to identify cryptic abnormalities in patients with a normal G-karyotype. In summary, M-FISH is a reliable tool for diagnostic applications, and results can be obtained in +info)

Association of chromosome territories with the nuclear matrix. Disruption of human chromosome territories correlates with the release of a subset of nuclear matrix proteins. (6/191)

To study the possible role of the nuclear matrix in chromosome territory organization, normal human fibroblast cells are treated in situ via classic isolation procedures for nuclear matrix in the absence of nuclease (e.g., DNase I) digestion, followed by chromosome painting. We report for the first time that chromosome territories are maintained intact on the nuclear matrix. In contrast, complete extraction of the internal nuclear matrix components with RNase treatment followed by 2 M NaCl results in the disruption of higher order chromosome territory architecture. Correlative with territorial disruption is the formation of a faint DNA halo surrounding the nuclear lamina and a dispersive effect on the characteristically discrete DNA replication sites in the nuclear interior. Identical results were obtained using eight different human chromosome paints. Based on these findings, we developed a fractionation strategy to release the bulk of nuclear matrix proteins under conditions where the chromosome territories are maintained intact. A second treatment results in disruption of the chromosome territories in conjunction with the release of a small subset of acidic proteins. These proteins are distinct from the major nuclear matrix proteins and may be involved in mediating chromosome territory organization.  (+info)

Spatial relationship between transcription sites and chromosome territories. (7/191)

We have investigated the spatial relationship between transcription sites and chromosome territories in the interphase nucleus of human female fibroblasts. Immunolabeling of nascent RNA was combined with visualization of chromosome territories by fluorescent in situ hybridization (FISH). Transcription sites were found scattered throughout the territory of one of the two X chromosomes, most likely the active X chromosome, and that of both territories of chromosome 19. The other X chromosome territory, probably the inactive X chromosome, was devoid of transcription sites. A distinct substructure was observed in interphase chromosome territories. Intensely labeled subchromosomal domains are surrounded by less strongly labeled areas. The intensely labeled domains had a diameter in the range of 300-450 nm and were sometimes interconnected, forming thread-like structures. Similar large scale chromatin structures were observed in HeLa cells expressing green fluorescent protein (GFP)-tagged histone H2B. Strikingly, nascent RNA was almost exclusively found in the interchromatin areas in chromosome territories and in between strongly GFP-labeled chromatin domains. These observations support a model in which transcriptionally active chromatin in chromosome territories is markedly compartmentalized. Active loci are located predominantly at or near the surface of compact chromatin domains, depositing newly synthesized RNA directly into the interchromatin space.  (+info)

Gain of chromosomes 15 and 19 is frequent in both mouse hepatocellular carcinoma cell lines and primary tumors, but loss of chromosomes 4 and 12 is detected only in the cell lines. (8/191)

Chromosomal alterations were investigated in hepatocellular carcinoma cell lines, primary tumors and liver epithelial cell lines derived from normal livers of C57BL/6JxC3H/HeJ F(1) and C3H/HeJxC57BL/6J F(1) mice. In the primary tumors, non-random gain of chromosomes 15 and 19 was found in seven and five of 14 hepatocellular carcinomas, respectively. On the other hand, in the cases of both liver epithelial and hepatocellular carcinoma cell lines, frequent changes were loss of chromosomes 4 (4/9 cell lines) and 12 (3/9) as well as gain of chromosomes 15 (5/9) and 19 (4/9). These results indicate that the chromosomal gain is associated with both in vivo carcinogenesis and establishment of cell lines, while the loss is specific for the latter. PCR analysis using polymorphic microsatellite DNA markers revealed that the loss of chromosome 12 as well as chromosome 4 was much more frequent for the C57BL/6J hepatocarcinogenesis-resistant rather than the susceptible C3H/HeJ strain.  (+info)