Sites Of Chromosome Breakage Research Articles

Genome analyses of the new model protist Euplotes vannus focusing on genome rearrangement and resistance to environmental stressors.

As a model organism for studies of cell and environmental biology, the free-living and cosmopolitan ciliate Euplotes vannus shows intriguing features like dual genome architecture (i.e., separate germline and somatic nuclei in each cell/organism), "gene-sized" chromosomes, stop codon reassignment, programmed ribosomal frameshifting (PRF) and strong resistance to environmental stressors. However, the molecular mechanisms that account for these remarkable traits remain largely unknown. Here we report a combined analysis of de novo assembled high-quality macronuclear (MAC; i.e., somatic) and partial micronuclear (MIC; i.e., germline) genome sequences for E.vannus, and transcriptome profiling data under varying conditions. The results demonstrate that: (a) the MAC genome contains more than 25,000 complete "gene-sized" nanochromosomes (~85Mb haploid genome size) with the N50 ~2.7kb; (b) although there is a high frequency of frameshifting at stop codons UAA and UAG, we did not observe impaired transcript abundance as a result of PRF in this species as has been reported for other euplotids; (c) the sequence motif 5'-TA-3' is conserved at nearly all internally-eliminated sequence (IES) boundaries in the MIC genome, and chromosome breakage sites (CBSs) are duplicated and retained in the MAC genome; (d) by profiling the weighted correlation network of genes in the MAC under different environmental stressors, including nutrient scarcity, extreme temperature, salinity and the presence of ammonia, we identified gene clusters that respond to these external physical or chemical stimulations, and (e) we observed a dramatic increase in HSP70 gene transcription under salinity and chemical stresses but surprisingly, not under temperature changes; we link this temperature-resistance to the evolved loss of temperature stress-sensitive elements in regulatory regions. Together with the genome resources generated in this study, which are available online at Euplotes vannus Genome Database (http://evan.ciliate.org), these data provide molecular evidence for understanding the unique biology of highly adaptable microorganisms.

The incidence and origin of segmental aneuploidy in human oocytes and preimplantation embryos.

What is the incidence, origin and clinical significance of segmental aneuploidy in human oocytes and preimplantation embryos? Segmental aneuploidy occurs at a considerable frequency in preimplantation embryos with a majority being mitotic in origin. In recent years, accurate techniques for the detection of aneuploidy in single cells have been developed. Research using such methods has confirmed that aneuploidy is a common feature of human oocytes and preimplantation embryos. However, thus far research has mainly focused on loss or gain of whole chromosomes. We utilized sensitive molecular methods to study another important form of cytogenetic abnormality at the earliest stages of human development, namely segmental aneuploidy. Chromosomal copy number data was obtained from oocytes and embryos of 635 IVF patients, who requested chromosome screening for various reasons, most commonly for advanced maternal age or previously unsuccessful IVF treatments. A total of 3541 samples comprising of 452 human oocytes, 1762 cleavage stage and 1327 blastocyst stage embryos were investigated in the present study. Whole genome amplification (Sureplex, Illumina) was performed on cells biopsied from oocytes and embryos of IVF patients who requested chromosome screening. The samples were subsequently processed and analyzed for their chromosome complement using microarray comparative genomic hybridization (aCGH), (Illumina, Cambridge, UK). Segmental abnormalities, involving loss or gain of chromosomal fragments in excess of 15 Mb, were found to occur at a high frequency. The incidence of such abnormalities was 10.4% in oocytes, but this increased dramatically during the first 3 days of embryonic development (24.3%), before starting to decline as embryos reached the final (blastocyst) stage of preimplantation development (15.6%). While some segmental errors were clearly of meiotic origin, most appear to arise during the first few mitoses following fertilization. The reduction in frequency at the blastocyst stage suggests that many cells/embryos affected by segmental abnormalities are eliminated (e.g. via arrest of the affected embryos or apoptosis of abnormal cells). Interestingly, sites of chromosome breakage associated with segmental aneuploidy were not entirely random but tended to occur within distinct chromosomal regions. Some of the identified hotspots correspond to known fragile sites while others may be considered novel and may be specific to gametogenesis and/or embryogenesis. The cytogenetic analysis was performed on biopsies of embryos, which might not be representative of the true incidence of mosaic segmental aneuploidy of the entire embryo. The findings of this study are valuable for understanding the origin of subchromosomal duplications and deletions, a clinically important class of abnormalities that are a common cause of congenital abnormalities and miscarriage. Furthermore, the results provide additional evidence that control of the cell cycle is more relaxed during the first few mitotic divisions following fertilization, permitting DNA double-strand breaks to occur and persist through cell division. The data are also of great relevance for preimplantation genetic testing, where the detection of segmental aneuploidy is currently considered problematic for embryo diagnosis and patient counseling. This study was supported by institutional funding (Reprogenetics UK). Additionally, DW is supported by the National Institute for Health Research (NIHR) Oxford Biomedical Research Centre Programme. DB was supported by the University of Oxford's Clarendon funding. No conflict of interests to declare.

Tetrahymena Pot2 Is a Developmentally Regulated Paralog of Pot1 That Localizes to Chromosome Breakage Sites but Not to Telomeres

Tetrahymena telomeres are protected by a protein complex composed of Pot1, Tpt1, Pat1, and Pat2. Pot1 binds the 3' overhang and serves multiple roles in telomere maintenance. Here we describe Pot2, a paralog of Pot1 which has evolved a novel function during Tetrahymena sexual reproduction. Pot2 is unnecessary for telomere maintenance during vegetative growth, as the telomere structure is unaffected by POT2 macronuclear gene disruption. Pot2 is expressed only in mated cells, where it accumulates in developing macronuclei around the time of two chromosome processing events: internal eliminated sequence (IES) excision and chromosome breakage. Chromatin immunoprecipitation (ChIP) demonstrated Pot2 localization to regions of chromosome breakage but not to telomeres or IESs. Pot2 association with chromosome breakage sites (CBSs) occurs slightly before chromosome breakage. Pot2 did not bind CBSs or telomeric DNA in vitro, suggesting that it is recruited to CBSs by another factor. The telomere proteins Pot1, Pat1, and Tpt1 and the IES binding factor Pdd1 fail to colocalize with Pot2. Thus, Pot2 is the first protein found to associate specifically with CBSs. The selective association of Pot2 versus Pdd1 with CBSs or IESs indicates a mechanistic difference between the chromosome processing events at these two sites. Moreover, ChIP revealed that histone marks characteristic of IES processing, H3K9me3 and H3K27me3, are absent from CBSs. Thus, the mechanisms of chromosome breakage and IES excision must be fundamentally different. Our results lead to a model where Pot2 directs chromosome breakage by recruiting telomerase and/or the endonuclease responsible for DNA cleavage to CBSs.

Regression of Replication Forks Stalled by Leading-strand Template Damage

Stalled replication forks are sites of chromosome breakage and the formation of toxic recombination intermediates that undermine genomic stability. Thus, replication fork repair and reactivation are essential processes. Among the many models of replication fork reactivation is one that invokes fork regression catalyzed by the strand exchange protein RecA as an intermediate in the processing of the stalled fork. We have investigated the replication fork regression activity of RecA using a reconstituted DNA replication system where the replisome is stalled by collision with leading-strand template damage. We find that RecA is unable to regress the stalled fork in the presence of the replisome and SSB. If the replication proteins are removed from the stalled fork, RecA will catalyze net regression as long as the Okazaki fragments are sealed. RecA-generated Holliday junctions can be detected by RuvC cleavage, although this is not a robust reaction. On the other hand, extensive branch migration by RecA, where a completely unwound product consisting of the paired nascent leading and lagging strands is produced, is observed under conditions where RuvC activity is suppressed. This branch migration reaction is inhibited by SSB, possibly accounting for the failure of RecA to generate products in the presence of the replication proteins. Interestingly, we find that the RecA-RuvC reaction is supported to differing extents, depending on the template damage; templates carrying a cyclopyrimidine dimer elicit more RecA-RuvC product than those carrying a synthetic abasic site. This difference could be ascribed to a higher affinity of RecA binding to DNAs carrying a thymidine dimer than to those with an abasic site.

Effect of high copy retrotransposon insertions on yeast genome stability

Retrotransposons are dispersed sequences that are able to copy themselves and reinsert throughout the genome. These mobile elements display preferences for the positions in the genome at which they insert and over 90% of retrotransposon insertions in Saccharomyces cerevisiae are within 2 kb upstream of tRNA genes. The yeast genome is comprised of only 3% retrotransposons as compared with 44% in humans; to use the yeast genome as a model to study insertion preferences in retrotransposon‐dense genomes, Retrotransposon Overdose (RO) strains were made by inserting a large number of Ty1 retrotransposons within the yeast genome. Our first goal was to analyze the genome sequences of these strains to determine the sites of Ty1 insertion. Analysis of the sequenced genomes revealed that as the number of retrotransposons increased, the number of insertions into less preferred target sites increased, including a significant number of insertions within essential genes, which may decrease either the quantity or size of transcription products. Our second goal was to analyze the targeting of Ty1 retrotransposons once all preferred sites in the genome were saturated. We have found that pairs of retrotransposons are frequently formed. Retrotransposons that are adjacent to each other in the genome are hypothesized to form fragile sites, which are sites of chromosome breakage that are associated with tumor formation in humans. RO strains contain numerous pairs of retrotransposons as well as higher order Ty1 multimers, despite the genome instability that might result from these retrotransposon arrangements.

Damage‐induced association of Rap1p with an internal binding site stimulates de novo telomere addition

At a low rate, sites of chromosome breakage in yeast are “healed” through telomere addition by the enzyme telomerase. While de novo telomere addition is normally repressed, it may serve to prevent chromosome loss in the face of irreparable damage. To address this possibility, we have sought factors that positively regulate telomerase action at chromosome breaks. Our study utilizes a “hotspot” for de novo telomere addition on yeast chromosome 5. This ~80 bp region contains TG‐rich sequences with similarity to yeast telomeric repeats. We find that function of the Rap1/Rif1/Rif2 complex is required for most de novo telomere addition within and flanking the hotspot. While this complex normally associates with telomeres to negatively regulate telomere length, Rap1/Rif1/Rif2 appears to stimulate telomere addition at ends with little or no telomeric sequence. Mutation of a putative Rap1p binding site at the centromere‐proximal end of the hotspot abolishes telomere addition, even though most of the TG‐rich sequences remain. Chromatin‐immunoprecipitation experiments demonstrate that Rap1p associates with hotspot DNA, but only after a chromosome break is induced telomere‐proximal to the hotspot. We suggest that Rap1p binds to internal chromosomal sites as a response to DNA damage and stimulates de novo telomere addition on a subset of broken ends to mediate damage tolerance.Funded by the National Science Foundation.

Replication Stress-Induced Chromosome Breakage Is Correlated with Replication Fork Progression and Is Preceded by Single-Stranded DNA Formation

Chromosome breakage as a result of replication stress has been hypothesized to be the direct consequence of defective replication fork progression, or “collapsed” replication forks. However, direct and genome-wide evidence that collapsed replication forks give rise to chromosome breakage is still lacking. Previously we showed that a yeast replication checkpoint mutant mec1-1, after transient exposure to replication impediment imposed by hydroxyurea (HU), failed to complete DNA replication, accumulated single-stranded DNA (ssDNA) at the replication forks, and fragmented its chromosomes. In this study, by following replication fork progression genome-wide via ssDNA detection and by direct mapping of chromosome breakage after HU exposure, we have tested the hypothesis that the chromosome breakage in mec1 cells occurs at collapsed replication forks. We demonstrate that sites of chromosome breakage indeed correlate with replication fork locations. Moreover, ssDNA can be detected prior to chromosome breakage, suggesting that ssDNA accumulation is the common precursor to double strand breaks at collapsed replication forks.

Non-B DNA structure-induced genetic instability and evolution.

Repetitive DNA motifs are abundant in the genomes of various species and have the capacity to adopt non-canonical (i.e., non-B) DNA structures. Several non-B DNA structures, including cruciforms, slipped structures, triplexes, G-quadruplexes, and Z-DNA, have been shown to cause mutations, such as deletions, expansions, and translocations in both prokaryotes and eukaryotes. Their distributions in genomes are not random and often co-localize with sites of chromosomal breakage associated with genetic diseases. Current genome-wide sequence analyses suggest that the genomic instabilities induced by non-B DNA structure-forming sequences not only result in predisposition to disease, but also contribute to rapid evolutionary changes, particularly in genes associated with development and regulatory functions. In this review, we describe the occurrence of non-B DNA-forming sequences in various species, the classes of genes enriched in non-B DNA-forming sequences, and recent mechanistic studies on DNA structure-induced genomic instability to highlight their importance in genomes.

Phosphorylation of SDT repeats in the MDC1 N terminus triggers retention of NBS1 at the DNA damage–modified chromatin

DNA double-strand breaks (DSBs) trigger accumulation of the MRE11–RAD50–Nijmegen breakage syndrome 1 (NBS1 [MRN]) complex, whose retention on the DSB-flanking chromatin facilitates survival. Chromatin retention of MRN requires the MDC1 adaptor protein, but the mechanism behind the MRN–MDC1 interaction is unknown. We show that the NBS1 subunit of MRN interacts with the MDC1 N terminus enriched in Ser-Asp-Thr (SDT) repeats. This interaction was constitutive and mediated by binding between the phosphorylated SDT repeats of MDC1 and the phosphate-binding forkhead-associated domain of NBS1. Phosphorylation of the SDT repeats by casein kinase 2 (CK2) was sufficient to trigger MDC1–NBS1 interaction in vitro, and MDC1 associated with CK2 activity in cells. Inhibition of CK2 reduced SDT phosphorylation in vivo, and disruption of the SDT-associated phosphoacceptor sites prevented the retention of NBS1 at DSBs. Together, these data suggest that phosphorylation of the SDT repeats in the MDC1 N terminus functions to recruit NBS1 and, thereby, increases the local concentration of MRN at the sites of chromosomal breakage.

The Highly Conserved Family of Tetrahymena thermophila Chromosome Breakage Elements Contains an Invariant 10-Base-Pair Core

As a typical ciliate, Tetrahymena thermophila is a unicellular eukaryote that exhibits nuclear dimorphism: each cell contains a diploid, germ line micronucleus (MICN) and a polyploid, somatic macronucleus (MACN). During conjugation, when a new MACN differentiates from a mitotic descendant of the diploid fertilization nucleus, the five MICN chromosomes are site-specifically fragmented into 250 to 300 MACN chromosomes. The classic chromosome breakage sequence (CBS) is a 15-bp element (TAAACCAACCTCTTT) reported to be necessary and sufficient for chromosome breakage. To determine whether a CBS is present at every site of chromosome fragmentation and to assess the range of sequence variation tolerated, 31 CBSs were isolated without preconception as to the sequence of the chromosome breakage element. Additional CBS-related sequences were identified in the whole-genome sequence by their similarities to the classic CBS. Forty CBS elements behaved as authentic chromosome breakage sites. The CBS nucleotide sequence is more diverse than previously thought: nearly half of the CBS elements identified by unbiased methods have a variant of the classic CBS. Only an internal 10-bp core is completely conserved, but the entire 15-bp chromosome breakage sequence shows significant sequence conservation. Our results suggest that any one member of the CBS family provides a necessary and sufficient cis element for chromosome breakage. No chromosome breakage element totally unrelated to the classic CBS element was found; such elements, if they exist at all, must be rare.

Genome-wide characterization of Tetrahymena thermophila chromosome breakage sites. II. Physical and genetic mapping.

The chromosomes of the macronuclear (expressed) genome of Tetrahymena thermophila are generated by developmental fragmentation of the five micronuclear (germline) chromosomes. This fragmentation is site specific, directed by a conserved chromosome breakage sequence (Cbs element). An accompanying article in this issue reports the development of a successful scheme for the genome-wide cloning and identification of functional chromosome breakage sites. This article reports the physical and genetic characterization of 30 functional chromosome breakage junctions. Unique sequence tags and physical sizes were obtained for the pair of macronuclear chromosomes generated by fragmentation at each Cbs. Cbs-associated polymorphisms were used to genetically map 11 junctions to micronuclear linkage groups and macronuclear coassortment groups. Two pairs of junctions showed statistically significant similarity of the sequences flanking the Cbs, suggestive of relatively recent duplications of entire Cbs junctions during Tetrahymena genome evolution. Two macronuclear chromosomes that lose at least one end in an age-related manner were also identified. The whole-genome shotgun sequencing of the Tetrahymena macronucleus has recently been completed at The Institute for Genome Research (TIGR). By providing unique sequence from natural ends of macronuclear chromosomes, Cbs junctions will provide useful sequence tags for relating macro- and micronuclear genetic, physical, and whole-genome sequence maps.

Genome-wide characterization of tetrahymena thermophila chromosome breakage sites. I. Cloning and identification of functional sites.

The chromosomes of the macronuclear (expressed) genome of Tetrahymena thermophila are generated by developmental fragmentation of the five micronuclear (germline) chromosomes. This fragmentation is site specific and directed by a conserved 15-bp chromosome breakage sequence (Cbs element). This article reports the construction of a library enriched for chromosome breakage junctions and the development of a successful scheme for the genome-wide isolation and characterization of functional Cbs junctions. Twenty-three new Cbs junctions were characterized and each was assigned to a specific micronuclear chromosome or chromosome arm. Two distinct previously unreported variant chromosome breakage sequences were found, each in two or more functional Cbs elements. Analysis of natural Cbs junctions confirmed that microheterogeneity in the macronuclear telomere addition site is associated with chromosome fragmentation. The physical and genetic characterization of these functional chromosome breakage junctions is reported in the accompanying article in this issue. The whole-genome shotgun sequencing and auto-assembly phase of the Tetrahymena Genome Initiative has recently been completed at The Institute for Genome Research (TIGR). By providing unique sequence from the natural ends of macronuclear chromosomes, Cbs junctions characterized in the work reported here will serve as useful sequence tags for relating macro- and micronuclear genetic, physical, and sequence maps.

The Tetrahymena thermophila genome sequence: 3. Completing macronuclear chromosome sequence scaffolds and ordering and aligning them along the micronuclear genome

In Ciliates, the diploid, mitotic, genetically silent micronucleus (MIC) is the cell's germline, while the macronucleus (MAC) is the exclusive site of gene expression. During (postzygotic) MAC differentiation in T. thermophila, several hundred chromosomes are generated by site specific fragmentation of the five germline‐derived chromosomes at highly conserved chromosome breakage sites. Telomeres are added to every newly generated end, and chromosomes are amplified to ∼45 N. The Tetrahymena thermophila macronuclear genome was recently whole‐genome‐shotgun sequenced to nine‐fold redundancy at TIGR. The sequence (publicly available at http://www.tigr.org/tdb/e2k1/ttg/) is assembled into contigs (contiguous segments of overlapping sequence reads) linked into scaffolds. Here, we report here the completion of the assembly of many scaffold ends by linking them to their corresponding telomere, using diverse bioinformatic or experimental approaches. 120 chromosomes are now assembled from telomere to telomere and 125 additional scaffolds are telomere capped at one end. The 120 complete scaffolds encompass ∼45% of the ∼105‐Mb assembly, suggesting a total of ∼290 MAC chromosomes. By using paired sequence tags flanking chromosome breakage sites, we are linking unambiguously ordered and oriented MAC chromosome DNA sequence into MIC genome assemblies.Supported by NIH grant RR02931. Preliminary sequence data were obtained from The Institute for Genomic Research website.

Adeno-associated virus vectors integrate at chromosome breakage sites.

Adeno-associated virus (AAV) vectors transduce cells by multiple pathways, including integration at nonhomologous chromosomal locations by an unknown mechanism. We reasoned that spontaneous chromosome breaks may facilitate vector integration and investigated this in cells containing a specific chromosomal double-strand break created by the endonuclease I-SceI or multiple breaks created by treatment with etoposide or gamma-irradiation. Vector proviruses were found at I-SceI cleavage sites, and sequencing of vector-chromosome junctions detected microhomologies, deletions and insertions that were similar when integration occurred spontaneously at random locations or at induced double-strand breaks. Infection with AAV vectors did not increase mutation rates in normal human cells. Our results establish a mechanism for integration and suggest that AAV vectors can integrate at existing chromosome breaks rather than causing breaks themselves, which has implications for their clinical use.

The Protein Kinase CK2 Facilitates Repair of Chromosomal DNA Single-Strand Breaks

CK2 was the first protein kinase identified and is required for the proliferation and survival of mammalian cells. Here, we have identified an unanticipated role for CK2. We show that this essential protein kinase phosphorylates the scaffold protein XRCC1 and thereby enables the assembly and activity of DNA single-strand break repair protein complexes in vitro and at sites of chromosomal breakage. Moreover, we show that inhibiting XRCC1 phosphorylation by mutation of the CK2 phosphorylation sites or preventing CK2 activity using a highly specific inhibitor ablates the rapid repair of cellular DNA single-strand breaks by XRCC1. These data identify a direct role for CK2 in the repair of chromosomal DNA strand breaks and in maintaining genetic integrity.

Jumping Translocations Involving 11q in a Non-Hodgkin Lymphoma

This paper presents the results of a cytogenetic analysis in an 11-year-old boy with non-Hodgkin lymphoma. The investigation was performed on slides obtained from short-term culture of lymph node cells. The analyses revealed an abnormal clone with loss of the Y, gain of an X chromosome, t(3;22), trisomy 11, and three cytogenetically-related subclones with jumping translocations involving 11q13 as the common breakpoint region. This region is an unusual site of chromosome breakage in jumping translocations, and has not been reported thus far. Contrary to most published reports, the jumping translocation in our patient is associated with long survival.

Nonrandom Cytogenetic Alterations in Hepatocellular Carcinoma from Transgenic Mice Overexpressing c-Myc and Transforming Growth Factor-α in the Liver

Identification of specific and primary chromosomal alterations during the course of neoplastic development is an essential part of defining the genetic basis of cancer. We have developed a transgenic mouse model for liver neoplasia in which chromosomal lesions associated with both the initial stages of the neoplastic process and the acquisition of malignancy can be analyzed. Here we analyze chromosomal alterations in 11 hepatocellular carcinomas from the c-myc/TGF-alpha double-transgenic mice by fluorescent in situ hybridization with whole chromosome probes, single-copy genes, and 4'-6-diamidino-2-phenylindole (DAPI-) and G-banded chromosomes and report nonrandom cytogenetic alterations associated with the tumor development. All tumors were aneuploid and exhibited nonrandom structural and numerical alterations. A balanced translocation t(5:6)(G1;F2) was identified by two-color fluorescent in situ hybridization in all tumors, and, using a genomic probe, the c-myc transgene was localized near the breakpoint on derivative chromosome der 6. Partial or complete loss of chromosome 4 was observed in all tumors with nonrandom breakage in band C2. Deletions of chromosome 1 were observed in 80% of the tumors, with the most frequent deletion at the border of bands C4 and C5. An entire copy of chromosome 7 was lost in 80% of the tumors cells. Eighty-five percent of the tumor cells had lost one copy of chromosome 12, and the most common breakpoint on chromosome 12 occurred at band D3 (28%). A copy of chromosome 14 was lost in 72%, and band 14E1 was deleted in 32% of the tumor cells. The X chromosome was lost in the majority of the tumor cells. The most frequent deletion on the X chromosome involved band F1. We have previously shown that breakages of chromosomes 1, 6, 7, and 12 were observed before the appearance of morphologically distinct neoplastic liver lesions in this transgenic mouse model. Thus breakpoints on chromosome 4, 9, 14, and X appear to be later events in this model of liver neoplasia. This is the first study to demonstrate that specific sites of chromosomal breakage observed during a period of chromosomal instability in early stages of carcinogenesis are later involved in stable rearrangements in solid tumors. The identification of the 5;6 translocation in all of the tumors has a special significance, being the first balanced translocation reported in human and mouse hepatocellular carcinoma and having the breakpoint near a tumor susceptibility gene and myc transgene site of integration. Moreover, its early occurrence indicates that this is a primary and relevant alteration to the initiation of the neoplastic process. In addition, the concordance between the breakpoints observed during the early dysplastic stage of hepatocarcinogenesis and the stable deletions of chromosomes 1, 4, 6, 7, 9, and 12 in the tumors provides evidence for preferential site of genetic changes in hepatocarcinogenesis.

Expansion and length-dependent fragility of CTG repeats in yeast.

Expansion of DNA trinucleotide repeats (TNRs) is the causative mutation in a growing number of human genetic diseases. Large expansions of a CTG tract were obtained and shown by genetic and physical assays to be length-dependent sites of chromosome breakage in Saccharomyces cerevisiae. Deletion of RAD27, which encodes a nuclease involved in Okazaki fragment processing, caused length-dependent destabilization of CTG tracts and a substantial increase in expansion frequency. The genetic assay described here can be used to evaluate other factors that induce TNR expansion or chromosome fragility in humans.

Ribosomal DNA is a site of chromosome breakage in aneuploid strains of Neurospora.

In wild-type strains of Neurospora crassa, the rDNA is located at a single site in the genome called the nucleolus organizer region (NOR), which forms a terminal segment on linkage group (LG) V. In the quasiterminal translocation strain T(I;V)AR190, most of the right arm of LG I moved to the distal tip of the NOR, and one or a few rDNA repeat units are moved to the truncated right arm of LG I. I report here that, in partial diploid strains derived from T(I;V)AR190, large terminal deletions result from chromosome breakage in the NOR. In most of these partial diploids, chromosome breakage is apparently frequent and the breakpoints occur in many parts of the NOR. The rDNA ends resulting from chromosome breakage are "healed" by the addition of new telomeres. Significantly, the presence of ectopic rDNA creates a new site of chromosome breakage in the genome of partial diploids. These results raise the possibility that, under certain conditions, rDNA is a region of fragility in eukaryotic chromosomes.

Cytogenetic changes at 11q11, 11q23, and 17q11 in myelodysplastic syndrome

Cytogenetic studies were performed on two patients with myelodysplastic syndromes. One patient was a 68 year old Japanese male in whose bone marrow cells two translocations were established, i.e., t(4;11)(q13;q23) and t(11;17)(q11?;q11), as well as other karyotypic changes (−6, −18,15p +). The other patient was a 74 year old white male whose bone marrow cells showed six marker chromosomes, i.e., der(5), t(5;17)( q12; q11), der(6), t(6;5)( q27; q22). der(8), t(8;11;?)( q11→ q23;?), der(11), t(11;?), an isochromosome of the long arm of chromosome #8, and a small G-group sized marker chromosome of unknown origin. Though the translocation patterns in the abnormal cells in these two cases were different, the breakpoints of the marker chromosomes were almost the same, i.e., 11q11, 11q23, and 17q11. Also, changes of chromosome #6 were observed; the first case showed monosomy 6 and the second a 6q + marker chromosome. In these two cases of myelodysplastic syndromes, common sites of chromosome breakage and reunion of 11q23 and 17q11 were close to recently established sites of human cellular oncogene homologs, c- ets (11q23) and c- erbA (17q21→24). These associations draw attention to a possible relationship between chromosome changes in myelodysplastic syndromes and oncogene (or other gene) activation and/or dysfunction.