Mechanisms of ring chromosome formation in 11 cases of human ring chromosome 21

We studied the mechanism of ring chromosome 21 (r(21)) formation in 13 patients (11 unique r(21)s), consisting of 7 from five families with familial r(21) and 6 with de novo r(21). The copy number of chromosome 21 sequences in the rings of these patients was determined by quantitative dosage analyses for 13 loci on 21q. Nine of 11 r(21)s, including the 5 familial r(21)s, showed no evidence for duplication of 21q sequences but did show molecular evidence of partial deletion of 21q. These data were consistent with the breakage and reunion of short- and long-arm regions to form the r(21), resulting in deletion of varying amounts of 21q22.1 to 21qter. The data from one individual who had a Down syndrome phenotype were consistent with asymmetric breakage and reunion of 21q sequences from an intermediate isochromosome or Robertsonian translocation chromosome as reported by Wong et al. Another patient, who also exhibited Down syndrome, showed evidence of a third mechanism of ring formation. The likely initial event was breakage and reunion of the short and long arms, resulting in a small r(21), followed by a sister-chromatid exchange resulting in a double-sized and symmetrically dicentric r(21). The phenotype of patients correlated well with the extent of deletion or duplication of chromosome 21 sequences. These data demonstrate three mechanisms of r(21) formation and show that the phenotype of r(21) patients varies with the extent of chromosome 21 monosomy or trisomy.     

Prenatal detection of an unstable ring 21 chromosome

Summary An unstable ring chromosome 21 detected through prenatal studies was associated at birth with an apparently normal male phenotype. At 14 months of age, examination indicated only minor developmental delay. The majority of cells examined from amniocyte, fibroblast, and lymphocyte cultures contained an asymmetrical dicentric ring 21 chromosome which was larger than a normal chromosome 21. This ring is presumed to be a duplication for most of chromosome 21 and a deletion of part of the terminal regions. The karyotype is described as mos45, XY,-21/46,XY,r(21)(p13q22.3). The child is monosomic for part of the sub-band 21q22.3 in every cell and trisomic for the remainder of the chromosome in most of his cells. The terminal deletion does not appear to have been severely detrimental to the phenotype and the effective trisomy present in many cells studies was insufficient to cause the Down syndrome.     

Unique genomic structure and distinct mitotic behavior of ring chromosome 21 in two unrelated cases

A ring chromosome replacing a normal chromosome could involve variable structural rearrangements and mitotic instability. However, most previously reported cases lacked further genomic characterization. High-resolution oligonucleotide array comparative genomic hybridization with single-nucleotide polymorphism typing (aCGH+SNP) was used to study 2 unrelated cases with a ring chromosome 21. Case 1 had severe myopia, hypotonia, joint hypermobility, speech delay, and dysmorphic features. aCGH detected a 1.275-Mb duplication of 21q22.12-q22.13 and a 6.731-Mb distal deletion at 21q22.2. Case 2 showed severe growth and developmental retardations, intractable seizures, and dysmorphic features. aCGH revealed a contiguous pattern of a 3.612- Mb deletion of 21q22.12-q22.2, a 4.568-Mb duplication of 21q22.2-q22.3, and a 2.243-Mb distal deletion at 21q22.3. Mitotic instability was noted in 13, 30, and 76% of in vitro cultured metaphase cells, interphase cells, and leukocyte DNA, respectively. The different phenotypes of these 2 cases are likely associated with the unique genomic structure and distinct mitotic behavior of their ring chromosome 21. These 2 cases represent a subtype of ring chromosome 21 probably involving somatic dicentric ring breakage and reunion. A cytogenomic approach is proposed for characterizing the genomic structure and mitotic instability of ring chromosome abnormalities.     

Submicroscopic duplication of chromosome 21 and trisomy 21 phenotype (Down syndrome)

Summary A patient with the phenotype of trisomy 21 (Down syndrome) was found to have a normal karyotype in blood lymphocytes and fibroblasts. Assessment of the chromosome 21 markers SOD1, CBS, ETS2, D21S11, and BCEI showed partial trisomy by duplication of a chromosome segment carrying the SOD1, CBS, and ETS2 loci and flanked by the BCEI and D21S11 loci, which are not duplicated. This submicroscopic duplication at the interface of 21q21 and 21q22.1 reduces to about 2000–3000kb the critical segment the trisomy of which is responsible for the phenotype of trisomy 21.     

Uniparental isodisomy due to duplication of chromosome 21 occurring in somatic cells monosomic for chromosome 21.

Uniparental disomy has been recently recognized as an important phenomenon in non-Mendelian inheritance of human genetic disorders. Several mechanisms for uniparental disomy, i.e., the presence of two homologous chromosomes derived from one parent, have been proposed. We studied two independent cases of abnormalities of chromosome 21 in which there were abnormal karyotypes at birth but blood cells with normal karyotype predominated later in life, and the cells with abnormalities disappeared. Uniparental isodisomy was observed in the normal cells in these individuals. The uniparental disomy in these families was the result of duplication of a chromosome in mitosis after the loss of the homologous abnormal chromosome. The duplication can be seen as mechanism for cell survival and is called here "compensatory" isodisomy, which provided a selective advantage for the cell population with the normal number of chromosomes 21.     

Cytogenetic and molecular analysis of a de novo tandem duplication of chromosome 21

Summary We have characterised by cytogenetic and molecular analysis a de novo tandem duplication of chromosome 21. High resolution chromosome examination of lymphocytes revealed the following karyotype in 90% of the cells: 46,XY,dir dup (21)(pterq22.300::q11.205 qter). Of these cells, 10% showed a normal karyotype. Gene dosage of chromosome 21 sequences by a slot blot method indicated that the duplication extends from D21S16 to D21S55. In situ hybridization with probes close to the borders of the duplicated segment confirmed the gene dosage data and gave results consistent with a true tandem duplication of chromosome 21. Pulsed field gel electrophoresis of the patient's DNA showed an abnormal restriction band common to D21S55 and D21S16, confirming that the junction point between the two homologous parts of the tandem chromosome brings these two sequences into proximity. Restriction fragment length polymorphism analysis indicated that the abnormal chromosome was maternal in origin and that the rearrangement of chromosome 21 could not have occurred at a post-zygotic stage of development but resulted from a recombination event during maternal gametogenesis. The possible mechanisms of formation of the abnormal chromosome are discussed, as is the presence of cells with normal chromosomes 21, in the patient.     

Rearrangement of chromosome 21 in Alzheimer's disease

We have analyzed the DNA from four patients suffering from a dementia of the Alzheimer type and of four controls of identical age. Estimates of the copy numbers of two genes located on chromosome 21, SOD1 and ETS2, gave the following results: in all the cases there was duplication of ETS2 whereas SOD1 was normal. These preliminary results indicate that the segment of chromosome 21(q21----q22.1) is rearranged in all the four patients who were investigated.     

Evolutionary breakpoints on human chromosome 21.

Segments of the long arm of human chromosome 21 are conserved, centromere to telomere, in mouse chromosomes 16, 17, and 10. There have been 28 genes identified in human chromosome 21 between TMPRSS2, whose orthologue is the most distal gene mapped to mouse chromosome 16, and PDXK, whose orthologue is the most proximal gene mapped to mouse chromosome 10. Only 6 of these 28 genes have been mapped in mouse, and all are located on chromosome 17. To better define the chromosome 17 segment and the 16 to 17 transition, we used a combination of mouse radiation hybrid panel mapping and physical mapping by mouse: human genomic sequence comparison. We have determined the mouse chromosomal location of an additional 12 genes, predicted the location of 7 more,and defined the endpoints of the mouse chromosome 17 region. The mouse chromosome 16/chromosome 17 evolutionary breakpoint is between human genes ZNF295 and UMODL1, showing there are seven genes in the chromosome 16 segment distal to Tmprss2. The chromosome 17/chromosome 10 breakpoint seems to have involved a duplication of the gene PDXK, which on chromosome 21 lies immediately distal to the KIAA0179 gene. These data suggest that there may be as few as 21 functional genes in the mouse chromosome 17 segment. This information is important for defining existing and constructing more complete mouse models of Down syndrome.     

Twofold CuZnSOD activity suggesting homozygous submicroscopic duplication of chromosome 21 in the orangutan

A twofold CuZnSOD activity was observed in an orangutan as compared to man and a chimpanzee. This suggests that a submicroscopic duplication of chromosome 21 (similar to those demonstrated in patients with the trisomy 21 phenotype but with a normal karyotype) has occurred in the homozygous state during evolution of the orangutan phylum. Such duplications could be important evolutionary mechanisms, together with visible chromosome rearrangements and classical gene mutations.     

Ring chromosome 21 in a normal female

In the present paper we report a phenotypically normal woman with a ring chromosome 21 in her karyotype, who sought genetic counseling because of her previous reproductive failure and whose third, prenatally diagnosed, pregnancy resulted in the delivery of a healthy and karyotypically normal boy.     

Retained heterodisomy for chromosome 12 in atypical lipomatous tumors: implications for ring chromosome formation

Atypical lipomatous tumor (ALT) is an intermediate malignant mesenchymal tumor that is characterized by supernumerary ring chromosomes and/or giant rod-shaped marker chromosomes (RGMC). Fluorescence in situ hybridization (FISH) and molecular genetic analyses have disclosed that the RGMCs always contain amplified sequences from the long arm of chromosome 12. Typically, RGMCs are the sole clonal changes and so far no deletions or other morphologic aberrations of the two normal-appearing chromosomes 12 that invariably are present have been detected. The mechanisms behind the formation of the RGMCs are unknown, but it could be hypothesized that RGMC formation is preceded by trisomy 12 or, alternatively, that ring formation of one chromosome 12 is followed by duplication of the remaining homolog. The latter scenario would always result in isodisomy for the two normal-appearing chromosomes 12, whereas the former would yield isodisomy in one-third of the cases. In order to investigate these possible mechanisms behind ring formation, we studied polymorphic loci on chromosome 12 in 14 cases of ALT showing one or more supernumerary ring chromosomes and few or no other clonal aberrations at cytogenetic analysis. The molecular genetic analyses showed that the tumor cells always retained both parental copies of chromosome 12, thus refuting the trisomy 12 and duplication hypotheses.     

Fluorescence in situ hybridization and Y ring chromosome

Summary Investigations by fluorescence in situ hybridization and a Y-specific probe (Y190) of a male patient with a Y ring chromosome, 46,X,r(Y) showed four bright fluorescent spots within the ring. Thus, using this technique, it is possible to suggest that the ring originates from the duplication of the short arms of the Y chromosome.     

Juxta-centromeric region of human chromosome 21 is enriched for pseudogenes and gene fragments

A physical map including four pseudogenes and 10 gene fragments and spanning 500 kb in the juxta-centromeric region of the long arm of human chromosome 21 is presented. cDNA fragments isolated from a selected cDNA library were characterized and mapped to the 831B6 YAC and to two BAC contigs that cover 250 kb of the region. An 85 kb genomic sequence located in the proximal region of the map was analyzed for putative exons. Four pseudogenes were found, including psiIGSF3, psiEIF3, psiGCT-rel whose functional copies map to chromosome 1p13, chromosome 2 and chromosome 22q11, respectively. The TTLL1 pseudogene corresponds to a new gene whose functional copy maps to chromosome 22q13. Ten gene fragments represent novel sequences that have related sequences on different human chromosomes and show 97-100% nucleotide identity to chromosome 21. These may correspond to pseudogenes on chromosome 21 and to functional genes in other chromosomes. The 85 kb genomic sequence was analyzed also for GC content, CpG islands, and repetitive sequence distribution. A GC-poor L isochore spanning 40 kb from satellite 1 was observed in the most centromeric region, next to a GC-rich H isochore that is a candidate region for the presence of functional genes. The pericentric duplication of a 7.8 kb region that is derived from the 22q13 chromosome band is described. We showed that the juxta-centromeric region of human chromosome 21 is enriched for retrotransposed pseudogenes and gene fragments transferred by interchromosome duplications, but we do not rule out the possibility that the region harbors functional genes also.     

Mirror image duplications of chromosome 21. Three new cases and discussion of the mechanisms of origin

Summary Three new cases of mirror image duplication of a chromosome 21 are studied. In the first case the extra chromosome derived from a single paternal chromosome 21 by intrachromosomal interchange. The analysis of similar published cases suggests that this mechanism may predominate, but interchromosomal interchange, para- and pericentric inversion are likely as well.     

No significant effect of monosomy for distal 21q22.3 on the Down syndrome phenotype in “Mirror” duplications of chromosome 21

Three Down syndrome patients for whom karyotypic analysis showed a "mirror" (reverse tandem) duplication of chromosome 21 were studied by phenotypic, cytogenetic, and molecular methods. On high-resolution R-banding analysis performed in two cases, the size of the fusion 21q22.3 band was apparently less than twice the size of the normal 21q22.3, suggesting a partial deletion of distal 21q. The evaluation of eight chromosome 21 single-copy sequences of the 21q22 region--namely, SOD1, D21S15, D21S42, CRYA1, PFKL, CD18, COL6A1, and S100B--by a slot blot method showed in all three cases a partial deletion of 21q22.3 and partial monosomy. The translocation breakpoints were different in each patient, and in two cases the rearranged chromosome was found to be asymmetrical. The molecular definition of the monosomy 21 in each patient was, respectively, COL6A1-S100B, CD18-S100B, and PFKL-S100B. DNA polymorphism analysis indicated in all cases a homozygosity of the duplicated material. The duplicated region was maternal in two patients and paternal in one patient. These data suggest that the reverse tandem chromosomes did not result from a telomeric fusion between chromosomes 21 but from a translocation between sister chromatids. The phenotypes of these patients did not differ significantly from that of individuals with full trisomy 21, except in one case with large ears with an unfolded helix. The fact that monosomy of distal 21q22.3 in these patients resulted in a phenotype very similar to Down syndrome suggests that the duplication of the genes located in this part of chromosome 21 is not necessary for the pathogenesis of the Down syndrome features observed in these patients, including most of the facial and hand features, muscular hypotonia, cardiopathy of the Fallot tetralogy type, and part of the mental retardation.     

Human chromosome 21-encoded cDNA clones

We have employed two strategies to isolate random cDNA clones encoded by chromosome 21. In the first approach, a cDNA library representing expressed genes of WA17, a mouse-human somatic cell hybrid carrying chromosome 21 as its sole human chromosome, was screened with total human DNA to identify human chromosome 21-specific cDNAs. The second approach utilized previously characterized single-copy genomic fragments from chromosome 21 as probes to retrieve homologous coding sequences from a human fetal brain cDNA library. Six cDNA clones on chromosome 21 were obtained in this manner. Two were localized to the proximal long arm of chromosome 21, two to the distal portion of the long arm, and one to the region of 21q22 implicated in the pathology of Down syndrome.     

The ring chromosome 13 syndrome

Summary A study of the ring chromosome 13 syndrome is presented with detailed clinical and cytogenetic features of three new unrelated cases. The clinical limits of this syndrome can now be defined. An analysis of these cases together with those in the literature indicates that the syndrome forms a continuous spectrum, and no further taxonomic subdivision is possible at this stage of knowledge. The chromosome breakpoints in the first two cases are 13p11 and 13q32 and in the third case 13p11 and 13q33 or 13q34. All described cases of the ring 13 syndrome have breakpoints within the region bounded by bands 13q21 to 13q34. All rings are negative for silver banding. Peripheral blood cultures showed an average of 88% of metaphases to be 46.XX,r(13), with the remaining 12% manifesting either random loss or ring duplication. The rings vary in size and show a variable number of centromeres. An estimate of the birth incidence of this condition in the Anglo-Saxon population is 1 in 58,000. Parents of affected children are clincally and cytogenetically normal, the rings in affected offspring being meiotic in origin.     

Ring chromosome 21 in healthy persons: different consequencies in females and in males

Summary A stable ring chromosome 21 was found in an azoospermic man with an otherwise normal phenotype. Meiotic studies in another known azoospermic male with r(21) had indicated that breakdown of spermatogenesis resulted from pairing failure of chromosome 21, followed by degenerative changes in the chromosomes, before the cells had completed the first meiotic division. While primary sterility was a constant feature in the three adult males, eight healthy females with r(21) were fertile. However, they were at risk for Down syndrome and spontaneous abortions.     

Formation of novel CENP-A domains on tandem repetitive DNA and across chromosome breakpoints on human chromosome 8q21 neocentromeres

Endogenous human centromeres form on megabase-sized arrays of tandemly repeated alpha satellite DNA. Human neocentromeres form epigenetically at ectopic sites devoid of alpha satellite DNA and permit analysis of centromeric DNA and chromatin organization. In this study, we present molecular cytogenetic and CENP-A chromatin immunoprecipitation (ChIP) on CHIP analyses of two neocentromeres that have formed in chromosome band 8q21 each with a unique DNA and CENP-A chromatin configuration. The first neocentromere was found on a neodicentric chromosome 8 with an inactivated endogenous centromere, where the centromeric activity and CENP-A domain were repositioned to band 8q21 on a large tandemly repeated DNA. This is the first example of a neocentromere forming on repetitive DNA, as all other mapped neocentromeres have formed on single copy DNA. Quantitative fluorescent in situ hybridization (FISH) analysis showed a 60% reduction in the alpha satellite array size at the inactive centromere compared to the active centromere on the normal chromosome 8. This neodicentric chromosome may provide insight into centromere inactivation and the role of tandem DNA in centromere structure. The second neocentromere was found on a neocentric ring chromosome that contained the 8q21 tandemly repeated DNA, although the neocentromere was localized to a different genomic region. Interestingly, this neocentromere is composed of two distinct CENP-A domains in bands 8q21 and 8q24, which are brought into closer proximity on the ring chromosome. This neocentromere suggests that chromosomal rearrangement and DNA breakage may be involved in neocentromere formation. These novel examples provide insight into the formation and structure of human neocentromeres.     

Regional assignment of six polymorphic DNA sequences on chromosome 21 by in situ hybridization to normal and rearranged chromosomes.

We have assigned six polymorphic DNA segments to chromosomal subregions and have established the physical order of these sequences on the long arm of chromosome 21 by in situ hybridization of cloned probes to normal metaphase chromosomes and chromosomes 21 from individuals with three different structural rearrangements: an interstitial deletion, a ring chromosome, and a reciprocal translocation involving four different breakpoints in band 21q22. Segments D21S1 and D21S11 map to region 21q11.2----q21, D21S8 to 21q21.1----q22.11, and D21S54 to 21q21.3----q22.11; D21S23 and D21S25 are both in the terminal subband 21q22.3, but they are separated by a chromosomal breakpoint in a ring 21 chromosome, a finding that places D21S23 proximal to D21S25. The physical map order D21S1/D21S11-D21S8-D21S54-D21S23-D21S25 agrees with the linkage map, but genetic distances are disproportionately larger toward the distal end of 21q.