Clinical and cytogenetic studies of the Prader-Willi syndrome: Evidence of phenotype-karyotype correlation

Summary Twenty-seven patients with the presumed diagnosis of Prader-Willi syndrome (PWS) were studied clinically and cytogenetically. The patients were classified into three study groups on the basis of their clinical pictures: group 1 with 12 children meeting the strict diagnostic criteria for PWS; group 2 with nine floppy infants and young children, aged 3 years or less, without obesity and hyperphagia; and group 3 with six older children in whom some characteristic features of the syndrome were absent. High-resolution GTG banding of prometaphase chromosomes revealed del(15)(q11.1;q12) in eleven and t(15;15)(qterp11.2::q12qter) in one of the twelve group 1 patients. In group 2, four patients had del(15)(q11.1;q12), one had t(15;15)(qterp11.1::q13qter), and the remaining four had normal karyotypes. The deleted segment common to the 17 patients with the chromosome aberrations was confined to subband 15q11.2. On the other hand, all six group 3 patients had normal karyotypes. These findings indicated that when strictly defined PWS is absolutely correlated with chromosome 15 aberrations, i.e, there is a positive phenotype-karyotype correlation, and that the aberrations are etiologically related to the syndrome. Parental origin of the deleted chromosome was determined in seven patients, with OFQ-heteromorphisms being used as hereditary markers. The deleted chromosome originated from the paternal chromosome 15 in six patients and from the maternal 15 in one.     

Investigations with fluorescence in situ hybridization (FISH) demonstrate loss of the telomeres on the reciprocal chromosome in three unbalanced translocations involving chromosome 15 in the Prader-Willi and Angelman syndromes

Two patients with classical features of Angelman syndrome (AS) and one with Prader-Willi syndrome (PWS) had unbalanced reciprocal translocations involving the chromosome 15 proximal long arm and the telomeric region of chromosomes 7, 8 and 10. Fluorescence isitu hybridization (FISH) was used for the detection of chromosome 15(q11-13) deletions (with probes from the PWS/AS region) and to define the involvement of the telomere in the derivative chromosomes (with library probes and telomere-specific probes). The 15(q11-13) region was not deleted in one patient but was deleted in the other two. The telomere on the derivative chromosomes 7, 8 and 10 was deleted in all three cases. Thus, these are true reciprocal translocations in which there has been loss of the small satellited reciprocal chromosome (15) fragment.     

Characterization of an interstitial deletion del(13)(q22q32) using microdissection and sequential FISH and G-banding

The objective of this study was to delineate a chromosome 13 abnormality and establish its clinical correlation by using molecular cytogenetics procedures. A newborn boy presented with clinical findings, including mild symmetric intrauterine growth retardation (IUGR), small ears with thickened helices, a scalp lesion, short fifth fingers, missing toes, and talipes equinovarus. Routine G-banding of cultured peripheral blood cells revealed that the patient had one abnormal and shortened chromosome 13, but uncertainty remained as to whether the abnormality was the result of an interstitial deletion or a translocation. Thirteen copies of G-banded abnormal chromosomes 13 were isolated with microdissection and amplified with PCR using degenerate oligonucleotide primers. Fluorescence in situ hybridization (FISH) of the PCR product to normal metaphases showed one pair of acrocentrics hybridized, more or less uniformly, along the length of the long arm with an unhybridized gap in the distal region, indicative of an interstitial deletion. Sequential FISH and G-banding of the same chromosome preparations conclusively demonstrated that the deleted segment was 13q22-q32. Four cases of del(13)(q22q32) have been previously reported. The common findings in all five cases, including the present one, are psychomotor and growth retardation, as well as hand and foot anomalies.     

Langer-Giedion syndrome,in a child with complex structural aberration of chromosome 8

A patient with typical features of the Langer-Giedion syndrome (tricho-rhino-phalangeal syndrome, type II) is described. In the karyotype an interstitial deletion of the long arm of chromosome 8 (band 8q22) was observed as the result of a complex rearrangement of chromosomes 1 and 8: 46,XY inv(8)(q23 leads to q242), del(8)(q221 leads to q223), ins(8;1) (q221;p321 p341;q242). Previously reported cases of Langer-Giedion syndrome with deletion of 8q are compared with the present one.     

Chromosomal rearrangements and their effects in spontaneous immortalization and transformation of rat embryo cells in vitro

G-banding analysis of LRec-1 and LRec-3, spontaneously immortalized cell lines from rat embryo fibroblast, revealed diploid karyotypes with specific clonal structural rearrangements of chromosomes 7 and 19 - del(7)(q11.2q22.1), t(7;19)(q11.1;q12) in malignant stage. Both clonal abnormalities of chromosomes 7 and 19 were also revealed in LRec-1k clone and LRec-1 sf cell line. Previous study of LRec-1 and LRec-3 cells showed the presence of karyotypes with pseudodiploid modal chromosome number, partial trisomy of chromosome 7 and same clonal structural rearrangements of chromosomes 7 and 19 in immortalized stage. In malignant stage, the trisomy 6 and new clonal structural rearrangements of chromosomes 1, 2, 11, 15, 18, 19 and of chromosomes 10, 20 were also found in LRec-1 sf and LRec-1 cells, accordingly. There were no new clonal structural chromosome rearrangements in LRec-1 k and LRec-3 cells. We compared locies of chromosomes involved in rearrangements with mapped genes on these chromosomes according to RATMAP. Supposedly these genes are involved in spontaneous immortalization of rat embryo fibroblast and malignant transformation of LRec-1 and LRec-3 cells and rearrangements of chromosomes 1, 2, 11, 15 and 18 facilitate expression of growth factors of LRec-1 sf cells.     

Somatic cell hybrids, sequence-tagged sites, simple repeat polymorphisms, and yeast artificial chromosomes for physical and genetic mapping of proximal 17p.

Somatic cell hybrids retaining the deleted chromosome 17 from 15 unrelated Smith-Magenis syndrome (SMS) [del(17)(p11.2p11.2)] patients were obtained by fusion of patient lymphoblasts with thymidine kinase-deficient rodent cell lines. Seventeen sequence-tagged sites (STSs) were developed from anonymous markers and cloned genes mapping to the short arm of chromosome 17. The STSs were used to determine the deletion status of these loci in these and four previously described human chromosome 17-retaining hybrids. Ten STSs were used to identify 28 yeast artificial chromosomes (YACs) from the St. Louis human genomic YAC library. Four of the 17 STSs identified simple repeat polymorphisms. The order and location of deletion breakpoints were confirmed and refined, and the regional assignment of several probes and cloned genes were determined. The cytogenetic band locations and relative order of six markers on 17p were established by fluorescence in situ hybridization mapping to metaphase chromosomes. The latter data confirmed and supplemented the somatic cell hybrid results. Most of the hybrids derived from [del(17)(p11.2p11.2)] patients demonstrated a similar pattern of deletion for the marker loci and were deleted for D17S446, D17S258, D17S29, D17S71, and D17S445. However, one of them demonstrated a unique pattern of deletion. This patient is deleted for several markers known to recognize a large DNA duplication associated with Charcot-Marie-Tooth (CMT) disease type 1A. These data suggest that the proximal junction of the CMT1A duplication is close to the distal breakpoint in [del(17)(p-11.2p11.2)] patients.     

Partial trisomy 14q

Summary A dysmorphic female born with partial trisomy of the proximal segment of the long arm of chromosome 14 had 47 chromosomes. The extra one was acrocentric, smaller than the D group, and bigger than the G-chromosome group. By GTG banding it was identified as a deleted chromosome 14, the karyotype being 47,XX,+del 14(q24). Chromosome analysis of the parents was normal.     

"Cri-du-chat" syndrome in a patient born to a mother with a paracentric inversion of chromosome 5q

We report the case of a female child presented at birth with hypotonia, growth retardation and respiratory distress. Chromosome study from peripheral blood showed a 46,XX,del(5)(p14pter) karyotype. Parental chromosome studies revealed that the mother carried an apparently balanced paracentric inversion of long arms of one chromosome 5, giving the karyotype 46,XX,inv(5)(q12q32), whereas paternal karyotype was normal. The maternal abnormality was confirmed by fluorescence in situ hybridization (FISH) and was not present in the daughter's metaphases. Microsatellite analysis in the proposita and her parents permitted us to conclude that the deleted chromosome 5 was paternal in origin, as usually described. Therefore, as might have been expected, maternal paracentric inversion of chromosome 5q and "cri-du-chat syndrome" presented by the daughter were not related.     

Comparative chromosome analysis of nine squamous cell carcinoma lines from tumors of the head and neck

The chromosomes of nine cell lines from human squamous cell carcinomas of the head and neck were examined. Five of the lines were derived from patients who had received chemotherapy and radiotherapy, one line was derived from a lymph-node metastasis from one of these patients, and three lines were from patients who had not been treated. The cell lines from the untreated patients had lower modal chromosome numbers than those of all but one of the lines from the treated patients. A structural abnormality involving chromosome 1 with breakpoints located in p13, p22, q21, or q32 was found in all cases. The NRAS and SK oncogenes are located in or near some of these segments. In four of the cell lines the nucleolus organizer regions (NORs) were 10-30 times larger than the NORs of the normal human chromosomes. Double minutes (DM) and homogeneously staining regions (HSR) were found in four of the lines.     

Unbalanced translocation t(15;22) in "severe" Prader-Willi syndrome

A 13-year-old girl with an unbalanced karyotype 45,XX,-15,der(22)t(15;22)(q13;q13.3) de novo had Prader-Willi syndrome (PWS), (score 13.5), but with features of mental and physical retardation more severe than usually seen in PWS. The clinical diagnosis of PWS was confirmed by methylation analysis that showed absence of the paternal band. With GTG banding, the cytogenetic breakpoint on chromosome 15q13, with 15q14 intact, encompassed the PWS region, while the breakpoint on 22q was terminal. Investigations with FISH utilised ten different probes/combinations, namely SNRPN/PML, TUPLE1/22q13.3, TUPLE/ARSA, GABRB3, three YAC clones and one cosmid for specific regions within chromosome 15q, painting probes for the long arm of chromosomes 15 and 22 and a pantelomere probe. Deletion of SNRPN,TYAC 9 (at 15q11-12), TYAC19 (at 15q13) and GABRB3 (within the PWS locus), was evident on the derivative (22) chromosome, while TYAC10 (at 15q22), cos15-5 (at 15q22) and PML (15q22) were not deleted. On the der(22), 22q13.3 and ARSA were not deleted, but the most distal non specific pantelomeric probe was deleted. Thus, the severe phenotype could be attributable to deletion on chromosome 15q extending beyond q13 to q14, (further than the usual chromosome 15q deletion (q11-13) in PWS), or be related to loss of the very terminal 22q region (from ARSA to the pantelomere) or be due to genetic factors elsewhere in the genome.     

An unexpected recurrence of Angelman syndrome suggestive of maternal germ-line mosaicism of del(15)(q11q13) in a Finnish family

Angelman syndrome is a neuro-developmental disorder caused by genetic abnormalities affecting the maternal gene expression in the chromosome region 15q11-q13. In a study group of 45 Finnish Angelman patients, a recurrence of a del(15)(q11q13) was detected in one family. The mother's chromosomes 15 were structurally normal, whereas the patients and their unaffected brother shared an identical maternally derived haplotype outside the deletion region. These findings are suggestive of maternal germ-line mosaicism of del(15)(q11q13).     

FISH studies on the telomeric regions of the T-cell acute lymphoblastic leukemia cell line CCRF-CEM

So far, the problem of an influence of translocations on the telomeres of the involved chromosomes has not been addressed yet in human cells. Therefore, the telomeres of a karyotypically rather well characterized T-cell acute lymphoblastic leukemia (T-ALL) cell line (CCRF-CEM) with several marker chromosomes were examined using peptide nucleic acid (PNA) telomere FISH probes to compare the telomere length of these markers with that of the chromosome arms of their origin. In addition, chromosome libraries, centromeric probes, and subtelomeric DNA probes were used to further define the marker chromosomes. Two markers could be newly defined and a concise karyotype of the cell line could be obtained by these detailed examinations: 42-47,X,-X,del(5) (q35?),t(5;15)(q14;q13.2),t(8;9)(p11;p24),del(9)(:p13-->qter)/inv(9)(pter-->p12::q21-->p12::q21-->qter),+13,+20,+der(22)(p+ [HSR?])[cp]. The relative telomere length of all chromosomes showed considerable interchromosomal, intercellular, and inter-passage variation. However, it could be shown, that in four different passages of the examined cell line the observed differences between relative telomere lengths of the markers and the chromosomes of their origin, with two exceptions (short arms of del/inv9 and der22), were not significant. On the other hand, because of its mentioned variability, telomere length alone is not sufficient to reliably define the derivation of markers.     

Increased sister chromatid exchanges in human cell lines characterized by monosomy X or structural abnormalities of the X chromosome

Summary In the present investigation we test the hypothesis that deficiencies in the X chromosome affect sister chromatid exchange (SCE) frequencies in human fibroblast cell lines. Our results show increased mean SCE frequencies in cell lines with abnormalities of the X chromosome: 45,X; 46,X,del(X) (q13), 46,X,del(X)(p11), and 46,X,i(Xq); control cell lines were 46,XX. In only one abnormal line [46,X,del(X)(p11)] was the increase not significant after correcting for multiple comparisons. If SCE formation is replication-dependent, the increased SCE frequencies might merely reflect the prolonged cell cycle we reported previously in cell lines with X chromosome abnormalities (Simpson and LeBeau 1981). Other explanations for differences between cell lines are possible, e.g., that deleted loci on the X chromosome affect cellular uptake of bromodeoxyuridine (BrDU). However, specific mechanisms were not explored directly.     

Dicentric chromosomes and 20q11.2 amplification in MDS/AML with apparent monosomy 20

FISH analysis of 41 previously karyotyped cases of MDS and AML with apparent monosomy of chromosome 20 revealed a variety of dicentric abnormalities involving chromosome 20. These usually, but not always, involved a breakpoint in the long arm of chromosome 20 and loss of the common deleted region at 20q12. Not one case of true monosomy 20 was confirmed. We found evidence for dicentric chromosome formation in 21 of 24 unbalanced translocations containing chromosome 20 and that were studied in more detail. Subsequent loss of one of the centromeres had occurred in eight of these 24 cases, and was more frequent than centromere inactivation as a means of resolving the inherent instability of a dicentric chromosome. In the three cases with dicentric chromosomes from which proximal 20q had been excised along with the 20 centromere, the excised segment was retained, and in two of these it was amplified. Proximal 20q was clearly retained in all but three cases, and present in three or more copies in 17 of 41 cases. The retention and amplification of proximal 20q provides support for the hypothesis that there is an oncogene located in this region of 20q that is activated in cases of MDS/AML with del(20q). Apparent monosomy 20 in MDS/AML should be treated as evidence of unidentified chromosome 20 abnormalities, and familiarity with the typical G-banded morphology of these derivatives can help with their identification. The reported incidence of dicentric chromosomes is clearly an under-estimate but is increasing in myeloid disorders as more cases are studied with methods allowing their detection.     

Two sisters with a distal deletion at the Xq26/Xq27 interface: DNA studies indicate that the gene locus for factor IX is present

Summary Two sisters with premature menopause and a small deletion of the long arm of one of their X chromosomes [del (X)(pterq26.3:)] were investigated with polymorphic DNA probes near the breakpoint. The deleted chromosome retained the factor IX (F9) locus and the loci DXS51 (52A) and DXS100 (pX45h), which are proximal to F9. However, the factor VIII (F8) locus was not present, nor were two loci tightly linked to this locus, DXS52 (St14) and DXS15 (DX13) This deletion refines the location of the F9 locus to Xq26 or to the interface Xq26/Xq27, thus placing it more proximally than has been previously reported. The DNA obtained from these patients should be valuable in the mapping of future probes derived from this region of the X chromosome.     

Sister chromatid exchanges and chromosome aberrations in fibroblasts from patients with retinoblastoma

Summary The frequencies of sister chromatid exchanges (SCEs) and chromosome breaks were investigated in five diploid fibroblast strains derived from three patients with deletion 13[del(13)] retinoblastoma, one patient with a hereditary form of retinoblastoma, and one trisomy 13. The fibroblasts with del(13)(q14q22) showed slightly increased SCEs (at a P level of 5–10%), but the others, including del(13)(q12q14), the hereditary form of retinoblastoma, and trisomy 13, did not have increased SCEs as compared to normal controls. No increase in chromosome breaks was found in these fibroblasts. The results suggest that retinoblastoma is not associated with spontaneous increased chromosomal instability.     

18q部分单体患儿的细胞和分子遗传学研究

临床发现1例智力低下伴轻度发育迟缓的女性患儿,对患儿进行G显带高分辨染色体核型分析, 发现18q21→qter缺失, 经多色荧光原位杂交和双色荧光原位杂交证实, 确定其核型为46,XX,del(18)(pter→q21:),ish del(18)(D18Z1+, qter-)。用DNA多态性方法分析, 该患儿从18q22.1至18qter区域内至少有8.7 Mbp丢失, 有MBP基因和GALNR基因缺失。缺失的18号染色体源自父亲。患者的智力低下和生长发育迟缓是18q21→qter缺失的结果, 或许与MBP基因和GALNR基因的缺失有关。     

Molecular analysis of a case of meiotic recombination leading to cri-du-chat syndrome

This paper describes a molecular investigation of a woman with an apparent large pericentric inversion of chromosome 5, inv(5)(p14;q35), and one normal chromosome 5 and her child, who was born with cri-du-chat syndrome. The four chromosome 5 homologs from the proband and his mother were isolated in somatic cell hybrids, and their haplotypes were determined at nine loci polymorphic for restriction enzyme sites. The deleted chromosome in the proband was shown to carry alleles from both maternal homologs, verifying molecularly that a meiotic recombination event in the mother gave rise to her son's deleted chromosome 5. The single crossover was presumably near the centromere.     

A complex mosaicism 45,X/46,X,del(Xq)/46,X,idic(Xq) in a patient with secondary amenorrhea

A complex mosaicism involving the X chromosome was found in a 35-year-old female affected by secondary amenorrhea and short stature. Her karyotype was: 45,X[20]/46,X,del(X)(pter-->q26::qter)[15]/46,X,idic(X)(pter-->q26::q26-->pter)[9]. No cell contained both abnormal X chromosomes. This observation would suggest a possible mechanism underlying the formation of isodicentric chromosomes.