Familial DiGeorge syndrome and associated partial monosomy of chromosome 22

Summary Partial monosomy of 22q due to an unbalanced 4;22 translocation was seen in a 2-month-old male with Type I truncus arterious, dysmorphic features, and T-cell abnormalities. The family history revealed a previous sib with Type I truncus arteriosus, thymic aplasia, and parathyroid hypoplasia noted on postmortem examination, consistent with DiGeorge syndrome. Evaluation of the asymptomatic mother of these two patients revealed partial T-cell deficiency and the same unbalanced translocation with deletion of proximal 22qll. These findings provide further evidence that some cases of complete or partial DiGeorge syndrome are associated with monosomy of the proximal long arm of chromosome 22, and they may explain many, if not all, familial cases of the syndrome.Supported in part by National Foundation-March of Dimes Grant No. 2-161/C-331. Funds from the Texas Department of Health through PL94-278 National Genetic Diseases Act, from the Robert J. Kleberg, Jr. Center for Human Genetics, and USPHS Grant No. RR-05425.     

In situ hybridization and translocation breakpoint mapping. II. Two unusual t(21;22) translocations

Using a combination of banding techniques, we examined two atypical 21;22 translocations, 46,XX or XY,t(21;22)(p11;q11). In situ chromosomal hybridization of a probe for the constant region of the lambda light chain locus demonstrated that the 22q11 breakpoints of both rearrangements were proximal to the C lambda gene cluster. These studies permitted us to distinguish the 22q11 breakpoints of these translocations from the breakpoint of the 22q--chromosome of chronic myelogenous leukemia.     

Partial monosomy 22 as result of an X/22 translocation in a newborn with DiGeorge syndrome

The case of a neonate with clinical symptoms of DiGeorge syndrome is reported. During pregnancy the measurements by ultrasonography revealed already a significant growth retardation of the fetus, for the first time obvious in the 20th week. The child died immediately after birth. A de novo translocation X/22 was observed with the translocation chromosome being late replicating in all mitoses analysed. The own observation is discussed regarding other cases with DiGeorge syndrome and taking the differential diagnoses into account. count.     

Recurrent t(11;22) breakpoint mapping by chromosome flow sorting and spot-blot hybridization

Summary The breakpoint of the recurrent t(11;22) translocation, one of the most frequent chromosome anomalies encountered in human population, always involves bands 11q23.2 and 22q11.2. The involvement of the C locus of the immunoglobulin gene cluster on chromosome 22 has been suggested: however, in situ hybridization experiments have yielded conflicting results. In order to solve these discrepancies by another approach, we have used bivariate flow sorting to separate the chromosomes of interest and to map the specific breakpoints by direct spot-blot hybridization with the gene-specific radiolabelled DNA probes, Alu, V, ets. The results showed unambiguously that in the t(11;22) patient analysed, a set of C and V genes was translocated to the der(11) chromosome. Since V genes are situated proximally to C genes, we demonstrate that, in the case studied here, the chromosome 22 breakpoint is not located within or even immediately close to the C region.Presented at the 7th International Congress of Human Genetics, Berlin, September 22–26, 1986     

Routine diagnosis of DiGeorge syndrome by fluorescent in situ hybridization

In a series of ten patients affected by DiGeorge syndrome, we screened, by high resolution banding and fluorescent in situ hybridization of a cosmid probe, for microdeletions associated with this syndrome. In the ten patients, a microdeletion was demonstrated by in situ hybridization, but suspected only in two patients by high resolution banding.     

Hypomagensemia with secondary hypocalcemia in a female with balanced X;9 translocation: mapping of the Xp22 chromosome breakpoint

Magnesium-dependent hypocalcaemia (HSH), a rare inherited disease, is caused by selective disorders of magnesium absorption. Both X-linked and autosomal recessive modes of inheritance have been reported for HSH; this suggests a genetically heterogeneous condition. A balanced de novo t(X;9)(p22;q12) translocation has been reported in a female manifesting hypomagnesemia with secondary hypocalcemia. In a lymphoblastoid cell line, derived from this patient, the normal X chromosome is preferentially inactivated, suggesting that the patient's phenotype is caused by disruption of an HSH gene in Xp22. In an attempt to define more precisely the position of the X breakpoint, we have constructed a hybrid cell line retaining the der(X)(Xqter-Xp22.2::9q12-9qter) in the absence of the der(9) and the normal X chromosome. Southern blot analysis of this hybrid and in situ hybridization on metaphase chromosomes have localized the breakpoint between DXS16 and the cluster (DXS207, DXS43), in Xp22.2. Thus, if a gene involved in HSH resides at or near the translocation breakpoint, our findings should greatly facilitate its isolation.     

Lowe oculocerebrorenal syndrome in a female with a balanced X;20 translocation: mapping of the X chromosome breakpoint.

A Hispanic girl with Lowe oculocerebrorenal syndrome (OCRL), an X-linked recessive condition characterized by cataracts, glaucoma, mental retardation, and proteinuria, is reported. A balanced X;20 chromosomal translocation with the X chromosome breakpoint at q26.1 was found with high-resolution trypsin-Giemsa banding. Somatic cell hybridization was used to separate the X chromosome derivative and the chromosome 20 derivative in order to position, with respect to the translocation breakpoint, several DNA loci that are linked to the Lowe syndrome locus (Xq24-q26). DXS10 and DXS53 were found to be distal to the breakpoint, whereas DXS37 and DXS42 were located proximal to it. These studies suggest that the OCRL locus lies in the region between these probes. The translocation chromosome originated from an unaffected male without a visible translocation, indicating that the most likely cause of OCRL in this patient is the de novo translocation that disrupted the OCRL locus.     

Molecular genetic study of the frequency of monosomy 22q11 in DiGeorge syndrome

It is well established that DiGeorge syndrome (DGS) may be associated with monosomy of 22q11-pter. More recently, DNA probes have been used to detect hemizygosity for this region in patients with no visible karyotypic abnormality. However, DGS has also been described in cases where the cytogenetic abnormality does not involve 22q11; for instance, four cases of 10p- have been reported. In this study we have prospectively analyzed patients, by using DNA markers from 22q11, to assess the frequency of 22q11 rearrangements in DGS. Twenty-one of 22 cases had demonstrable hemizygosity for 22q11. Cytogenetic analysis had identified interstitial deletion in 6 of 16 cases tested; in 6 other cases no karyotype was available. When these results are combined with those from our previous studies, 33 of 35 DGS patients had chromosome 22q11 deletions detectable by DNA probes.     

Deletion mapping by FISH with BACs in patients with partial monosomy 22q13

Patients with deletions in 22q13 are known to have phenotypic features that include normal or accelerated growth, large hands and feet, hypotonia, delayed psychomotor development and mild facial dysmorphism. To date, very few cases have been investigated by detailed molecular genetic analysis. We have analyzed three new patients with terminal deletions in 22q. We compared the cytogenetic observations with molecular data assessed by fluorescence in situ hybridization and an array of characterized bacterial artificial chromosome recombinants. The shortest region of deletion overlap is localized in 22q13.2–qter distal to the marker D22S94, but the telomeric repeat in the deleted chromosome appears to remain intact. When parental alleles were investigated in two of the three patients, the aberrant homolog was found to be of paternal origin in both cases. Although the deleted region still spans >20 cM, molecular analysis of additional patients and screening for new genes might help in elucidating candidate genes connected with the dysmorphisms defined by deletions of 22q13. Received: 14 August 1997 / Accepted: 27 January 1998     

Mapping of human chromosome 22 by in situ hybridization

The second smallest chromosome of the human karyotype, i.e., chromosome 22, is involved in many congenital or acquired structural aberrations. This variety can be taken advantage of to determine the exact linear order, from centromere to telomere, of cloned probes and chromosomal breakpoints. Eleven probes were localized with respect to breakpoints of 11 der(22) of independent cell lines using in situ hybridization on metaphasic spreads. The deduced order of the tested probes and that of the breakpoints are in complete agreement with the published genetic map and the karyotypic analysis, respectively. This approach enables a correlation of the genetic map with the chromosomal banding.     

In situ hybridization and chromosome banding in mammalian species

Chromosome banding is often required in conjunction with fluorescent in situ hybridization of labelled probes for chromosome painting, satellite DNA and low-copy sequences to allow identification of chromosomes and simultaneous probe localization. Here, we present a method that reveals both patterns with only one observation step. The band pattern is produced by restriction-enzyme digestion of chromosomes, followed by fixation with paraformaldehyde in PBS, a short chromosome denaturation step in hybridization solution, and then standard in situ hybridization, washing and detection protocols. Using a range of different mammalian species, chromosome-banding patterns were immediately recognizable, although synchronisation procedures normally required for high- resolution G-banding were not needed. Unlike other methods available, only one round of observation is required using a conventional fluorescence microscope, the method works without modification in many species, and in situ hybridization is not used for chromosome identification (allowing multiple targets and minimizing background). The banding pattern is probably generated by a combination of DNA dissolution and heterochromatin reorganisation after enzyme digestion, followed by paraformaldehyde fixation of the new chromatin structure and incomplete denaturation. The method is of widespread utility in comparative genomics and genome organization programmes.     

Isolation and mapping of cosmid markers on human chromosome 22, including one within the submicroscopically deleted region of DiGeorge syndrome

A genomic cosmid library was constructed from a Chinese hamster/human hybrid cell containing human intact chromosome 22 as its only human component. Of 1000 cosmids with inserts derived from human chromosome 22, 191 were tested for restriction fragment length polymorphisms (RFLPs). As a result, 64 clones detected RFLPs, including five variable number of tandem repeats systems. Of the remaining 127 cosmids, 111 detected a single copy sequence on human chromosome 22. Five somatic cell hybrids allowed us to assign all of the 64 polymorphic cosmids and 44 non-polymorphic cosmids to four different regions of human chromosome 22. In two patients with DiGeorge syndrome, one of the cosmids that had been sublocalized to 22pter-q11 detected hemizygosity. These 108 cosmid markers regionally assigned to human chromosome 22 should be useful for the construction of long-range physical maps and the identification of genetic alterations on the chromosome.     

Zaprionus tuberculatus: chromosome map and gene mapping by DNA in situ hybridization.

The genus Drosophila has long been used as a model of karyotype evolution, demonstrating change by paracentric inversion and occasional centric fusion of an ancestral karyotype of five rod-shaped and one "dot" chromosome. This study shows, by mapping D. melanogaster probes hybridized to polytene chromosomes of Zaprionus tuberculatus, that this ancestral pattern extends beyond the genus Drosophila. A formal polytene chromosome map of Z. tuberculatus is presented.     

Comparative mapping of bovine chromosome 13 by fluorescence in situ hybridization

We present chromosomal fluorescence in situ hybridization (FISH) results that both extend the HSA20/BTA13 comparative map as well as cytogenetically anchor two microsatellite markers. A bovine bacterial artificial chromosome (BAC) library was screened for conserved genes (type I loci) previously assigned to HSA10 or HSA20 and BTA13, and for microsatellites selected from two published BTA13 linkage maps. Clones from six out of nine comparative loci and both microsatellites were found represented in the BAC library. These BAC clones were used as probes in single colour FISH to determine the chromosome band position of each locus. As predicted by the human/bovine comparative map, all type I loci mapped to BTA13. Because single colour FISH analysis revealed that the loci were clustered within the distal half of BTA13, dual colour FISH was used to confirm the locus order. Established order was centromere- PRNP-(SODIL/AVP/OXT)-(BL42/GNAS1)-HCK-CSSM30 . The findings confirm the presence of a conserved HSA20 homologous synteny group on BTA13 distal of a HSA10 homologous segment.     

Partial monosomy 22 as the result of an unbalanced translocation 5:22 in a patient with cri-du-chat syndrome

Summary Properdin factor B phenotypes were determined in 7 Bantu speaking Negroid populations, 1 Indian, and 1 Colored population of South Africa in a total of 1258 individuals. In the Negroid populations allele frequencies were: Bf^F 0.655, Bf^S 0.282, Bf^RARE 0.063, in the Indian population: Bf^F 0.322, Bf^S 0.645, Bf^RARE 0.033, and in the Colored population: Bf^F 0.513, Bf^S 0.435, Bf^RARE 0.052. In addition, 2 so far unknown F alleles and possibly 1 new S allele were discovered.     

In situ hybridization with fluoresceinated DNA.

We have used fluorescein-11-dUTP in a nick-translation format to produce fluoresceinated human nucleic acid probes. After in situ hybridization of fluoresceinated DNAs to human metaphase chromosomes, the detection sensitivity was found to be 50-100 kb. The feasibility and the increase in detection sensitivity of microscopic imaging of in situ hybridized, fluoresceinated DNA with an integrating solid state camera for rapid cosmid mapping is illustrated. Combination of fluoresceinated DNA with biotinated and digoxigeninated DNAs allowed easy performance of triple fluorescence in situ hybridization. The potential of these techniques for DNA mapping, cytogenetics and biological dosimetry is briefly discussed.     

Der(22) syndrome and velo-cardio-facial syndrome/DiGeorge syndrome share a 1.5-Mb region of overlap on chromosome 22q11

Derivative 22 (der[22]) syndrome is a rare disorder associated with multiple congenital anomalies, including profound mental retardation, preauricular skin tags or pits, and conotruncal heart defects. It can occur in offspring of carriers of the constitutional t(11;22)(q23;q11) translocation, owing to a 3:1 meiotic malsegregation event resulting in partial trisomy of chromosomes 11 and 22. The trisomic region on chromosome 22 overlaps the region hemizygously deleted in another congenital anomaly disorder, velo-cardio-facial syndrome/DiGeorge syndrome (VCFS/DGS). Most patients with VCFS/DGS have a similar 3-Mb deletion, whereas some have a nested distal deletion endpoint resulting in a 1.5-Mb deletion, and a few rare patients have unique deletions. To define the interval on 22q11 containing the t(11;22) breakpoint, haplotype analysis and FISH mapping were performed for five patients with der(22) syndrome. Analysis of all the patients was consistent with 3:1 meiotic malsegregation in the t(11;22) carrier parent. FISH-mapping studies showed that the t(11;22) breakpoint occurred in the same interval as the 1.5-Mb distal deletion breakpoint for VCFS. The deletion breakpoint of one VCFS patient with an unbalanced t(18;22) translocation also occurred in the same region. Hamster-human somatic hybrid cell lines from a patient with der(22) syndrome and a patient with VCFS showed that the breakpoints occurred in an interval containing low-copy repeats, distal to RANBP1 and proximal to ZNF74. The presence of low-copy repetitive sequences may confer susceptibility to chromosome rearrangements. A 1.5-Mb region of overlap on 22q11 in both syndromes suggests the presence of dosage-dependent genes in this interval.