Molecular study of parental origin of extra chromosome 21 in regular and de novo translocation trisomies.

The parental origin of the extra chromosome 21 (or extra 21q) was determined in seven informative families with a Down syndrome (DS) child by using molecular polymorphisms. Five DS patients had regular trisomy, one a de novo 14/21 translocation and another a de novo 21/21 translocation or isochromosome 21q. In four families with regular trisomy, the extra chromosome was of maternal origin, and in one family it was paternally derived. In the two families with a de novo aberration, both the 14/21 translocation and 21/21 rearrangement originated during maternal meiosis. For a better evaluation of the stage of meiotic error and the occurrence of crossovers between nondisjoined chromosomes, the regional map position of four of the nine informative DNA markers, used in this study, was refined, leading to useful localizations in both centromeric and distal regions. Recombination events were found in two families with regular trisomy, one occurring between chromosomes 21 that failed to disjoin at maternal meiosis I, the other prior to a paternal meiosis II nondisjunction.

Potential gene sequence isolation and regional mapping in human chromosome 21.

The transcription start sites of many genes are associated with CpG-rich DNA regions (CpG islands) containing clusters of rare cutting, methylation-sensitive restriction enzyme sites [Bird, 1986]. To detect gene sequences from human chromosome 21, we have screened cloned DNA fragments from a chromosome 21-specific cosmid library for the presence of such restriction sites. Several DNA fragments containing rare cutter sites, including Sac II, were isolated and five of them partially characterized. The average insert size of the fragments was 38.4 kb. By using a panel of somatic cell hybrids, one insert was assigned to the distal part of region 21q21, three fragments to the region 21q22.1, and one sequence to the segment 21q22.2-22.3. Restriction mapping showed clusters of rare cutter sites in at least three of the cloned fragments, suggesting the presence of CpG islands. These fragments are thus good candidates for carriers of coding sequences.

Pericentric inversion of chromosome 9: prevalence in 300 Down syndrome families and molecular studies of nondisjunction.

The incidence of Down syndrome (DS) families where one of the parents is an heterozygous carrier of pericentric inversion of the heterochromatic region of chromosome 9-inv(9) (qh) - was determined in 3 independent groups of 100 families each. The total number of 17 such families found in the sample is significantly greater than the expected number of 5.73 for a sample of non-DS families of equal size. Consequently, the statistical association of the presence of inv (9) (qh) in one parent with the birth of a DS offspring, and the correlative 3-fold increased risk of a DS child for such families, seem to be demonstrated. A study of the origin of nondisjunction, using restriction fragment length polymorphism (RFLP) segregation analysis with a sufficient number of chromosome 21 specific probes, has provided complete information in 7 of 8 available families. Although the statistical interpretation of the results is not straightforward, due to the small size of the sample, the observed data do not contradict the assumption that the presence of inv (9) (qh) in a parent increases, by a factor of about 3, the chance that the offspring will inherit an extra chromosome 21 from that parent. Nevertheless, gathering further data appears desirable because stronger evidence would have relevance both for clinical implications and for the understanding of the function of heterochromatin, particularly with respect to meiotic and mitotic processes.

Coincident maternal meiotic nondisjunction of chromosomes X and 21 without evidence of autosomal asynapsis.

A family in which the proband showed phenotypic signs of both the Turner and Down syndromes was studied cytogenetically and with restriction fragment length polymorphisms. The proband's karyotype was 46,X,+21, showing double aneuploidy without any signs of mosaicism. The single X and one chromosome 21 were of paternal origin while two chromosome 21 were of maternal origin. The nondisjunction of chromosome 21 took place in maternal meiosis II. If it is assumed that the absence of mosaicism renders postzygotic mitotic loss of the X chromosome unlikely, then the X chromosome would have been lost in maternal meiosis I or II. Recombination had occurred between the nondisjoined chromosomes 21. We conclude that double nondisjunction took place in one patient and that asynapsis was not a prerequisite for the autosomal nondisjunction.

Use of restriction fragment length polymorphic probes in the analysis of Down's syndrome trisomy.

Restriction fragment length polymorphic probes are being used more frequently in the molecular analysis of Down's syndrome and in the origin of nondisjunction in the syndrome. The type of information gained from RFLPs overlaps but differs from the information from cytogenetic heteromorphisms. From the allele frequencies of commonly available probes we have derived the expected frequencies of all matings in the population. Each mating has been defined and partitioned to show the genotypes and phenotypes expected, with numerical values based on studies with heteromorphisms. From this we show how the various phenotypes can be used to calculate the origin of nondisjunctions and their expected frequencies. Further, an alternative method is outlined for mapping the distance between a probe and its centromere based on the distortion, caused by crossing-over, of the expected 1st to 2nd division nondisjunction ratio. Finally, we discuss prospects for various uses of probes in the analysis of Down's syndrome.

On the origin of recurrent trisomy 21: determination using chromosomal and DNA polymorphisms.

A family is described in which two cases of trisomy 21 occurred in, respectively, a newborn infant and a prenatally diagnosed fetus. Using fluorescent chromosomal polymorphisms, it was established that in both cases the extra chromosome resulted from a first meiotic division error in the mother and that the father contributed the same centromeric region to both children. RFLP-associated probes were used to examine the genetic content of the chromosomes. It was noted that the polymorphism patterns of the chromosomes 21 which both children inherited from their parents were identical for three, but not identical for one of the probes studied. This difference must be the result of recombination. This result is discussed in relation to the suggestion that the increased recurrence rate in mothers with a trisomic child could be due to a reduced recombination rate.

The X chromosome shows less genetic variation at restriction sites than the autosomes.

Using a standard technique, 122 single-copy probes were screened for their ability to detect restriction fragment length polymorphisms (RFLPs) in the human genome. The use of a standardized RFLP screening enables the introduction of statistical methods in the analysis of differences in RFLP content between chromosomes and enzymes. RFLPs were detected from panels containing at least 17 unrelated chromosomes, digested with TaqI, MspI, BglII, HindIII, EcoRI, and PstI. Forty autosomal probes, representing a sample of 2,710 base pairs (bp) per haploid genome, were tested, and 24 RFLPs were found. With 82 X-chromosomal probes, 17 RFLPs were found in 6,228 bp per haploid genome. The frequency of X-chromosomal RFLPs is three times less than that of the autosomes; this difference is highly significant (P = less than .001). The frequency of RFLPs revealed by various restriction enzymes and the possibility that the X chromosome is a "low mutation" niche in the human genome are discussed.

Construction and analysis of an EMBL-3 phage library containing partially digested human chromosome 21-specific DNA inserts (15-20 kb).

In the mouse-human hybrid cell line SCC 16-5, chromosome 21 is the only human chromosome present. Fractions highly enriched for this chromosome were obtained by applying the chromosome velocity sedimentation technique to this cell line. DNA prepared from these chromosomal fractions was partially digested with Mbo I, size fractionated on an NaCl gradient, and cloned in the EMBL-3 phage vector. The phage library thus prepared was highly enriched for human chromosome 21-specific recombinant DNA sequences 15-20 kb long. Of the approximately 21,000 phage clones obtained, at least 99% were recombinant. Following phage plaque filter hybridization and Southern blotting, it was found that half of the recombinants were positive for human repetitive DNA. Almost all phages harbored highly or middle repetitive human or mouse DNA sequences owing to the large size of the recombinant inserts. In this library, the human chromosome 21 is represented approximately four times. All human recombinants studied thus far contained DNA inserts originating from chromosome 21 only. The employed cloning strategy is discussed with regard to utility, purity, quality, and completeness of chromosome-specific recombinant DNA libraries.

Bifunctionality and polarized infidelity at the hisB locus of Aspergillus nidulans.

The histidine (hisB) locus of Aspergillus nidulans is unusual in two ways. Firstly, it is bifunctional; besides coding for imidazole glycerol phosphate (IGP) dehydrase, it is required for the production of ascospores (fertility). It appears, therefore, to be partly homologous to the hisB locus of Salmonella typhimurium, which codes for IGP dehydrase and histidinol phosphate phosphatase. Secondly, during meiosis it is often inaccurately transmitted to the progeny (infidelity). This phenomenon may be akin to the aberrant recombination events which cause Bar reversion in Drosophila, "selfing" in Salmonella and Neurospora, and gene fusions of the haemoglobin lepore type. A molecular model is proposed to account for the results.