Copy number variation burden does not predict severity of neurodevelopmental phenotype in children with a sex chromosome trisomy

Sex chromosome trisomies (SCTs) (XXX, XXY, and XYY karyotypes) are associated with an elevated risk of neurodevelopmental disorders. The range of severity of the phenotype is substantial. We considered whether this variable outcome was related to the presence of copy number variants (CNVs)—stretches of duplicated or deleted DNA. A sample of 125 children with an SCT were compared with 181 children of normal karyotype who had been given the same assessments. First, we compared the groups on measures of overall CNV burden: number of CNVs, total span of CNVs, and likely functional impact (probability of loss‐of‐function intolerance, pLI, summed over CNVs). Differences between groups were small relative to within‐group variance and not statistically significant on overall test. Next, we considered whether a measure of general neurodevelopmental impairment was predicted by pLI summed score, SCT versus comparison group, or the interaction between them. There was a substantial effect of SCT/comparison status but the pLI score was not predictive of outcomes in either group. We conclude that variable presence of CNVs is not a likely explanation for the wide phenotypic variation in children with SCTs. We discuss methodological challenges of testing whether CNVs are implicated in causing neurodevelopmental problems.     

Isolation of DNA from the centromere of human chromosome 7 by microdissection

Centromeres remain the least characterized regions of human chromosomes because they have a very high content of repetitive DNA. Here, we describe a micro-dissection library from the centromeric region of human chromosome 7 and its use for generating sequence tagged sites (STSs). The library contains about 1500 clones with an average insert size of 150 bp and only about 15% of the clones harbour repetitive human DNA. Seven clones hybridizing to alphoid DNA were found to correspond to a fragment of the D7Z2 alphoid array on chromosome 7, thus confirming the origin of the library. A number of clones not containing known repetitive DNA were used to generate STSs that identified yeast artificial chromosomes (YACs) and in turn allowed the STSs to be placed on the physical map. One STS is located between the two Genethon genetic markers closest to the centromere on the q side. Another STS was located 3–4 cM away in 7q11.2, while a third identified YACs containing both low-copy and alphoid sequences that are not yet mapped but are clearly centromeric. The library therefore comprises a collection of sequences from the centromeric region of chromosome 7 that can be used to generate STSs and to map the entire centromeric region.This revised version was published online in November 2005 with corrections to the Cover Date.     

The frequency of chromosome anomalies in human preimplantation embryos after in-vitro fertilization

Previous studies have reported chromosome aberrations in humanpre-embryos after in-vitro fertilization (IVF). Although thereason for these abnormalities is not clear, there is evidencethat they can arise during gametogenesis, fertilization or cleavage.The present study has examined further the incidence of chromosomeabnormalities in human pre-embryos after IVF, using oocytesrecovered from normal volunteer women and from women undergoinginfertility treatment in an embryo-replacement programme. Chromosomepreparations were performed for 75 pre-embryos. Of these 35(47%) gave at least one metaphase in which analysis was possible.The overall incidence of abnormal pre-embryos was 40% (14/35).The absolute frequency of aberrations was 9% for trisomies,3% for polyploidies, 26% for structural anomalies and 3% forhypodiploidies. Five pre-embryos were found to be mosaics, threeof which had each one trisomic metaphase. In five of the pre-embryosmultiple anomalies were found. In 13 of the 14 abnormal pre-embryosthe aberrations were found in only one metaphase. The presentstudy demonstrates that trisomic mosaicism may not be a rareevent in human pre-embryos. Further evidence is provided thatmitotic non-disjunction is important for the production of aberrationsin human pre-embryos     

Preaxial acrofacial dysostosis (Nager syndrome) associated with an inherited and apparently balanced X;9 translocation: Prenatal and postnatal late replication studies

We report on an infant with preaxial acrofacial dysostosis (Nager syndrome) who was diagnosed prenatally as having an apparently balanced X/autosome translocation [46,X,t(X;9)(p22.1;q32)mat] inherited from a previously diagnosed mosaic translocation carrier mother [46,XX/46,X,t(X;9)(p22.1;q32)]. Replication studies on amniocytes showed the normal X chromosome to be late replicating while the same studies repeated on the infant's lymphocytes showed the translocated X chromosome to be late replicating in most cells. Late replication studies of the mother's lymphocytes demonstrated that the normal X chromosome was late replicating in most cells. The presence of Nager syndrome in this infant may be the result of critical break-points and/or position effects on chromosome 9, inducing expression of a gene responsible for the syndrome. © 1993 Wiley-Liss, Inc.     

Nature of telomere dimers and chromosome looping in human spermatozoa

Specific and well-organized chromosome architecture in human sperm cells is supported by the prominent interactions between centromeres and between telomeres. The telomere-telomere interactions result in telomere dimers that are positioned at the nuclear periphery. It is unknown whether composition of sperm telomere dimers is random or specific. We now report that telomere dimers result from specific interactions between the two ends of each chromosome. FISH using pairs of subtelomeric DNA probes that correspond to the small and long arms of seven human chromosomes demonstrates that subtelomeres of one chromosome are brought together. Statistical analysis confirmed that telomere associations could not result from the random proximity of DNA sequences. Therefore, chromosomes in human sperm nuclei adopt a looped conformation. This higher-order chromosome structure is most likely required for chromosome withdrawal/decondensation during the early fertilization events leading to zygote formation. These individuals contributed equally to the work     

Genetic damage detected in CD-1 mouse pups exposed perinatally to 3'-azido-3'-deoxythymidine or dideoxyinosine via maternal dosing, nursing, and direct gavage: II. Effects of the individual agents compared to combination treatment

We previously reported extraordinary increases in micronucleated erythrocytes in CD-1 mouse pups exposed to 3'-azido-3'-deoxythymidine (AZT) and dideoxyinosine (ddI; 50/250, 75/375, 150/750 mg/kg/day AZT/ddI) by gavage throughout gestation and lactation, followed by direct pup dosing beginning postnatal day (PND) 4 (Bishop et al. [2004]: Environ Mol Mutagen 43: 3-9). That study was conducted to explore the potential for genetic damage in newborns exposed perinatally to antiretrovirals in order to reduce maternal-infant transmission of HIV-1. Because dramatic increases in frequencies of micronucleated erythrocytes were seen in exposed pups, additional studies were conducted to clarify the relative contribution of each drug to the observed damage. Pregnant CD-1 mice were administered AZT (50, 75, 150 mg/kg/day) or ddI (250, 375, 750 mg/kg/day) by gavage twice daily in equal fractions beginning prior to mating and continuing throughout gestation and lactation. Direct pup dosing (same regimens) began on PND 4. Peripheral blood erythrocytes of male pups were screened for micronuclei on PNDs 1, 4, 8, and 21. Significant increases in micronucleated erythrocytes were observed in pups and dams exposed to AZT at all doses and sampling times. The highest micronucleus levels were observed in pups on PND 8 after the initiation of direct dosing. In contrast, effects seen in pups and dams treated with ddI were minimal. These results demonstrate that AZT, a component of many anti-HIV combination therapies, induces chromosomal damage in perinatally exposed neonatal mice. Comparison of micronucleated cell frequencies induced by AZT alone or in combination with ddI suggests that ddI potentiates AZT-induced chromosomal damage following direct exposure.     

FISH deletion mapping defines a single location for the Y chromosome stature gene, GCY

At least 1 in 1000 males lacks part of the long arm of the Y chromosome. This chromosomal aberration is often associated with short stature and infertility. Deletion mapping and genotype-phenotype analysis have previously defined two non-overlapping critical regions for growth controlling gene(s), GCY(s), on the euchromatic portion of the Y chromosome long arm. These initial mapping assignments were based on the analysis of patients carrying a pure 46,XYq- karyotype as defined by classical cytogenetic karyotyping. Four genes have been assigned to the distal one of the two critical regions. To determine whether one or both of these two critical regions harbours GCY and whether one of the four genes assigned to the distal region is involved in determination of stature, nine adult patients with Yq chromosomal abnormalities were studied in detail. By PCR and FISH analysis, we showed that all patients with a previously defined pure 46,XYq- karyotype are actually mosaics with cells containing an idic(Y) or ring(Y) chromosome in association with 45,X0 cells. This leads us to conclude that (1) FISH is an absolute prerequisite for the correct identification of Y chromosomal rearrangements and (2) only patients with interstitial Y deletions are reliable predictors for the physical location of stature gene(s) on Yq. Our molecular analyses of chromosomes from patients with interstitial Yq deletions finally establishes the proximal interval between markers DYZ3 and DYS11 as the only GCY critical interval. No functional gene has so far been identified in this region adjacent to the centromere.     

Tandem chromosome fusions in karyotypic evolution of Muntiacus: evidence from M. feae and M. gongshanensis

The muntjacs (Muntiacus, Cervidae) are famous for their rapid and radical karyotypic diversification via repeated tandem chromosome fusions, constituting a paradigm for the studies of karyotypic evolution. Of the five muntjac species with defined karyotypes, three species (i.e. Muntiacus reevesi, 2n = 46; M. m. vaginalis, 2n = 6/7; and M. crinifrons, 2n = 8/9) have so far been investigated by a combined approach of comparative chromosome banding, chromosome painting and BAC mapping. The results demonstrated that extensive centromere–telomere fusions and a few centric fusions are the chromosomal mechanisms underlying the karyotypic evolution of muntjacs. Here we have applied the same approach to two additional muntjac species with less well-characterized karyotypes, M. feae (2n = 14♂) and M. gongshanensis (2n = 8♀). High-resolution G-banded karyotypes for M. feae and M. gongshanensis are provided. The integrated analysis of hybridization results led to the establishment of a high-resolution comparative map between M. reevesi, M. feae, and M. gongshanensis, proving that all tandem fusions underpinning the karyotypic evolution of these two muntjac species are also centromere–telomere fusions. Furthermore, the results have improved our understanding of the karyotypic relationships of extant muntjac species and provided compelling cytogenetic evidence that supports the view that M. crinifrons, M. feae, and M. gongshanensis should each be treated as a distinct species.     

The kinase haspin is required for mitotic histone H3 Thr 3 phosphorylation and normal metaphase chromosome alignment

Post-translational modifications of conserved N-terminal tail residues in histones regulate many aspects of chromosome activity. Thr 3 of histone H3 is highly conserved, but the significance of its phosphorylation is unclear, and the identity of the corresponding kinase unknown. Immunostaining with phospho-specific antibodies in mammalian cells reveals mitotic phosphorylation of H3 Thr 3 in prophase and its dephosphorylation during anaphase. Furthermore we find that haspin, a member of a distinctive group of protein kinases present in diverse eukaryotes, phosphorylates H3 at Thr 3 in vitro. Importantly, depletion of haspin by RNA interference reveals that this kinase is required for H3 Thr 3 phosphorylation in mitotic cells. In addition to its chromosomal association, haspin is found at the centrosomes and spindle during mitosis. Haspin RNA interference causes misalignment of metaphase chromosomes, and overexpression delays progression through early mitosis. This work reveals a new kinase involved in composing the histone code and adds haspin to the select group of kinases that integrate regulation of chromosome and spindle function during mitosis and meiosis.     

The origin of human chromosome 2 analyzed by comparative chromosome mapping with a DNA microlibrary

Fluorescencein situ hybridization (FISH) of microlibraries established from distinct chromosome subregions can test the evolutionary conservation of chromosome bands as well as chromosomal rearrangements that occurred during primate evolution and will help to clarify phylogenetic relationships. We used a DNA library established by microdissection and microcloning from the entire long arm of human chromosome 2 for fluorescencein situ hybridization and comparative mapping of the chromosomes of human, great apes (Pan troglodytes, Pan paniscus, Gorilla gorilla, Pongo pygmaeus) and Old World monkeys (Macaca fuscata andCercopithecus aethiops). Inversions were found in the pericentric region of the primate chromosome 2p homologs in great apes, and the hybridization pattern demonstrates the known phylogenetically derived telomere fusion in the line that leads to human chromosome 2. The hybridization of the 2q microlibrary to chromosomes of Old World monkeys gave a different pattern from that in the gorilla and the orang-utan, but a pattern similar to that of chimpanzees. This suggests convergence of chromosomal rearrangements in different phylogenetic lines.