Population cytogenetics of folate-sensitive fragile sites

Summary The location and frequency of folate-sensitive common fragile sites (CFS) were studied in three populations: (1) 111 mentally retarded children of school age, (2) 240 mentally subnormal children attending special schools, and (3) 85 healthy children attending normal schools. Common fragile sites were found at 54 chromosomal bands including also the band Xq27, where gaps and breaks were detected in 4% of the children. The most frequent CFS were FRA3B (at 3p14.2), FRA6E (at 6q26), and FRA16D (at 16q23) seen in 73%, 65%, and 58% of the individuals totally studied. The frequencies of CFS-positive individuals did not differ among the populations. The variation found in the distribution of CFS among the populations was primarily assumed to be due to sampling differences and study method. The rate of expression of the most frequent CFS varied significantly among the individuals, seeming to suggest that polymorphism exists at those CFS.     

Population cytogenetics of aphidicolin-induced fragile sites

Summary Chromosome fragile sites are inducible by aphidicolin in cultured human lymphocytes. To assess the frequency and distribution of these common fragile sites in the general population, a cytogenetic survey was performed on 126 subjects, 59 males and 67 females, whose age ranged from 1 day to 72 years. Common fragile sites, induced by aphidicolin, were widespread and showed a remarkably different sensitivity among individuals; age influenced the overall frequency of fragile sites. Moreover, both age and sex seemed to modulate the expression of specific fragile sites. In our population, the most common fragile sites were: 3p14, 16q23, Xp22, 6q26, 1p31, 4q31, 1p22, 7q22, 2q33, 3q27, 2q31, 7q32, 14q24, 10q22, 5q31, 2q37, 6p21.     

Population cytogenetics of rare fragile sites in Japan

Summary A population cytogenetic study of three groups of rare fragile sites defined in Human Gene Mapping 8 (HGM8, Berger et al. 1985) has been conducted using peripheral blood lymphocytes of healthy Japanese subjects. We have examined 1,022 blood donors for folate-sensitive and bromodeoxyuridine (BrdU)-requiring, and 845 for distamycin A-inducible fragile sites. Out of 17 rare autosomal fragile sites defined in HGM8, the following six were identified in Japan; folate-sensitive fra(2)(q11), fra(11)(q13) and fra(11)(q23), distamycin A-inducible fra(16)(q22) and fra(17)(p12), and BrdU-requiring fra(10)(q25). The incidences of distamycin A-inducible fra(16)(q22) (1.42%) and fra(17)(p12) (3.08%) were considerably higher than those of the other sites in Japan. Furthermore, a folate-sensitive fra(17)(p12) and a distamycin A-inducible fra(8)(q24.1) have been newly found in the present study. Their incidences were 0.10% (1/1,022) and 0.71% (6/845), respectively. Since the expression of this fra(17)(p12) was induced by fluorodeoxyuridine, supressed by thymidine, but not induced by distamycin A, it can be classified as a folate-sensitive site. The expression of the new distamycin A-inducible fra(8)(q24.1) was also enhanced by treatment with Hoechst 33258, berenil and 4,6-diamidino-2-phenylindole (DAPI). This fragile site fulfils all four classical criteria suggested by Sutherland (1979) and also new criteria for a rare fragile site defined in HGM8 (Berger et al. 1985).     

Expression of folate-sensitive fragile sites in lymphocyte chromosomes

Summary The expression of folate-sensitive fragile sites (FS) was analyzed using MTX as a fragility inducer in seven normal subjects [four unrelated persons and three members of one family (father, mother, and son)]; a woman heterozygous for fra Xq27.3 with a 47,XXX karyotype; and her son, affected by the fra-X syndrome. The mean expression of chromosome lesions (CL) other than Xq27.3 was 70.1% (686CL in 978 metaphases), and the coincidence between CL and FS was 68.9%. We propose six new c-fra sites: bands 4q33 and 11q22 because they were found in two members of the same family; band 13q32 because it had a frequency of expression of 3% of metaphases; and bands 3p13, 8q21, and Xq21 because they were observed in four of the nine individuals studied.     

Expression of the autosomal folate-sensitive fragile sites in ten kindreds with Martin-Bell syndrome

The authors analyse the expression of all the folate-sensitive fra sites in a sample of 24 male patients with Martin-Bell syndrome (MBS) and their 12 mothers distributed in 10 kindreds. The cytogenetic results are compared with that of a control group, constituted by 8 unrelated normal subjects. Except for the fra Xq27, there was no autosomal folate-sensitive fra site significantly more expressed in patients with MBS than in the control group. On the basis of the present cytogenetic sample of about 6 500 R-banded mitoses, a list of all the in vitro folate-sensitive fra sites and their relative frequencies is given.     

Heritable fragile sites on human chromosomes. IX. Population cytogenetics and segregation analysis of the BrdU-requiring fragile site at 10q25

The frequencies of the bromodeoxyuridine (BrdU)-requiring fragile site at 10q25 in 1,026 unselected neonates, 901 patients referred for chromosome studies, and 87 institutionalized retardates were not significantly different from each other. The gene frequency was .013, and the population was in Hardy-Weinberg equilibrium. Segregation analysis confirmed that the fragile site followed codominant inheritance. This fragile site and its nonfragile allelomorph can be considered to constitute the first true chromosomal polymorphism to be described in man.     

Population cytogenetics

Chromosome variants are well established as useful genetic markers and integral components both of the genetic structure of populations, and in speciation. The current explosion of molecular techniques is facilitating the localization of many DNA sequences, while a reassessment of the fitness of chromosome mutants challenges some classical views on polymorphism and polytypy. Recent experiments on hybrid zones and mathematical modelling have greatly clarified the formation and evolution of chromosome races.     

Heritable fragile sites on human chromosomes. X. New folate-sensitive fragile sites: 6p23, 9p21, 9q32, and 11q23

Four new folate-sensitive fragile sites are documented at 6p23, 9p21, 9q32, and 11q23. These have all been shown to be heritable except for the one at 9p21, which has been seen only in a single individual. As with the other autosomal fragile sites, these appear to be innocuous in heterozygotes.     

Heritable fragile sites on human chromosomes II. Distribution, phenotypic effects, and cytogenetics.

Individuals and families have been documented in which there are a number of fragile sites on chromosomes. These include sites at 2q11, 10q23, 11q13, 16p124, 16q22, 20p11, and Xq27 or 28. Fragile sites reported in the literature are compiled. The cytogenetics of the sites is discussed. The phenotypic effects of the sites are considered, and it is speculated that homozygosity of the autosomal sites might be deleterious as is hemizygosity of the site on Xq. These sites are used in the previous report which documents the effect of tissue medium components on their expression.     

Common fragile sites

Aphidicolin-induced common fragile sites are site-specific gaps or breaks seen on metaphase chromosomes after partial inhibition of DNA synthesis. These fragile sites were first recognized during the early studies of the fragile X syndrome and are induced by the same conditions of folate or thymidylate stress used to induce the fragile X site. Common fragile sites are normally stable in cultured human cells. However, following induction with replication inhibitors, they display a number of characteristics of unstable and highly recombinogenic DNA. From the many studies that have cloned and characterized fragile sites, it is now known that these sites extend over large regions, are associated with genes, exhibit late or delayed replication, and contain regions of high flexibility but are otherwise unremarkable in sequence. Studies showing that fragile sites and their associated genes are frequently deleted or rearranged in cancer cells have clearly demonstrated their importance in genome instability in tumorigenesis. Yet until recently, very little was known about the molecular mechanisms involved in their stability. Recent findings showing that the key checkpoint genes ATR and BRCA1 are critical for genome stability at fragile sites have shed new light on these mechanisms and on the biological significance of common fragile sites.     

Autosomal fragile sites

It is possible to distribute the 17 autosomic fragile sites presently known in three categories according to their sensitivity: BrdU-sensitive sites (10q25, 16q22, 17p12), distamycin A-sensitive sites (16q22, 17p12) and folate- and thymidilate-sensitive sites (2q11-q14, 3p14, 6p23, 7p11, 8q22, 9p21, 9q32, 10q23, 11q13, 11q23, 12q13, 16p12, 16q23, 17p12, 20p11). Four fundamental problems are discussed, first the relation between the presence of a fragile site and the phenotype, secondly the incidence of autosomic sites, third the origin of fragility (particularity of DNA structure, defect of the DNA/proteins binding and abnormal arrangement of chromatin, abnormality of the metaphasic scaffold) and fourth the localization of fragile sites.     

Rare fragile sites

Rare folate-sensitive fragile sites are the archetypal trinucleotide repeats. Although the CAG repeat in the androgen receptor, associated with spinobulbar muscular atrophy, was the first to be published in 1991, it was the publication in the same year of the molecular basis of fragile X that focused much attention on trinucleotide repeat expansion as a mutational mechanism. A number of rare fragile sites have had their repeat elements characterised since that time. The so-called "folate-sensitive" fragile sites are likely to be all CCG repeat expansions similar to the fragile X. The folate insensitive fragile sites have more complex longer repeat elements. Only two rare fragile sites (FRAXA and FRAXE) are of unequivocal clinical significance in that they are associated with intellectual disability.     

Population cytogenetics of Atractomorpha similis

Atractomorpha similis (2 n=19 ♂, 20 ♀) is a hygrophilous, tropical to temperate, species of pyrgomorphine grasshopper. We have sampled 70 populations covering the known distributional range of this species within Australia. All of them proved to be polymorphic for heterochromatin content as revealed by C-band analysis of embryonic neuroblasts. This polymorphism affects all ten members of the basic haploid set and includes variants involving differences in either the presence or the amount of procentric, interstitial and terminal C-blocks, as well as variation in the occurrence and nature of short arms on otherwise telocentric chromosomes. A majority of these variants appear to result from heterochromatin addition since the presumptive sibling, Atractomorpha australis, like other species of the genus that have been C-banded, is generally depauperate in heterochromatin. The net result of this extraordinary polymorphism is that each chromosome of A. similis exists in 10–50 distinct morphs. Consequently, there is a high level of chromosomal heterozygosity in all populations in terms of the number of heterozygous pairs present within a complement and an even higher level in terms of the total range of karyomorph patterns. There is also a wide range of total heterochromatin content, as measured by the percent of the total chromosome area occupied by C-band material, with values ranging from 13% to 44%. Specific marker chromosomes which predominate in particular geographical areas serve to distinguish six major cytotypes within A. similis. The two most southerly of these cytotypes show a narrower range of heterochromatin content but with higher values which reflect the more general occurrence of substantial terminal C-blocks within them. Finally, the populations from Fraser Island constitute a particularly distinctive cytotype characterised by the least number of morphs, the lowest level of chromosomal heterozygosity and a restricted range of heterochromatin content confined to the lower end of the known distributional spectrum.     

Population cytogenetics of Atractomorpha similis

A 537 bp Taq 1 restriction fragment was cloned from the satellite-1 DNA of the Dee Why population of Atractomorpha similis from New South Wales. Tritiumlabelled-cRNA copies of this sat-1 probe, or else of a related Sau3A fragment from the same source, were used as in situ hybridisation probes to characterise the molecular organisation of the distal C-bands, which form a permanent and distinctive feature of the chromosomes of this species. Both probes were shown to be uniformly represented throughout all the distal C-bands not only of the Dee Why population itself but additionally in two other Australian populations where the bands are either less numerous (Fraser Island, Queensland) or smaller in size (Fogg Dam, Northern Territory). The same result was found in a population from Morehead, Papua New Guinea, which has a banding pattern similar to that of Fogg Dam. This holds whether the bands are single or multiple, terminal or subterminal. The probes were, however, consistently absent from all the proximal C-bands, whether centric or paracentromeric, as well as from the short arms which are sometimes present in the otherwise telocentric chromosomes. The results show that all the distal C-bands contain tandem blocks of highly repeated DNA from the same family of sequences. Moreover, the numerous polymorphisms which are present in the distal bands of all ten members of the basic mitotic set can be accounted for by differences in the amount of the sat-1 DNA present in a given pair of homologues. Since there is evidence to indicate that the size of the distal C-bands has increased subsequent to the introduction of the species into Australia there are good grounds for concluding that this increase has involved the amplification of the highly repeated sequence DNA present within the C-bands.     

DAPI-inducible common fragile sites

DAPI, a compound specific for the AT bases of DNA, causes gaps and breaks in three human chromosome sites, at the 1q41-1q42 interface, 2q31, and 7p22. It also induces undercondensation of a chromosome site at the 13q21-13q22 interface. The first three sites have the characteristics of "common fragile sites" and are present as gaps and breaks on the chromosomes of seven individuals.