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.     

Segregation analysis of rare autosomal fragile sites

Summary Segregation analyses were performed on pedigrees with rare autosomal fragile sites. The results of the analysis of pedigrees with folate sensitive fragile sites, including 2q1, 6p23, 7p11, 8q22, 9q32, 10q23, 11q13, 11q23, 12q13, 16p12, and 20p11, suggested that expression of the gene depended on the carrier parent: it was only 50% penetrant when transmitted by a carrier father, but fully penetrant when transmitted by a carrier mother. Pedigrees with the bromodeoxyuridine (BrdU) fragile site, fra(10)(q25), showed the same trend but the results were not statistically significant. In addition, 38 of the 44 probands with folate sensitive or BrdU-sensitive fragile sites received the gene from their carrier mother and only six received it from their father. In contrast, the analysis of pedigrees with the distamycin A-inducible site, fra(16)(q22), gave the results expected for a simple codominant trait with complete penetrance. Probands with this fragile site received the gene equally from mothers or fathers. The genetic implications of these results are discussed.     

Heritable fragile sites on human chromosomes. XI. Factors affecting expression of fragile sites at 10q25, 16q22, and 17p12.

The fragile sites at 10q25, 16q22, and 17p12 can all be induced in lymphocyte culture by BrdU or BrdC added 6-12 hrs prior to harvest. Without induction, fra(10)(q25) is rarely expressed spontaneously, whereas fra(16)(q22) is frequently expressed spontaneously. Fra(17)(p12) is frequently expressed spontaneously but is probably expressed only after induction in some individuals. Distamycin A, netropsin, and Hoechst 33258 induced high levels of expression of fra(16)(q22) and fra(17)(p12) but did not enhance expression of fra(10)(q25). The mechanisms of induction of fra(16)(q22) by BrdU and distamycin A appear to be different, since the time of induction by BrdU reaches a maximum about 12 hrs prior to harvest whereas induction by distamycin A requires much longer exposure. The fragile sites at 10q25 and 16q22 were both induced in fibroblast culture by BrdU. Fra(17)(p12) is accepted as a fragile site because preliminary studies show that it behaves similarly in lymphocyte culture to fra(16)(q22); however, there is only limited evidence for fragility at 17p12.     

Studies on three rare fragile sites

Summary Three fragile sites 2q13, 12q13, and 17p12 were found in one family. In the index case, who was first investigated in 1969 for low birth weight and bilateral inguinal hernia, three tissues were examined, blood, marrow, and skin. Three of the family have been reinvestigated after 17 years. Cultures for sister chromatid exchange (SCE) and the effects of aphidicolin, fluorodeoxyuridine (FUdR), bromodeoxyuridine (BrdU), and methotrexate on the frequency of the fragilities were studied. The mother of the index case who is an obligate carrier for the fragile 2q13 does not express it in folate/thymidine deficient medium. Further studies on her using a lymphoblastoid cell line, showed that there was a reduced level of fragility of 12q13 and 17p12 in B-lymphocytes compared to T-lymphocytes. Excess thymidine and FUdR when added to the lymphoblastoid cell line did not induce the 2q13. These studies also confirm the induction of a range of common fragile sites by treatment with aphidicolin, showing in addition homozygosity for at least 3p14, 6q26, 16q23, and Xp22. There were no detectable increases in the SCE rate between individuals with fragile sites and the five controls tested. There was no history of cancer or phenotypic abnormalities in the family.     

Nonrandom distribution of methotrexate-induced aberrations on human chromosomes. Detection of further folic acid sensitive fragile sites

Summary Eleven folic acid sensitive fragile sites (3p14, 7p13, 7q31.1, 7q32, 9q32, 11p13, 14q23, 15q22, 16q23, Xp22.2, Xq22) were detected in one individual, eight of them previously unknown. These sites seem to bear each its specific sensitivity to folic acid deficiency. Six of the sites were observed simultaneously on both homologous chromosomes in at least one cell. Each of these 11 sites was also found in at least one among 12 individuals further examined. Some of these individuals showed six of these 11 sites. The fragile site 3p14 was detected in all individuals examined. The homologous sites 3p14 of one individual differed from each other in their frequency of lesions induced by methotrexate as well as fluorodeoxyuridine. This observation suggests that folic acid sensitivity is a property inherent in the chromatin of an individual chromosome at the site involved in fragility. This property seems to be responsible for the nonrandom fragility at that site and also for the individual sensitivity of each chromosomal site.     

Induction of distamycin A-inducible rare fragile sites and increased sister chromatid exchanges at the fragile site

Summary Expression of distamycin A-inducible rare fragile sites by AT-specific DNA-ligands was examined in lymphoblastoid cell lines derived from heterozygous carriers for the fra(8)(q24), fra(16)(pl2), and fra(16)(q22) sites. The sensitivity of fragile site expression to the inducers was different at these fragile sites. The expression of fra(8)(q24) was induced markedly by Hoechst 33258, but not by distamycin A or berenil. An increased expression of fra(16)(p12) was found following treatment with Hoechst 33258 or berenil, but not with distamycin A. At fra(16)(q22), distamycin A markedly induced the fragile site, but Hoechst 33258 and berenil did not. Since their response to the different inducers was similar to that found in cultured lymphocytes, lymphoblastoid cell lines appear to retain their inherent properties. Although BrdUrd alone did nto induce any fragile sites, concomitant treatment with BrdUrd plus the inducer was synergistically effective in inducing all the fragile sites. An increased frequency of sister chromatid exchanges was observed at fra(16)(p12) following simultaneous treatment with BrdUrd and berenil, mainly when the site was expressed as an isochromatid gap. Thus, the induced fra (16)(pl2) site is a hot spot for the formation of sister chromatid exchanges, as found in other reported fragile sites.     

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).     

Chromosomal fragile sites in schizophrenic patients

Schizophrenia is a common and complex mental disorder. Cytogenetic and molecular studies have shown that genetic factors play an important role in the etiology of schizophrenia. As a preliminary step in the search for chromosomal location of a susceptible gene predisposing to schizophrenia, cytogenetic screening patients might be useful. Therefore, this report is aimed at studying the relationship between chromosomal fragile sites (FS: gaps, breaks, triradial figures, and several rearrangements) and the etiology of schizophrenia. Because of this, we were compared the frequencies of folate-sensitive FS from schizophrenic patients and normal individuals in short-term whole blood cultures. The rate of FS expression in the patients was considerably higher than in the controls. We determined 15 common FS (cFS) (1q21, 1q32, 2q21, 2q31, 3p14, 4q31, 5q31, 6q21, 6q26, 7q22, 7q32, 10q22, 13q32, Xp22 and Xq22), 6 rare FS (rFS) (6p21, 8q22, 11q23, 12q24, 16q22, and Xq26) and 2 previously unknown FS (3p25 and 5q22). Among these expressed FS, there was a significantly higher frequency of 12 FS at 2q31, 3p25, 3p14, 5q31, 6q21, 7q22, 7q32, 10q22, 11q23, 12q24, Xq22 and Xq26 in patient group than in controls by chi2 test (P = between 0.0001 to 0.036). Sites 3p14, 5q31 and 7q22 were also the most frequently observed cFS. Males exhibited twice as many FS as females, but no age effects were observed. The potential relationship between increased FS frequency and the occurrence of schizophrenia in these patients is discussed.     

Chromosomal fragile sites in schizophrenic patients

Schizophrenia is a common and complex mental disorder. Cytogenetic and molecular studies have shown that genetic factors play an important role in the etiology of schizophrenia. As a preliminary step in the search for chromosomal location of a susceptible gene predisposing to schizophrenia, cytogenetic screening of patients might be useful. Therefore, this report is aimed at studying the relationship between chromosomal fragile sites (FS) (gaps, breaks, triradial figures, and several rearrangements) and the etiology of schizophrenia. Because of this, we compared the frequencies of folate-sensitive FS from schizophrenic patients and normal individuals in short-term whole-blood cultures. The rate of FS expression in the patients was considerably higher than in the controls. We determined 15 common FS (cFS) (1q21, 1q32, 2q21, 2q31, 3p14, 4q31, 5q31, 6q21, 6q26, 7q22, 7q32, 10q22, 13q32, Xp22, and Xq22), six rare FS (rFS) (6p21, 8q22, 11q23, 12q24, 16q22, and Xq26), and two previously unknown FS (3p25 and 5q22). Among these expressed FS, there was a significantly higher frequency of 12 FS at 2q31, 3p25, 3p14, 5q31, 6q21, 7q22, 7q32, 10q22, 11q23, 12q24, Xq22, and Xq26 in patient group than in controls by x ²-test (P between 0.0001 to 0.036). Sites 3p14, 5q31, and 7q22 were also the most frequently observed cFS. Males exhibited twice as many FS as females, but no age effects were observed. The potential relationship between increased FS frequency and the occurrence of schizophrenia in these patients is discussed. The text was submitted by the authors in English.     

Exact localization of several fragile sites remains uncertain. The example of fra(10) sensitive to folate

The localization of the folate-sensitive fragile site first proposed to be at 10q24.2 was further assigned to 10q23. The study of one case using several banding techniques, and of 12 other unpublished cases studied with R-banding confirm the original localization, at the junction between 10q24.1 and 10q24.2. Because many reports propose that fragile sites induce further anomalies, we suggest that a re-study of the exact location of these sites with accurate methods is in order if misinterpretations are to be avoided.     

Heritable fragile sites on human chromosomes

Summary A kindred is described in which six members have a fragile site at 12q13. This fragile site was found to be suppressed by folic acid and thymidine in lymphocyte culture. An updated classification of known fragile sites is presented.     

Chromosomal fragile site expression in lymphocytes from patients with schizophrenia

Schizophrenia is a common complex mental disorder. The lifetime prevalence of this disease is about 1% across different populations. The etiology is still unknown despite decades of intensive study. This report is aimed at studying the relationship between chromosomal fragile sites and the etiology of schizophrenia. Lymphocytes of 72 schizophrenic patients and 66 healthy controls were cultured in M medium, which is deficient in folic acid, and in medium RPMI1640 with distamycin A. G-banding was carried out on 100 metaphases of each individual. Fragile sites were characterized as specific chromosomal bands that exhibit nonrandom gaps or breaks. Culture in M medium resulted in significant differences in the total number of chromosomal lesions and the total number of cells with chromosomal lesions between patients and controls (P<0.001), while no difference was noted after exposure to distamycin A. In the case of M medium, 17 bands in both patients and controls were recognized as expressing fragile sites nonrandomly using a statistical method based on the relationship of the binomial and F distributions. Further analysis using Fisher’s exact test revealed a significant excess of expression of a rare fragile site at 2q11.2 among patients compared with controls (P<0.05). In the case of distamycin A induction, 13 bands were identified as having nonrandom expression of fragile sites using the same statistical method. A significant excess expression of a fragile site at 9q12 was identified among patients compared with controls by applying Fisher’s exact test (P<0.001). Thus, our data suggest that chromosomal bands 2q11.2 and 9q12 are interesting regions that may harbor important genes associated with schizophrenia. Received: 21 July 1998 / Accepted: 19 September 1998     

A male with two different familial autosomal fragile sites and a cytogenetically abnormal offspring

During the course of routine prenatal cytogenetics, a male with 2 different autosomal fragile sites (FS) was detected. The FS were at 9p21 and 12q13 and his sister also had both fragile sites, inherited from an obligate carrier father. He was the father of a foetus with an abnormal chromosome complement 46,XY/47,XY,+frag. The origin of the fragment could not be determined.     

Aberrant breakpoints in chronic myelogenous leukaemia; oncogenes and fragile sites

Summary Three hundred and twenty-five aberrant breakpoints in chronic myelogenous leukaemia (CML) with Philadelphia chromosome variant were reviewed. Eight chromosomal bands (3p21, 6p21, 7p22, 11q13, 12p13, 17p13, 17q21, and 17q25) were found to be highly involved. Apart from 17q25, all these bands correspond to oncogenes sites and/or sites involved as primary breakpoints in cancer.This work was presented in part at the Congrès National d'Hématologie et de Transfusion Sanguine, March 1985 (Nouv Rev Fr Hématol, 1985, 27:72) and at the American Association for Cancer Research Meeting (Proceeding of the AACR 1986, 27:148)     

Family study of common fragile sites

Summary The frequency of folate-sensitive common fragile sites (1p31, 1q44, 3p14, 3q26.2, 6q26, 16q23, Xp22.3) was determined in 19 healthy individuals from four families. The individuals consisted of 12 males and 7 females from 1 to 59 years of age. The frequency showed intrafamilial variation, but we were unable to demonstrate that the frequency was inherited in a Mendelian codominant fashion. In eight subjects whose chromosome 3 homologues could be distinguished by Q-band polymorphism, breakages at 3p14 occurred with equal frequencies on the homologues. Our study suggests that common fragile sites are a part of normal chromosome structure, and the frequency of their expression largely depends on environmental factors.     

Chromosomal expression and localization of aphidicolin-induced fragile sites in the standard karyotype of river buffalo (Bubalus bubalis)

The present study reports on the chromosomal expression and localization of aphidicolin-induced fragile sites in the standard karyotype of river buffalo (Bubalus bubalis, 2n = 50) with the aim of establishing a 'fragile site map' of the species. Totally, 400 aphidicolin-induced breakages were analyzed from eight young and clinically healthy animals, four males and four females; these breakages were localized in 106 RBG-negative chromosome bands or at the band-interband regions. The number of breakages per chromosome did not vary statistically 'among' the animals investigated but the differences among individual chromosomes were highly significant thus indicating that the chromosomal distribution of the breakages is not random and appears only partially related to chromosome length. Fragile sites were statistically determined as those chromosomal bands showing three or more breakages. In the river buffalo karyotype, 51 fragile sites were detected and localized on the standardized ideogram of the species. The most fragile bands were as follows: 9q213 with 24 breakages out of 400; 19q21 with 16, 17q21 and inacXq24 with 15, 15q23 with 13 and 13q23 with 12 breaks, respectively. Previous gene mapping analysis in this species has revealed that the closest loci to these fragile sites contain genes such as RASA1 and CAST (9q214), NPR3 and C9 (19q19), PLP and BTK (Xq24-q25), OarCP09 (15q24), and EDNRB (13q22) whose mutations are responsible for severe phenotypic malformations and immunodeficiency in humans as well as in mice and meat quality in pigs. Further cytogenetic and molecular studies are needed to fully exploit the biological significance of the fragile sites in karyotype evolution of domestic animals and their relationships with productive and reproductive efficiency of livestock.     

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.     

Heritable fragile sites on human chromosomes

Summary The chromosomal gaps associated with fragile sites at 2q11, 10q23, 11q13, 16q22, 20p11 and Xq27 do not stain with silver nitrate as do NOR regions of the acrocentric chromosomes.     

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.     

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.