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Sequence of the chromosomal breakpoint shows truncated chromosome 18 sequence followed by telomeric repeats. The telomeric repeat is indicated by horizontal arrows.
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Fragile sites are specific genomic loci that form gaps, constrictions and breaks on chromosomes exposed to replication stress conditions. In the father of a patient with Beckwith-Wiedemann syndrome and a pure truncation of 18q22-qter, a new aphidicolin-sensitive fragile site on chromosome 18q22.2 (FRA18C) is described. The region in 18q22 appears h...
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... 0.7% agarose gel, and after denaturation and neutralisation transferred to Hybond N + membranes. Hybridisations were performed at 65 ̊ C using a single PCR probe specific for each candidate repeat. Sequences of the primers, used to generate probes, and the appropriate restriction enzymes can be obtained from the authors on request. Previously, expression of the paternal fragile site at 18q22 in the father I.1 (fig 1) was found in 3/200 cells, analysed in two different experiments, grown in a folate-deficient medium. 18 However, we could not observe FRA18C expression under low folate conditions and moreover the 18q22 sequence appeared to be AT-rich—a feature of common fragile sites. Therefore we cultivated lymphoblastoid cells in the presence of 0.4 m M aphidicolin to activate common fragile site expression, and observed 18q22 fragile sites in 5/100 metaphases scored (fig 2). No fragile sites on chromosome 18 were found when cells were cultured in the absence of aphidicolin. This indicates that the fragile site can be induced by aphidicolin and is probably previously undescribed aphidicolin-induced fragile site, which was assigned the name FRA18C. Since aphidicolin-induced fragile sites are enriched with highly flexible AT-dinucleotide rich sequences, a flexibility analysis of the entire 18q22 region (chr18: 59 800 000–71 300 000) (http:// genome.ucsc.edu/cgi-bin/hgGateway), to which the fragile site was mapped, was performed. 7 8 For this, we used the Twistflex program—a computer program that identifies DNA sequences with a potential high flexibility at the twist angle. 8 This region is composed of three sub-bands: 18q22.1 (G band), 18q22.2 (R band) and 18q22.3 (G band). We previously showed that R and G bands differ in the mean number of flexibility islands. 8 Genomic regions mapped to G bands have significantly higher number of flexibility islands relative to R bands, hence each of the 18q22 sub-bands were analysed separately. As can be seen in figure 3, the G band 18q22.1 and most of G band 18q22.3 are highly enriched in flexibility islands (an average of 6.4 and 6.1 islands per 100 kb, respectively), relative to non-fragile regions mapped to G bands (an average of 3.3 islands per 100 kb), as well as relative to other aphidicolin-induced fragile sites mapped to this band type (an average of 5.7 islands per 100 kb). 8 The R band 18q22.2 is also highly enriched in flexibility islands (an average of 4.2 islands per 100 kb), both relative to non-fragile (an average of 1.8 islands per 100 kb) and fragile regions (an average of 3.4 islands per 100 kb) mapped to this band type. These results suggest that the flexibility of a substantial part of 18q22 is characteristic for common fragile sites. The chromosome breakpoint in the proband II.1 has been mapped previously to the interval between markers D18S477 and D18S61, a region of approximately 2.3 Mb. 18 By analysing additional markers in this family, we refined the candidate region to a 181 kb interval between markers D18S1092 and D18S61 and confirmed that the breakpoint occurred on the paternal chromosome (fig 1). To investigate whether the 18q22- qter deletion in the patient II.1 is a pure truncation or rather part of a more complex chromosomal rearrangement, we performed fluorescent in situ hybridisation experiments on metaphase spreads in combination with whole chromosome 18 painting using an 18q subtelomeric probe. The paint stained both copies of chromosome 18 in the father I.1 and the intact chromosome 18 as well as the derivative chromosome 18 in the patient II.1. The subtelomeric probe was detectable on the intact chromosome 18, but not on the derivative chromosome 18 in the patient II.1 (fig 4A). By analysing additional newly developed polymorphic CA- repeat markers in the family, the candidate region for chromosome breakage in patient II.1 was narrowed to the 157 kb interval between markers D18S61 and the newly developed marker 18q22_di 34, located proximal to D18S61. The breakage region was further refined to a 2.2 kb interval by performing a quantitative PCR-based fluodosage assay using 20 primersets developed in the D18S61–18q22_di34 interval. Using Southern blots of genomic DNA of the father I.1 and the patient II.1 digested with restriction endonucleases and hybridised with a probe specific for the region, we further refined the location of the breakpoint. Using probe 2, we detected an additional Hind II fragment of 4.8 kb in patient II.1 in addition to the fragment of 5.5 kb present in controls and the father I.1, suggesting that the breakpoint is situated approximately 700 bp from the Hind II site. The sequence of the breakpoint was determined after PCR amplification with a primer designed ¡ 500 bp from the expected breakpoint and a telomere-specific primer. 21 The sequence showed the expected, pure 18q sequence, up until base position 15 185 171 in contig NT_025028.13 (), immediately followed by the repetitive telomeric sequence (TTAGGG) n (fig 5). The breakpoint disrupts the DOK6 gene involved in the activation of the receptor tyrosine kinases. 22 The breakpoint is situated in intron 4 of the DOK6 gene. According to the NCBI database (), the entire DOK6 gene (441 kb) maps to 18q22.2 indicating that the breakpoint is positioned in 18q22.2 and not in 18q22.1 as suggested previously. 18 To investigate a possible association between the novel fragile site FRA18C in the father I.1 and the chromosomal breakpoint in the daughter II.1 of our family, fluorescent in situ hybridisation analysis on FRA18C-expressing cells of the father I.1 was performed with bacterial artificial chromosome clone RP11-64C15, spanning the breakpoint in combination with the reference probes CEP18 and GS-964-M9, located in the centromeric and telomeric region of chromosome 18, respectively. The results showed the colocalisation of the paternal fragile site with the chromosomal breakpoint in the progeny (II.1) in five of five analysed fragile sites (fig 2B). As shown in figure 3, the 18q22.2 region is highly enriched in flexibility islands. Interestingly, the breakpoint occurred in a genomic region of 300 kb within the R band 18q22.2, which shows even higher DNA flexibility (21 islands/300 kb) than the rest of this sub-band. Together, the results suggest that the breakpoint occurred within a fragile site region, which has high DNA flexibility. Since in vivo chromosome truncation at fragile sites has so far only been reported in case of a single, rare, folate-sensitive fragile site as a result of repeat expansion, 11 23 we wanted to investigate whether repeat expansions may occur within the FRA18C region. We searched a 400 kb interval surrounding the chromosomal breakpoint for repeats using the computer program Tandem Repeat Finder. 24 The breakpoint region appeared enriched in AT-rich repeats and no CGG-repeats were identified. To test whether any of the repeats were expanded in the father I.1, the 29 longest repeats were selected (see supplementary table 1, available at supplemental). They were first tested for heterozygosity using PCR with primers flanking the repeats. For six of these repeats (1, 2, 4, 6, 14 and 25) the father I.1 showed two alleles in the normal size range, excluding the possibility of an expansion in one of the alleles. The remaining 23 repeats were analysed using Southern blotting and hybridisation with probes specific for the region. Although the majority of the repeats were polymorphic, no repeat expansions were detected. This paper provides evidence that FRA18C is a previously undescribed aphidicolin-inducible fragile site at chromosome 18q22.2. When cells of the father I.1 were cultured in the presence of aphidicolin to activate the expression of common fragile sites, we observed fragile site expression in 5% of metaphases, but none were observed when cells were cultured without aphidicolin. We have no explanation why FRA18C expression is so low. However this does not seem to be unique for this site; Leversha et al 25 observed the common fragile site FRA6E in up to 3.5% of metaphases. We analysed the DNA flexibility of 18q22 and found that the region is relatively enriched in flexibility islands—characteristics of molecularly cloned common fragile sites. 8 26 Although this is the only case of FRA18C so far, it shows features of a common fragile site. Although common fragile sites are considered to be present in all individuals, there is little information concerning their distribution among the individuals. Several reports claim that the frequency of common fragile sites varies among different individuals. 27–29 Some common fragile sites, like FRA3B and FRA16D, are expressed in almost 100% of individuals, whereas others like FRA13A are less frequent and are only observed in a minority of individuals. 28 The latter group is sometimes referred to as the low-frequency common fragile sites. It seems plausible that FRA18C belongs to this group. We do not know why FRA18C was not detected previously. Assuming it has not been overlooked in the past, there are several possibilities why FRA18C is expressed in the father I.1, but not in others. First, the regional DNA composition of the father I.1 may be different as hypothesised by Savelyeva et al , 30 who suggested that the presence of sequence polymorphisms only in individuals who express the fragile site can explain the interindividual variability. However, we did not find any evidence for this possibility: all primer sets and probes developed by us on the basis of the published sequence of the human genome gave amplification products in the father as in controls, indicating absence of gross sequence differences between the father and the published sequence. In addition, no proof for repeat expansion or exceptional repeat composition in the region was found. A second possible explanation for the low occurrence of FRA18C is a variation in one of the enzymes responsible for the cell cycle checkpoints ...
Citations
... Irony-Tur Sinai, et al. integrated a 3.4 kb AT-rich sequence derived from FRA16C into a stable chromosomal region in the human genome, and showed that this integration drives fragile site formation under conditions of replication stress [16]. Importantly, the recurrent breakpoints found in cancer show significant overlaps with the AT-rich sequences at CFSs [43,51,52]. These data strongly suggest that CFS-ATs are one of the important elements contributing to CFS instability. ...
Abstract Common fragile sites (CFSs) are large chromosomal regions that exhibit breakage on metaphase chromosomes upon replication stress. They become preferentially unstable at the early stage of cancer development and are hotspots for chromosomal rearrangements in cancers. Increasing evidence has highlighted the complexity underlying the instability of CFSs, and a combination of multiple mechanisms is believed to cause CFS fragility. We will review recent advancements in our understanding of the molecular mechanisms underlying the maintenance of CFS stability and the relevance of CFSs to cancer-associated genome instability. We will emphasize the contribution of the structure-prone AT-rich sequences to CFS instability, which is in line with the recent genome-wide study showing that structure-forming repeat sequences are principal sites of replication stress.
... 123 At FRA18C, a paternal aphidicolin-sensitive fragile site coincided with a chromosome truncation in the daughter, and the breakpoint was in a region that was highly enriched in AT-rich sequences. 139 This case implicates in vivo fragility at AT/TA dinucleotide repeat sequences in genome rearrangements. ...
Alternative non‐B form DNA structures, also called secondary structures, can form in certain DNA sequences under conditions that produce single‐stranded DNA, such as during replication, transcription, and repair. Direct links between secondary structure formation, replication fork stalling, and genomic instability have been found for many repeated DNA sequences that cause disease when they expand. Common fragile sites (CFSs) are known to be AT‐rich and break under replication stress, yet the molecular basis for their fragility is still being investigated. Over the past several years, new evidence has linked both the formation of secondary structures and transcription to fork stalling and fragility of CFSs. How these two events may synergize to cause fragility and the role of nuclease cleavage at secondary structures in rare and CFSs are discussed here. We also highlight evidence for a new hypothesis that secondary structures at CFSs not only initiate fragility but also inhibit healing, resulting in their characteristic appearance.
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... These sequence and structural fea- tures have been validated experimentally in subsequent studies that reported replication fork arrest at AT-DRSs together with data highlight- ing the crucial role of proteins that can resolve DNA secondary structures in maintaining CFS stability. A direct indication of the role of AT-DRSs in DNA instability in vivo was found when recurrent cancer breakpoints occurring in CFSs were shown to overlap with AT-DRSs.[51][52][53]A recent paper by Tubbs et al. has found that large homopoly- meric (dA/dT) tracts are also preferential sites of replication fork stal- ling and collapse within a class of early replicating fragile sites (ERFSs), ...
Common fragile sites (CFSs) are specific genomic regions in normal chromosomes that exhibit genomic instability under DNA replication stress. Since replication stress is an early feature of cancer development, CFSs are involved in the signature of genomic instability found in malignant tumors. The landscape of CFSs is tissue‐specific and differs under different replication stress inducers. Nevertheless, the features underlying CFS sensitivity to replication stress are shared. Here we review the events generating replication stress and discuss the unique characteristics of CFS regions and the cellular responses aimed to stabilizing these regions.
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... 7 FRA18C was first identified in a child with Beckwith-Wiedemann syndrome, and her father. 8 Both had a pure truncation of 18q22-qter, which disrupted DOK6 in intron 4. Hemizygous deletion of FRA18C was found among 746 human cancer cell lines derived from 31 different tumor types. 9 Copy number loss at FRA18C was observed in Barrett's esophagus. ...
Chromosomal rearrangements are common in cancer. More than 50% occur in common fragile sites and disrupt tumor suppressors. However, such rearrangements are not known in gastric cancer. Here we report recurrent 18q2 breakpoints in 6 of 17 gastric cancer cell lines. The rearranged chromosome 18, t(9;18), in MKN7 cells was flow sorted and identified by reverse chromosome painting. High-resolution tiling array hybridization mapped breakpoints to DOK6 (docking protein 6) intron 4 in FRA18C (18q22.2) and an intergenic region in 9q22.2. The same rearrangement was detected by FISH in 22% of 99 primary gastric cancers. Intron 4 truncation was associated with reduced DOK6 transcription. Analysis of The Cancer Genome Atlas stomach adenocarcinoma cohort showed significant correlation of DOK6 expression with histological and molecular phenotypes. Multiple oncogenic signaling pathways (gastrin-CREB, NGF-neurotrophin, PDGF, EGFR, ERK, ERBB4, FGFR1, RAS, VEGFR2 and RAF/MAP kinase) known to be active in aggressive gastric cancers were strikingly diminished in gastric cancers with low DOK6 expression. Median survival of patients with low DOK6-expressing tumors was 2100 days compared with 533 days in patients with high DOK6-expressing tumors (log-rank P = 0.0027). The level of DOK6 expression in tumors predicted patient survival independent of TNM stage. These findings point to new functions of human DOK6 as an adaptor that interacts with diverse molecular components of signaling pathways. Our data suggest that DOK6 expression is an integrated biomarker of multiple oncogenic signals in gastric cancer and identify FRA18C as a new cancer-associated fragile site.
... To explain direct dup/del rearrangements we recall the mechanism proposed in maize for formation of a tandem direct duplication (Zhang et al. 2013) following a terminal deletion. In short, this hypothetic mechanism would imply a double-strand break at 18q22.3 (described as fragile site by Debacker et al. 2007) leaving both sister chromatids with 'exposed' ends; in turn, the illegitimate rejoining of one 'exposed' end with the sister chromatid at 18q12.2 would lead to a new recombinant chromatid with both the tandem direct duplication and the terminal deletion. Finally, given the risk of DNA erosion inherent to telomereless ends (Courtens et al. 1998;Heard et al. 2009), we assume healing of the broken 18q by formation of a neotelomere ( figure 3). ...
IntroductionAbout seven critical regions along the whole 18q have been proposed for Edwards syndrome (ES)/trisomy 18 (T18) phenotype (Boghosian-Sell et al. 1994; Nguyen-Minh et al. 2013). In contrast, hemizygosity for 18q22.3-18qter region has been linked to most of the features of the 18q− syndrome (Feenstra et al. 2007; Cody et al. 2009). Moreover, combination of both a partial gain and deletion within the same 18q seems to be fairly rare. Here we report on the first case of de novo 18q direct duplication (∼34.9 Mb) / deletion (∼8.8 Mb) in a girl child with combined phenotypes related to both syndromes as seen in other duplication/deletion complex rearrangements (Neira et al. 2012). Since these complex rearrangements usually exhibit an inv dup/del configuration derived from a U-type exchange, our findings and observations suggest variations of that mechanism for leading to the configuration observed here.Material and methodsClinical reportThe girl child is the third child of a G3P2C1 ...
... Chromosome breakage at or near the rare fragile site FRA11B has been implicated in Jacobsen syndrome and an aphidicolin-inducible fragile site, FRA 18C, was identified in the father of a patient with an 18q22.2-qter truncation and the Beckwith-Wiedemann syndrome [8,9]. Given the paucity of data indicating a role for other fragile sites in the formation of constitutional rearrangements as well as a study reporting on the lack of association between fragile sites and constitutional chromosome breakpoints [10], it was thought that the involvement of CFSs might be minimal in constitutional aberrations. ...
Chromosomal fragile sites (CFSs) are loci or regionssusceptible to spontaneous or induced occurrence of gaps, breaks and rearrangements. In this work, we studied the data of 4535 patients stored at DECI- PHER (Database of Chromosomal Imbalance and Phenotype in Humans Using Ensembl Resources). We mapped fragile sites to chromosomal bands and di- vided the 23 chromosomes into fragile and non-fragile sites. The frequency of rearrangements at the chro- mosomal location of clones found to be deleted or du- plicated in the array/CGH analysis, provided by DE- CIPHER, was compared in Chromosomal Fragile Sites vs. non-Fragile Sites of the human genome. The POSSUM Web was used to complement this study. The results indicated 1) a predominance of rear- rangements in CFSs, 2) the absence of statistically significant difference between the frequency of rear- rangements in common CFSs vs. rare CFSs, 3) a predominance of deletions over duplications in CFSs. These results on constitutional chromosomal rear- rangements are evocative of the findings previously reported by others relatively to cancer supporting the current line of evidence and suggesting that a com- mon mechanism can underlie the generation of con- stitutional and somatic rearrangements. The combi- nation of insights obtained from our results and their interrelationships can indicate strategies by which the mechanisms can be targeted with preventive medical interventions.
... FRA2H is about 2.5-fold enriched in clusters of flexibility peaks, defined as at least three flexibility peaks between which the distance of any two adjacent peaks is B5 kb (Supplementary Figure 1a). As the mean flexibility value for non-fragile G-band sequences has been defined as 3.3 flexibility islands/ 100 kb (Debacker et al. 2007), the number of flexibility peaks is increased 3.9-fold in FRA2H and 2.7-fold in its centromeric flanking sequence. At a twist angle of 16°, however, the values do not appear to differ substantially. ...
Common fragile sites (cFSs) are non-random chromosomal regions that are prone to breakage under conditions of replication stress. DNA damage and chromosomal alterations at cFSs appear to be critical events in the development of various human diseases, especially carcinogenesis. Despite the growing interest in understanding the nature of cFS instability, only a few cFSs have been molecularly characterised. In this study, we fine-mapped the location of FRA2H using six-colour fluorescence in situ hybridisation and showed that it is one of the most active cFSs in the human genome. FRA2H encompasses approximately 530 kb of a gene-poor region containing a novel large intergenic non-coding RNA gene (AC097500.2). Using custom-designed array comparative genomic hybridisation, we detected gross and submicroscopic chromosomal rearrangements involving FRA2H in a panel of 54 neuroblastoma, colon and breast cancer cell lines. The genomic alterations frequently involved different classes of long terminal repeats and long interspersed nuclear elements. An analysis of breakpoint junction sequence motifs predominantly revealed signatures of microhomology-mediated non-homologous recombination events. Our data provide insight into the molecular structure of cFSs and sequence motifs affected by their activation in cancer. Identifying cFS sequences will accelerate the search for DNA biomarkers and targets for individualised therapies.
... The Xq27-q28 band has already been recognized as having a high repeat density and high recombination frequency, indicating that it is a hot spot region for the generation of aberrant X recombination (Fimiani et al., 2006;Carvalho et al., 2009;Fusco et al., 2009) also in unbalanced translocations with different autosomal chromosomes (Caiulo et al., 1989;Yatsenko et al., 2004;Guo et al., 2009). Similarly, the surrounding areas of the 18q22 breakpoint are full of unstable sequences spanning a cluster of breakpoints for balanced and unbalanced translocations (Boghosian-Sell et al., 1996;Cuker et al., 2004;Riegel et al., 2005;Netzer et al., 2006;Bache et al., 2007;Debacker et al., 2007). Furthermore, the 18q22 breakpoint coincides with the FRA18C region, an aphidicolin-sensitive fragile site (Debacker et al., 2007). ...
... Similarly, the surrounding areas of the 18q22 breakpoint are full of unstable sequences spanning a cluster of breakpoints for balanced and unbalanced translocations (Boghosian-Sell et al., 1996;Cuker et al., 2004;Riegel et al., 2005;Netzer et al., 2006;Bache et al., 2007;Debacker et al., 2007). Furthermore, the 18q22 breakpoint coincides with the FRA18C region, an aphidicolin-sensitive fragile site (Debacker et al., 2007). ...
Diminished ovarian reserve (DOR) is a heterogeneous disorder causing infertility, characterized by a decreased number of oocytes, the genetic cause of which is still unknown.
We describe a family with a new unbalanced X;18 translocation der(X) associated with either fully attenuated or DOR phenotype in the same family. Cytogenetics and array comparative genomic hybridization (aCGH) studies have revealed the same partial Xq monosomy and partial 18q trisomy in both the 32-year-old female with DOR and the unaffected mother. The genetic analysis has defined a subtelomeric deletion spanning 13.3 Mb from Xq27.3 to -Xqter, which covers the premature ovarian failure locus 1 (POF1); and a duplication spanning 13.4 Mb, from 18q22.1 to 18qter. From a parental-origin study, we have inferred that the rearranged X chromosome is maternally derived. The Xq27 and 18q22 breakpoint regions fall in a region extremely rich in long interspersed nuclear element, a class of retrotransposons able to trigger mispairing and unusual crossovers. X-inactivation studies reveal a skewing of der(X) both in the mother and the proband. Therefore, the phenotypic expression of der(X) is fully attenuated in the fertile mother and partially attenuated in the DOR daughter.
We report on an unbalanced maternally derived translocation (X;18)(q27;q22) with different intra-familial reproductive performances, ranging from fertility to DOR. Skewed X-inactivation seems to restore the unbalanced genetic make-up, fully silencing the 18q22 trisomy and at least in part the Xq27 monosomy. The chromosomal abnormality observed in this family supports the presence of a DOR susceptibility locus in the distal Xq region and targets the POF1 region for further investigation.
... The flexibility of the FRA2Ctel and FRA2Ccen sequences was analyzed using the TwistFlex software. Both cFS are highly enriched in flexibility peaks relative to nonfragile regions, which exhibit on average 3.3 peaks/100 kb in a G-band and 1.8 peaks/100 in an R-band kb (43). In particular, FRA2Ccen is 3.18-fold more enriched (10.8 peaks per 100 kb, Table 2) in predicted flexibility peaks even relative to other characterized cFS sequences, which on average hold 3.4 peaks/100 kb in an R-band. ...
Common fragile sites (cFS) represent chromosomal regions that are prone to breakage after partial inhibition of DNA synthesis. Activation of cFS is associated with various forms of DNA instability in cancer cells, and is thought to be an initiating event in the generation of DNA damage in early-stage tumorigenesis. Only a few cFS have been fully characterized despite the growing interest in cFS instability in cancer genomes. In this study, six-color fluorescence in situ hybridization revealed that FRA2C consists of two cFS spanning 747 kb FRA2Ctel and 746 kb FRA2Ccen at 2p24.3 and 2p24.2, respectively. Both cFS are separated by a 2.8 Mb non-fragile region containing MYCN. Fine-tiling array comparative genomic hybridization of MYCN amplicons from neuroblastoma (NB) cell lines and primary tumors revealed that 56.5% of the amplicons cluster in FRA2C. MYCN amplicons are either organized as double minutes or as homogeneously stained regions in addition to the single copy of MYCN retained at 2p24. We suggest that MYCN amplicons arise from extra replication rounds of unbroken DNA secondary structures that accumulate at FRA2C. This hypothesis implicates cFS in high-level gene amplification in cancer cells. Complex genomic rearrangements, including deletions, duplications and translocations, which originate from double-strand breaks, were detected at FRA2C in different cancers. These data propose a dual role for cFS in the generation of gross chromosomal rearrangements either after DNA breakage or by inducing extra replication rounds, and provide new insights into the highly recombinogenic nature of cFS in the human cancer genome.
... Additionally, differences in the observable FS-frequency within different individuals are well known (11,12). Besides these 88 official, database-annotated common FS others were observed and reported, including six recently reported new sites, particularly FRA4F (13), FRA7K (14), FRA6H (15), FRA9G (16), FRA13E (15) and FRA18C (17). ...
... Even more complicating is the fact that 15 of these sites map at the same cytogenetic region but have different names because of different ways of induction (NCBI genome browser, Fig. 3). Additionally, 52 identified and in parts mapped FS were already published, but not included in the current genome browser versions (13,14,15,16,17,19,23,34). Most of these sites were confirmed in this study which underlines their eligibility to be appreciated as FS. ...
Since the first description of human fragile sites (FS) more than 40 years ago, a variety of substances were reported to induce chromosomal breaks at non-random, breakage-prone regions. According to information available from human genome browsers aphidicolin, an inhibitor of DNA replication induces 77 of 88 known common FS. However, in the literature additional FS are reported, which are also, at least in part, inducible by aphidicolin. To the best of our knowledge, here we present the first and largest ever done systematic, whole genome-directed and comprehensive screening for aphidicolin-inducible breakage-prone regions. The study was performed on stimulated peripheral blood lymphocytes of 3 unrelated healthy individuals. Twenty-five thousand metaphase spreads were analyzed and overall 22,537 FS located in 230 different loci were recorded. Sixty-one of those FS were never observed before and 52 were already previously reported but not included in genome browsers and yet verified. Interestingly, aphidicolin was able to induce all types of rare and common FS, suggesting that these breakage-prone regions are less dependent on the inducing chemicals than originally supposed. Overall, we provide the first comprehensive genome wide map for FS and studied possible correlations of chromosome length and GTG-banding level with FS-frequency. To handle FS better in future, an extension of the already existing alphabetical nomenclature for FS on single chromosomes is suggested.
