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. 2000 Jun;20(12):4420-7.
doi: 10.1128/MCB.20.12.4420-4427.2000.

Replication delay along FRA7H, a common fragile site on human chromosome 7, leads to chromosomal instability

A Hellman  1 , A Rahat, S W Scherer, A Darvasi, L C Tsui, B Kerem
Affiliations

Replication delay along FRA7H, a common fragile site on human chromosome 7, leads to chromosomal instability

A Hellman et al. Mol Cell Biol. 2000 Jun.

Abstract

Common fragile sites are specific chromosomal loci that show gaps, breaks, or rearrangements in metaphase chromosomes under conditions that interfere with DNA replication. The mechanism underlying the chromosomal instability at fragile sites was hypothesized to associate with late replication time. Here, we aimed to investigate the replication pattern of the common fragile site FRA7H, encompassing 160 kb on the long arm of human chromosome 7. Using in situ hybridization on interphase nuclei, we revealed that the replication of this region is initiated relatively early, before 30% of S phase is completed. However, a high fraction ( approximately 35%) of S-phase nuclei showed allelic asynchrony, indicating that the replication of FRA7H is accomplished at different times in S phase. This allelic asynchrony is not the result of a specific replication time of each FRA7H allele. Analysis of the replication pattern of adjacent clones along FRA7H by using cell population and two-color fluorescent in situ hybridization analyses showed significant differences in the replication of adjacent clones, under normal growth condition and upon aphidicolin treatment. This pattern significantly differed from that of two nonfragile regions which showed a coordinated replication under both conditions. These results indicate that aphidicolin is enhancing an already existing difference in the replication time along the FRA7H region. Based on our replication analysis of FRA7H and on previous analysis of the common fragile site FRA3B, we suggest that delayed replication is underlying the fragility at aphidicolin-induced common fragile sites.

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Figures

FIG. 1
FIG. 1
In situ hybridization pattern along the FRA7H region. (a) A cosmid map covering the FRA7H region. The SV40 and the ZNF131 pseudogene integration sites are marked by arrows. (b) Percentage of in situ hybridization signals in nuclei from the entire cell cycle, upon treatment with the fragility inducer (+aphidicolin). (c) The same analysis in nuclei grown under normal growth conditions (−aphidicolin). The percentage of alleles showing D signals (DD plus one-half SD nuclei) are indicated below each cosmid clone.
FIG. 2
FIG. 2
In situ hybridization pattern along nonfragile control regions. (a) Top, a cosmid map encompassed ∼160 kb from the CFTR region. The vertical bar indicates exon 1 of the CFTR gene. Bottom, percentage of in situ hybridization signals in nuclei from the entire cell cycle grown under normal growth conditions (−aphidicolin) and upon aphidicolin treatment (+aphidicolin). (b) Same analysis as in panel a of ∼140 kb from the AML1 region. The shading designations for SS, SD, and DD are the same as in Fig. 1.
FIG. 3
FIG. 3
Regression analysis of the correlation between the percentage of replicated alleles (D%) and the distance from the clone showing the highest level of replicated alleles along the FRA7H (●), the CFTR (□), and the AML1 (▵) regions. The distances were estimated from the center of each clone.
FIG. 4
FIG. 4
Two-color FISH analysis of the replication pattern of adjacent clones along the FRA7H region. Upper panel, number and percentage of FRA7H alleles showing differential FISH signals between cosmid clone 72c11 (red) and either 141e11 or 170g6 (green). The number of alleles that were scored until reaching 50 alleles showing differential signals is also indicated. Bottom panel, examples of nuclei showing differential in situ hybridization signals between clones 72c11 and 141e11 (a and b) and between clones 72c11 and 170g6 (c and d).
FIG. 5
FIG. 5
Replication pattern of FRA7H alleles in GM00847 cell line, carrying a duplicated SV40-marked chromosome 7. (a) Illustrations of the two possible type of nuclei: type 1, nuclei in which at least one nonmarked FRA7H allele, marked by an arrow, replicated later than at least one of the SV40-marked alleles; type 2, nuclei in which at least one nonmarked FRA7H allele, marked by an arrow, preceded the replication of at least one of the marked alleles. The percentage of nuclei of each type is indicated. (b) Examples of hybridization signals of type 1 nuclei. (c) Examples of hybridization signals of type 2 nuclei.
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References

    1. Austin M J, Collins J M, Corey L A, Nance W E, Neale M C, Schieken R M, Brown J A. Aphidicolin-inducible common fragile-site expression: results from a population survey of twins. Am J Hum Genet. 1992;50:76–83. - PMC - PubMed
    1. Boggs B A, Chinault A C. Analysis of replication timing properties of human X-chromosomal loci by fluorescence in situ hybridization. Proc Natl Acad Sci USA. 1994;91:6083–6087. - PMC - PubMed
    1. Chess A, Simon I, Cedar H, Axel R. Allelic inactivation regulates olfactory receptor gene expression. Cell. 1994;78:823–834. - PubMed
    1. Coquelle A, Pipiras E, Toledo F, Buttin G, Debatisse M. Expression of fragile sites triggers intrachromosomal mammalian gene amplification and sets boundaries to early amplicons. Cell. 1997;89:215–225. - PubMed
    1. Djalali M, Adolph S, Steinbach P, Winking H, Hameister H. A comparative mapping study of fragile sites in the human and murine genomes. Hum Genet. 1987;77:157–162. - PubMed

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