Trends in Genetics
Volume 28, Issue 6, June 2012, Pages 295-302
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Review
The complex basis underlying common fragile site instability in cancer

https://doi.org/10.1016/j.tig.2012.02.006Get rights and content

Common fragile sites (CFSs) were characterized almost 30 years ago as sites undergoing genomic instability in cancer. Recently, in vitro studies have found that oncogene-induced replication stress leads to CFS instability. In vivo, CFSs were found to be preferentially unstable during early stages of cancer development and to leave a unique signature of instability. It is now increasingly clear that, along the spectrum of replication features characterizing CFSs, failure of origin activation is a common feature. This and other features of CFSs, together with the replication stress characterizing early stages of cancer development, lead to incomplete replication that results in genomic instability preferentially at CFSs. Here, we review the shared and unique characteristics of CFSs, their underlying causes and their implications, particularly with respect to the development of cancer.

Section snippets

The involvement of CFSs in cancer

CFSs were first described in 1984 as gaps and constrictions in metaphase chromosomes of cells grown under mild replication stress conditions [1]. The expression level of a CFS is measured by the frequency of the gaps and constrictions at that site on metaphase chromosomes. CFSs are present in all individuals and are considered to be part of the normal chromosomal structure. For some CFSs in humans, an ortholog has been found in mice 2, 3, 4, 5 and primates [6]. Several studies suggested that

Sequence characteristics of CFSs

For many years, no specific DNA sequences characterizing CFSs were found. However, examination of the DNA sequence and structural characteristics showed that they are enriched in sequences with high DNA helix flexibility in the twist angle 27, 28. These flexible sequences were found to comprise interrupted AT-dinucleotide repeats (AT-rich) with a potential to form stable DNA secondary structures that might lead to inhibition of DNA replication [29] (Box 1). Recent studies described below in

Replication timing studies

Numerous mechanisms underlying CFS sensitivity to replication stress have been proposed years ago, including late timing of replication, widely separated origins and DNA sequences that are unusually difficult to replicate [36]. Analyzing the unique replication pattern of CFSs is important for understanding the replication features leading to their sensitivity to replication stress, which results in their instability. The replication pattern of several CFSs was initially analyzed using

FRA3B

FRA3B is mapped over a large genomic region exceeding 3 Mb. Within this region lies the large tumor suppressor gene, fragile histidine triad (FHIT), extending over 1.5 Mb. DNA combing analysis of 1.6 Mb overlapping FHIT demonstrated that the replication rate and fork stalling frequency along this site are similar to that along the whole genome [45]. A correlation between the origin pattern and the expression level of FRA3B was found. Analysis in lymphocyte cells, where FRA3B is highly

FRA16C

A recent DNA combing study analyzing the replication pattern along 600 kb within the CFS FRA16C identified four AT-rich flexibility sequences over a length of 400 bp [35]. The analysis revealed a slower replication rate and shorter origin distance at the fragile site region compared with the whole genome under normal growth conditions, indicating that additional origins are activated. Because dormant origins are normally activated following replication stress, the replication pattern of FRA16C

FRA6E

FRA6E harbors the large parkinson protein 2 (PARK2) gene, which extends over 1.3 Mb. The replication pattern of approximately 1 Mb of FRA6E containing part of PARK2 was analyzed using DNA combing [49]. The analyzed region of this fragile site contains relatively long AT-rich sequences (400–1383 bp), along which the replication rate of the fragile site was slower, the origin distance was shorter and a higher frequency of fork arrests was found compared with the whole genome. A comparison between

Mechanisms leading to CFS instability

Several lines of evidence suggest that no single mechanism can account for the instability of all CFSs. Different potential characteristics of CFSs have been proposed over the years, but none are common to all of the cloned CFSs (Figure 1).

One characteristic of CFSs contributing to instability is colocalization with very large genes. Some of the large CFS genes [such as FHIT and WW domain containing oxidoreductase (WWOX)] play important roles in cancer development. It has been proposed that

Genome instability of CFSs during cancer development

Molecular mapping of fragile sites reveals that genomic instability in CFSs alters the expression of important cancer genes in different cancer types. FRA3B, the most studied CFS, maps to intron 5 of the tumor suppressor gene FHIT [52]. This CFS is altered in a variety of human cancers, including lung, breast, head and neck, stomach and pancreatic carcinomas [53]. Interestingly, integrations of human papilloma virus (HPV) in cervical cancer cells were also found in FRA3B [54]. FRA16D also

CFSs are preferentially unstable during early stages of cancer development

To shed light on the events leading to genomic instability in cancer, several studies focused on the early stages of cancer development. Large-scale LOH and comparative genome hybridization analyses during early stages of different cancers revealed that the instability in CFSs precedes the instability in other genomic regions 25, 59. Similar results were obtained in genome-wide analyses in precancerous experimental models [75]. These findings suggest a new model in which replication stress is

Concluding remarks

Recent advances in technologies enabling whole-genome instability and DNA replication analyses have led to a breakthrough in understanding the mechanisms leading to the instability of CFSs during the early stages of cancer development. These analyses revealed that replication stress that occurs during cancer development leads to preferential instability along the replication-sensitive CFS regions. Instability in CFSs in cancer may lead to deletions, translocations, amplifications and

References (102)

  • E. Ozeri-Galai

    Failure of origin activation in response to fork stalling leads to chromosomal instability at fragile sites

    Mol. Cell

    (2011)
  • M. Giacca

    Mapping replication origins by quantifying relative abundance of nascent DNA strands using competitive polymerase chain reaction

    Methods

    (1997)
  • A. Helmrich

    Collisions between replication and transcription complexes cause common fragile site instability at the longest human genes

    Mol. Cell

    (2011)
  • L.V. O’Keefe et al.

    Common chromosomal fragile sites and cancer: focus on FRA16D

    Cancer Lett.

    (2006)
  • D.G. Albertson

    Gene amplification in cancer

    Trends Genet.

    (2006)
  • F. Pelliccia

    Breakages at common fragile sites set boundaries of amplified regions in two leukemia cell lines K562 – molecular characterization of FRA2H and localization of a new CFS FRA2S

    Cancer Lett.

    (2010)
  • W. Paradee

    A 350-kb cosmid contig in 3p14.2 that crosses the t(3;8) hereditary renal cell carcinoma translocation breakpoint and 17 aphidicolin-induced FRA3B breakpoints

    Genomics

    (1996)
  • M. Ohta

    The FHIT gene, spanning the chromosome 3p14.2 fragile site and renal carcinoma-associated t(3;8) breakpoint, is abnormal in digestive tract cancers

    Cell

    (1996)
  • A. Coquelle

    Expression of fragile sites triggers intrachromosomal mammalian gene amplification and sets boundaries to early amplicons

    Cell

    (1997)
  • A.M. Casper

    ATR regulates fragile site stability

    Cell

    (2002)
  • A.M. Casper

    Chromosomal instability at common fragile sites in Seckel syndrome

    Am. J. Hum. Genet.

    (2004)
  • M.F. Arlt et al.

    Inhibition of topoisomerase I prevents chromosome breakage at common fragile sites

    DNA Repair

    (2010)
  • G. Sozzi

    The FHIT gene 3p14.2 is abnormal in lung cancer

    Cell

    (1996)
  • Y.-H. Wang

    Chromatin structure of human chromosomal fragile sites

    Cancer Lett.

    (2006)
  • T.W. Glover

    DNA polymerase alpha inhibition by aphidicolin induces gaps and breaks at common fragile sites in human chromosomes

    Hum Genet.

    (1984)
  • T.W. Glover

    The murine Fhit gene is highly similar to its human orthologue and maps to a common fragile site region

    Cancer Res.

    (1998)
  • T. Shiraishi

    Sequence conservation at human and mouse orthologous common fragile regions, FRA3B/FHIT and Fra14A2/Fhit

    Proc. Natl. Acad. Sci. U.S.A.

    (2001)
  • K.A. Krummel

    The common fragile site FRA16D and its associated gene WWOX are highly conserved in the mouse at Fra8E1

    Genes Chromosomes Cancer

    (2002)
  • L. Rozier

    Characterization of a conserved aphidicolin-sensitive common fragile site at human 4q22 and mouse 6C1: possible association with an inherited disease and cancer

    Oncogene

    (2004)
  • A. Ruiz-Herrera

    Conservation of aphidicolin-induced fragile sites in Papionini (Primates) species and humans

    Chromosome Res.

    (2004)
  • R.S. Cha et al.

    ATR homolog Mec1 promotes fork progression, thus averting breaks in replication slow zones

    Science

    (2002)
  • A. Admire

    Cycles of chromosome instability are associated with a fragile site and are increased by defects in DNA replication and checkpoint controls in yeast

    Genes Dev.

    (2006)
  • K. Mrasek

    Global screening and extended nomenclature for 230 aphidicolin-inducible fragile sites, including 61 yet unreported ones

    Int. J. Oncol.

    (2010)
  • C.H. Cheng et al.

    DNA polymerase epsilon: aphidicolin inhibition and the relationship between polymerase and exonuclease activity

    Biochemistry

    (1993)
  • S. Ikegami

    Aphidicolin prevents mitotic cell division by interfering with the activity of DNA polymerase-alpha

    Nature

    (1978)
  • F. Hecht et al.

    Fragile sites and chromosome breakpoints in constitutional rearrangements I. Amniocentesis

    Clin. Genet.

    (1984)
  • D. Kotzot

    Parental origin and mechanisms of formation of cytogenetically recognisable de novo direct and inverted duplications

    J. Med. Genet.

    (2000)
  • C.T. Miller

    Genomic amplification of MET with boundaries within fragile site FRA7G and upregulation of MET pathways in esophageal adenocarcinoma

    Oncogene

    (2006)
  • E.C. Thorland

    Common fragile sites are preferential targets for HPV16 integrations in cervical tumors

    Oncogene

    (2003)
  • E.C. Thorland

    Human papillomavirus type 16 integrations in cervical tumors frequently occur in common fragile sites

    Cancer Res.

    (2000)
  • R. Di Micco

    Oncogene-induced senescence is a DNA damage response triggered by DNA hyper-replication

    Nature

    (2006)
  • J. Bartkova

    DNA damage response as a candidate anti-cancer barrier in early human tumorigenesis

    Nature

    (2005)
  • V.G. Gorgoulis

    Activation of the DNA damage checkpoint and genomic instability in human precancerous lesions

    Nature

    (2005)
  • D. Mishmar

    Molecular characterization of a common fragile site (FRA7H) on human chromosome 7 by the cloning of a simian virus 40 integration site

    Proc. Natl. Acad. Sci. U.S.A.

    (1998)
  • E. Zlotorynski

    Molecular basis for expression of common and rare fragile sites

    Mol. Cell. Biol.

    (2003)
  • S.N. Shah

    DNA structure and the Werner protein modulate human DNA polymerase delta-dependent replication dynamics within the common fragile site FRA16D

    Nucleic Acids Res.

    (2010)
  • K. Ried

    Common chromosomal fragile site FRA16D sequence: identification of the FOR gene spanning FRA16D and homozygous deletions and translocation breakpoints in cancer cells

    Hum. Mol. Genet.

    (2000)
  • K. Debacker

    FRA18C: a new aphidicolin-inducible fragile site on chromosome 18q22, possibly associated with in vivo chromosome breakage

    J. Med. Genet.

    (2007)
  • M.M. Le Beau

    Replication of a common fragile site, FRA3B, occurs late in S phase and is delayed further upon induction: implications for the mechanism of fragile site induction

    Hum. Mol. Genet.

    (1998)
  • L. Wang

    Allele-specific late replication and fragility of the most active common fragile site, FRA3B

    Hum. Mol. Genet.

    (1999)
  • Cited by (0)

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