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Evidence for replicative repair of DNA double-strand breaks leading to oncogenic translocation and gene amplification.

Difilippantonio MJ, Petersen S, Chen HT, Johnson R, Jasin M, Kanaar R, Ried T, Nussenzweig A - J. Exp. Med. (2002)

Bottom Line: Subsequent DNA repair events juxtaposing IgH and c-myc are mediated by a break-induced replication pathway.Cycles of breakage-fusion-bridge result in amplification of IgH/c-myc while chromosome stabilization occurs through telomere capture.Thus, mice deficient in NHEJ provide excellent models to study the etiology of unbalanced translocations and amplification events during tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Genetics Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA. difilipm@mail.nih.gov

ABSTRACT
Nonreciprocal translocations and gene amplifications are commonly found in human tumors. Although little is known about the mechanisms leading to such aberrations, tissue culture models predict that they can arise from DNA breakage, followed by cycles of chromatid fusion, asymmetric mitotic breakage, and replication. Mice deficient in both a nonhomologous end joining (NHEJ) DNA repair protein and the p53 tumor suppressor develop lymphomas at an early age harboring amplification of an IgH/c-myc fusion. Here we report that these chromosomal rearrangements are initiated by a recombination activating gene (RAG)-induced DNA cleavage. Subsequent DNA repair events juxtaposing IgH and c-myc are mediated by a break-induced replication pathway. Cycles of breakage-fusion-bridge result in amplification of IgH/c-myc while chromosome stabilization occurs through telomere capture. Thus, mice deficient in NHEJ provide excellent models to study the etiology of unbalanced translocations and amplification events during tumorigenesis.

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Related in: MedlinePlus

FISH mapping of the chromosome 12 and 15 breakpoints in 11 pro-B cell lymphomas. Top central panel illustrates the locations of the IgH Cα and c-myc loci, two overlapping BAC clones spanning the breakpoint in tumor PKT2 (96F18 and 212H11), and the hybridization site of a chromosome 15 subtelomeric probe (15T) on normal chromosomes. In the bottom panels, the breakpoints and rearrangements in the pro-B cell lymphomas are shown. The normal chromosomes appear to the far left (Chr. 12) and right (Chr. 15) of each box while the derivative chromosomes are illustrated between them. Arrows on the normal chromosomes indicate the location of the rearrangements on the derivative chromosomes.
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fig5: FISH mapping of the chromosome 12 and 15 breakpoints in 11 pro-B cell lymphomas. Top central panel illustrates the locations of the IgH Cα and c-myc loci, two overlapping BAC clones spanning the breakpoint in tumor PKT2 (96F18 and 212H11), and the hybridization site of a chromosome 15 subtelomeric probe (15T) on normal chromosomes. In the bottom panels, the breakpoints and rearrangements in the pro-B cell lymphomas are shown. The normal chromosomes appear to the far left (Chr. 12) and right (Chr. 15) of each box while the derivative chromosomes are illustrated between them. Arrows on the normal chromosomes indicate the location of the rearrangements on the derivative chromosomes.

Mentions: Sequence analysis and localization of fusion points and rearrangements in the IgH Ca and c-myc loci. (A) The fusion points of three Ku80−/−p53−/− pro-B cell lymphomas (PKT no. 2, 7, and 13) were cloned by digestion circularization PCR [der(15)a]. Sequence analysis of the three breakpoints from scenario A demonstrate microhomology (yellow) at the junction of chromosomes 12 (red) and 15 (green) in two cases (PKT2 and PKT7), but none in tumor PKT13. Homologous bases outside of this region are also indicated in uppercase letters. (B) Sequence analysis implied that the breakpoint was the V(D)J recombination signal sequence (RSS; blue) flanking IgH J1 in PKT2 and IgH J4 in PKT7 and PKT13 (top panel); however, exonuclease digestion of the cleaved ends resulted in loss of 45, 208, and 189 bp, respectively, from the original site of the RAG-mediated DSB to the fusion point (arrows). Approximate localization of the breakpoints on chromosome 15 (second panel) was determined by FISH hybridization with two BAC clones in this region (96F18 and 212H11) as illustrated in Fig. 5. Tumors 1–4, 7, and 13 are from Ku80−/−p53−/− mice, tumors *1–*3 are from Ku70−/−p53−/− mice and tumors N9984 and N10527 are from Ku80−/−p53−/−RAD54−/− mice. The resulting der(15)a and der(12)b chromosomes are illustrated in the bottom panels. In scenario A a portion of the IgH locus is copied to chromosome 15 downstream of c-myc and in the reverse orientation. In scenario B it appears that the copying of chromosome 15 to the der(12)b initiates centromeric of the c-myc locus and continues along toward the telomere.


Evidence for replicative repair of DNA double-strand breaks leading to oncogenic translocation and gene amplification.

Difilippantonio MJ, Petersen S, Chen HT, Johnson R, Jasin M, Kanaar R, Ried T, Nussenzweig A - J. Exp. Med. (2002)

FISH mapping of the chromosome 12 and 15 breakpoints in 11 pro-B cell lymphomas. Top central panel illustrates the locations of the IgH Cα and c-myc loci, two overlapping BAC clones spanning the breakpoint in tumor PKT2 (96F18 and 212H11), and the hybridization site of a chromosome 15 subtelomeric probe (15T) on normal chromosomes. In the bottom panels, the breakpoints and rearrangements in the pro-B cell lymphomas are shown. The normal chromosomes appear to the far left (Chr. 12) and right (Chr. 15) of each box while the derivative chromosomes are illustrated between them. Arrows on the normal chromosomes indicate the location of the rearrangements on the derivative chromosomes.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196056&req=5

fig5: FISH mapping of the chromosome 12 and 15 breakpoints in 11 pro-B cell lymphomas. Top central panel illustrates the locations of the IgH Cα and c-myc loci, two overlapping BAC clones spanning the breakpoint in tumor PKT2 (96F18 and 212H11), and the hybridization site of a chromosome 15 subtelomeric probe (15T) on normal chromosomes. In the bottom panels, the breakpoints and rearrangements in the pro-B cell lymphomas are shown. The normal chromosomes appear to the far left (Chr. 12) and right (Chr. 15) of each box while the derivative chromosomes are illustrated between them. Arrows on the normal chromosomes indicate the location of the rearrangements on the derivative chromosomes.
Mentions: Sequence analysis and localization of fusion points and rearrangements in the IgH Ca and c-myc loci. (A) The fusion points of three Ku80−/−p53−/− pro-B cell lymphomas (PKT no. 2, 7, and 13) were cloned by digestion circularization PCR [der(15)a]. Sequence analysis of the three breakpoints from scenario A demonstrate microhomology (yellow) at the junction of chromosomes 12 (red) and 15 (green) in two cases (PKT2 and PKT7), but none in tumor PKT13. Homologous bases outside of this region are also indicated in uppercase letters. (B) Sequence analysis implied that the breakpoint was the V(D)J recombination signal sequence (RSS; blue) flanking IgH J1 in PKT2 and IgH J4 in PKT7 and PKT13 (top panel); however, exonuclease digestion of the cleaved ends resulted in loss of 45, 208, and 189 bp, respectively, from the original site of the RAG-mediated DSB to the fusion point (arrows). Approximate localization of the breakpoints on chromosome 15 (second panel) was determined by FISH hybridization with two BAC clones in this region (96F18 and 212H11) as illustrated in Fig. 5. Tumors 1–4, 7, and 13 are from Ku80−/−p53−/− mice, tumors *1–*3 are from Ku70−/−p53−/− mice and tumors N9984 and N10527 are from Ku80−/−p53−/−RAD54−/− mice. The resulting der(15)a and der(12)b chromosomes are illustrated in the bottom panels. In scenario A a portion of the IgH locus is copied to chromosome 15 downstream of c-myc and in the reverse orientation. In scenario B it appears that the copying of chromosome 15 to the der(12)b initiates centromeric of the c-myc locus and continues along toward the telomere.

Bottom Line: Subsequent DNA repair events juxtaposing IgH and c-myc are mediated by a break-induced replication pathway.Cycles of breakage-fusion-bridge result in amplification of IgH/c-myc while chromosome stabilization occurs through telomere capture.Thus, mice deficient in NHEJ provide excellent models to study the etiology of unbalanced translocations and amplification events during tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Genetics Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA. difilipm@mail.nih.gov

ABSTRACT
Nonreciprocal translocations and gene amplifications are commonly found in human tumors. Although little is known about the mechanisms leading to such aberrations, tissue culture models predict that they can arise from DNA breakage, followed by cycles of chromatid fusion, asymmetric mitotic breakage, and replication. Mice deficient in both a nonhomologous end joining (NHEJ) DNA repair protein and the p53 tumor suppressor develop lymphomas at an early age harboring amplification of an IgH/c-myc fusion. Here we report that these chromosomal rearrangements are initiated by a recombination activating gene (RAG)-induced DNA cleavage. Subsequent DNA repair events juxtaposing IgH and c-myc are mediated by a break-induced replication pathway. Cycles of breakage-fusion-bridge result in amplification of IgH/c-myc while chromosome stabilization occurs through telomere capture. Thus, mice deficient in NHEJ provide excellent models to study the etiology of unbalanced translocations and amplification events during tumorigenesis.

Show MeSH
Related in: MedlinePlus