DNA double-strand breaks (DSB) are considered to be critical primary lesions in the formation of chromosomal aberrations. DSB may be induced by exogenous agents, such as ionizing radiation, but also occur spontaneously during cellular processes at quite significant frequencies. To repair this potentially lethal damage, eukaryotic cells have evolved a variety of repair pathways related to homologous and illegitimate recombination, also called non-homologous DNA end joining, which may induce small scale mutations and chromosomal aberrations. In this paper we review the major cellular sources of spontaneous DSB and the different homologous and illegitimate recombination repair pathways, with particular focus on their potential to induce chromosomal aberrations.

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Mechanisms of DNA double-strand break repair and their potential to induce chromosomal aberrations.

This paper presents an analysis of the utility of fluorescence in situ hybridization (FISH) with whole-chromosome probes for measurement of the genomic frequency of translocations found in the peripheral blood of individuals exposed to ionizing radiation. First, we derive the equation: Fp = 2.05fp(1-fp)FG, relating the translocation frequency, Fp, measured using FISH to the genomic translocation frequency, FG, where fp is the fraction of the genome covered by the composite probe. We demonstrate the validity of this equation by showing that: (a) translocation detection efficiency predicted by the equation is consistent with experimental data as fp is changed; (b) translocation frequency dose-response curves measured in vitro using FISH agree well with dicentric frequency dose-response curves measured in vitro using conventional cytogenetic procedures; and (c) the genomic translocation frequencies estimated from FISH measurements for 20 Hiroshima A-bomb survivors and four workers exposed to ionizing radiation during the Y-12 criticality accident are approximately the same as the translocation frequencies measured using G-banding. We also show that translocation frequency dose response curves estimated using FISH are similar for Hiroshima A-bomb survivors and for first division lymphocytes irradiated in vitro. We conclude with a discussion of the potential utility of translocation frequency analysis for assessment of the level of acute radiation exposure independent of the time between analysis and exposure.

Rapid translocation frequency analysis in humans decades after exposure to ionizing radiation.

Fluorescence in situ hybridization: applications in cytogenetics and gene mapping

This paper describes the four cytogenetic endpoints most frequently used in hazard identification assays as the first step in the risk assessment process. These are structural chromosome aberrations, micronuclei, aneuploidy, and sister chromatid exchanges. The biological mechanisms involved in the formation of the alterations observed in each assay are briefly discussed. Variations in and recent improvements to each assay are described, with an emphasis on the use of molecular techniques to improve the sensitivity of the assay, and to allow for detection of specific alterations that are, or could be, associated with cancer induction. This, in turn, will make the data obtained in the cytogenetic assays more useful in cancer and genetic risk assessment. Thus, the aim of this paper is to encourage cytogeneticists to design their experiments in such a way that the data obtained will be of maximum possible benefit for characterizing and quantifying adverse human health effects, particularly cancer.

Chromosome aberrations, micronuclei, aneuploidy, sister chromatid exchanges, and cancer risk assessment.

Chromosomal aberrations (CA) are the microscopically visible part of a wide spectrum of DNA changes generated by different repair mechanisms of DNA double strand breaks (DSB). The method of fluorescence in situ hybridisation (FISH) has uncovered unexpected complexities of CA and this will lead to changes in our thinking about the origin of CA. The inter- and intrachromosomal distribution of breakpoints is generally not random. CA breakpoints occur preferentially in active chromatin. Deviations from expected interchromosomal distributions of breakpoints may result from the arrangement of chromosomes in the interphase nucleus and/or from different sensitivities of chromosomes with respect to the formation of CA. Telomeres and interstitial telomere repeat like sequences play an important role in the formation of CA. Subtelomeric regions are hot spots for the formation of symmetrical exchanges between homologous chromatids and cryptic aberrations in these regions are associated with human congenital abnormalities.

Chromosomal aberrations: formation, identification and distribution

SummaryWe have used in situ hybridization of repeat-sequence DNA probes, specific to the paracentromeric locus 1q12 and the telomeric locus 1p36, to fluorescently stain regions that flank human chromosome 1p. This procedure was used for fast detection of structural aberrations involving human chromosome 1p in two separate experiments. In one, human lymphocytes were irradiated with 0, 0·8, 1·6, 2·4 and 3·2 Gy of 137Cs γ-rays. In the other, human lymphocytes were irradiated with 0, 0·09, 0·18, 2·0, 3·1 and 4·1 Gy of 60Co γ-rays. The frequencies (per cell) of translocations and dicentrics with one breakpoint in 1p and one elsewhere in the genome were determined for cells irradiated at each dose point. These frequencies both increased with dose, D, in a linear-quadratic manner. The δ, α, and β coefficients resulting from a fit of the equation f(D) = δ + αD + βD2 to the translocation frequency dose-response data were 0·0025, 0·0027 and 0·0037 for 137Cs γ-rays, and 0·0010, 0·0041, and 0·0057 for 60Co γ-rays. Th...

D. Pinkel

Papers

Rapid human chromosome aberration analysis using fluorescence in situ hybridization.