Search

The chromosome-type exchange aberrations induced by ionizing radiation during the G 0 /G 1 phase of the cell cycle are believed to be the result of illegitimate rejoining of chromosome breaks. From numerous studies using chromosome painting, it has emerged that even after a moderate dose of radiation, a substantial fraction of these exchanges is complex. Most of them are derived from the free interaction between the ends of three or more breaks. Other studies have demonstrated that chromosomes occupy distinct territories in the interphase nucleus. Since breaks that are in close proximity have an enhanced interaction probability, it seems likely that after ionizing radiation many of the interacting breaks will be present within one chromosome or chromosome arm. Unfortunately, the majority of these intrachanges remain undetected, even when sophisticated molecular cytogenetic detection methods (i.e. mFISH) are applied to paint all chromosome pairs in distinct colors. In the present paper, we evaluate the limitations of full-color painting for the detection of complex exchanges and the correct interpretations of break interactions.

The chromosome-type exchange aberrations induced by ionizing radiation during the G 0 /G 1 phase of the cell cycle are believed to be the result of illegitimate rejoining of chromosome breaks. From numerous studies using chromosome painting, it has emerged that even after a moderate dose of radiation, a substantial fraction of these exchanges is complex. Most of them are derived from the free interaction between the ends of three or more breaks. Other studies have demonstrated that chromosomes occupy distinct territories in the interphase nucleus. Since breaks that are in close proximity have an enhanced interaction probability, it seems likely that after ionizing radiation many of the interacting breaks will be present within one chromosome or chromosome arm. Unfortunately, the majority of these intrachanges remain undetected, even when sophisticated molecular cytogenetic detection methods (i.e. mFISH) are applied to paint all chromosome pairs in distinct colors. In the present paper, we evaluate the limitations of full-color painting for the detection of complex exchanges and the correct interpretations of break interactions.

Euchromatic and heterochromatic regions are easily distinguished in Chinese hamster sex chromosomes, hence offering the possibility of studying the role of chromatin structure in the induction, processing and persistence of radiation-induced chromosome damage. X-ray (4Gy)-induced breaks in the euchromatic Xp and in the heterochromatic Xq were analysed immediately and 4h after irradiation by premature chromosome condensation (PCC) in combination with either FISH using chromosome arm-specific probes or Giemsa staining. The study, performed with female Chinese hamster splenocytes, was extended to a 34h recovery followed by arm-specific FISH in metaphase. A significant over-involvement of the heterochromatic Xq in radiation-induced breakage was observed at all sampling times (p<0.001). However, the heterochromatic state had little effect on the processing of the induced lesions. In a second experiment, the persistence of radiation-induced chromosome aberrations (CAs) involving Xp, Xq and Y chromosome was studied with cultured Chinese hamster male splenocytes sampled 30, 56 and 96h after irradiation (4Gy). A higher involvement of the heterochromatic regions (Xq and Y) in radiation-induced CAs was again observed in the first sampling time (p<0.001), suggesting that Chinese hamster heterochromatin could be more radiosensitive than euchromatin. Cells with CAs involving heterochromatin were apparently less persistent than those with lesions involving euchromatin. This observation could be attributable to either the distribution of CA per cell or to the fraction of potentially stable exchanges.

Ionizing radiation and restriction endonucleases are very efficient in inducing chromosomal aberrations (CAs). These aberrations are mainly consequences of misrepair of DNA double-strand breaks (DSBs). The fast repairing component of DSBs induced by ionizing radiation seems to be responsible for exchange aberration. Use of premature chromosome condensation technique in combination with DNA repair inhibitors such as ara A has given valuable information on the assessment of the frequencies of initial chromosome breaks and the kinetics of their repair following low LET radiation. The recently developed ‘‘chromosome painting’’ technique using chromosome-specific libraries has also increased considerably the resolution of identifying and scoring of CAs. After low LET radiation, stable chromosome exchanges (translocations) are induced more frequently than unstable chromosome exchanges (dicentrics). Fluorescence in situhybridization employing telomeric probe has made it possible to score efficiently exchange aberrations involving the acrocentric chromosomes of mouse. Chinese hamster cells have several intercalary telomeric sequences present in most of the chromosomes. These telomeric blocks have been found to be associated with chromosomal aberrations induced by restriction endonucleases and short wave UV and evidence has been obtained for apparent amplification of telomeric sequences at the break points.

Frequencies of chromosomal aberrations were estimated in X-irradiated (0; 0.5; 1; 2; 3 and 4 Gy) human lymphocytes using arm-specific probes for chromosome 1 in combination with a pancentromeric probe in a triple colour FISH procedure. This allowed the construction of dose-response curves for both interchanges as well as interarm intrachanges. We found that 11.7-18.4% of aberrant chromosomes contained interarm intrachanges. Assuming a free interaction of randomly induced exchange breakpoints throughout the whole genome, interarm intrachanges occur about 8.7 times more frequently than expected. The close proximity of the two arms of a chromosome in the interphase nucleus appears to promote the formation of interarm intrachanges. Convincing evidence was obtained, by comparing the frequency of colour junctions that occur between 1p-1q and 1p-3q (the arms 1q and 3q are of similar size). With this approach, we observed 8.8 time more colour junctions between 1p-1q than between 1p-3q, illustrating the importance of proximity in the formation of chromosomal exchange aberrations. At doses higher than 2 Gy, a saturation of simple reciprocal aberrations, both for interchanges as well as for interarm intrachanges was observed. Furthermore, we found that symmetrical and assymmetrical simple interchanges as well as interarm intrachanges occur with equal frequencies in human chromosome 1.