Adaptive response to DNA and chromosomal damage induced by X-rays in human blood lymphocytes.

Nucleoid sedimentation, single-cell gel electrophoresis (comet assay) and premature chromosome condensation (PCC) technique were utilized to estimate the involvement of DNA strand breaks and chromosomal damage in radio-adaptive response of stimulated human lymphocytes. Conditioning of cells with 0.02 Gy X-rays rendered them more resistant to single- and double-strand DNA breaks produced by 1 Gy challenging treatment as revealed by the sedimentation behaviour of the nucleoids and the comet assay. Nucleoid sedimentation also demonstrated that adaptive reaction towards X-ray-induced DNA damage is favoured in the presence of oxygen. A concomitant decrease in the amount of interphase chromosomal breaks visualized by PCC under the same experimental conditions was observed. Data indicate that adaptation of human lymphocytes to X-rays is tightly linked to the reduced susceptibility towards generation of DNA and chromosomal breaks. It is proposed that the very persistence of DNA strand discontinuities might serve as a triggering signal for the adaptation of human lymphocytes against ionizing radiation exposure.

HaeIII induces position-dependent chromosomal breakage in barley (Hordeum vulgare L.).

The pattern of localized chromosomal breakage induced by the restriction endonuclease HaeIII in reconstructed barley karyotypes T-1586 and T-21 was investigated. It was found that nucleolus organizing regions (NORs) of chromosomes 6 and 7 (segments 46 and 38, respectively), containing actively transcribed ribosomal (r)DNA, as well as segments 39 and 47, both containing condensed rDNA repeats, are the most pronounced aberration hot-spots in T-1586. The number of aberrations observed in these segments was three to five times higher than theoretically expected. The intrachromosomal distribution of chromatid aberrations in karyotype T-21, where the NOR-bearing segments in chromosomes 6 and 7 change their position, revealed a substantial difference in the aberration hot-spot behaviour. A position-specific increase in aberration clustering was observed, most pronounced in segments 38 and 47. On the other hand, segment 46 retained its initial sensitivity, while segment 39 in the new position lost its previous status as a mutation hot-spot. The data are indicative of the expressivity of aberration hot-spots generated after treatment with this restriction endonuclease being influenced by their distinct chromosomal location.

Restriction endonucleases induce chromosomal aberrations in barley.

The clastogenic ability of the restriction endonucleases (MspI, HpaII and HaeIII) in germinating seeds of reconstructed barley karyotype was assessed. An effective induction of chromosomal aberrations after restrictase treatment was observed. The frequency, types and cell-cycle dependence of the observed abnormalities are discussed in relation to the distinct characteristics of the enzymes and the features of the plant genome. The capacity to induce aberrations was not significantly influenced by the nature of the double-strand breaks (blunt- or cohesive-ended); however, it was dependent on the methylation status of the plant DNA. The restriction enzymes displayed an S-independent mode of action revealing the transition between G1 and S as the most sensitive stage of the cell cycle in barley for induction of chromosomal damage.

Caffeine potentiates or protects against radiation-induced DNA and chromosomal damage in human lymphocytes depending on temperature and concentration.

The effect of caffeine on radiation-induced chromosomal aberrations and DNA strand breaks in unstimulated human lymphocytes was investigated. When present prior to and during the radiation exposure, caffeine treatment was found to cause either potentiation or protection against induction of chromosomal aberrations depending on the concentration and temperature. When the nucleoid sedimentation technique was applied, enhancement or reduction of radiation-induced DNA strand breaks by caffeine was also found to be dependent on temperature and caffeine concentration. It is proposed that caffeine, in addition to its suspected ability to influence DNA repair, can also influence the induction of DNA damage, leading to alterations in the yield of chromosomal aberrations.

Matrix-associated DNA from maize is enriched in repetitive sequences.

In order to elucidate some features of the topological organization of DNA within the plant nucleus, DNA fragments involved in the attachment of the DNA loops to the nuclear matrix in maize were studied. The matrix-associated DNA from dry embryo and meristematic cells after extensive digestion with DNase I and high salt treatment was about 2% of the total DNA, sized within the range of 50 and 250 bp. This DNA was found to be enriched in repetitive DNA sequences, both for nuclei from dry embryo and meristematic cells. The loop size of the DNA in cells of Zea mays appeared to be between 5 and 25 kbp.

Supercoils in plant DNA: nucleoid sedimentation studies.

Plant nuclei have been studied with respect to the three-dimensional structure of DNA. Nucleoids derived from nuclei by non-ionic detergent and high salt treatment were analysed by sedimentation in a series of sucrose gradients containing increasing amounts of the intercalating agent ethidium bromide. In addition the nucleoid sedimentation behaviour was investigated following gamma irradiation. The results show that plant DNA is supercoiled, as is the DNA from the other eukaryotes studied, and contains approximately the same concentration of superhelical turns but probably relatively fewer DNA superhelical loops. The plant nuclear populations in all cases studied give rise to two distinct nucleoid bands. These have been characterized by electron microscopy and by their DNA and protein content. The possible origin of the two bands is discussed.

DNA repair precedes replicative synthesis during early germination in maize.

DNA synthesis was studied during germination by following the rate of incorporation of radioactive thymidine into high molecular weight DNA. A peak of DNA synthesis was observed between the 8th and the 12th hour, i.e. before the beginning of the semi-conservative replication of genomic DNA, accompanied by an increase in the DNA content of the embryo. By the use of nucleoid sedimentation and nick-translation it was shown that, during the first hours of germination, extensive repair occurs of the DNA single-strand breaks present in the dry embryo. As a result, the DNA of the 16-h-germinated embryo acquires the conformation typical of that of the root meristemic cells active in transcription and replication.In addition we have shown that cytoplasmic organelle (most probably mitochondrial) DNA synthesis is very active during the prereplicative state which confirms earlier microscopic data on mitochondrial biogenesis during early germination.