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M Kozubek

Publications and source records attributed to M Kozubek.

22 records · Page 2Linked to original sources
PubMed1997

Distribution of ABL and BCR genes in cell nuclei of normal and irradiated lymphocytes.

Using dual-color fluorescence in situ hybridization (FISH) combined with two-dimensional (2D) image analysis, the locations of ABL and BCR genes in cell nuclei were studied. The center of nucleus-to-gene and mutual distances of ABL and BCR genes in interphase nuclei of nonstimulated and stimulated lymphocytes as well as in lymphocytes stimulated after irradiation were determined. We found that, after stimulation, the ABL and BCR genes move towards the membrane, their mutual distances increase, and the shortest distance between heterologous ABL and BCR genes increases. The distribution of the shortest distances between ABL and BCR genes in the G0 phase of lymphocytes corresponds to the theoretical distribution calculated by the Monte-Carlo simulation. Interestingly, the shortest ABL-BCR distances in G1 and S(G2) nuclei are greater in experiment as compared with theory. This result suggests the existence of a certain regularity in the gene arrangement in the G1 and S(G2) nuclei that keeps ABL and BCR genes at longer than random distances. On the other hand, in about 2% to 8% of lymphocytes, the ABL and BCR genes are very close to each other (the distance is less than approximately 0.2 to 0.3 microm). For comparison, we studied another pair of genes, c-MYC and IgH, that are critical for the induction of t(8;14) translocation that occurs in the Burkitt's lymphoma. We found that in about 8% of lymphocytes, c-MYC and IgH are very close to each other. Similar results were obtained for human fibroblasts. gamma-Radiation leads to substantial changes in the chromatin structure of stimulated lymphocytes: ABL and BCR genes are shifted to the nuclear center, and mutual ABL-BCR distances become much shorter in the G1 and S(G2) nuclei. Therefore, we hypothesize that the changes of chromatin structure in the irradiated lymphocytes might increase the probability of a translocation during G1 and S(G2) stages of the cell cycle. The fact that the genes involved in the t(8;14) translocation are also located close together in a certain fraction of cells substantiates the hypothesis that physical distance plays an important role in the processes leading to the translocations that are responsible for oncogenic transformation of cells.

Cell Cycle↗
PubMed1997

Localisation and distance between ABL and BCR genes in interphase nuclei of bone marrow cells of control donors and patients with chronic myeloid leukaemia.

Quantitative measurements of the nuclear localisation of the ABL and BCR genes and the distance between them were performed in randomly oriented bone marrow cells of control donors and patients with chronic myeloid leukaemia (CML). Most ABL and BCR genes (75%) are located at a distance of 20-65% of the local radius from the nuclear centre to the nuclear membrane. A chimeric BCR-ABL gene located on a derivative chromosome 22 resulting from t(9;22)(q34;q11) [the so-called Philadelphia (Ph) chromosome] as well as the intact ABL and BCR genes of patients suffering from chronic myeloid leukaemia are also located mostly in this region, which has a mean thickness of 2 microns in bone marrow cells. We have not found any significant differences in the location of the two genes in the G1 and G2 phases of the cell cycle, nor between bone marrow cells and stimulated lymphocytes. Irradiation of lymphocytes with a dose of 5 Gy of gamma-rays results in a shift of both genes to the central region of the nucleus (0-20% of the radius distant from the nuclear centre) in about 15% of the cells. The minimum distance between one ABL and one BCR gene is less than 1 micron in 47.5% of bone marrow cells of control donors. Such a small distance is found between homologous ABL and between homologous BCR genes in only 8.1% and 8.4% of cells, respectively. It is possible that the relative closeness of nonhomologous ABL and BCR genes in interphase nuclei of bone marrow cells could facilitate translocation between these genes. In 16.4% of bone marrow cells one ABL and one BCR gene are juxtaposed (the distance between them varies from 0-0.5 micron) and simulate the Ph chromosome. This juxtaposition is the result of the projection of two genes located one above another into a plane, as follows from the probability calculation.

Adult↗
PubMed1996

Efficient real-time confocal microscopy with white light sources.

The main advantage of confocal microscopes over their conventional counterparts arises from their ability to optically 'section' nearly transparent materials; the thin image slices thus obtained can be used to reconstruct three-dimensional images, a capability which is particularly useful for the study of biological specimens. Confocal microscopes have previously used either a single laser-illuminated point-source and single point-detector (which are scanned in tandem across the object) or white-light illumination with multiple point-sources and detectors. Single-point-source systems, however, do not usually form images in real time and are restricted to using available laser wavelengths. Multiple-point-source systems, on the other hand, produce images in real time but use light very inefficiently--typically 1% or less is used for imaging. Here we demonstrate a white-light, multiple-point-source method which can in principle produce images in real time with light efficiencies as high as 50%. This system is likely to find broad practical application, particularly in the imaging of weakly reflecting or weakly fluorescent specimens.

Animals↗