Spatial Organization, Dynamics, and Associations of Telomeresin Healthy, Telomerase-Positive and ALT Human Cell Lines
2007
The main goal of this study was to compare the telomere
localizations, dynamics, and occurrences of telomere
associations (TAs) in healthy cells, in tumor cells with active
telomerase, and in tumor cells with mechanism of alternative
lengthening of telomeres (ALT). These objectives were pursued
by time-lapse microscopy of living cells and by visualizing
telomeres with subtelomere DNA probes in repeated double-color
fluorescence in situ hybridization (FISH) on 2D fixed nuclei.
Samples were observed by high-resolution cytometry, and
followed by image and statistical analysis of acquired data. As
models for in situ experiments were chosen following lung
tissue cell lines: cell line WI-38 (healthy lung fibroblasts),
tumor cell line HT-1080 with active telomerase, and cell line
WI-38 VA-13/2RA (VA-13) with ALT mechanism. For these studies
were chosen telomeres of all acrocentric chromosomes (13q, 14q,
15q, 21q a 22q) and also p- and q-telomeres of similarly sized
chromosomes 18 a 19, that differ in their localization in cell
nucleus and in content of transcriptionaly active genes. As
models for living-cells experiments were chosen following cell
lines: U-2 OS (osteosarcoma ALT cell line) and HeLa (cervical
carcinoma cell line with active telomerase). Distributions of
radial distances of all in situ analyzed telomeres for all
assessed cell lines are comparable in most cases with only
several exceptions. Among these exceptions belongs shift of
localization of telomere of chromosome 13 towards nuclear
periphery in cell line VA-13 or localization of telomere of
chromosome 15 in cell line HT-1080, which radial distribution
is significantly shifted into the interior of cell nucleus in
comparison to other cell lines. All telomeres of chromosomes 18
and 19 were found to exhibit higher occurrence of telomere
associations in cells with ALT mechanism in comparison to
health cells or cells with active telomerase as was expected.
This conclusion was not found to be valid for telomeres of
acrocentric chromosomes, probably due to influence of binding
of these chromosomes to nucleolus. After calculation of average
occurrence of associations of certain telomere with other
telomeres of acrocentric chromosomes was found that these
average occurrences are the highest in tumor cells HT-1080 with
active telomerase. We also analyzed and compared in unsurpassed
detail the telomere dynamics in living U-2 OS and HeLa cell
lines varying in mechanism of telomere length maintenance in 3D
space by time-lapse fluorescent confocal microscopy.
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