The position of centromeres in interphase nuclei of human leukemic cells during myeloid differentiation and after gamma-irradiation
| Authors | |
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| Year of publication | 1999 |
| Type | Article in Proceedings |
| Conference | Fluorescence microscopy and fluorescent probes |
| MU Faculty or unit | |
| Citation | |
| Field | Biophysics |
| Keywords | centromeres |
| Description | Centromeres are frequently associated with heterochromatin and other chromatin components that cause the epigenetic transcriptional repression of genes. Heterochromatin is often found in close association with the nuclear periphery and tends to replicate late in the S phase of the cell cycle. Centromere-induced gene silencing also interferes with chromatin segregation, proving a critical link between chromatin structure and centromere function. The aim of this study is to analyse the spatial organisation of centromeres by means of a newly developed automated fluorescence microscopy system (Brno high resolution cytometer) in combination with Fast-Fluorescence in situ Hybridisation (FAST-FISH). The latter has been recently developed as an easy to handle technique especially for centromere labelling. In the experiments the positions of some human centromeres during myeloid differentiation and after gamma-irradiation were detected and analysed. Methodologically, the results indicate that due to high reproducibility and signal intensity, FAST-FISH was useful for automated microscopy and image analysis by the Brno high resolution cytometer. The human leukemic cell line U937 blocked on the level of promyelocytes was differentiated in granulocytes with all-trans retinoic acid (ATRA) and dimethyl sulfoxide (DMSO) and then into monocytes-macrophages with phorbol esters (PMA). The U937 cells were also irradiated with 5 Gy gamma rays (60Co) to determine the topology of some centromeres after radiation treatment. The positions of the centromeres of the human chromosomes 8, 9, 12, 17, 18 and X were determined and compared to the untreated control. The proximity of homologous and heterologous centromeres was detected. Generally, during granulocytic differentiation the positions of centromeres did not change significantly, but in monocytic differentiation some centromeres were closer to each other and closer to the nuclear centre. Changes in the nuclear topology of human centromeres were found after gamma irradiation, the centromeres were repositioned closer to the nuclear centre (repositioning about 9-19% of nuclear radius). |
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