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Journal of Cell Science, Vol 111, Issue 22 3275-3285, Copyright © 1998 by Company of Biologists
JOURNAL ARTICLES |
N Itoh and N Shimizu
Faculty of Integrated Arts and Sciences, Hiroshima University, Higashi-hiroshima, Japan.
Double minutes (DMs) seen in a substantial fraction of human tumors are the cytogenetic manifestation of gene amplification which renders the tumor cells advantageous for growth and survival. DMs are acentric and atelomeric chromatin composed of circular DNA. In this study, we found they showed a remarkable relocation inside the nucleus which was spatially and temporally coupled to DNA replication. Using the human COLO 320DM tumor line, we detected DMs by fluorescence in situ hybridization followed by confocal examination. The location of multi-copy DMs was evaluated statistically by an easy method developed in this study. By examination of a synchronized culture, we found that DMs preferentially located at the nuclear periphery during G1-phase of the cell cycle, which is consistent with the location at M-phase. The peripheral DMs were in contact with the nuclear lamina which was shown by the simultaneous detection of DMs and lamin protein. The peripheral location persisted until the cells reached the G1/S-boundary, then the DMs relocated promptly to inward once the DNA replication started. The relocation was obvious using two different probes that detect DMs, or using two different methods for the cell fixation. Furthermore, the simultaneous detection of DMs and the site of DNA replication suggested that the inward relocation of peripheral DMs initiated just prior to the onset of DNA replication at the periphery. On the other hand, if the same amplified sequences were placed in a chromosome as an homogeneously staining region, they did not show any significant relocation during S-phase. From these and reported results, there may exist a generalized inward motion of some kind of chromatin that precedes the replication of their DNA. DMs might magnify the motion by their acentric, atelomeric or small circular nature.
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