Spatial and temporal dynamics of DNA replication sites in mammalian cells

被引:246
作者
Ma, H
Samarabandu, J
Devdhar, RS
Acharya, R
Cheng, PC
Meng, CL
Berezney, R [1 ]
机构
[1] SUNY Buffalo, Dept Biol Sci, Buffalo, NY 14260 USA
[2] SUNY Buffalo, Dept Comp Sci & Engn, Buffalo, NY 14260 USA
[3] SUNY Buffalo, Dept Elect Engn, Buffalo, NY 14260 USA
关键词
replication sites; replication timing; cell nucleus; chromosomes; computer image segmentation;
D O I
10.1083/jcb.143.6.1415
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
Fluorescence microscopic analysis of newly replicated DNA has revealed discrete granular sites of replication (RS). The average size and number of replication sites from early to mid S-phase suggest that each RS contains numerous replicons clustered together. We are using fluorescence laser scanning confocal microscopy in conjunction with multidimensional image analysis to gain more precise information about RS and their spatial-temporal dynamics. Using a newly improved imaging segmentation program, we report an average of similar to 1,100 RS after a 5-min pulse labeling of 3T3 mouse fibroblast cells in early 5-phase. Pulse-chase-pulse double labeling experiments reveal that RS take similar to 45 min to complete replication. Appropriate calculations suggest that each RS contains an average of 1 mbp of DNA or similar to 6 average-sized replicons. Double pulse-double chase experiments demonstrate that the DNA sequences replicated at individual RS are precisely maintained temporally and spatially as the cell progresses through the cell cycle and into subsequent generations. By labeling replicated DNA at the G(1)/S borders for two consecutive cell generations, we show that the DNA synthesized at early S-phase is replicated at the same time and sites in the next round of replication.
引用
收藏
页码:1415 / 1425
页数:11
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