Figure 4
Replication timing profile of human telomeres is conserved between individuals and during immortalization.
(A) ReD-FISH on TP metaphases. This hybridization uses two different telomere probes (CCCTAA-Cy3, TTAGGG-FITC). The chromosome in the white box was enlarged as an example. It has a detargeted telomere on the p-arm, (one probe per chromatid) and a non-detargeted telomere on the q-arm, (the two probes per chromatid), signifying that the telomere of the p-arm was replicated after BrdU addition unlike the telomere of the q-arm which was replicated before. (B) Replication timing of every single telomeres in TP cells. Horizontal bars represent the percentage of replication during each of four periods (Early-S, Early-middle-S, Late-middle-S and Late-S) and are normalized to 100%. Single telomeres are listed in ascending order of mrt (right). (C) Significant correlation between mrt of single telomeres in TP cells and IMR90 cells (left) and HCA2T cells (right), indicating the global conservation of replication timing of human telomeres. Mrt values of IMR90 and HCA2T correspond to those previously published14 (Spearman’s rank correlation coefficient). (D) Summary of telomere classification as a function of their replication. Telomeres were classified into six categories, represented by the color code, according to their mrt (in hour). As the length of the different sub-phases was not constant during immortalization, the categories were not equal between the different population doublings. (E) Pattern of single telomere replication during immortalization (TP as a control and PD64, 263 and 523 of TP15.5). The single telomeres studied for this study are those of both arms of chromosomes 1, 2, 3, 8, 9, 13, 16, 19 and 21. Telomeres are classified according their mrt as described in D. The percentage change of replication timing in each PD compared to the control TP is indicated below. (F) Significant correlation between the mrt of single telomeres in TP and of those of the different PD studied of TP 15.5 (PD64, PD263 and PD523 in red, green and purple respectively) for the 18 telomeres studied indicating that the relative order of telomere replication timing is not modified during immortalization (Spearman’s rank correlation coefficient).
