J.M. Bridger, S. Boyle, I.R. Kill, W.A. Bickmore Current Biology

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Re-modelling of nuclear architecture in quiescent and senescent human fibroblasts J.M. Bridger, S. Boyle, I.R. Kill, W.A. Bickmore Current Biology Volume 10, Issue 3, Pages 149-152 (February 2000) DOI: 10.1016/S0960-9822(00)00312-2

Figure 1 Chromosome position in proliferating, quiescent and senescent fibroblasts. (a–c) Mean percentage hybridisation signal, normalised to the percentage of the DAPI signal, of HSA18 (grey) and HSA19 (white) in each shell of nuclei from (a) proliferating 1HD fibroblasts, (b) G0 (quiescent or serum starved) 1HD fibroblasts, and (c) senescent (passage 38) 2DD fibroblasts. The data were analysed by erosion [2] of five concentric shells of 50 nuclei; shells were numbered 1–5 from the edge to the centre. Error bars show the SEM. (d–i) Representative images showing the position of HSA18 and 19 territories (green) in DAPI-stained (blue) nuclei. The scale bar represents 2 μm. Current Biology 2000 10, 149-152DOI: (10.1016/S0960-9822(00)00312-2)

Figure 2 Three-dimensional analysis of nuclear positioning in proliferating, quiescent and senescent cells. Spatial positioning of HSA18 and 19 (marked with a chromosome paint detected with fluorescein-isothiocyanate, green) relative to (a–c) the nuclear periphery (as defined by propidium iodide staining, red), or (e–g) the nucleolus, delineated by either anti-pKi-67 or anti-fibrillarin staining (red), in 3D preparations of (a,e) proliferating, (b,f) quiescent and (c,g) senescent fibroblasts. Slides were sectioned at 1 μm intervals by CLSM, but only confocal midsections are shown here. (d) Measurements were made of the mean distance (in μm ± SEM) from the centre of chromosome territories to the closest edges of the nucleus in the x (apical), y (lateral) and z (top–bottom) planes (n=20) . The scale bar represents 2 μm. Current Biology 2000 10, 149-152DOI: (10.1016/S0960-9822(00)00312-2)

Figure 3 Dynamics of chromosome re-positioning after serum stimulation of G0 cells. Mean percentage hybridisation signals, normalised to DAPI staining, of HSA18 (grey) and 19 (white) in nuclei of (a) proliferating 1HD fibroblasts, 1HD fibroblasts in G0, and 1HD cells at 8, 24 and 32 h after serum stimulation and (b) 1HD fibroblasts in G1 36 h after re-stimulation. Cell cycle stage was judged by BrdU incorporation and the mitotic index, and only BrdU-positive cells were analysed at 24 and 32 h. The data were analysed by erosion [2] of five concentric shells of 50 nuclei; shells were numbered 1–5 from the edge to the centre. Error bars show the SEM. In (b) the stage of G1 (type Ia, early; type Ib, early/mid; or type II, mid/late) was assessed from the pKi-67 staining pattern [6]. (c) Representative images of HSA18 or 19 territories (green) in DAPI-stained (blue) nuclei of cells with a type Ia, Ib, or II distribution of pKi-67 (red). The scale bar represents 2 μm. Current Biology 2000 10, 149-152DOI: (10.1016/S0960-9822(00)00312-2)