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Figure 5


Fig. 5. Reduced lamin B1 expression correlates with altered location of chromosome territories. CTs were analysed in lamin-B1-depleted cells using: (A) FISH to chromosome 19 (wide-field imaging); (B) BrdU, by indirect immunofluorescence (high-resolution 3D imaging); (C) Alexa Fluor 488 dUTP, in living cells (confocal sections). (B,C) Unsynchronised cells were labelled and grown for 5-7 days so that most labelled cells retained 1-3 discrete, labelled territories. RNAi vectors were transfected 48 hours before analysis. CTs were assigned to nuclear zones of equal area (Materials and Methods) using at least 100 territories/sample (A-C). Changes were analysed using two-tailed Student's t test as follows: (A) control vs lamin A/C depletion (t=2.396, P<0.0172*); control vs lamin B1 depletion (t=9.203. P<1.567e-17***); (B) control vs lamin A/C depletion (t=0.478, P<0.634); control vs lamin B1 depletion (t=5.780, P<2.973e-8***); (C) control vs lamin B1 depletion (t=6.442, P<1.996e-09***). Marginalisation of CTs (D, left panel, shows a single confocal section of CTs labelled with BrdU in green) correlated with collapse of open chromatin (D, centre panel, DsRed-histone H2B in red) and a corresponding expansion of the inter-chromatin compartment. Scale bars: 10 µm (A); 5 µm (C,D).





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