Depletion of p150CAF-1 in ES Cells Results in a Severe Alteration of Heterochromatin Organization
(A) Strategy used to deplete p150CAF-1 by RNAi in ES cells. The siRNA expression vector includes a puromycin selection cassette (Puro), the mouse H1 promoter, and the siRNA encoding sequence. ES cells were kept under puromycin selection during 48 h following transfection.
(B) Abnormal heterochromatin organization in p150CAF-1-depleted ES cells. Immunodetection of p150CAF-1 (green) and HP1α (red) in ES cells transfected with control (cont) and p150CAF-1 siRNA. siRNA expression results in efficient p150CAF-1 depletion. The right-hand image shows the merge between HP1α fluorescence and DAPI-stained DNA in blue. Scale bar = 10 μm.
(C) Heterochromatin organization is not altered in p150CAF-1-depleted MEFs. Immunodetection of p150CAF-1 (green) and HP1α (red) in MEFs transfected with control (cont) or p150CAF-1 siRNA. The right-hand image shows the merge between HP1α fluorescence and DAPI-stained DNA in blue.
(D) PML bodies are not altered in p150CAF-1-depleted ES cells. Immunodetection of PML (red) in ES cells transfected with control (cont) or p150CAF-1 siRNA. The right-hand image shows the merge between PML fluorescence and DAPI-stained DNA in blue.