Fig. 8. Orc1p* focalization survives TSA and RNase treatment. (A) NIH-3T3 cells were treated with 0, 0.1 and 0.5 µg/ml TSA for 6 hours. Fixed cells were stained with the anti-HP1 antibody. Heterochromatin was stained with DAPI. Dispersal of HP1 from DAPI foci is already detectable at 0.1 µg/ml TSA. (B) Cells expressing Orc1-Flag were incubated in the presence of 0.5 µg/ml TSA as in panel A. Fixed cells were co-stained with the anti-Flag and anti-HP1 antibodies. Heterochromatin was stained with DAPI. In contrast to HP1, Orc1-Flag still colocalizes with DAPI foci. (C) Cells expressing Orc1-Flag were incubated with RNase A as described in Materials and Methods. Orc1-Flag was visualized with the anti-Flag antibody; heterochromatin was stained with DAPI visualized in green for a better resolution of the merged image. Confocal laser images are shown. Bar, 10 µm.