Figure 7.

Heterochromatin protein 1 accumulation at centromeres is impaired in cells briefly exposed to TSA. (A) Immunofluorescence detection of HP1α and kinetochore proteins by using 2HP 1H5 antibody and CREST serum on PtK1 metaphase cells from cultures treated for 7 h with 0.1% DMSO (–TSA) or 500 ng/ml TSA (+TSA). DNA was counterstained using DAPI (blue). The right column shows an overlay of CREST, HP1, and DAPI staining. HP1 is detected at centromeres in a region wider than the one defined by the CREST antibody in the control cell (top, middle panel). In the TSA-treated cell the centromeric accumulation is lost (bottom, middle panel). Bar, 5 μm. (B) Quantitative analysis of HP1 fluorescence on MRC-5 and PtK1 cells treated for 7 h with 0.1% DMSO (red bars) or receiving 500 ng/ml TSA for 1 h (white bars) or 7 h (gray bars). The values shown are the mean + SE of kinetochore fluorescence intensities (obtained as described in MATERIALS AND METHODS). Numbers above bars represent numbers of kinetochores measured per each experimental condition. p < 0.001 when TSA-treated samples were compared with their respective controls by the Student's t test.