Figure 3.

Colocalization of HDAC4 and 5 with ANKRA2. (A) COS cells were transfected with an expression vector encoding Myctagged ANKRA2 in the absence or presence of vectors for FLAG-tagged HDAC5 or HDAC4. As indicated, some cells also received a plasmid encoding constitutively active CaMKI. The subcellular localization of ANKRA2 and HDACs was determined by indirect immunofluorescence (red, ANKRA2; green, HDAC4 or 5). (B) Effects of HDAC5 and CaMKI overexpression on ANKRA2 subcellular distribution were quantified by microscopic examination of greater than 100 cells per condition. N, exclusive staining of ANKRA2 in the nucleus; N>C, nuclear ANKRA2 staining greater than cytoplasmic staining; C≥N, cytoplasmic ANKRA2 staining greater than or equal to nuclear staining; C, exclusive staining of ANRKA2 in the cytoplasm. (C) COS cells were cotransfected with expression vectors for ANKRA2 and HDAC5 in the absence or presence of a plasmid encoding activated CaMKI. Forty-eight hours after transfection, cells were exposed to leptomycin B (LMB; 10 nM) for 2 h. The subcellular localization of ANKRA2 and HDAC5 was determined by indirect immunofluorescence.