Class IIa histone deacetylases (HDACs) form complexes with a class of transcriptional repressors in the nucleus. While screening for compounds that could block the association of HDAC4 with the BTB domain-containing transcriptional repressor Bach2. we discovered that phorbol 12-myristate 13-acetate (PMA) induced the cytoplasmic retention of HDAC4 mutants lacking a nuclear export signal (NES). Although PMA treatment and PKD overexpression has been proposed to facilitate the nuclear export Of class IIa HDACs by Creating 14-3-3 binding sites containing phosphoserines, our experiments using HDAC Mutants demonstrated that PMA greatly reduces nuclear import. PMA treatment repressed the NLS activity in a manner dependent on 14-3-3 binding. These results suggest that nuclear HDAC4 is not tethered in the nucleus, but instead shuttles between the nucleus and the cytoplasm. Phosphorylation-induced 14-3-3 binding biases the balance Of nucleo-cytoplasmic shuttling toward the cytoplasm by inhibiting nuclear import. (C) 2008 Elsevier Inc. All rights reserved.