Histone deacetylases (HDACs) are a growing family of enzymes implicated in transcriptional regulation by affecting the acetylation state of core histones in the nucleus of cells. HDACs are known to have key roles in the regulation of cell proliferation [Brehm, Miska, McCance, Reid, Bannister and Kouzarides (1998) Nature (London) 391, 597–600], and aberrant recruitment of an HDAC complex has been shown to be a key step in the mechanism of cell transformation in acute promyelocytic leukaemia [Grignani, De Matteis, Nervi, Tomassoni, Gelmetti, Cioce, Fanelli, Ruthardt, Ferrara, Zamir et al. (1998) Nature (London) 391, 815–818; Lin, Nagy, Inoue, Shao, Miller and Evans (1998), Nature (London) 391, 811–814]. Here we present the complete nucleotide sequence of a cDNA clone, termed HDAC8, that encodes a protein product with similarity to the RPD3 class (I) of HDACs. The predicted 377-residue HDAC8 product contains a shorter C-terminal extension relative to other members of its class. After expression in two cell systems, immunopurified HDAC8 is shown to possess trichostatin A- and sodium butyrate-inhibitable HDAC activity on histone H4 peptide substrates as well as on core histones. Expression profiling reveals the expression of HDAC8 to various degrees in every tissue tested and also in several tumour lines. Mutation of two adjacent histidine residues within the predicted active site severely decreases activity, confirming these residues as important for HDAC8 enzyme activity. Finally, linkage analysis after radiation hybrid mapping has localized HDAC8 to chromosomal position Xq21.2–Xq21.3. These results confirm HDAC8 as a new member of the HDAC family.