PTEN is a lipid and protein phosphatase that, by dephosphorylating phosphatidylinositol 3,4,5-trisphosphate, decreases signaling through the phosphoinositide 3-kinase pathway, which activates the kinase Akt. In a screen for proteins that interacted with PTEN, Okumura et al. identified the histone acetyltransferase PCAF (p300/CBP-associated factor), and this interaction was confirmed by coimmunoprecipitation of endogenous proteins from 293T cells. Although most of PTEN is cytosolic, deconvolution microscopy of cells transfected with tagged versions of PTEN and PCAF showed that when coexpressed, a population of PTEN was present in the nucleus, which is the location of PCAF. A PTEN mutant that failed to interact with PCAF did not appear in the nucleus when coexpressed with tagged PCAF. Using antibodies that recognized acetyl-lysine residues, acetylated endogenous PTEN was detected, and when histone deacetylases were inhibited pharmacologically or PCAF was overexpressed, acetylation of PTEN increased. PCAF directly acetylated PTEN in an in vitro assay, and this was prevented if lysines 125 and 128 of PTEN were mutated. Acetylation of PTEN inhibited its lipid phosphatase activity in an in vitro assay. Akt phosphorylation was increased in cells coexpressing PTEN and PCAF, and this was not observed when the PTEN acetylation-mimicking mutants were used. Pten–/– mouse embryo fibroblasts in which PTEN or PTEN mutants were expressed with or without PCAF showed that expression of PTEN caused G1 cell cycle arrest and suggested that acetylation of PTEN by PCAF alleviated this cell cycle arrest. Growth factors or serum increased the interaction between PTEN and PCAF and increased the abundance of acetylated PTEN, suggesting that acetylation of PTEN by PCAF may be a dynamic, regulated mechanism controlling PTEN activity. Because the expression of histone acetylases is commonly increased in cancerous cells, the acetylation of PTEN may provide a mechanism by which these enzymes contribute to overriding the normal controls on the cell cycle.