The promyelocytic leukemia (PML) tumor suppressor is essential for the formation of PML nuclear bodies (NBs). PML and PMLNBs have been implicated in the regulation of growth inhibition, senescence and apoptosis. PML is activated in response to stress signals and is downregulated in certain human cancers. However, the factors mediating PML stability are incompletely understood. Here we demonstrate that a catalytically active form of the mammalian E3 ligase E6AP (HPV E6-associated protein) acts to reduce the half-life of the PML protein by promoting its degradation in the proteasome. E6AP mediates the ubiquitination of PML in an in vitro ubiquitination assay. E6AP and PML interact at physiological levels and colocalize in PML-NBs. Importantly, PML protein expression is elevated in multiple organs and cell types from E6AP null mice and in lymphoid cells is associated with increased number and intensity of PML-NBs. This PML elevation is enhanced in response to DNA damage. Our results identify E6AP as an important regulator of PML and PML-NBs. The promyelocytic leukemia (PML) tumor suppressor is implicated in the regulation of cell-cycle progression, premature senescence (triggered by oncogenic Ras) and apoptosis (reviewed by Bernardi and Pandolfi 1 ). Deregulation of PML can be oncogenic. PML-RARa contributes to acute promyelocytic leukemia (APL) 2,3 and downregulation of PML was observed in multiple human cancers. 4 PML knockout (KO) mice are resistant to lethal doses of ionizing irradiation (IR), and exhibit genomic instability and enhanced susceptibility to tumorigenesis upon exposure to carcinogens 1,5 or in the context of additional oncogenic events (e.g., a loss of Pten 6 ). The myriad of PML functions have been linked to its function in PML nuclear body (PML-NB) formation. 7 These are dynamic structures whose configuration and composition are modified in response to specific stress signals. 8 However, their mechanisms of action are only partially understood and are believed to be mediated by key proteins that are recruited to these structures, including pRb, SUMO, Daxx and p53. 9 The regulation of certain proteins, such as p53, is associated with the PML-NBs. 9 Stress stimuli that have been identified as activators of PML and PML-NBs include interferon (a and g), DNA damage, oncogenic stress and viral infection (reviewed in references 1,8,10 ). In contrast, exposure of cells to arsenic trioxide (As 2 O 3 ) or retinoic acid promotes PML degradation. 11 In contrast, the overexpression of specific cellular ubiquitin ligases, such as the Siah protein, 12 or certain viral ubiquitin ligases, such as the ICP0 regulatory protein of herpesvirus-1, 10 promotes the proteasomal degradation of PML and PML-RARa. However, the nature of the underlying mechanism of this degradation and whether it occurs under physiological conditions is not known. Recently, a function for CK2-mediated phosphorylation of PML (on serine 517) in the control of its protein stability was demonstrated. 13 In addition to phosphorylation, the covalen...