ABSTRACT Proteasome-catalyzed proteolysis is a central regulatory element of most essential eukaryotic cellular processes. Proteasome function is regulated at many levels including the assembly of multiple proteasome holoenzymes, alterations of the relative and absolute amount of holoenzymes, and direct control of holoenzyme activity. Proteasome holoenzymes are composed of a protease core complex (20S proteasome) interchangeably bound to any of multiple regulatory complexes. The goals of this research are to discover the physiologic roles of PI31, the most poorly understood and least studied proteasome regulator, and to determine the molecular and cellular basis of its function and regulation. The project is based on the premise that PI31 mediates cellular changes in proteasome-dependent protein degradation during cellular stress. We will test the general hypothesis that PI31 exerts this role by regulating proteasome content, composition, and/or function in response to stress. We also will test the hypothesis that PI31 action is regulated by reversible PI31 posttranslational modifications cued by specific stress-induced signals. We will employ a complementary suite of cellular and molecular approaches to define the mechanisms by which PI31 accomplishes its role, the regulation of these mechanisms by PI31 posttranslational modifications, and the physiologic significance of these effects. The pervasive role of proteasome-catalyzed protein degradation in normal cellular function and regulation explains why many diseases are associated with abnormal proteasome function or how diseased cells exploit normal proteasome action to their advantage. Such knowledge is the foundation of current proteasome-based therapies to treat cancer. A deeper understanding of normal proteasome function and regulation will significantly advance understanding of the molecular basis of human diseases and provide new strategies for their treatment.