Kodadek, Thomas J.2010-07-122010-07-122008-09-12https://hdl.handle.net/2152.5/310The Ubiquitin-Proteasome Pathway plays both proteolytic and non-proteolytic roles in the regulation of transcription. We recently reported that the ATPases of the 26S proteasome can destabilize activator-DNA complexes in a non-proteolytic manner that requires direct interactions between the Rpt4 and 6 subunits with the activation domain of the activator. Remarkably, mono-ubiquitylation of the activator blocks this repressive activity. In this study, we probe the mechanism of this protective effect. Using novel label transfer and chemical cross-linking techniques, we show that ubiquitin contacts the ATPase complex directly, apparently via Rpn1 and/or Rpt1, and that this interaction results in the dissociation of the activation domain-ATPase complex via an allosteric process. We also provide in vivo evidence demonstrating the importance of monoubiquitylation in inhibition of activator-DNA destabilization. A model is proposed in which activator mono-ubiquitylation serves to limit the lifetime of the activator-ATPase complex interaction and thus the ability of the ATPases to unfold the activator and dissociate the protein-DNA complex.Electronicapplication/pdfenUbiquitinProteolysisTranscription FactorsThesisborn digital297228246