In eukaryotic cells, proteins are targeted to the proteasome for degradation by polyubiquitination. These proteins bind to ubiquitin receptors, are engaged and unfolded by proteasomal ATPases, and are processively degraded. The factors determining to what extent the proteasome can successfully unfold and degrade a substrate are still poorly understood. We find that the architecture of polyubiquitin chains attached to a substrate affects the ability of the proteasome to unfold and degrade the substrate, with K48-or mixed-linkage chains leading to greater processivity than K63-linked chains. Ubiquitin-independent targeting of substrates to the proteasome gave substantially lower processivity of degradation than ubiquitin-dependent targeting. Thus, even though ubiquitin chains are removed early in degradation, during substrate engagement, remarkably they dramatically affect the later unfolding of a protein domain. Our work supports a model in which a polyubiquitin chain associated with a substrate switches the proteasome into an activated state that persists throughout the degradation process.

ATP-dependent proteases are present in all organisms, where they are responsible for much of intracellular protein degradation. Most proteins are processively unfolded and degraded into small peptides; however, in a few so-called slippery substrates, the protease stalls at a folded domain and releases a large protein fragment. In this review, we describe the properties of physiological slippery substrates that are processed in this manner by ATPdependent proteases and the recent advances that have been made in understanding the mechanism underlying their partial degradation.

The ubiquitin-proteasome system (UPS) is responsible for the bulk of protein degradation in eukaryotic cells, but the factors that cause different substrates to be unfolded and degraded to different extents are still poorly understood. We previously showed that polyubiquitinated substrates were degraded with greater processivity (with a higher tendency to be unfolded and degraded than released) than ubiquitin-independent substrates. Thus, even though ubiquitin chains are removed before unfolding and degradation occur, they affect the unfolding of a protein domain. How do ubiquitin chains activate the proteasome’s unfolding ability? We investigated the roles of the three intrinsic proteasomal ubiquitin receptors - Rpn1, Rpn10 and Rpn13 - in this activation. We find that these receptors are required for substrate-mediated activation of the proteasome’s unfolding ability. Rpn13 plays the largest role, but there is also partial redundancy between receptors. The architecture of substrate ubiquitination determines which receptors are needed for maximal unfolding ability, and, in some cases, simultaneous engagement of ubiquitin by multiple receptors may be required. Our results suggest physical models for how ubiquitin receptors communicate with the proteasomal motor proteins.

Nuclear receptor coregulators are master regulators of transcription and selectively interact with the vitamin D receptor (VDR) to modulate cell differentiation, cell proliferation and calcium homeostasis. Herein, we report the syntheses and evaluation of highly potent and selective VDR–coactivator inhibitors based on a recently identified 3-indolylmethanamine scaffold. The most active compound, PS121912, selectively inhibited VDR-mediated transcription among eight other nuclear receptors tested. PS121912 is also selectively disrupting the binding between VDR and the third nuclear receptor interaction domain of the coactivator SRC2. Genetic studies revealed that PS121912 behaves like a VDR antagonist by repressing 1,25-(OH)2D3 activated gene transcription. In addition, PS121912 induced apoptosis in HL-60.