Autoubiquitination of the 26S proteasome on Rpn13 regulates breakdown of ubiquitin conjugates.

Degradation rates of most proteins in eukaryotic cells are determined by their rates of ubiquitination. However, possible regulation of the proteasome's capacity to degrade ubiquitinated proteins has received little attention, although proteasome inhibitors are widely used in research and cancer treatment. We show here that mammalian 26S proteasomes have five associated ubiquitin ligases and that multiple proteasome subunits are ubiquitinated in cells, especially the ubiquitin receptor subunit, Rpn13.

The ATP costs and time required to degrade ubiquitinated proteins by the 26 S proteasome.

The degradation of ubiquitinated proteins by 26 S proteasomes requires ATP hydrolysis. To investigate if the six proteasomal ATPases function independently or in a cyclic manner, as proposed recently, we used yeast mutants that prevent ATP binding to Rpt3, Rpt5, or Rpt6. Although proteasomes contain six ATPase subunits, each of these single mutations caused a 66% reduction in basal ATP hydrolysis, and each blocked completely the 2-3-fold stimulation of ATPase activity induced by ubiquitinated substrates.

Ubiquitinated proteins activate the proteasomal ATPases by binding to Usp14 or Uch37 homologs.

Degradation of ubiquitinated proteins by 26 S proteasomes requires ATP hydrolysis, but it is unclear how the proteasomal ATPases are regulated and how proteolysis, substrate deubiquitination, degradation, and ATP hydrolysis are coordinated. Polyubiquitinated proteins were shown to stimulate ATP hydrolysis by purified proteasomes, but only if the proteins contain a loosely folded domain. If they were not ubiquitinated, such proteins did not increase ATPase activity.

ATP-dependent steps in the binding of ubiquitin conjugates to the 26S proteasome that commit to degradation.

Eukaryotic cells target proteins for degradation by the 26S proteasome by attaching a ubiquitin chain. Using a rapid assay, we analyzed the initial binding of ubiquitinated proteins to purified 26S particles as an isolated process at 4°C. Subunits Rpn10 and Rpn13 contribute equally to the high-affinity binding of ubiquitin chains, but in their absence, ubiquitin conjugates bind to another site with 4-fold lower affinity.

Ubiquitinated proteins activate the proteasome by binding to Usp14/Ubp6, which causes 20S gate opening.

In eukaryotic cells, ubiquitination of proteins leads to their degradation by the 26S proteasome. We tested if the ubiquitin (Ub) chain also regulates the proteasome's capacity for proteolysis. After incubation with polyubiquitinated proteins, 26S proteasomes hydrolyzed peptides and proteins 2- to 7-fold faster.

Getting to first base in proteasome assembly.

Assembly of complex structures such as the eukaryotic 26S proteasome requires intricate mechanisms that ensure precise subunit arrangements. Recent studies have shed light on the pathway for ordered assembly of the base of the 19S regulatory particle of the 26S proteasome by identifying new precursor complexes and four dedicated chaperones involved in its assembly.

Downregulation of COP9 signalosome subunits differentially affects the CSN complex and target protein stability.

Background: The COP9 signalosome (CSN) is a conserved protein complex in eukaryotic cells consisting of eight subunits (CSN1 to CSN8). Recent data demonstrate that the CSN is a regulator of the ubiquitin (Ub) proteasome system (UPS). It controls substrate ubiquitination by cullin-RING Ub ligases (CRLs), a process that determines substrate specificity of the UPS.

The COP9 signalosome-mediated deneddylation is stimulated by caspases during apoptosis.

In concert with the ubiquitin (Ub) proteasome system (UPS) the COP9 signalosome (CSN) controls the stability of cellular regulators. The CSN interacts with cullin-RING Ub ligases (CRLs) consisting of a specific cullin, a RING protein as Rbx1 and substrate recognition proteins. The Ub-like protein Nedd8 is covalently linked to cullins and removed by the CSN-mediated deneddylation.

Ubiquitin-dependent proteolysis of the microtubule end-binding protein 1, EB1, is controlled by the COP9 signalosome: possible consequences for microtubule filament stability.

The COP9 signalosome (CSN) is a regulatory particle of the ubiquitin (Ub) proteasome system (UPS) consisting of eight subunits (CSN1-CSN8). We show that the CSN stabilizes the microtubule end-binding protein 1 (EB1) towards degradation by the UPS. EB1, the master regulator of microtubule plus ends, controls microtubule growth and dynamics.