The Proteasome Lid Triggers COP9 Signalosome Activity during the Transition of <i>Sachharomyces cerevisiae</i> Cells into Quiescence

Abstract

The class of Cullin−RING E3 ligases (CRLs) selectively ubiquitinate a large portion of proteins targeted for proteolysis by the 26S proteasome. Before degradation, ubiquitin molecules are removed from their conjugated proteins by deubiquitinating enzymes, a handful of which are associated with the proteasome. The CRL activity is triggered by modification of the Cullin subunit with the ubiquitin-like protein, NEDD8 (also known as Rub1 in Saccharomyces cerevisiae). Cullin modification is then reversed by hydrolytic action of the COP9 signalosome (CSN). As the NEDD8−Rub1 catalytic cycle is not essential for the viability of S. cerevisiae, this organism is a useful model system to study the alteration of Rub1−CRL conjugation patterns. In this study, we describe two distinct mutants of Rpn11, a proteasome-associated deubiquitinating enzyme, both of which exhibit a biochemical phenotype characterized by high accumulation of Rub1-modified Cdc53−Cullin1 (yCul1) upon entry into quiescence in S. cerevisiae. Further characterization revealed proteasome 19S-lid-associated deubiquitination activity that authorizes the hydrolysis of Rub1 from yCul1 by the CSN complex. Thus, our results suggest a negative feedback mechanism via proteasome capacity on upstream ubiquitinating enzymes.

Keywords

• 26S proteasome
• proteasome lid
• Rpn11
• Cdc53
• Cullin
• SCF (Skp, Cullin, F-box containing complex)
• NEDD8 (neural precursor cell expressed developmentally down-regulated 8)
• Rub1 (Related ubiquitin 1)
• CSN (COP9 signalosome)
• <i>Saccharomyces cerevisiae</i>
• diauxic shift
• budding yeast