Increasing the Unneddylated Cullin1 Portion Rescues the csn Phenotypes by Stabilizing Adaptor Modules To Drive SCF Assembly

Abstract

The dynamic SCF (Skp1–cullin1–F-box protein) assembly is controlled by cycles of cullin neddylation/deneddylation based on the deneddylation activity of the COP9 signalosome (CSN) and global sequestration of cullins by CAND1. However, acceptance of this prediction was hampered by the results of recent studies, and the regulatory mechanism and key players remain to be identified. We found that maintaining a proper Cul1 Nedd8 /Cul1 ratio is crucial to ensure SCF functions. Reducing the high Cul1 Nedd8 /Cul1 ratios in csn mutants through ectopic expression of the nonneddylatable Cul1 K722R proteins or introducing the endogenous cul1 K722R point mutation significantly rescues their defective phenotypes. In vivo protein degradation assays revealed that the large portion of the unneddylated Cul1 contributes to F-box protein stabilization. Moreover, the unneddylated Cul1 tends to associate with adaptor modules, and disruption of Cul1–Skp-1 binding fails to restore the csn phenotypes. Taking the data together, we propose that unneddylated Cul1 is another central player involved in maintenance of the adaptor module pool through the formation of Cul1–Skp-1–F-box complexes and promotion of rapid SCF assembly.