Degradation of p53 by adenovirus E4orf6 and E1B55K proteins occurs via a novel mechanism, involving a Cullin-containing complex

Although MDM2 plays a major role in regulating the stability of the p53 tum or suppressor protein, other poorly understood MDM2-independent pathways al so exist. Human adenoviruses have evolved strategies to regulate p53 functi on and stability to permit efficient viral replication. One mechanism invol ves adenovirus E1B55K and E4orf6 proteins, which collaborate to target p53 for, degradation. To determine the mechanism of this process, a multiprotei n E4orf6-associated complex was purified and shown to contain a novel Culli n-containing E3 ubiquitin ligase that is (1) composed of Cullin family memb er Cul5, Elongins B and C, and the RING-H2 finger protein Rbx1(ROC1); (2) r emarkably similar to the von Hippel-Lindau tumor suppressor and SCF (Skp1-C ul1/Cdc53-F-box) E3 ubiquitin ligase complexes; and (3) capable of stimulat ing ubiquitination of p53 in vitro in the presence of E1/E2 ubiquitin-activ ating and -conjugating enzymes. Cullins are activated by NEDD8 modification ; therefore, to determine whether Cullin complexes are required for adenovi rus-induced p53 degradation, studies were conducted in ts41 Chinese hamster ovary cells that are temperature sensitive for the NEDD8 pathway. E4orf6/E 1B55K failed to induce the degradation of p53 at the nonpermissive temperat ure. Thus, our results identify a novel role for the Cullin-based machinery in regulation of p53.