A Nedd8 conjugation pathway is essential for proteolytic targeting of p27Kip1 by ubiquitination

Vladimir N. Podust, James E. Brownell, Tatiana B. Gladysheva, Rong Shu Luo, Chunhua Wang, Michael B. Coggins, Jacqueline W. Pierce, Eric S. Lightcap, Vincent Chau

    Research output: Contribution to journalArticlepeer-review

    231 Scopus citations

    Abstract

    Temporal control of p27Kip1 (p27) degradation imposes periodicity in its activity during cell cycle progression and its accumulation during cell cycle exit. Degradation of p27 is initiated by phosphorylation of p27 at Thr-187, which marks the protein for ubiquitination by SCFSkp2 and subsequent proteolysis by the 26S proteasome. Here we show that the p27 ubiquitination activity in cell extracts depends on the presence of the ubiquitin-like protein Nedd8 and enzymes that catalyze Nedd8 conjugation to proteins. Moreover, we show that reconstitution of the p27 ubiquitination activity of recombinant SCFSkp2 also requires Nedd8 conjugation pathway components. Inactivation of the Nedd8 conjugation pathway by a dominant negative mutant of the Nedd8-conjugating enzyme Nce1/Ubc12 blocks the ubiquitination and degradation of p27 in cell extracts. Consistent with a role in cell-cycle progression, Nedd8 is expressed in proliferating cells and is itself down-regulated upon cellular differentiation. These results suggest that the Nedd8 conjugation pathway may regulate the turnover of p27Kip1, independently of p27 phosphorylation, and further establishes the identity of protein components involved in p27 ubiquitination. Finally, these findings provide a direct demonstration of a function for Nedd8 in a biological process.

    Original languageEnglish (US)
    Pages (from-to)4579-4584
    Number of pages6
    JournalProceedings of the National Academy of Sciences of the United States of America
    Volume97
    Issue number9
    DOIs
    StatePublished - Apr 25 2000

    All Science Journal Classification (ASJC) codes

    • General

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