Ubiquitin- and MDM2 E3 ligase-independent proteasomal turnover of nucleostemin in response to GTP depletion

Dorothy Lo, Mu Shui Dai, Xiao Xin Sun, Shelya X. Zeng, Hua Lu

Research output: Contribution to journalArticle

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Abstract

Nucleostemin (NS) is a nucleolar GTP-binding protein essential for ribosomal biogenesis, proliferation, and animal embryogenesis. It remains largely unclear how this protein is regulated. While working on its role in suppression of MDM2 and activation of p53, we observed that NS protein (but not mRNA) levels decreased drastically in response to GTP depletion. When trying to further elucidate the molecular mechanism(s) underlying this unusual phenomenon, we found that NS was degraded independently of ubiquitin and MDM2 upon GTP depletion. First, depletion of GTP by treating cells with mycophenolic acid decreased the level of NS without apparently affecting the levels of other nucleolar proteins. Second, mutant NS defective in GTP binding and exported to the nucleoplasm was much less stable than wild-type NS. Although NS was ubiquitinated in cells, its polyubiquitination was independent of Lys-48 or Lys-63 in the ubiquitin molecule. Inactivation of E1 in E1 temperature-sensitive mouse embryonic fibroblast (MEF) cells failed to prevent the proteasomal degradation of NS. The proteasomal turnover of NS was also MDM2-independent, as its half-life in p53/MDM2 double knock-out MEF cells was the same as that in wild-type MEF cells. Moreover, NS ubiquitination was MDM2-independent. Mycophenolic acid or doxorubicin induced NS degradation in various human cancerous cells regardless of the status of MDM2. Hence, these results indicate that NS undergoes a ubiquitin- and MDM2-independent proteasomal degradation when intracellular GTP levels are markedly reduced and also suggest that ubiquitination of NS may be involved in regulation of its function rather than stability.

Original languageEnglish (US)
Pages (from-to)10013-10020
Number of pages8
JournalJournal of Biological Chemistry
Volume287
Issue number13
DOIs
StatePublished - Mar 23 2012

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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