Nuclear export is required for degradation of endogenous p53 by MDM2 and human papillomavirus E6.
The MDM2 oncoprotein targets the p53 tumor suppressor protein for degradation when the two proteins are expressed in cells. The regulation of p53 levels by MDM2 requires the ability of MDM2 to be exported from the nucleus by utilizing its nuclear export signal (NES). The drug leptomycin B (LMB) blocks the formation of nuclear export complexes consisting of CRM1, RanGTP, and NES-containing proteins. It is predicted that LMB should inhibit nuclear-cytoplasmic shuttling by MDM2 and subsequently stabilize p53. This communication demonstrates that LMB treatment of various cell lines led to an increase in the steady-state levels of the p53 protein as a result of an increase in its stability. The stabilized p53 protein localized to the nucleus and was an active transcription factor. These results indicate that the low steady-state levels of p53 in the absence of DNA damage result from p53's nuclear export for cytoplasmic degradation. LMB also led to p53 stabilization in cell lines that contain human papillomavirus (HPV) DNA and express HPV E6, a protein that targets p53 for degradation. MDM2 is not necessary for E6-dependent degradation of p53, as evidenced by the observation that E6 promoted p53 degradation in cells lacking endogenous MDM2. In addition, LMB reduced E6's ability to degrade p53 in the absence of MDM2, demonstrating that complete degradation of p53 by E6 requires nuclear export and therefore likely occurs in cytoplasmic proteasomes. These data suggest that the nuclear export of p53 to the cytoplasm for degradation is a general mechanism for regulating p53 levels.
Freedman
D A
DA
Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544, USA.
Levine
A J
AJ
D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002467:Cell Nucleus / Q000378:metabolism*; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry; D009687:Nuclear Proteins; D027383:Papillomaviridae / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051736:Proto-Oncogene Proteins c-mdm2; D014158:Transcription, Genetic / Q000235:genetics; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D005231:Fatty Acids, Unsaturated; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D016159:Tumor Suppressor Protein p53; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; C038753:leptomycin B