An inhibitor of nuclear export activates the p53 response and induces the localization of HDM2 and p53 to U1A-positive nuclear bodies associated with the PODs.
Leptomycin B is a cytotoxin which directly interacts with and inhibits the action of CRM1, an essential mediator of the nuclear exit of proteins containing nuclear export signals (NES) of the HIV1 REV type. We show that addition of leptomycin B to human primary fibroblasts increased the levels of the p53 tumor suppressor protein. This was accompanied by the induction of p53-dependent transcriptional activity in cultured cells and an increase in the levels of the products of two p53-responsive genes, the p21(CIP1/WAF1) and HDM2 proteins. Leptomycin B induced the accumulation of p53 and HDM2 in the nucleus and the appearance of discrete nuclear aggregates containing both proteins. It has been reported that the transcriptional activity of p53 is modulated by its interaction with the HDM2 protein which also targets p53 for rapid degradation. Using a model cell line conditionally expressing MDM2, the murine analogue of HDM2, we present evidence indicating that leptomycin B abrogates MDM2's role in p53 degradation and that the accumulation of p53 in distinct nuclear bodies is mediated by MDM2. Since HDM2 has recently been shown to contain a functional NES of the REV type, the most likely explanation for our results is that the effect of leptomycin B on HDM2 and p53 is due to the inhibition of nuclear export. The ability to visualize sites where p53 and HDM2 colocalize provides a new approach to study the association between the two proteins in vivo. These p53/HDM2-positive nuclear foci were found to also contain the U1A snRNP A and to be juxtaposed to the PML oncogenic domains.
Laín
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Department of Biochemistry, University of Dundee, United Kingdom.
Midgley
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Sparks
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Lane
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Lane
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D001692:Biological Transport / Q000187:drug effects; D002467:Cell Nucleus / Q000378:metabolism*; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000302:isolation & purification; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology; D006801:Humans; D008297:Male; D009363:Neoplasm Proteins / Q000302:isolation & purification; D009687:Nuclear Proteins; D000071156:Promyelocytic Leukemia Protein; D011518:Proto-Oncogene Proteins / Q000302:isolation & purification*; D051736:Proto-Oncogene Proteins c-mdm2; D017412:Ribonucleoprotein, U1 Small Nuclear / Q000302:isolation & purification*; D014157:Transcription Factors / Q000302:isolation & purification; D014158:Transcription, Genetic; D016159:Tumor Suppressor Protein p53 / Q000302:isolation & purification*; D025521:Tumor Suppressor Proteins
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D005231:Fatty Acids, Unsaturated; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D000071156:Promyelocytic Leukemia Protein; D011518:Proto-Oncogene Proteins; D017412:Ribonucleoprotein, U1 Small Nuclear; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; C070576:PML protein, human; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; C038753:leptomycin B