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Geldanamycin prevents nuclear translocation of mutant p53
1Department of Cell and Tumor Biology, City of Hope National Medical Center, Duarte, California 91010-3000, USA.
Abstract:
p53 is a tumor suppressor protein that acts in the nucleus to effect cell cycle arrest and apoptosis. In some cells p53 is located in the cytoplasm, perhaps as a means of downregulating its activity. We recently showed that hsp90 forms a complex with the cytoplasmically localized mutant p53 (TSp53vall35) within transformed cells (Sepehrnia et al., J. Biol. Chem. 271, 15084, 1996). The present study was undertaken to determine the p53 conformation bound to hsp90 and the role of hsp90 in p53 nuclear translocation. We show that hsp90 binds both a native and a denatured form of p53 as determined by conformation-specific antibodies. hsp90 does not bind p53 in a spatial-specific manner because it remains bound to p53 when induced to translocate to the nucleus by the protein synthesis inhibitor cycloheximide (CHX). Treatment of transformed cells with geldanamycin (GA), a small molecule that binds hsp90, causes a rapid destabilization of p53 by 50%. Residual p53 that survives GA treatment is incapable of translocating to the nucleus. GA does not destabilize p53 in cells where p53 is genotypically wild type. Although GA appears to dramatically alter the translocating properties of mutant p53 it does not dissociate the p53-hsp90 complex. We suggest that a second chaperone protein, called p23, which we show also binds p53, may play an important role in these GA-mediated effects.
Insights
Heat shock protein 90 (hsp90) binds both native and denatured p53, influencing its nuclear translocation. Geldanamycin treatment destabilizes mutant p53 but doesn't disrupt the hsp90-p53 complex, suggesting p23's role.
Area of Science:
- Molecular Biology
- Cell Biology
- Oncology
Background:
- p53 is a tumor suppressor protein regulating cell cycle arrest and apoptosis.
- Cytoplasmic localization of p53 can downregulate its activity.
- Heat shock protein 90 (hsp90) forms complexes with cytoplasmic mutant p53 in transformed cells.
Purpose of the Study:
- To determine the conformation of p53 bound to hsp90.
- To investigate the role of hsp90 in p53 nuclear translocation.
- To explore the effects of geldanamycin (GA) on the p53-hsp90 interaction and p53 function.
Main Methods:
- Utilized conformation-specific antibodies to assess p53 conformation bound to hsp90.
- Induced nuclear translocation of p53 using cycloheximide (CHX).
- Treated cells with geldanamycin (GA) to inhibit hsp90 and observed effects on p53 stability and translocation.
Main Results:
- hsp90 binds both native and denatured forms of p53.
- hsp90 remains bound to p53 during cycloheximide-induced nuclear translocation.
- Geldanamycin treatment destabilizes mutant p53 by 50% and inhibits its nuclear translocation, without dissociating the p53-hsp90 complex.
- GA does not destabilize wild-type p53.
- p23, another chaperone, also binds p53 and may be involved in GA-mediated effects.
Conclusions:
- hsp90 binds p53 in various conformations and is not spatially restricted in its interaction.
- Geldanamycin disrupts mutant p53 function by destabilizing it, rather than by dissociating it from hsp90.
- The chaperone p23 may play a critical role in the geldanamycin-induced alterations of p53 behavior.