Energy-dependent nucleolar localization of p53 in vitro requires two discrete regions within the p53 carboxyl terminus.
The p53 tumor suppressor is a nucleocytoplasmic shuttling protein that is found predominantly in the nucleus of cells. In addition to mutation, abnormal p53 cellular localization is one of the mechanisms that inactivate p53 function. To further understand features of p53 that contribute to the regulation of its trafficking within the cell, we analysed the subnuclear localization of wild-type and mutant p53 in human cells that were either permeabilized with detergent or treated with the proteasome inhibitor MG132. We, here, show that either endogenously expressed or exogenously added p53 protein localizes to the nucleolus in detergent-permeabilized cells in a concentration- and ATP hydrolysis-dependent manner. Two discrete regions within the carboxyl terminus of p53 are essential for nucleolar localization in permeabilized cells. Similarly, localization of p53 to the nucleolus after proteasome inhibition in unpermeabilized cells requires sequences within the carboxyl terminus of p53. Interestingly, genotoxic stress markedly decreases the association of p53 with the nucleolus, and phosphorylation of p53 at S392, a site that is modified by such stress, partially impairs its nucleolar localization. The possible significance of these findings is discussed.
Karni-Schmidt
O
O
Department of Biological Sciences, Columbia University, New York, USA.
Friedler
A
A
Zupnick
A
A
McKinney
K
K
Mattia
M
M
Beckerman
R
R
Bouvet
P
P
Sheetz
M
M
Fersht
A
A
Prives
C
C
D000255:Adenosine Triphosphate / Q000378:metabolism; D015153:Blotting, Western; D002466:Cell Nucleolus / Q000378:metabolism*; D002478:Cells, Cultured; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004249:DNA Damage / Q000187:drug effects; D003902:Detergents / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D006801:Humans; D066298:In Vitro Techniques; D007976:Leupeptins / Q000494:pharmacology; D010539:Permeability; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000378:metabolism*
D015853:Cysteine Proteinase Inhibitors; D003902:Detergents; D007976:Leupeptins; D016159:Tumor Suppressor Protein p53; D000255:Adenosine Triphosphate; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde