The stabilization mechanism of mutant-type p53 by impaired ubiquitination: the loss of wild-type p53 function and the hsp90 association.
Mutant-type p53 (mt p53) is largely accumulated in cancer cells due to its increased stability. To elucidate the mechanism of mt p53 stabilization, we analysed the turnover of p53 mutated at codon 248 whose alteration is most frequently found in human cancers. Proteasome inhibition induced the accumulation of ubiquitinated mt p53, indicating that the ubiquitinated forms were essentially unstable and degraded by the proteasome. The presence of a small amount of the ubiquitinated mt p53 relative to the abundant non-ubiquitinated form suggested that the mt p53 ubiquitination was a rate-limiting process in the slow turnover. Two phenomena destabilizing mt p53 via the ubiquitin-proteasome degradation were proved to be independent. First, the coexpression of wild-type p53 (wt p53) promoted mt p53 destabilization as feedback regulation. Second, geldanamycin also induced mt p53 destabilization through the dissociation of the protein from hsp90 but not through the restoration of wt p53 function. Neither the mutant-specific conformation nor the N-terminal phosphorylation seemed to contribute directly to the mt p53 stabilization. Further, a two-dimensional gel electrophoresis revealed that most of the post-translationally modified mt p53 was equally subjected to ubiquitination and subsequent proteasomal degradation. These findings are evidence that mt p53 stabilization depends on the impaired ubiquitination due to both the loss of wt p53 function and the hsp90 association.
Nagata
Y
Y
Department of Tumor Genetics and Biology, Kumamoto University School of Medicine, 2-2-1 Honjo, Kumamoto 860-0811, Japan.
Anan
T
T
Yoshida
T
T
Mizukami
T
T
Taya
Y
Y
Fujiwara
T
T
Kato
H
H
Saya
H
H
Nakao
M
M
D000595:Amino Acid Sequence; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D016227:Benzoquinones; D001859:Bone Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D047029:Lactams, Macrocyclic; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000378:metabolism; D009154:Mutation; D012516:Osteosarcoma / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex; D011487:Protein Conformation; D011809:Quinones / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D014452:Ubiquitins / Q000378:metabolism*
D000903:Antibiotics, Antineoplastic; D016227:Benzoquinones; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D009097:Multienzyme Complexes; D011809:Quinones; D016159:Tumor Suppressor Protein p53; D014452:Ubiquitins; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C001277:geldanamycin