Cytoplasmically sequestered wild-type p53 protein in neuroblastoma is relocated to the nucleus by a C-terminal peptide.
Cytoplasmic sequestration of wild-type p53 protein occurs in a subset of primary human tumors including breast cancer, colon cancer, and neuroblastoma (NB). The sequestered p53 localizes to punctate cytoplasmic structures that represent large protein aggregates. One functional consequence of this blocked nuclear access is impairment of the p53-mediated G1 checkpoint after DNA damage. Here we show that cytoplasmic p53 from NB cells is incompetent for specific DNA binding, probably due to its sequestration. Importantly, the C-terminal domain of sequestered p53 is masked, as indicated by the failure of a C-terminally directed antibody to detect p53 in these structures. To determine (i) which domain of p53 is involved in the aggregation and (ii) whether this phenotype is potentially reversible, we generated stable NB sublines that coexpress the soluble C-terminal mouse p53 peptide DD1 (amino acids 302-390). A dramatic phenotypic reversion occurred in five of five lines. The presence of DD1 blocked the sequestration of wild-type p53 and relocated it to the nucleus, where it accumulated. The nuclear translocation is due to shuttling of wild-type p53 by heteroligomerization to DD1, as shown by coimmunoprecipitation. As expected, the nuclear heterocomplexes were functionally inactive, since DD1 is a dominant negative inhibitor of wild-type p53. In summary, we show that nuclear access of p53 can be restored in NB cells.
Ostermeyer
A G
AG
Department of Pathology, State University of New York at Stony Brook 11794-8691, USA.
Runko
E
E
Winkfield
B
B
Ahn
B
B
Moll
U M
UM
D000818:Animals; D001665:Binding Sites; D001932:Brain Neoplasms; D001943:Breast Neoplasms; D002453:Cell Cycle; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D003110:Colonic Neoplasms; D003593:Cytoplasm / Q000378:metabolism; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D006801:Humans; D051379:Mice; D009447:Neuroblastoma; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*
D004273:DNA, Neoplasm; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments; D011994:Recombinant Proteins; D016159:Tumor Suppressor Protein p53