Cooperation of a single lysine mutation and a C-terminal domain in the cytoplasmic sequestration of the p53 protein

J Biol Chem. 1998 Jul 31;273(31):19817-21. doi: 10.1074/jbc.273.31.19817.

Abstract

Cytoplasmic sequestration of the p53 tumor suppresser protein has been proposed as a mechanism involved in abolishing p53 function. However, the mechanisms regulating p53 subcellular localization remain unclear. In this report, we analyzed the possible existence of cis-acting sequences involved in intracellular trafficking of the p53 protein. To study p53 trafficking, the jellyfish green fluorescent protein (GFP) was fused to the wild-type or mutated p53 proteins for fast and sensitive analysis of protein localization in human MCF-7 breast cancer, RKO colon cancer, and SAOS-2 sarcoma cells. The wild-type p53/GFP fusion protein was localized in the cytoplasm, the nucleus, or both compartments in a subset of the cells. Mutagenesis analysis demonstrated that a single amino acid mutation of Lys-305 (mt p53) caused cytoplasmic sequestration of the p53 protein in the MCF-7 and RKO cells, whereas the fusion protein was distributed in both the cytoplasm and the nucleus of SAOS-2 cells. In SAOS-2 cells, the mutant p53 was a less efficient inducer of p21/CIP1/WAF1 expression. Cytoplasmic sequestration of the mt p53 was dependent upon the C-terminal region (residues 326-355) of the protein. These results indicated the involvement of cis-acting sequences in the regulation of p53 subcellular localization. Lys-305 is needed for nuclear import of p53 protein, and amino acid residues 326-355 can sequester mt p53 in the cytoplasm.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Cell Nucleus / metabolism
  • Conserved Sequence / genetics
  • Cytoplasm / metabolism
  • Gene Expression Regulation, Neoplastic / genetics
  • Genes, Reporter / genetics
  • Green Fluorescent Proteins
  • Humans
  • Luminescent Proteins / metabolism
  • Microscopy, Fluorescence
  • Molecular Sequence Data
  • Mutagenesis / genetics
  • Mutation / genetics
  • Recombinant Fusion Proteins / metabolism
  • Transfection / genetics
  • Tumor Cells, Cultured
  • Tumor Suppressor Protein p53 / genetics
  • Tumor Suppressor Protein p53 / metabolism*

Substances

  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • Tumor Suppressor Protein p53
  • Green Fluorescent Proteins