CYTOPLASMIC P53 POLYPEPTIDE IS ASSOCIATED WITH RIBOSOMES

Our previous finding that the tumor suppressor p53 is covalently linke d to 5.8S rRNA suggested functional association of p53 polypeptide wit h ribosomes. p53 polypeptide is expressed at low basal levels in the c ytoplasm of normal growing cells in the G(1) phase of the cell cycle. We report here that cytoplasmic wild-type p53 polypeptide from both ra t embryo fibroblasts and MCF7 cells and the A135V transforming mutant p53 polypeptide were found associated with ribosomes to various extent s. Treatment of cytoplasmic extracts with RNase or puromycin in the pr esence of high salt, both of which are known to disrupt ribosomal func tion, dissociated p53 polypeptide from the ribosomes. In immunoprecipi tates of p53 polypeptide-associated ribosomes, 5.8S rRNA was detectabl e only after proteinase K treatment, indicating all of the 5.8S rRNA i n p53-associated ribosomes is covalently linked to protein. while 5.88 rRNA linked to protein was found in the immunoprecipitates of either wild-type or A135V mutant p53 polypeptide associated with ribosomes, l ittle 5.8S rRNA was found in the immunoprecipitates of the slowly sedi menting p53 polypeptide, which was not associated with ribosomes. In c ontrast, 5.8S rRNA was liberated from bulk ribosomes by 1% sodium dode cyl sulfate, without digestion with proteinase K, indicating that thes e ribosomes contain 5.8S rRNA, which is not linked to protein. Immunop recipitation of p53 polypeptide coprecipitated a small fraction of rib osomes. p53 mRNA immunoprecipitated with cytoplasmic p53 polypeptide, while GAPDH mRNA did not. These results show that cytoplasmic p53 poly peptide is associated with a subset of ribosomes, having covalently mo dified 5.8S rRNA.