RNA-p53 interactions in vitro.
The tumor suppressor protein p53 is mutated in over half of human cancers. Despite 25 years of study, the complex regulation of this protein remains unclear. After serendipitously detecting RNA binding by p53 in the yeast three-hybrid system (Y3H), we are exploring the specificity and function of this interaction. Electrophoretic mobility shift assays show that full-length p53 binds equally to RNAs that are strongly distinguished in the Y3H. RNA binding blocks sequence-specific DNA binding by p53. The C-terminus of p53 is necessary and sufficient for strong RNA interaction in vitro. Mouse and human C-terminal p53 peptides have different affinities for RNA, and an acetylated human p53 C-terminal peptide does not bind RNA. Circular dichroism spectroscopy of p53 peptides shows that RNA binding does not induce a structural change in the p53 C-terminal peptide, and C-terminal peptides do not detectably affect the structure of RNA. These results demonstrate that p53 binds RNA with little sequence specificity, RNA binding has the potential to regulate DNA binding, and RNA-p53 interactions can be regulated by acetylation of the p53 C-terminus.
Riley
Kasandra J-L
KJ
Department of Biochemistry and Molecular Biology, Mayo Clinic College of Medicine, Rochester, Minnesota 55905, USA.
Ramirez-Alvarado
Marina
M
Maher
L James
LJ
D000818:Animals; D001483:Base Sequence; D002942:Circular Dichroism; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D012313:RNA / Q000737:chemistry / Q000378:metabolism*; D013056:Spectrophotometry, Ultraviolet; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D016159:Tumor Suppressor Protein p53; D012313:RNA