Thyroid hormone promotes serine phosphorylation of p53 by mitogen-activated protein kinase

L-Thyroxine (T-4) nongenomically promotes association of mitogen-activated protein kinase (MAPK) and thyroid hormone receptor TR beta1 (TR) in the cel l nucleus, leading to serine phosphorylation of the receptor. The oncogene suppressor protein, p53, is serine phosphorylated by several kinases and is known to interact with TR beta1. We studied whether association of p53 and TR is modulated by T-4 and involves serine phosphorylation of p53 by MAPK. TR-replete 293T human kidney cells were incubated with a physiological con centration of T-4 for 10-90 min. Nuclear fractions were immunoprecipitated and the resulting proteins separated and immunoblotted for co-immunoprecipi tated proteins. Activated MAPK immunoprecipitates of nuclei from T-4-treate d cells accumulated p53 in a time-dependent manner; T-4 and T-4-agarose wer e more effective than T-3. T-4-induced nuclear complexing of p53 and MAPK w as inhibited by PD 98059 (PD) and U0126, two MAPK kinase (MEK) inhibitors, and was absent in cells treated with MEK antisense oligonucleotide and in d ominant negative Ras cells. T-4 also caused nuclear co-immunoprecipitation of TR beta1 and p53, an effect also inhibited by PD. Nuclear complexing of p53 and MAPK also occurred in HeLa cells, which lack functional TR. Constit utively activated MAPK caused phosphorylation of a recombinant p53-GST fusi on protein in vitro; thus, p53 is a substrate for MAPK. An indicator of p53 transcriptional activity, accumulation of the immediate-early gene product , c-Jun, was inhibited by T-4. This T-4 effect was reversed by PD, indicati ng that the transcriptional activity of p53 was altered by T-4-directed MAP K-p53 interaction.