p53 in human embryonal carcinoma: identification of a transferable, transcriptional repression domain in the N-terminal region of p53.
Testicular germ cell tumors (TGCTs) arise despite possessing high levels of wild-type p53, suggesting p53 latency. We have previously shown that p53 repression in TGCT-derived human embryonal carcinoma (EC) is relieved upon treatment with all-trans retinoic acid (RA), resulting in enhanced p53 transactivation activity. To further investigate p53 repression in EC, a series of gal4-p53 truncation constructs were generated. Deletion of the core DNA-binding region, residues 117-274, had no effect on basal or RA-induced p53 activity. Progressively, larger truncations were made in the C- or N-terminal direction. Deletion of residues toward the C-terminus of p53 as far as residue 354 did not affect either the basal or RA-inducible activity of gal4-p53. When a small region in the N-terminus was deleted (residues 105-116), relief of the basal repression of p53 activity characteristic of EC was observed. Fusion of this region to the VP16 activation domain (VPAD) resulted in a 10-20-fold repression of VPAD activity in NT2/D1 human EC cells, indicating that this region acts as a heterologous repressor. Owing to its location in the N-terminal half of p53, we have named this region the p53 N-terminal Repression Domain (p53-NRD). The p53-NRD mediated repression in a variety of cell lines, with the most prominent repression observed in human EC cells. While RA alone had no effect on p53-NRD activity, cotreatment with RA and the histone deacetylase inhibitor trichostatin-A (TSA) completely relieved p53-NRD-mediated repression. In contrast, NRD-mediated repression was not sensitive to RA and TSA in a derived RA-resistant cell line with a retinoic acid receptor gamma (RARgamma) defect, but sensitivity could be restored with transfection of RARgamma. These data indicate that a unique repressor domain resides in p53 at residues 90-116 whose activity can be modulated in the presence of 'differentiation therapy' and 'transcription therapy' agents.
Curtin
Joshua C
JC
Department of Pharmacology and Toxicology, Dartmouth Medical School, Hanover, NH 03755, USA.
Spinella
Michael J
MJ
D000595:Amino Acid Sequence; D000818:Animals; D001943:Breast Neoplasms; D016466:CHO Cells; D019556:COS Cells; D018236:Carcinoma, Embryonal / Q000235:genetics*; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D006224:Cricetinae; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008969:Molecular Sequence Data; D014158:Transcription, Genetic / Q000235:genetics*; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry / Q000235:genetics*
D016159:Tumor Suppressor Protein p53; D014212:Tretinoin