Fig. 3. xp53 does not require de novo protein synthesis to induce Xhox3 and Mix.1. (A) Schematic representation of xp53 constructs used in this study. xp53 contains an N-terminal transactivation domain (TAD), a central DNA-binding domain (DBD), a tetramerization domain (TD) and a C-terminal regulatory domain (RD). xp53:GR contains the hormone binding domain of the glucocorticoid hormone receptor (GRHBD) at the C terminus. (B) Conditional activation of xp53:GR by dexamethasone (DEX). Animal caps expressing xp53:GR were prepared at blastula stages (stage 9) and cultured in the absence or presence of 20 µM DEX. The expression of molecular markers was detected by RT-PCR at the stages indicated. Animal caps expressing xE2F(1-88):GR are used as a control for expression of GRHBD (Suzuki and Hemmati-Brivanlou, 2000). (C) xp53 does not require de novo protein synthesis to induce target gene expression. Animal caps expressing xp53:GR were treated with cycloheximide (CHX) for 30 minutes, and then transferred into medium containing both CHX and DEX to activate xp53:GR. The expression of marker genes was detected by RT-PCR after 3 hours of DEX treatment (equivalent to stage 11).

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