Hormone-induced progesterone receptor phosphorylation consists of sequential DNA-independent and DNA-dependent stages: analysis with zinc finger mutants and the progesterone antagonist ZK98299

摘要: Human progesterone receptors (hPRs) are phosphorylated at multiple serine residues, first in a basal step and then hormone-induced step. To determine whether phosphorylation precedes or follows the interaction of hPRs with DNA two strategies were used. (i) binding was prevented altered site-specific mutants A form hPR; (ii) wild-type hPR forms B antagonist ZK98299. Two hPRA constructed: DBDCys, which lacks critical cysteine residue zinc finger, DBDsp, is mutated three discriminatory amino acids to change its specificity from response element an estrogen element. Receptors transiently expressed PR-negative cells intranuclear. DBDCys did not bind vitro DBDsp bound only Transiently showed upward shift electrophoretic mobility characteristic phosphorylation; it absent DBDCys. Hormone-induced [32P] orthophosphate incorporation into reduced 60% compared but eliminated. ZK98299 binds prevents their DNA. Compared R5020, hPRB T47D breast cancer by totally shift. We conclude that includes DNA-independent DNA-dependent stages sites contribute