Inhibition of DNA binding and transcriptional activity of a nuclear receptor transcription factor by aurothiomalate and other metal ions.
The antirheumatic gold salt aurothiomalate (AuTM) has cellular actions that are consistent with modulation of gene expression. We have tested the hypothesis that an important mode of action of AuTM is inhibition of binding of certain transcription factors to regulatory elements in DNA. The chemistry of transcription factors containing the zinc finger motif makes them candidates for such an interaction with AuTM. In this regard, the interaction of a steroid hormone receptor, the progesterone receptor (PR), with its DNA response element (PRE) was chosen as a suitable model. Nuclear extracts of T-47D human breast cancer cells rich in PR were incubated with radiolabeled PRE, and binding was determined by gel retardation assay. Preincubation of nuclear extract with AuTM caused a concentration-dependent inhibition of binding of PR to PRE (IC50, approximately 3 microM). Other metal ions inhibited binding at higher concentrations, in a rank order correlating with their binding affinity for thiols. Thiomalic acid had no effect in the absence of gold in this system. To test the effect of AuTM on PR-mediated transcription, we transfected the progestin-inducible expression vector pMSG-CAT into T-47D cells. Transfected cells were incubated in the absence or presence of AuTM and treated with the synthetic progestin ORG2058, to induce chloramphenicol acetyl transferase (CAT) activity. With 10 and 100 microM AuTM, there was inhibition to 67 +/- 3% (p = 0.012) and 42 +/- 8% (p = 0.008) of CAT specific activity, respectively, compared with controls. These results demonstrate that AuTM can regulate gene expression and that inhibition of binding of a transcription factor to its response element is a likely mechanism. This provides a molecular model for further study of the antirheumatic action of gold salts.
Handel
M L
ML
Cancer Biology Division, Garvan Institute of Medical Research, Sydney, N.S.W., Australia.
deFazio
A
A
Watts
C K
CK
Day
R O
RO
Sutherland
R L
RL
D002467:Cell Nucleus / Q000378:metabolism; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis / Q000235:genetics; D004247:DNA / Q000378:metabolism*; D005260:Female; D006052:Gold Sodium Thiomalate / Q000494:pharmacology*; D006801:Humans; D011372:Progestins / Q000494:pharmacology; D011980:Receptors, Progesterone / Q000378:metabolism; D013870:Thiomalates / Q000494:pharmacology; D014157:Transcription Factors / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects*; D015032:Zinc / Q000494:pharmacology
D011372:Progestins; D011980:Receptors, Progesterone; D013870:Thiomalates; D014157:Transcription Factors; D006052:Gold Sodium Thiomalate; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase; D015032:Zinc