We have studied the effects of Fos and Fos/Jun on glucocorticoid induction of hormone-sensitive gene expression. In NIH3T3 cells overexpression of Fos or Fos/Jun by transfection of pSV2-fos and pSV2-jun inhibited glucocorticoid-dependent expression of MMTV LTR-CAT. Expression of p39v-mos had a similar effect on glucocorticoid-dependent reporter gene expression which is most likely mediated by simulation of endogenous Fos. In both cases, this inhibition could be overcome by overexpression of the glucocorticoid receptor (GR) from a transiently transfected expression vector. In receptor deficient CV-1 cells glucocorticoid-dependent reporter gene expression was induced by a range of functional GR truncation mutants. It was established that the C/D domain of the receptor was a sufficient target for inhibition by Fos and Fos/Jun. The C/D domain encompasses the DNA-binding domain, a dimerisation domain and a weak transactivational domain of the GR. When present simultaneously in the cell nucleus Fos and Jun were shown to form a specific and stable protein/protein complex with the glucocorticoid receptor. Finally, it was demonstrated that the GR interacts physically with both Fos and Jun when cotranslated simultaneously in vitro. We propose that this interaction may be the mechanism by which Fos or Fos/Jun bring about inhibition of GR function.