Cyclic AMP-induced forkhead transcription factor, FKHR, cooperates with CCAAT/enhancer-binding protein β in differentiating human endometrial stromal cells

Decidual transformation of human endometrial stromal (ES) cells requires sustained activation of the protein kinase A (PKA) pathway. In a search for novel transcriptional mediators of this process, we used differential display PCR analysis of undifferentiated primary ES cells and cells stimulated with 8-bromo-cAMP (8-Br-cAMP). We now report on the role of forkhead homologue in rhabdomyosarcoma (FKHR), a recently described member of the forkhead/winged-helix transcription factor family, as a mediator of endometrial differentiation. Sustained 8-Br-cAMP stimulation resulted in the induction and nuclear accumulation of FKHR in differentiating ES cells. Immunohistochemical studies revealed that endometrial stromal expression of FKHR in vivo is confined to decidualizing cells during the late secretory phase of the cycle and coincides with the expression of CCAAT/enhancer-binding protein 13 (C/EBPbeta). Reporter gene studies showed that FKHR potently enhances PKA-dependent activation of the tissue-specific decidual prolactin (dPRL) promoter, a major differentiation marker in human ES cells. Transcriptional augmentation by FKHR was effected through functional cooperation with C/EBPbeta and binding to a composite FKHR-C/EBPbeta response unit in the proximal promoter region. Furthermore, FKHR and C/EBPbeta were shown to interact directly in a glutathione S-transferase pull-down assay. These results provide the first evidence of regulated expression of FKHR and demonstrate that FKHR has an integral role in PKA-dependent endometrial differentiation through its ability to bind and functionally cooperate with C/EBPbeta.