Within the nuclear receptor family, Nur77 (also known as NGFI-B) distinguishes itself by its ability to bind a target sequence (the NBRE) as a monomer and by its role in T-cell receptor (TCR)-induced apoptosis in T cells. We now report on a novel mechanism of Nur77 action that is mediated by homodimers. These dimers bind a Nur77 response element (NurRE), which has been identified as a target of CRH-induced Nur77 in the pro-opiomelanocortin (POMC) gene promoter. Both halves of the palindromic NurRE are required for responsiveness to physiological signals, like CRH in pituitary-derived AtT-20 cells. Similarly, in T-cell hybridomas, TCR activation induced NurRE but not NBRE reporters. The in vivo signaling function of Nur77 thus appears to be mediated by dimers acting on a palindromic response element of unusual spacing between its half-sites. This mechanism may represent the biologically relevant paradigm of action for this subfamily of orphan nuclear receptors.

We have recently shown that the orphan nuclear receptor Nur77 (NGFI-B) is most active in transcription when it is interacting with a cognate DNA sequence as a homodimer. Further, we have shown that the target for Nur77 dimers, the Nur response element (NurRE), is responsive to physiological stimuli in both endocrine and lymphoid cells, whereas other DNA targets of Nur77 action are not. The Nur77 subfamily also includes two related receptors, Nur-related factor 1 (Nurr1) and neuron-derived orphan receptor 1 (NOR-1). Often, more than one member of this subfamily is induced in response to extracellular signals. We now show that Nur77 and Nurr1 form heterodimers in vitro in the presence or absence of NurRE, and we have documented interactions between these proteins in vivo by using a two-hybrid system in mammalian cells. These heterodimers synergistically enhance transcription from NurRE reporters in comparison to that seen with homodimers. The naturally occurring NurRE from the pro-opiomelanocortin gene preferentially binds and activates transcription in the presence of Nur77 homo-or heterodimers, while a consensus NurRE sequence does not show this preference. Taken together, the data indicate that members of the Nur77 subfamily are most potent as heterodimers and that different dimers exhibit target sequence preference. Thus, we propose that a combinatorial code relying on specific NurRE sequences might be responsible for the activation of subsets of target genes by one of the members of the Nur77 subfamily of transcription factors.

The NGFI-B (Nur77) subfamily of orphan nuclear receptors (NRs), which also includes Nurr1 and NOR1, bind the NurRE regulatory element as either homo-or heterodimers formed between subfamily members. These NRs mediate the activation of pituitary proopiomelanocortin (POMC) gene transcription by the hypothalamic hormone corticotropin-releasing hormone (CRH), an important link between neuronal and endocrine components of the hypothalamo-pituitary-adrenal axis. CRH effects on POMC transcription do not require de novo protein synthesis. We now show that CRH signals activate Nur factors through the cyclic AMP/protein kinase A (PKA) pathway. CRH and PKA rapidly increase nuclear DNA binding activity of NGFI-B dimers but not monomers. Accordingly, CRH-or PKA-activated Nur factors enhance dimer (but not monomer) target response elements. We also show that p160/SRC coactivators are recruited to Nur dimers (but not to monomers) and that coactivator recruitment to the NurRE is enhanced in response to CRH. Moreover, PKA-and coactivator-induced potentiation of NGFI-B activity are primarily exerted through the N-terminal AF-1 domain of NGFI-B. The TIF2 (SRC-2) glutamine-rich domain is required for this activity. Taken together, these results indicate that Nur factors behave as endpoint effectors of the PKA signaling pathway acting through dimers and AF-1-dependent recruitment of coactivators.