Dimer-Specific Potentiation of NGFI-B (Nur77) Transcriptional Activity by the Protein Kinase A Pathway and AF-1-Dependent Coactivator Recruitment

Abstract

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.

ACKNOWLEDGMENTS

We thank Vincent Giguère, Sylvain Meloche, and other colleagues in our laboratory for critical comments on the manuscript. We thank Jeffrey Milbrandt for providing NGFI-B antisera, Thomas Perlman for providing antisera against Nurr1, Pierre Chambon for providing the TIF2 antibody, and Yoko Hirata for providing the NGFI-B phospho-Ser316 antibody. We are grateful to Vincent Giguère, O. Conneeley, and N. Ohkura for providing the NGFI-B, Nurr1, and NOR-1 cDNAs, respectively. S. McKnight provided the expression plasmids for PKAc and PKI, and Pierre Chambon provided the TIF2 expression vector and mutant constructs. Amin Ismael and Véronique Montplaisir helped with the EMSA, and Yoko Schreiber helped with transient transfection. We thank Lise Laroche for expert secretarial assistance.

This work was funded in part by the Canadian Institutes of Health Research (CIHR), and M. Maira is the recipient of a doctoral research award from CIHR.