Mynard Vanessa, Latchoumanin Olivier, Guignat Laurence, Devin-Leclerc Jocelyne, Bertagna Xavier, Barré Benjamin, Fagart Jerome, Coqueret Olivier, Catelli Maria Grazia
Institut Cochin, Département d'Endo-crinologie, 24 rue du Faubourg Saint Jacques, 75014 Paris, France.
Mol Endocrinol. 2004 Dec;18(12):2997-3010. doi: 10.1210/me.2003-0417. Epub 2004 Aug 19.
Leukemia inhibitory factor (LIF) cooperates with CRH at the pituitary level to induce POMC gene transcription, resulting in activation of the pituitary-adrenal axis. However, the underlying molecular mechanisms remain elusive. Here, we show that the NurRE-signal transducers and activators of transcription (STAT) composite element of the POMC promoter was the predominant target of the LIF-CRH synergy. Whereas NurRE or STAT sites alone conferred synergy, the maximal response was found with the NurRE-STAT reporter, suggesting that direct DNA binding of both transcription factors is required for an optimal synergy. During LIF-CRH stimulation, Nur77 and activated STAT1-3 were bound to the composite element, and the binding of each factor was abolished by appropriate mutations. CREB was also detected in this complex in a stimulation-dependent and DNA binding-independent manner. Nur77 and STAT1-3 bound to the NurRE-STAT site were each sufficient for CREB recruitment. Recombinant CREB directly interacted with recombinant Nur77 or STAT1-3. Moreover, CREB-Nur77 interaction was increased by CREB phosphorylation at Ser-133 and the dominant-negative mutant CREB-M1 efficiently inhibited the synergistic LIF-CRH response. This synergism was also inhibited after transfection of CREB-small interfering RNA. We conclude that both CREB phosphorylation at Ser-133 and level of CREB expression are crucial in LIF-CRH synergism where CREB, without direct DNA binding, could improve the stability of Nur77 and STAT1-3 binding to POMC promoter and facilitate the recruitment of coactivators. This novel intrapituitary signaling mechanism may have more general implications in cross talks between cAMP-protein kinase A and Janus kinase-STAT pathways.
白血病抑制因子（LIF）在垂体水平与促肾上腺皮质激素释放激素（CRH）协同作用，诱导阿黑皮素原（POMC）基因转录，从而激活垂体-肾上腺轴。然而，其潜在的分子机制仍不清楚。在此，我们表明POMC启动子的NurRE-信号转导和转录激活因子（STAT）复合元件是LIF-CRH协同作用的主要靶点。虽然单独的NurRE或STAT位点可产生协同作用，但NurRE-STAT报告基因显示出最大反应，这表明两种转录因子的直接DNA结合对于最佳协同作用是必需的。在LIF-CRH刺激过程中，Nur77和活化的STAT1-3与复合元件结合，并且每个因子的结合可通过适当的突变消除。CREB也以刺激依赖且不依赖DNA结合的方式在该复合物中被检测到。与NurRE-STAT位点结合的Nur77和STAT1-3各自足以募集CREB。重组CREB直接与重组Nur77或STAT1-3相互作用。此外，CREB在Ser-133处的磷酸化增加了CREB-Nur77相互作用，并且显性负性突变体CREB-M1有效地抑制了LIF-CRH的协同反应。转染CREB小干扰RNA后，这种协同作用也受到抑制。我们得出结论，Ser-133处的CREB磷酸化和CREB表达水平在LIF-CRH协同作用中至关重要，其中CREB无需直接结合DNA，可提高Nur77和STAT1-3与POMC启动子结合的稳定性，并促进共激活因子的募集。这种新的垂体信号转导机制可能在环磷酸腺苷-蛋白激酶A和Janus激酶-STAT途径之间的相互作用中具有更广泛的意义。
