Yeagley David, Quinn Patrick G
The Pennsylvania State University, College of Medicine, Department of Cellular and Molecular Physiology, C4718, 500 University Drive, Hershey, Pennsylvania 17033, USA.
Mol Endocrinol. 2005 Apr;19(4):913-24. doi: 10.1210/me.2004-0281. Epub 2004 Dec 16.
Phosphoenolpyruvate carboxykinase (PEPCK) transcription is induced by cAMP/protein kinase A (PKA) and glucocorticoids [dexamethasone (Dex)] and is inhibited by insulin to regulate blood glucose. Recent reports suggested that CCAAT enhancer binding protein (C/EBP) binding to the PEPCK cAMP response element (CRE) plays a role in Dex induction and that insulin-induces inhibitory forms of C/EBPbeta to inhibit transcription. Here, we assessed the roles of CRE-binding protein (CREB) and C/EBP factors in mediating hormone-regulated transcription. Neither cAMP nor insulin regulated the phosphorylation of C/EBP. Cycloheximide did not block insulin inhibition, indicating that alternate translation of C/EBPbeta is not required. Dominant-negative CREB or C/EBP blocked induction by PKA, but neither affected regulation by Dex or insulin. Tethering the activation domains of CREB or C/EBP to a CRE-->Gal4 (G4) site mediated varying extents of basal and PKA-inducible activity, but neither activation domain affected induction by Dex or inhibition by insulin. Surprisingly, synergistic induction by PKA and Dex did not require the CRE and was unaffected by dominant-negative CREB or C/EBP. PKA and Dex also synergistically induced a minimal 3 x glucocorticoid response element promoter, but inhibited Dex induction of the mouse mammary tumor virus and IGF-binding protein 1 promoters, even though PKA alone did not regulate these promoters. These results suggest that PKA modifies the activity of other factors involved in Dex induction to mediate synergistic induction or inhibition in a promoter-specific manner. Our data indicate that the roles of CREB and C/EBP are restricted to mediating PEPCK induction by PKA, and that other factors mediate PEPCK induction by Dex, synergism between PKA and Dex, and inhibition by insulin.
磷酸烯醇式丙酮酸羧激酶（PEPCK）的转录受环磷酸腺苷/蛋白激酶A（PKA）和糖皮质激素[地塞米松（Dex）]诱导，并受胰岛素抑制以调节血糖。最近的报道表明，CCAAT增强子结合蛋白（C/EBP）与PEPCK环磷酸腺苷反应元件（CRE）的结合在Dex诱导中起作用，并且胰岛素诱导C/EBPβ的抑制形式以抑制转录。在此，我们评估了CRE结合蛋白（CREB）和C/EBP因子在介导激素调节转录中的作用。环磷酸腺苷和胰岛素均未调节C/EBP的磷酸化。放线菌酮未阻断胰岛素抑制作用，表明不需要C/EBPβ的交替翻译。显性负性CREB或C/EBP阻断了PKA的诱导作用，但两者均不影响Dex或胰岛素的调节作用。将CREB或C/EBP的激活域连接到CRE→Gal4（G4）位点介导了不同程度的基础活性和PKA诱导活性，但两个激活域均不影响Dex诱导或胰岛素抑制作用。令人惊讶的是，PKA和Dex的协同诱导不需要CRE，并且不受显性负性CREB或C/EBP的影响。PKA和Dex还协同诱导了最小的3x糖皮质激素反应元件启动子，但抑制了Dex对小鼠乳腺肿瘤病毒和IGF结合蛋白1启动子的诱导，尽管单独的PKA不调节这些启动子。这些结果表明，PKA修饰参与Dex诱导的其他因子的活性，以启动子特异性方式介导协同诱导或抑制。我们的数据表明，CREB和C/EBP的作用仅限于介导PKA诱导的PEPCK，而其他因子介导Dex诱导的PEPCK、PKA和Dex之间的协同作用以及胰岛素的抑制作用。
