The Cyclic AMP-GEF/C/EBP Pathway; a New PKA-independent Route for the Control of Gene Expression by Cyclic AMP

循环AMP级联是第一个被描述的解释激素如何在细胞内发挥功能变化的细胞内机制，并且仍然是典型的信号系统。直到最近，人们还认为，细胞内环状AMP浓度的增加仅仅是通过酶‘PKA’对转录因子‘CREB’的事先磷酸化而改变基因的活性。然而，现在已经发现了一种新的途径，通过这种途径，环状AMP可以激活称为RAS和Rap的蛋白质。它们参与控制细胞存活、细胞周期控制和细胞黏附。然而，为了激活RAS和Rap，环状AMP必须首先直接与另一类称为环状AMP GEF的酶相互作用。两个这样的环状AMP GEF是EPAC或CNrasGlobal。这个项目的目的是确定环状AMP-GEF的激活可以导致依赖于环状AMP的、独立于PKA的基因表达变化的机制。为了做到这一点，我们将使用SoCS-3基因作为范例。SOCS-3是一个基因，它的诱导导致细胞因子和生长因子受体信号的抑制。我们最近发现，SoCS-3基因也受到环状AMP的正调控，通过EPAC和Rap1。我们的工作表明，这可能是通过称为C/EBP的转录因子实现的。此外，我们还发现了一条不依赖于PKA的途径，导致生长调节酶ERK从环状AMP中被激活。ERK是环状AMP诱导SOCS-3所必需的，并以环状AMP依赖、PKA不依赖的方式磷酸化一类C/EBPs。因此，我们将描绘这一新的途径，从激活环状AMP-GEF，如EPAC或CNrasGlobal，通过ERK到C/EBPS，并确定其激活在人脐静脉内皮细胞(HUVECs)中的生物学后果。这个细胞系统已经被证明表达环状AMP诱导的SOCS-3，并且呈现的优点是，通过SOCS-3介导的细胞因子信号的抑制来确定基因诱导的生物学意义，可以很容易地被测量。由于环状AMP-环境基金-C/EBP途径是环状AMP激活的信号通路的第一个例子，可以独立于经典的PKA/CREB途径来控制基因的表达，因此这些研究是迫切的科学优先事项。因此，我们的研究将对典型的环状AMP信号系统如何调控基因表达提供一个至关重要的新理解。我们在研究环AMP信号的分子和细胞基础方面的联合专业知识意味着我们有能力开展这些研究。

The cyclic AMP cascade was the first intracellular mechanism described to explain how hormones exert functional changes inside cells and remains the archetypical signalling system. Until very recently in was thought that increases in the intracellular concentration of cyclic AMP exerted changes in the activity of genes solely through prior phosphorylation of the transription factor 'CREB' by the enzyme 'PKA'. However, a new route has now been discovered whereby cyclic AMP can activate proteins called Ras and Rap. These are involved in the control of cell survival, the cell cycle control and cell adhesion. However, in order to activate Ras and Rap, cyclic AMP must first directly interact with another class of enzymes called cyclic AMP GEFs. Two such cyclic AMP GEFs are EPAC or CNrasGEF. The aims of this project are to determine the mechanisms by which activation of cyclic AMP-GEFs can lead to changes in gene expression, in a cyclic AMP-dependent, independently of PKA. To do this we will use the SOCS-3 gene as a paradigm. SOCS-3 is a gene whose induction leads to the suppression of signaling from cytokine and growth factor receptors. We have recently found that the SOCS-3 gene is also positively regulated by cyclic AMP, through EPAC and Rap1. Our work shows that this is probably through transcription factors called C/EBP. Moreover, we have found that a PKA-independent pathway leads to the activation of the growth regulatory enzyme ERK from cyclic AMP. ERK is required for SOCS-3 induction by cyclic AMP and phosphorylates one class of C/EBPs, also in a cyclic AMP-dependent, PKA-independent fashion. We will therefore delineate this new pathway leading from activation of cyclic AMP-GEFs, like EPAC or CNrasGEF, through ERK to C/EBPs and determine the biological consequences of its activation in human umbilical cord endothelial cells (HUVECs). This cell system has been shown to express cyclic AMP-inducible SOCS-3 and presents the advantage that the biological significance of gene induction, as determined by SOCS-3 mediated inhibition of cytokine signalling, can be easily measured. Because the cyclic AMP-GEF-C/EBP pathway is the first example of a cyclic AMP-activated signalling pathway that can control gene expression independently of the classical PKA/CREB route, these studies are an urgent scientific priority. Our investigations will therefore provide a critical new understanding of how gene expression is regulated by the prototypical cyclic AMP signalling system. Our joint expertise in studying the molecular and cellular basis of cyclic AMP signalling means that we are well equipped to carry out these investigations.