Spatial control of cAMP signaling by Epacs

描述(申请人提供)：这项建议研究了一类新的cAMP调节蛋白的生理功能，即由cAMP激活的交换蛋白(EPacs)，并检验了新的假设，即每种异构体(ePac1和ERpac2)在细胞中执行不同的生理功能。这个蛋白家族直接与cAMP结合，代表着cAMP的主要细胞内靶标，而不是PKA本身。我们对EPAC激活小G蛋白Rap1的研究表明，交换器的亚细胞定位可以决定它们激活的G蛋白所利用的效应物的选择(Wang等人。2006年；刘等人。2008年)。我们已经在EPAC 1和Epac2中确定了不同的靶向结构域，称为RAS关联(RA)结构域(Liu等人。2008年)。我们认为，这些结构域允许特定的小G蛋白分别连接到Epac1和Epac2，激活不同的Rap1亚细胞池。我们认为Epac1和Epac2的RA结构域分别将cAMP信号与小G蛋白RAN和RAS联系起来。这可以解释RAN如何耦合到Rap1(通过Epac1交换器)以及RAS如何耦合到Rap1(通过Epac2交换器)。这一模型及其对Epac1和Epac2的生理影响在本提案中进行了检验。我们发现Epac1结合在核孔上的小G蛋白上。我们将检验这一假设，即Epac1的RA结构域将Ran-GTP偶联到Rap1，以调节蛋白质的进出细胞核的运输。这代表了RAN与细胞内信号级联的第一个链接，并确定了Epac1和Rap的新功能。这一发现的意义在于，通过RAN周期的蛋白质核转位控制着细胞的生长和分化，其失调是许多癌症的标志。我们发现Epac1调控癌蛋白2-连环蛋白的核功能。我们认为，这一点和Epac1的相关功能可能解释了cAMP的一些抗增殖作用。与Epac1不同，我们认为Epac2与质膜上激活的RAS结合，从而将cAMP和RAS与Rap1的激活联系起来。这种模型的Epac2通过RAS和cAMP作为信号的符合检测器，对糖尿病有重要的意义，因为Epas现在被认为是磺脲类药物的靶标。我们首次提出了RAS和cAMP信号在Epac2上汇聚以促进胰岛素释放的模型。如果这是真的，这将建立一种治疗糖尿病的创新方法，包括将RAS激活剂与胰岛素样激素和磺脲类药物结合起来。 与公共健康相关：药物疗法比任何其他途径更针对环磷酸腺苷(CAMP)信号通路。小G蛋白Rap的鸟嘌呤核苷酸交换器EPAC是cAMP信号通路中的一个新成员，由cAMP结合直接激活。这项建议验证了EPAC促进健康和糖尿病患者胰岛素释放的假设，并通过两个EPAC亚型Epac1和Epac2的非重叠作用调节蛋白质进入正常和癌细胞的细胞核。

DESCRIPTION (provided by applicant): This proposal examines the physiological function of a novel class of cAMP-regulated proteins, the exchange proteins activated by cAMP (Epacs) and tests the novel hypothesis that each isoform (Epac1 and ERpac2) carries out distinct physiological functions in the cell. This family of proteins binds cAMP directly and represents the major intracellular target of cAMP other than PKA itself. Our studies of Epac activation of small G protein Rap1 have demonstrated that subcellular localization of exchangers can dictate the choice of effectors utilized by the G proteins they activate (Wang et al. 2006; Liu et al. 2008). We have identified distinct targeting domains in both Epac 1 and Epac2 called Ras association (RA) domains (Liu et al. 2008). We propose that these domains allow specific small G proteins to link to Epac1 and Epac2, respectively, to the activation of distinct subcellular pools of Rap1. We propose that the RA domains of Epac1 and Epac2 link cAMP signaling to the small G proteins, Ran and Ras, respectively. This can explain how Ran couples to Rap1 (via the Epac1 exchanger) and how Ras can couple to Rap1 (via the Epac2 exchanger). This model and its physiological consequences for both Epac1 and Epac2 are examined in this proposal. We have discovered that Epac1 binds to the small G protein Ran at the nuclear pore. We will test the hypothesis that the RA domain of Epac1 couples Ran-GTP to Rap1 to regulate the transport of proteins in and out of the nucleus. This represents the first link of Ran to intracellular signaling cascades and identifies a novel function for both Epac1 and Rap. The significance of this finding is that the nuclear translocation of proteins via the Ran cycle governs cell growth and differentiation, and its dysregulation is a hallmark of many cancers. We show that Epac1 regulates the nuclear functions of the oncoprotein 2-catenin. We propose that this and related functions of Epac1 may explain some of the anti-proliferative effects of cAMP. In contrast to Epac1, we propose that Epac2 binds to activated Ras at the plasma membrane to link cAMP and Ras to Rap1 activation. This model of Epac2 function as a coincidence detector for signals through Ras and cAMP has significant implications for diabetes, as Epacs are now known to be targets of the sulfonylureas. We propose here for the first time a model that Ras and cAMP signals converge on Epac2 to potentiate insulin release. If true, this would establish an innovative approach to diabetes involving combining Ras activators with both insulinomimetic hormones and the sulfonylurea drugs. PUBLIC HEALTH RELEVANCE: Pharmacological therapeutics target the cyclic AMP (cAMP) signaling pathway more than any other pathway. The guanine nucleotide exchanger for the small G protein Rap called Epac is a new member of the cAMP signaling pathways and is directly activated by cAMP binding. This proposal tests the hypotheses that Epac promotes insulin release in both healthy and diabetic patients, and regulates the entry of proteins into the nucleus of normal and cancer cells through the non-overlapping actions of the two Epac isoforms, Epac1 and Epac2.