Dissecting the PDE4d orthologue Dunce isoform functions in Drosophila melanogaster

人类和果蝇一样，存在几种磷酸二酯酶（PDEs）。它们将cAMP水解为AMP并终止cAMP信号传导。与磷酸二酯酶功能相关的一个最重要的问题已经由Houslay在2010年提出：“为什么当所有的磷酸二酯酶都水解cAMP时，会产生这么多不同的同工异构体？”虽然越来越多的证据表明，由于靶向PDE活性和细胞中不同结构域的cAMP效应蛋白，cAMP的亚细胞微结构域参与了cAMP信号特异性的调节，但我们的知识还远远不够全面。我们拟以黑腹果蝇为研究对象，在亚细胞、神经元和行为水平上对Pde4d同源蛋白的功能进行剖析。我们想研究在神经元中是否需要不同的Pde4D同源基因Dunce的异构体来建立不同的cAMP微结构域。我们集中分析《蠢蛋》有几个原因。首先，Dunce是果蝇中唯一的cAMP特异性PDE，其次，我们之前已经表明，不同的Dunce亚型在不同的亚细胞区室中表达。第三，笨蛋突变体在神经元可塑性缺陷方面存在差异。为了测量神经元可塑性和功能的变化，我们将分析Dunce在联想厌恶嗅觉学习记忆和神经元形态中的功能。特别是，我们想利用行为、细胞和亚细胞水平上的差异来解决不同的Dunce亚型是否在不同亚细胞区域的相同神经元中起作用以调节cAMP信号。进一步，我们想确定相同的异构体是否在不同的神经元中调节类似形式的神经元可塑性。最后，我们将研究PDE活性的亚细胞变化是否局限于特定的细胞位点，或者这些变化是否扩展到整个神经元。为此，我们将使用遗传学、分子遗传学、想象技术和行为分析。所得结果将揭示果蝇细胞cAMP结构域是如何建立的，以及不同的Dunce异构体如何促进神经元可塑性的变化。

In humans as in the fruit fly Drosophila melanogaster several phosphodiesterases (PDEs) exit. They hydrolyze cAMP to AMP and terminate cAMP signaling. One of the most important questions related to phosphodiesterase function was already raised by Houslay in 2010: “Why are there so many different isoforms generated when all of them hydrolyze cAMP?” While there is emerging evidence that sub-cellular micro-domains of cAMP are involved in the regulation of cAMP signaling specificity due to targeted PDE activity and cAMP effector proteins to different domains in the cell, our knowledge is far from comprehensive. We propose to use Drosophila melanogaster to diessect the function of the Pde4d orthologue Dunce on subcellular, on neuronal and behavioral level. We want to investigate whether different isoforms of the Pde4D orthologue Dunce are required to set up distinct cAMP micro-domains in neurons. We focus our analysis on Dunce for serval reasons. First Dunce is the only cAMP specific PDE in Drosophila, second we have previosly shown that different Dunce isoforms are expressed in different sub-cellular compartments. Third dunce mutants differ in their defects in neuronal plasticity. To measure changes in neuronal plasticity and function we will analyze the function of Dunce in associative aversive olfactory learning and memory and neuron morphology. In particular we want to use the differences on behavioral, cellular und subcellular level to address whether different Dunce isoforms act in the same neurons in different subcellular domains to regulate cAMP signaling. Further we want to determine whether the same isoform regulates in different neurons similar forms of neuronal plasticity. Finally we will investigate whether subcellular changes of PDE activity are restricted to specific cellular sites or whether these changes extend over the whole neuron. Towards this end, we will use genetic, molecular-genetic, imagine techniques and behavioral analysis. The obtained results will shed light on how in Drosophila cellular cAMP domains are set up and how different Dunce isoforms contribute to changes of neuronal plasticity.