Functional Genomics of Drosophila Olfactory Memory

描述(申请人提供)：LTM形成的分子基础似乎在进化上是保守的。特别是，简单的学习形式似乎反映了神经元保守的细胞功能，而更复杂的行为任务反映了不同物种神经系统(解剖学)的多样性。尽管如此，对这些复杂任务的记忆似乎唤起了共同的潜在突触可塑性。因此，我们利用果蝇的规模经济和遗传工具的可用性，通过DNA微阵列和对具有记忆缺陷的单基因突变体的行为筛选相结合的方式，专注于发现LTM涉及的下游基因，我们已经确定了参与新转录的mRNAs的细胞运输和随后的局部易位的一种新的生物途径的几个遗传成分。重要的是，这些基因中的两个，Pumilio和Staufen已经实现了体内生物学验证。 我们建议通过以下具体目标重点关注这一途径： 1.Pumilio/Staufen途径中选定基因(Staufen、Oskar、moesin、Pumilio、orb、elF2g、elFSc)的分子遗传学验证：我们计划(I)与现有等位基因进行遗传互补，(Ii)时间和空间拯救，以确定这些基因在记忆形成中的作用。 2.Pumilio/Staufen通路中部分基因的蛋白表达研究：大多数发育基因在成年中枢神经系统中从未见过免疫染色的报道。我们将这样做，特别关注亚细胞共定位实验。 3.细胞纯化后的DNA芯片筛选：对嗅觉记忆神经底物的研究继续表明，焦点可能是蘑菇体的神经元。使用一种新的方法，我们计划从训练有素的果蝇中提纯MB神经元，用于DNA微阵列实验。我们预计，从这个纯化的细胞群体中分离出来的RNA中，来自转录反应基因的信号会更大，从而检测到Pumilio/Staufen途径的更多神经元参与者。 记忆形成的机制是非常保守的。因此，在动物模型系统中的工作可以极大地帮助我们理解人类认知功能障碍。我们将利用果蝇的规模经济和分子遗传学工具来建立这种记忆形成的新生物途径。我们在果蝇上的工作将推动脊椎动物同源物的研究，并最终将有助于我们理解人类各种形式的认知功能障碍。

DESCRIPTION (provided by applicant): The molecular basis of LTM formation appears to be evolutionarily conserved. In particular, simple forms of learning appear to reflect the conserved cellular functions of neurons, while more complex behavioral tasks reflect the diversity of neuronal systems (anatomy) across species. Nonetheless, memory of these complex tasks appears to invoke a common underlying synaptic plasticity. Consequently, we have capitalized on the economy-of-scale and the availability of genetic tools in Drosophila to focus on the discovery of the downstream genes involved in LTM Via a combination of DNA microarrays and a behavioral screen for single-gene mutants with defective memory, we have identified several genetic components of a novel biological pathway involved in the cellular transport of newly transcribed mRNAs and their subsequent local translocation. Importantly, in vivo biological validation has been achieved for two of these genes, pumilio and staufen. We propose to focus on this pathway via the following specific aims: 1. Molecular-genetic confirmation of selected genes in the pumilio/staufen pathway (staufen, oskar, moesin, pumilio, orb, elF2g, elFSc): We plan (i) genetic complementation with existing alleles and (ii) temporal and spatial rescue to establish a role for each of these genes in memory formation. 2. Protein expression studies of selected genes in the pumilio/staufen pathway: Immunostaining for most of these developmental genes never has been reported for the adult CNS. We will do so, with particular focus on subcellular co-localization experiments. 3. DNA microarray screen after cell purification: Studies focused on the neural substrates of olfactory memory continue to suggest the focus to be neurons of the mushroom bodies. Using a novel method, we plan to purify MB neurons from trained flies for DNA microarray experiments. We expect the signal from transcriptionally responsive genes to be greater in RNA isolated from this purified population of cells, thereby detecting additional neuronal participants of the pumilio/staufen pathway. Mechanisms of memory formation are remarkably conserved. Work in animal model systems can therefore greatly inform our understanding of human cognitive dysfunction. We will take advantage of the economy-of-scale and the molecular-genetic tools available in Drosophila to establish this novel biological pathway underlying memory formation. Our work in Drosophila will fuel studies of vertebrate homologs and eventually will contribute to our understanding of various forms of cognitive dysfunction in humans.