Functional Genomics of Drosophila Olfactory Memory

描述（由申请人提供）：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元系统（解剖学）的多样性。尽管如此，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，专注于发现涉及LTM的下游基因。通过DNA微阵列和记忆缺陷单基因突变体的行为筛选相结合，我们已经确定了一种新的生物学途径的几个遗传组分，该途径涉及新转录的mRNA的细胞运输及其随后的局部易位。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，eIF 2g，eIFSc）的分子遗传学确认：我们计划（i）与现有等位基因的遗传互补和（ii）时间和空间拯救，以确定这些基因中的每一个在记忆形成中的作用。 2. pumilio/staufen通路中选定基因的蛋白质表达研究：这些发育基因中的大多数从未在成人CNS中报道过免疫染色。我们将这样做，特别侧重于亚细胞共定位实验。 3.细胞纯化后的DNA微阵列筛选：对嗅觉记忆神经基质的研究继续表明重点是蘑菇体的神经元。使用一种新的方法，我们计划从训练的果蝇中纯化MB神经元用于DNA微阵列实验。我们预计，从转录响应基因的信号是从这个纯化的细胞群体中分离的RNA更大，从而检测pumilio/staufen途径的其他神经元参与者。 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告知我们对人类认知功能障碍的理解。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新的记忆形成的生物学途径。我们在果蝇中的工作将推动脊椎动物同源物的研究，并最终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.