Activity-dependent modulation of the Drosophila mushroom body function by FMRP

描述（由申请人提供）：脆性X智力迟钝蛋白（FMRP）是一种RNA结合翻译调节因子，与包括儿童癫痫、自闭症谱系障碍和脆性X综合征在内的几种发育性脑障碍有关。我们的实验室已经确定，人类FMRP功能在果蝇疾病模型中是完全保守的，并且已经多次证明与哺乳动物生物学直接相关60。果蝇FMRP同源物dFMRP 1的缺失的特征在于外周和中枢神经系统中的突触过度生长，包括蘑菇体（MB）学习和记忆中心11，13，58。与人类一样，缺乏dFMRP 1的果蝇在学习和记忆方面表现出明显的缺陷，并且不能诱导突触连接性的活动依赖性修剪12，54。虽然这些行为和细胞表型是很好地建立，这些表型背后的神经元活动是知之甚少。利用果蝇模型系统的遗传能力，我试图确定dFMRP 1在确定和调节MB学习和记忆中心的功能特性方面的要求。我假设1.）FMRP调节MB回路2中的感觉表征和记忆巩固。FMRP通过选择性地抑制果蝇成年神经发育早期神经元活动的靶mRNA转录物而在活动依赖性MB回路功能中起作用。3.）第三章FMRP与其他mRNA结合的翻译调节因子Pumilio和Staufen组合相互作用，以控制发育阶段特异性蛋白表达驱动电路组装。我将使用全细胞膜片钳记录、Ca 2+成像和光遗传学操作的组合来测试这些假设36 -41。我将把用于可视化MB Kenyon细胞的IR-DIC光学技术集成到我们现有的电生理记录设备中，并遵循现有的程序来记录KC及其神经投射神经元（PN）的气味诱发活动36，55，56。我将产生钙成像和光遗传学实验所需的遗传储备，以及FMRP和已知mRNA结合FMRP相互作用物stau和pum 47，48，64的反式杂合突变体。总之，这些实验承诺显着的贡献，了解脆性X相关的学习缺陷和感觉超敏反应的功能相关性，并提供基本的见解感觉信号转导，代表性和巩固的性质。 公共卫生相关性：脆性X综合征是遗传性智力残疾的主要形式，由单一基因产物脆性X智力迟钝蛋白（FMRP）的丢失引起。FMRP的缺失与学习和记忆受损、昼夜节律紊乱、感觉超敏和突触过度生长相关。这项研究计划利用强大的果蝇遗传系统来定义FMRP在蘑菇体学习和记忆中心的功能需求。

DESCRIPTION (provided by applicant): Fragile X Mental Retardation Protein (FMRP) is an RNA-binding translational regulator implicated in several developmental brain disorders including childhood epilepsy, autism spectrum disorder and Fragile X syndrome. Our lab has established that human FMRP function is completely conserved in the Drosophila disease model, and has repeatedly proven direct relevance to mammalian biology60. The loss of the drosophila FMRP homolog, dFMRP1, is characterized by synaptic overgrowth in both the peripheral and central nervous systems, including the mushroom body (MB) learning and memory center11,13,58. As in humans, Drosophila lacking dFMRP1 exhibit pronounced deficits in learning and memory and are unable to induce activity-dependent pruning of synaptic connectivity12,54. Though these behavioral and cellular phenotypes are well established, the neuronal activity underlying these phenotypes is poorly understood. Using the genetic power of the Drosophila model system I seek to determine the requirements of dFMRP1 in determining and modulating the functional properties of the MB learning and memory center. I hypothesize 1.) FMRP regulates sensory representation and memory consolidation in the MB circuit 2.) FMRP functions in activity- dependent MB circuit function by selectively repressing target mRNA transcripts in response to neuronal activity during the early-use period of Drosophila adult neurodevelopment. 3.) FMRP interacts combinatorially with other mRNA-binding translational regulators, Pumilio and Staufen, to control development stage-specific protein expression driving circuit assembly. I will test these hypotheses using a combination of whole-cell patch clamp recording, Ca2+ imaging, and optogenetic manipulations36-41. I will integrate the IR-DIC optic technology used visualize MB Kenyon cells into our existing electrophysiological recording equipment and follow existing procedures for recording odor evoked activity from both KCs and their innervating projection neurons (PN)36,55,56. I will generate the genetic stocks necessary for calcium imaging and optogenetics experiments as well as the trans-heterozygous mutants for both FMRP and known mRNA binding FMRP interactors stau and pum47,48,64. Together these experiments promise significant contributions to understanding the functional correlates of Fragile X associated learning deficits and sensory hypersensitivity, and offer fundamental insights into the nature of sensory signal transduction, representation, and consolidation. PUBLIC HEALTH RELEVANCE: Fragile X Syndrome is the leading form of inherited intellectual disability and is caused by the loss of a single gene product, the Fragile X Mental Retardation Protein (FMRP). Loss of FMRP is associated with impaired learning and memory, circadian dis-rhythmicity, sensory hypersensitivity, and synaptic overgrowth. This research proposal utilizes the powerful Drosophila genetic system to define the functional requirements of FMRP in the mushroom body learning and memory center.